Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders [1 ed.] 0323988180, 9780323988186

Citation preview

MOTOR SYSTEM DISORDERS, PART I: NORMAL PHYSIOLOGY AND FUNCTION AND NEUROMUSCULAR DISORDERS

HANDBOOK OF CLINICAL NEUROLOGY Series Editors

MICHAEL J. AMINOFF, FRANÇOIS BOLLER, AND DICK F. SWAAB VOLUME 195

MOTOR SYSTEM DISORDERS, PART I: NORMAL PHYSIOLOGY AND FUNCTION AND NEUROMUSCULAR DISORDERS Series Editors

MICHAEL J. AMINOFF, FRANÇOIS BOLLER, AND DICK F. SWAAB

Volume Editor

DAVID S. YOUNGER VOLUME 195 3rd Series

ELSEVIER Radarweg 29, PO Box 211, 1000 AE Amsterdam, Netherlands The Boulevard, Langford Lane, Kidlington, Oxford OX5 1GB, United Kingdom 50 Hampshire Street, 5th Floor, Cambridge, MA 02139, United States Copyright © 2023 Elsevier B.V. All rights reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency can be found at our website: www.elsevier.com/permissions. This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein). Notices Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary. Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility. With respect to any drug or pharmaceutical products identified, readers are advised to check the most current information provided (i) on procedures featured or (ii) by the manufacturer of each product to be administered, to verify the recommended dose or formula, the method and duration of administration, and contraindications. It is the responsibility of practitioners, relying on their own experience and knowledge of their patients, to make diagnoses, to determine dosages and the best treatment for each individual patient, and to take all appropriate safety precautions. To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein. ISBN: 978-0-323-98818-6 For information on all Elsevier publications visit our website at https://www.elsevier.com/books-and-journals

Publisher: Nikki Levy Editorial Project Manager: Kristi Anderson Production Project Manager: Punithavathy Govindaradjane Cover Designer: Matthew Limbert Typeset by STRAIVE, India

Handbook of Clinical Neurology 3rd Series Available titles Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol.

79, The human hypothalamus: Basic and clinical aspects, Part I, D.F. Swaab, ed. ISBN 9780444513571 80, The human hypothalamus: Basic and clinical aspects, Part II, D.F. Swaab, ed. ISBN 9780444514905 81, Pain, F. Cervero and T.S. Jensen, eds. ISBN 9780444519016 82, Motor neurone disorders and related diseases, A.A. Eisen and P.J. Shaw, eds. ISBN 9780444518941 83, Parkinson’s disease and related disorders, Part I, W.C. Koller and E. Melamed, eds. ISBN 9780444519009 84, Parkinson’s disease and related disorders, Part II, W.C. Koller and E. Melamed, eds. ISBN 9780444528933 85, HIV/AIDS and the nervous system, P. Portegies and J. Berger, eds. ISBN 9780444520104 86, Myopathies, F.L. Mastaglia and D. Hilton Jones, eds. ISBN 9780444518996 87, Malformations of the nervous system, H.B. Sarnat and P. Curatolo, eds. ISBN 9780444518965 88, Neuropsychology and behavioural neurology, G. Goldenberg and B.C. Miller, eds. ISBN 9780444518972 89, Dementias, C. Duyckaerts and I. Litvan, eds. ISBN 9780444518989 90, Disorders of consciousness, G.B. Young and E.F.M. Wijdicks, eds. ISBN 9780444518958 91, Neuromuscular junction disorders, A.G. Engel, ed. ISBN 9780444520081 92, Stroke – Part I: Basic and epidemiological aspects, M. Fisher, ed. ISBN 9780444520036 93, Stroke – Part II: Clinical manifestations and pathogenesis, M. Fisher, ed. ISBN 9780444520043 94, Stroke – Part III: Investigations and management, M. Fisher, ed. ISBN 9780444520050 95, History of neurology, S. Finger, F. Boller and K.L. Tyler, eds. ISBN 9780444520081 96, Bacterial infections of the central nervous system, K.L. Roos and A.R. Tunkel, eds. ISBN 9780444520159 97, Headache, G. Nappi and M.A. Moskowitz, eds. ISBN 9780444521392 98, Sleep disorders Part I, P. Montagna and S. Chokroverty, eds. ISBN 9780444520067 99, Sleep disorders Part II, P. Montagna and S. Chokroverty, eds. ISBN 9780444520074 100, Hyperkinetic movement disorders, W.J. Weiner and E. Tolosa, eds. ISBN 9780444520142 101, Muscular dystrophies, A. Amato and R.C. Griggs, eds. ISBN 9780080450315 102, Neuro-ophthalmology, C. Kennard and R.J. Leigh, eds. ISBN 9780444529039 103, Ataxic disorders, S.H. Subramony and A. Durr, eds. ISBN 9780444518927 104, Neuro-oncology Part I, W. Grisold and R. Sofietti, eds. ISBN 9780444521385 105, Neuro-oncology Part II, W. Grisold and R. Sofietti, eds. ISBN 9780444535023 106, Neurobiology of psychiatric disorders, T. Schlaepfer and C.B. Nemeroff, eds. ISBN 9780444520029 107, Epilepsy Part I, H. Stefan and W.H. Theodore, eds. ISBN 9780444528988 108, Epilepsy Part II, H. Stefan and W.H. Theodore, eds. ISBN 9780444528995 109, Spinal cord injury, J. Verhaagen and J.W. McDonald III, eds. ISBN 9780444521378 110, Neurological rehabilitation, M. Barnes and D.C. Good, eds. ISBN 9780444529015 111, Pediatric neurology Part I, O. Dulac, M. Lassonde and H.B. Sarnat, eds. ISBN 9780444528919 112, Pediatric neurology Part II, O. Dulac, M. Lassonde and H.B. Sarnat, eds. ISBN 9780444529107 113, Pediatric neurology Part III, O. Dulac, M. Lassonde and H.B. Sarnat, eds. ISBN 9780444595652 114, Neuroparasitology and tropical neurology, H.H. Garcia, H.B. Tanowitz and O.H. Del Brutto, eds. ISBN 9780444534903 115, Peripheral nerve disorders, G. Said and C. Krarup, eds. ISBN 9780444529022 116, Brain stimulation, A.M. Lozano and M. Hallett, eds. ISBN 9780444534972 117, Autonomic nervous system, R.M. Buijs and D.F. Swaab, eds. ISBN 9780444534910 118, Ethical and legal issues in neurology, J.L. Bernat and H.R. Beresford, eds. ISBN 9780444535016 119, Neurologic aspects of systemic disease Part I, J. Biller and J.M. Ferro, eds. ISBN 9780702040863 120, Neurologic aspects of systemic disease Part II, J. Biller and J.M. Ferro, eds. ISBN 9780702040870 121, Neurologic aspects of systemic disease Part III, J. Biller and J.M. Ferro, eds. ISBN 9780702040887 122, Multiple sclerosis and related disorders, D.S. Goodin, ed. ISBN 9780444520012 123, Neurovirology, A.C. Tselis and J. Booss, eds. ISBN 9780444534880 124, Clinical neuroendocrinology, E. Fliers, M. Korbonits and J.A. Romijn, eds. ISBN 9780444596024 125, Alcohol and the nervous system, E.V. Sullivan and A. Pfefferbaum, eds. ISBN 9780444626196 126, Diabetes and the nervous system, D.W. Zochodne and R.A. Malik, eds. ISBN 9780444534804 127, Traumatic brain injury Part I, J.H. Grafman and A.M. Salazar, eds. ISBN 9780444528926 128, Traumatic brain injury Part II, J.H. Grafman and A.M. Salazar, eds. ISBN 9780444635211

vi

AVAILABLE TITLES (Continued)

Vol. 129, The human auditory system: Fundamental organization and clinical disorders, G.G. Celesia and G. Hickok, eds. ISBN 9780444626301 Vol. 130, Neurology of sexual and bladder disorders, D.B. Vodušek and F. Boller, eds. ISBN 9780444632470 Vol. 131, Occupational neurology, M. Lotti and M.L. Bleecker, eds. ISBN 9780444626271 Vol. 132, Neurocutaneous syndromes, M.P. Islam and E.S. Roach, eds. ISBN 9780444627025 Vol. 133, Autoimmune neurology, S.J. Pittock and A. Vincent, eds. ISBN 9780444634320 Vol. 134, Gliomas, M.S. Berger and M. Weller, eds. ISBN 9780128029978 Vol. 135, Neuroimaging Part I, J.C. Masdeu and R.G. González, eds. ISBN 9780444534859 Vol. 136, Neuroimaging Part II, J.C. Masdeu and R.G. González, eds. ISBN 9780444534866 Vol. 137, Neuro-otology, J.M. Furman and T. Lempert, eds. ISBN 9780444634375 Vol. 138, Neuroepidemiology, C. Rosano, M.A. Ikram and M. Ganguli, eds. ISBN 9780128029732 Vol. 139, Functional neurologic disorders, M. Hallett, J. Stone and A. Carson, eds. ISBN 9780128017722 Vol. 140, Critical care neurology Part I, E.F.M. Wijdicks and A.H. Kramer, eds. ISBN 9780444636003 Vol. 141, Critical care neurology Part II, E.F.M. Wijdicks and A.H. Kramer, eds. ISBN 9780444635990 Vol. 142, Wilson disease, A. Członkowska and M.L. Schilsky, eds. ISBN 9780444636003 Vol. 143, Arteriovenous and cavernous malformations, R.F. Spetzler, K. Moon and R.O. Almefty, eds. ISBN 9780444636409 Vol. 144, Huntington disease, A.S. Feigin and K.E. Anderson, eds. ISBN 9780128018934 Vol. 145, Neuropathology, G.G. Kovacs and I. Alafuzoff, eds. ISBN 9780128023952 Vol. 146, Cerebrospinal fluid in neurologic disorders, F. Deisenhammer, C.E. Teunissen and H. Tumani, eds. ISBN 9780128042793 Vol. 147, Neurogenetics Part I, D.H. Geschwind, H.L. Paulson and C. Klein, eds. ISBN 9780444632333 Vol. 148, Neurogenetics Part II, D.H. Geschwind, H.L. Paulson and C. Klein, eds. ISBN 9780444640765 Vol. 149, Metastatic diseases of the nervous system, D. Schiff and M.J. van den Bent, eds. ISBN 9780128111611 Vol. 150, Brain banking in neurologic and psychiatric diseases, I. Huitinga and M.J. Webster, eds. ISBN 9780444636393 Vol. 151, The parietal lobe, G. Vallar and H.B. Coslett, eds. ISBN 9780444636225 Vol. 152, The neurology of HIV infection, B.J. Brew, ed. ISBN 9780444638496 Vol. 153, Human prion diseases, M. Pocchiari and J.C. Manson, eds. ISBN 9780444639455 Vol. 154, The cerebellum: From embryology to diagnostic investigations, M. Manto and T.A.G.M. Huisman, eds. ISBN 9780444639561 Vol. 155, The cerebellum: Disorders and treatment, M. Manto and T.A.G.M. Huisman, eds. ISBN 9780444641892 Vol. 156, Thermoregulation: From basic neuroscience to clinical neurology Part I, A.A. Romanovsky, ed. ISBN 9780444639127 Vol. 157, Thermoregulation: From basic neuroscience to clinical neurology Part II, A.A. Romanovsky, ed. ISBN 9780444640741 Vol. 158, Sports neurology, B. Hainline and R.A. Stern, eds. ISBN 9780444639547 Vol. 159, Balance, gait, and falls, B.L. Day and S.R. Lord, eds. ISBN 9780444639165 Vol. 160, Clinical neurophysiology: Basis and technical aspects, K.H. Levin and P. Chauvel, eds. ISBN 9780444640321 Vol. 161, Clinical neurophysiology: Diseases and disorders, K.H. Levin and P. Chauvel, eds. ISBN 9780444641427 Vol. 162, Neonatal neurology, L.S. De Vries and H.C. Glass, eds. ISBN 9780444640291 Vol. 163, The frontal lobes, M. D’Esposito and J.H. Grafman, eds. ISBN 9780128042816 Vol. 164, Smell and taste, Richard L. Doty, ed. ISBN 9780444638557 Vol. 165, Psychopharmacology of neurologic disease, V.I. Reus and D. Lindqvist, eds. ISBN 9780444640123 Vol. 166, Cingulate cortex, B.A. Vogt, ed. ISBN 9780444641960 Vol. 167, Geriatric neurology, S.T. DeKosky and S. Asthana, eds. ISBN 9780128047668 Vol. 168, Brain-computer interfaces, N.F. Ramsey and J. del R. Millán, eds. ISBN 9780444639349 Vol. 169, Meningiomas, Part I, M.W. McDermott, ed. ISBN 9780128042809 Vol. 170, Meningiomas, Part II, M.W. McDermott, ed. ISBN 9780128221983 Vol. 171, Neurology and pregnancy: Pathophysiology and patient care, E.A.P. Steegers, M.J. Cipolla and E.C. Miller, eds. ISBN 9780444642394 Vol. 172, Neurology and pregnancy: Neuro-obstetric disorders, E.A.P. Steegers, M.J. Cipolla and E.C. Miller, eds. ISBN 9780444642400 Vol. 173, Neurocognitive development: Normative development, A. Gallagher, C. Bulteau, D. Cohen and J.L. Michaud, eds. ISBN 9780444641502 Vol. 174, Neurocognitive development: Disorders and disabilities, A. Gallagher, C. Bulteau, D. Cohen and J.L. Michaud, eds. ISBN 9780444641489 Vol. 175, Sex differences in neurology and psychiatry, R. Lanzenberger, G.S. Kranz, and I. Savic, eds. ISBN 9780444641236 Vol. 176, Interventional neuroradiology, S.W. Hetts and D.L. Cooke, eds. ISBN 9780444640345 Vol. 177, Heart and neurologic disease, J. Biller, ed. ISBN 9780128198148 Vol. 178, Neurology of vision and visual disorders, J.J.S. Barton and A. Leff, eds. ISBN 9780128213773 Vol. 179, The human hypothalamus: Anterior region, D.F. Swaab, F. Kreier, P.J. Lucassen, A. Salehi and R.M. Buijs, eds. ISBN 9780128199756

AVAILABLE TITLES (Continued)

vii

Vol. 180, The human hypothalamus: Middle and posterior region, D.F. Swaab, F. Kreier, P.J. Lucassen, A. Salehi and R.M. Buijs, eds. ISBN 9780128201077 Vol. 181, The human hypothalamus: Neuroendocrine disorders, D.F. Swaab, R.M. Buijs, P.J. Lucassen, A. Salehi and F. Kreier, eds. ISBN 9780128206836 Vol. 182, The human hypothalamus: Neuropsychiatric disorders, D.F. Swaab, R.M. Buijs, F. Kreier, P.J. Lucassen, and A. Salehi, eds. ISBN 9780128199732 Vol. 183, Disorders of emotion in neurologic disease, K.M. Heilman and S.E. Nadeau, eds. ISBN 9780128222904 Vol. 184, Neuroplasticity: From bench to bedside, A. Quartarone, M.F. Ghilardi, and F. Boller, eds. ISBN 9780128194102 Vol. 185, Aphasia, A.E. Hillis and J. Fridriksson, eds. ISBN 9780128233849 Vol. 186, Intraoperative neuromonitoring, M.R. Nuwer and D.B. MacDonald, eds. ISBN 9780128198261 Vol. 187, The temporal lobe, G. Miceli, P. Bartolomeo, and V. Navarro, eds. ISBN 9780128234938 Vol. 188, Respiratory neurobiology: Physiology and clinical disorders, Part I, R. Chen and P.G. Guyenet, eds. ISBN 9780323915342 Vol. 189, Respiratory neurobiology: Physiology and clinical disorders, Part II, R. Chen and P.G. Guyenet, eds. ISBN 9780323915328 Vol. 190, Neuropalliative care: Part I, J.M. Miyasaki and B.M. Kluger, eds. ISBN 9780323850292 Vol. 191, Neuropalliative care: Part II, J.M. Miyasaki and B.M. Kluger, eds. ISBN 9780128245354 Vol. 192, Precision medicine in neurodegenerative disorders: Part I, A.J. Espay, ed. ISBN 9780323855389 Vol. 193, Precision medicine in neurodegenerative disorders: Part II, A.J. Espay, ed. ISBN 9780323855556 Vol. 194, Mitochondrial diseases, R. Horvath, M. Hirano, and P.F. Chinnery, eds. ISBN 9780128217511 All volumes in the 3rd Series of the Handbook of Clinical Neurology are published electronically, on Science Direct: http://www.sciencedirect.com/science/handbooks/00729752.

This page intentionally left blank

Foreword

It is a pleasure to introduce Volumes 195 and 196 of the Handbook of Clinical Neurology, devoted to the motor system. In Volume 195, there are sections on normal physiology and function, diagnostic approaches, and neuromuscular disorders occurring in patients of all ages. In Volume 196, the focus is on spinal cord disorders, progressive neurodegenerative diseases, nonprogressive cortical and subcortical disorders, and therapeutics. A detailed summary about the content of these various sections is provided in the Preface. It must be made clear, however, that these volumes cover more than is generally subsumed by the designation “motor system,” as they provide an account of virtually any disorder with motor manifestations through the perspective of many different subspecialties, thereby providing a truly comprehensive account of the subject matter. The editor, David S. Younger, is an Affiliated Professor of Neuroscience at the City University of New York School of Medicine and Attending Physician at White Plains Hospital, New York. He is a neuromuscular specialist, teacher, and clinical neurophysiologist who has authored studies on many different aspects of neurologic function. More than 20 years ago, he edited a book that provided a detailed and practical account of motor disorders, and he has continued to maintain an encyclopedic knowledge of these disorders ever since. We were therefore delighted when he accepted our invitation to edit a volume in the Handbook of Clinical Neurology series to provide a comprehensive account of these disorders and how they have been impacted by advances in the neurosciences. As he proceeded, however, it became apparent that two volumes would be required to cover the subject adequately, providing clinicians with an up-to-date summary of the investigation, diagnosis, management, and treatment of various motor disorders, and neuroscientists with an account of the many different approaches to determine their fundamental bases through the perspective of molecular biology, immunology, neurophysiology, imaging, and pharmacology. To do so, he has attracted an outstanding group of authors—many of whom are internationally recognized experts in the field—to contribute to these volumes, ensuring a consistently high standard throughout them. Moreover, as the sole volume editor, he has edited the chapters so that there is a pleasing uniformity of style and they fit together seamlessly. As series editors, we reviewed all the chapters and made suggestions for improvement, but it was clear that the volume editor and chapter authors had produced scholarly and comprehensive accounts of different aspects of the topic. We thank Dr. Younger and all the contributors for their work. It is our hope that the volumes will have wide appeal to clinicians and scientists alike. As always, it is a pleasure to thank Elsevier, our publisher, and in particular Nikki Levy and Kristi Anderson in San Diego, and Punithavathy Govindaradjane at Elsevier Global Book Production in Chennai, for their assistance in the development and production of the Handbook of Clinical Neurology. Michael J. Aminoff Franc¸ois Boller Dick F. Swaab

This page intentionally left blank

Preface

A striking aspect of the history of motor disorders has been the persistence of generations of neuroscientists and neurologists in searching for a more cohesive understanding of the motor system in health and disease. Remarkable progress has been achieved over the past 30 years in the classification, diagnosis, etiopathogenesis, and treatment of many peripheral and central nervous system motor disorders since the publication of the last handbook volume on this subject (Volume 59, edited by JMB V De Jong, published in 1991). These advances have occurred along the diverse lines of electrophysiology, immunology, molecular genetics, and ultrastructural analysis, as well as through technical advances in translational methods applicable to neurologic therapeutics, neurosurgery, and rehabilitative care. Such advances have, in turn, fostered more effective and innovative treatments confirmed to be efficacious in randomized clinical trials, showing improved outcomes and enhanced independence. My goal in developing the present work was to produce two classic volumes with unsurpassed depth and breadth along a list of interrelated topics on the motor system and its disorders. The first volume (Volume 195 of the Handbook) is divided into three sections. Section 1 covers normal physiology and function encompassing basic concepts of skeletal muscle function, upper and lower motor neuron control, the vestibular and autonomic nervous system and autonomic failure, and immunology and Coronavirus-2 illness. Section 2 overviews the topic of clinical and laboratory diagnosis, beginning with genetics, electrodiagnosis, neuromuscular pathology, autonomic physiology, imaging, sleep disorders, and the path to evidence-based therapy for neuromuscular diseases. Section 3 delves into the neuromuscular disorders, covering classic topics such as infant hypotonia, inflammatory myopathy, childhood muscular dystrophy, distal and congenital myopathies, channelopathies, mitochondrial encephalomyopathies, autoimmune and hereditary polyneuropathies, neuromuscular junction disorders, and adult and childhood vasculitis. The second volume (Volume 196) is divided into four sections. Section 1 covers spinal cord diseases, spinal muscular atrophy, hereditary spastic paraplegias and primary lateral sclerosis, transverse myelitis, and motor aspects of multiple sclerosis. Section 2 addresses progressive neurodegenerative diseases involving the cerebellum, extrapyramidal system, tauopathies, paraneoplastic disorders, motor neuron disease, and Alzheimer disease. Section 3 addresses nonprogressive cortical and subcortical disorders exemplified by motor seizure disorders, stroke, brainstem nuclei and cortical motor control disturbances, antineuronal antibodies, autoimmune encephalopathy, tremor, dystonia, frontal and parietal lobe syndromes, and bowel, bladder, and sexual disorders. Section 4 concludes with a comprehensive list of topics addressing neurologic therapeutics that includes the treatment of spasticity, use of botulinum toxin, enzyme replacement therapy, growth factors, applied strategies of neuroplasticity, tau-based immunotherapies, and novel uses of brain–computer interfaces. The invited authors, all experts in their fields, have been drawn from all parts of the United States and abroad, covering a multitude of traditions and heritages in neurology. As the sole editor of these two volumes, I was overjoyed at the prospect of instilling my own voice to assure consistency of style and readability. I appreciate the extreme tolerance of each of the contributors in allowing me to edit their work no matter how much it appeared to be at the point of finality so that there would be cohesiveness and consistency beyond just the apparent flow of titles. I thank the series editors for their tolerance and for their style setting and content-oriented benchmarks that made my job easier. I have been privileged to have been trained with, mentored by, or closely associated with many preeminent clinicians beginning at Columbia University’s Neurological Institute with Lewis P (“Bud”) Rowland, a former editor of the journal Neurology, whose unselfish guidance to become the best clinical investigator and author fostered the careers of many of my contemporaries, some of whom have contributed chapters to these two volumes. Most of all, I am continuously instructed and humbled by my patients, whose never-ending battles with debilitating illness are always fought with dignity.

xii

PREFACE

I am pleased to dedicate these two volumes to the spouses, children, and significant others of all of the contributors, and to my own family, who patiently sat by and were often displaced while the process of writing and editing took place. David S. Younger

Contributors

A.A. Amato Department of Neurology, Division of Neuromuscular Diseases, Neuropathology Laboratory, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, United States F. Binkofski Division of Clinical Cognitive Sciences, Medical Faculty of the RWTH Aachen University, Aachen, Germany T.H. Brannagan III Department of Neurology, Columbia University, Vagelos College of Physicians and Surgeons, New York, NY, United States M.B. Bromberg Department of Neurology, University of Utah, Salt Lake City, UT, United States S.V. Brooks Department of Molecular & Integrative Physiology; Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, United States G. Buccino Division of Neuroscience, University Vita Salute San Raffaele and IRCCS San Raffaele, Milan, Italy K.E. Cullen Departments of Biomedical Engineering, of Otolaryngology-Head and Neck Surgery, and of Neuroscience; Kavli Neuroscience Discovery Institute, Johns Hopkins University, Baltimore, MD, United States

Lisboa-Norte; Faculdade de Medicina-Instituto de Medicina Molecular-Centro de Estudos Egas Moniz, Universidade de Lisboa, Lisbon, Portugal U. De Girolami Department of Pathology, Neuropathology Laboratory, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, United States M.A. Ferrante Electromyography Laboratory, Department of Neurology, Veterans Administration Medical Center, University of Tennessee, Memphis, TN, United States N.E. Gilhus Department of Neurology, Haukeland University Hospital and Department of Clinical Medicine, University of Bergen, Bergen, Norway J. Guti
errez Department of Clinical Neurophysiology, Cuban Institute of Neurology and Neurosurgery; Department of Clinical Neurophysiology, Havana University of Medical Sciences, Havana, Cuba S.D. Guzman Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor, MI, United States K.G. Gwathmey Neuromuscular Division, Department of Neurology, Virginia Commonwealth University, Richmond, VA, United States

M.C. Dalakas Department of Neurology, Thomas Jefferson University, Philadelphia, PA, United States; Neuroimmunology Unit National and Kapodistrian University of Athens Medical School, Athens, Greece

M.G. Hanna Centre for Neuromuscular Disorders, Queen Square UCL Institute of Neurology, London, United Kingdom

M. de Carvalho Department of Neurosciences and Mental Health, Hospital de Santa Maria, Centro Hospitalar Universitário

D. Jayaseelan University College London Hospital, London, United Kingdom

xiv

CONTRIBUTORS

K.G. Johnson Department of Neurology; Institute for Healthcare Delivery and Population Science, University of Massachusetts Chan School of Medicine-Baystate, Springfield, MA, United States

Y.S. Ng Wellcome Centre for Mitochondrial Research, Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne, United Kingdom

M. Jokela Neuromuscular Research Center, Department of Neurology, Tampere University and University Hospital, Tampere; Division of Clinical Neurosciences, Department of Neurology, Turku University Hospital, Turku, Finland

C. Pisciotta Department of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy

J. Lee-Iannotti Department of Medicine, University of Arizona College of Medicine, Phoenix, AZ, United States Y. Mahjoub Department of Clinical Neurosciences, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada D. Martino Department of Clinical Neurosciences, Cumming School of Medicine; Hotchkiss Brain Institute, University of Calgary, Calgary, AB, Canada J.C. Masdeu Stanley H. Appel Department of Neurology, Houston Methodist Hospital, Houston, TX; Department of Neurology, Weill Cornell Medicine, New York, NY, United States R. McFarland NHS Highly Specialised Service for Rare Mitochondrial Disorders, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, United Kingdom C. Miranda Department of Neurology, Columbia University, Vagelos College of Physicians and Surgeons, New York, NY, United States M.O. Nakawah Stanley H. Appel Department of Neurology, Houston Methodist Hospital, Houston, TX; Department of Neurology, Weill Cornell Medicine, New York, NY, United States R. Naum Department of Neurology, Virginia Commonwealth University, Richmond, VA, United States

E. Rounis Chelsea and Westminster NHS Foundation Trust, West Middlesex University Hospital, Isleworth; Department of Brain Sciences, Faculty of Medicine, Imperial College, London, United Kingdom L.P. Ruiz Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor, MI, United States M. Savarese Folkh€alsan Research Center; Department of Medical Genetics, Medicum, University of Helsinki, Helsinki, Finland M.E. Shy Department of Neurology, University of Iowa Hospitals and Clinics, Iowa City, IA, United States M. Swash Faculdade de Medicina-Instituto de Medicina Molecular-Centro de Estudos Egas Moniz, Universidade de Lisboa, Lisbon, Portugal; Department of Neurology, Barts and London School of Medicine, Queen Mary University of London and Royal London Hospital, London, United Kingdom K. Takakusaki Department of Physiology, Division of Neuroscience, Asahikawa Medical University, Asahikawa, Japan B. Udd Folkh€alsan Research Center; Department of Medical Genetics, Medicum, University of Helsinki, Helsinki; Neuromuscular Research Center, Department of Neurology, Tampere University and University Hospital, Tampere; Department of Neurology, Vaasa Central Hospital, Vaasa, Finland V. Vivekanandam Centre for Neuromuscular Disorders, Queen Square UCL Institute of Neurology, London, United Kingdom

CONTRIBUTORS

xv

Y. Vorakunthada Department of Medicine, University of Arizona College of Medicine, Phoenix, AZ, United States

S.G. Wong Department of Medicine, University of Arizona College of Medicine, Phoenix, AZ, United States

E.L. Weil Department of Neurology, University of Michigan, Ann Arbor, MI; Stanley H. Appel Department of Neurology, Houston Methodist Hospital, Houston, TX, United States

D.S. Younger Department of Clinical Medicine and Neuroscience, CUNY School of Medicine, New York; and Department of Medicine, Section of Internal Medicine and Neurology, White Plains Hospital, White Plains, NY, United States

This page intentionally left blank

Contents Foreword ix Preface xi Contributors xiii SECTION 1

Normal physiology and function

1.

Skeletal muscle structure, physiology, and function S.V. Brooks, S.D. Guzman, and L.P. Ruiz (Ann Arbor, United States)

2.

Upper and lower motor neuron neurophysiology and motor control M. de Carvalho and M. Swash (Lisbon, Portugal and London, United Kingdom)

17

3.

Vestibular motor control K.E. Cullen (Baltimore, United States)

31

4.

Autonomic failure: Clinicopathologic, physiologic, and genetic aspects D.S. Younger (New York City, United States)

55

5.

Gait control by the frontal lobe K. Takakusaki (Asahikawa, Japan)

6.

Parietal control of hand movement 127 E. Rounis, G. Buccino, and F. Binkofski (Isleworth and London, United Kingdom, Milan, Italy, and Aachen, Germany)

7.

Immunology and microbiome: Implications for motor systems Y. Mahjoub and D. Martino (Calgary, Canada)

135

8.

COVID-19 (novel SARS-CoV-2) neurological illness D.S. Younger (New York City, United States)

159

SECTION 2

3

103

Clinical and laboratory diagnosis

9. Neurogenetic motor disorders D.S. Younger (New York City, United States)

183

10. Neuromuscular electrodiagnosis M.A. Ferrante (Memphis, United States)

251

11. Quantitative electrodiagnosis of the motor unit M.B. Bromberg (Salt Lake City, United States)

271

12. Neuromuscular pathology A.A. Amato and U. De Girolami (Boston, United States)

287

xviii

CONTENTS

13. Electrophysiological assessment of peripheral and central autonomic disorders J. Gutierrez (Havana, Cuba)

301

14. On the path to evidence-based therapy in neuromuscular disorders D.S. Younger (New York City, United States)

315

15. Advances in the neuroimaging of motor disorders E.L. Weil, M.O. Nakawah, and J.C. Masdeu (Ann Arbor, Houston and New York, United States)

359

16. Sleep-related motor disorders 383 S.G. Wong, Y. Vorakunthada, J. Lee-Iannotti, and K.G. Johnson (Phoenix and Springfield, United States) SECTION 3

Neuromuscular disorders

17. Neonatal and infantile hypotonia D.S. Younger (New York City, United States)

401

18. Autoimmune inflammatory myopathies M.C. Dalakas (Philadelphia, United States and Athens, Greece)

425

19. Childhood muscular dystrophies D.S. Younger (New York City, United States)

461

20. Distal myopathy M. Savarese, M. Jokela, and B. Udd (Helsinki, Tampere, Turku, and Vaasa, Finland)

497

21. Muscle channelopathies V. Vivekanandam, D. Jayaseelan, and M.G. Hanna (London, United Kingdom)

521

22. Congenital myopathies D.S. Younger (New York City, United States)

533

23. Mitochondrial encephalomyopathy Y.S. Ng and R. McFarland (Newcastle upon Tyne, United Kingdom)

563

24. Autoimmune polyneuropathies R. Naum and K.G. Gwathmey (Richmond, United States)

587

25. Hereditary neuropathy C. Pisciotta and M.E. Shy (Milan, Italy and Iowa City, United States)

609

26. Acute/chronic inflammatory polyradiculoneuropathy C. Miranda and T.H. Brannagan III (New York, United States)

619

27. Myasthenia gravis and congenital myasthenic syndromes N.E. Gilhus (Bergen, Norway)

635

28. Adult and childhood vasculitis D.S. Younger (New York City, United States)

653

29. Critical illness–associated weakness and related motor disorders D.S. Younger (New York City, United States)

707

Index

779

Section 1 Normal physiology and function

This page intentionally left blank

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00013-3 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 1

Skeletal muscle structure, physiology, and function SUSAN V. BROOKS1,2*, STEVE D. GUZMAN1, AND LLOYD P. RUIZ1 1

Department of Molecular & Integrative Physiology, University of Michigan, Ann Arbor, MI, United States 2

Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, United States

Abstract Contractions of skeletal muscles provide the stability and power for all body movements. Consequently, any impairment in skeletal muscle function results in some degree of instability or immobility. Factors that influence skeletal muscle structure and function are therefore of great interest scientifically and clinically. Injury, neuromuscular disease, and old age are among the factors that commonly contribute to impairments in skeletal muscle function. The goal of this chapter is to summarize the fundamentals of skeletal muscle structure and function to provide foundational knowledge for this Handbook volume. We examine the molecular interactions that provide the basis for the generation of force and movement, discuss mechanisms of the regulation of contraction at the level of myofibers, and introduce concepts of the activation and control of muscle function in vivo. Where appropriate, the chapter updates the emerging science that will increase understanding of muscle function.

INTRODUCTION Skeletal muscle comprises a little less than one-half of body mass. Muscles are linked to bones by tendons or tendon-like structures through which the forces and movements developed by muscle contractions are transmitted to the skeleton. Contractions of skeletal muscles provide the stability and power for all body movements. Thus, any impairment in skeletal muscle function results in at least some degree of instability or immobility. Muscle function may be impaired as a result of injury, disease, disuse, or old age. Impaired muscle function impacts quality of life at all ages, but particularly in the elderly, due to increased risk of severe injury, reduced participation in recreational activities, and ultimately impaired ability to perform activities of daily living and retain one’s independence. The importance of mobility to quality of life supports the great interest, both scientifically and clinically, in factors that influence skeletal muscle structure and function. The goal of this chapter is to provide basic information on skeletal muscle structure and

function to facilitate understanding of physiologic and pathophysiologic conditions described in subsequent chapters. Contraction is defined as the activation of muscle fibers with a tendency of the fibers to shorten (Faulkner, 2003). Contraction occurs when excitatory signals from presynaptic motor neuron terminals are transmitted to postsynaptic endplates located on muscle fibers across a chemical synapse referred to as the neuromuscular junction (NMJ). Muscle action potentials result in an increase in cytosolic calcium, which triggers a series of molecular events that includes the binding of calcium to the muscle-regulatory proteins, the interaction of myosin cross-bridges with actin filaments, and the production of the cross-bridge power stroke. The wide range of physical abilities possible by humans is achieved by the vast heterogeneity of the structural and functional properties of skeletal muscles. The functional properties of a given skeletal muscle are determined by the relative proportions and collective properties of the individual muscle fibers that

*Correspondence to: Susan V. Brooks, PhD, Department of Molecular & Integrative Physiology and Biomedical Engineering. 2029 BSRB, 109 Zina Pitcher Pl, Ann Arbor, MI 48109-2002, United States. Tel: +1-734-936-2147, Fax: +1-734-615-3292, E-mail: [email protected]

4

S.V. BROOKS ET AL.

make up the muscle, namely their contractile speed and ability to maintain force or power over time along with the size and orientation of the fibers. The control via the central nervous system of which muscle fibers are contracting at a given time and to what degree determines the variety of physical activities achieved by the organism. Each of these determinants of muscle function is considered in the upcoming sections.

SKELETAL MUSCLE STRUCTURE There are over 600 skeletal muscles in the human body with each muscle composed of hundreds to hundreds of thousands of elongated, parallel running, multinucleated cells called fibers (Fig. 1.1). Within individual fibers, the contractile proteins myosin and actin are incorporated into thick and thin filaments, respectively. The thick and thin filaments are parallel and overlapping and arrayed into longitudinally repeated banding patterns termed sarcomeres (Fig. 1.1). Sarcomeres in series form myofibrils, and many parallel myofibrils exist in each fiber. The number of myofibrils arranged in parallel determines the force-generating capability of the fiber. In mammals, the number of fibers in a given muscle is determined at birth and remains essentially constant

through to adulthood. In contrast, the number of myofibrils, and, consequently, muscle fiber cross-sectional area (CSA), can change dramatically, increasing with normal growth or hypertrophy induced by strength training and decreasing with atrophy associated with immobilization, inactivity, injury, disease, or old age. Muscle fiber shortening is achieved by the shortening of individual sarcomeres as thick and thin filaments slide past each other, mediated by cyclical interactions between projections from the myosin thick filaments, called crossbridges, and binding sites on the actin molecules of the thin filament. The cross-bridges are formed by the N-terminal region of the myosin molecule. Muscle myosin is a hexamer consisting of two so-called heavy chains and two pairs of light chains. A long length of the molecule starting at the COOH-terminal ends of the heavy chains form a coiled coil of two alpha helices that aggregates in the cell to create the backbone of the thick filaments. The remainder of the myosin molecule projects outward from the thick filament, forming the cross-bridge portion of the molecule to which the “essential” and “regulatory” light chains are noncovalently bound. Mild proteolytic treatment of thick filaments results in the generation of the light (LMM) and heavy meromyosin (HMM)

Fig. 1.1. Structural hierarchy of skeletal muscle. Skeletal muscle is composed of many elongated fibers, with the fiber volume dominated by cylindrical subcellular structures called myofibrils. The myofibrils display a repeating banding pattern that emerges from to the underlying array of overlapping myosin thick and actin thin filaments with a single repeat of the banding pattern, referred to as a sarcomere. An additional set of filaments (titin) filaments depicted form the basis for the passive elasticity of the fiber and act to stabilize sarcomere structure holding the thick filaments roughly mid-way between the z-discs. Created with BioRender.com.

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION

Fig. 1.2. Myosin S1 structure. (A) The ribbon representation is shown for the X-ray crystallographic structure of the myosin head. (B) The superimposed structures of S1 with nucleotide present and absent suggest that rotation of the lever arm about a fulcrum at the junction with the catalytic domain provides the basis for the cross-bridge power stroke. 3D protein structures were created using NIH iCn3D. PBD ID: 1SR6 (Risal et al., 2004).

fragments, and further proteolysis of HMM gives rise to two additional fragments, referred to as the S1 and S2 subfragments. The S1 subfragment, also referred to as the “head” of the cross-bridge, retains all the motor functions of the molecule, i.e., the ability to produce movement and force (R€ uegg et al., 2002). The crystallographic structure of myosin S1 (Fig. 2A) shows that the subfragment consists of a globular “catalytic domain” that contains both the actin-binding site and the nucleotide-binding pocket, which constitutes the active site for ATP hydrolysis. An a-helical “level arm domain” extends toward the tail of the molecule and appears to be stabilized by its interaction with the two light chains (Fig. 1.2A).

CONTRACTILE MECHANISMS A conformational change within S1 while tightly bound to actin generates the power stroke that underlies force production and movement (Vale and Milligan, 2000). The precise nature of the conformational change associated with the cross-bridge power stroke remains an active area of investigation using structural biology, molecular genetic, and biophysical approaches. Details of recent advances and remaining questions regarding contractile

5

mechanisms are nicely reviewed elsewhere (Månsson, 2016). Briefly, the cross-bridge power stroke appears to be produced by an angular swinging movement of the lever arm domain of S1 about a pivot point located at the junction with the catalytic domain (Fig. 1.2B). The level arm of S1 amplifies sub-nanometer movements associated with the closing and opening of the active site for ATP hydrolysis and release of the hydrolysis products, respectively. Ultimately, the swinging lever arm produces a displacement at the end of the neck of S1 that is on the order of 5–10 nm (Vale and Milligan, 2000; Geeves and Holmes, 2003) with little change in structure of the remainder of the molecule (Fig. 1.2B). With the base of the cross-bridge embedded in the thick filament, the swinging of the lever portion of S1 while the cross-bridge is attached to actin drives the displacement of the thin filament relative to the thick filament to generate shortening of muscle fibers. Similarly, under circumstances when there is resistance to filament sliding, the same molecular movement results in the deformation of the cross-bridge and the generation of force (Vale and Milligan, 2000; R€uegg et al., 2002). Strong support for the lever arm hypothesis of the power stroke was provided by ingenious and very elegant experiments that used molecular genetic techniques to alter the length of the lever arm portion of the myosin molecule, with the expectation that lever arm length would be related linearly to the speed at which myosin translates actin filaments in vitro (Uyeda et al., 1996) and to the magnitude of the displacement of the actin filaments during a single cross-bridge power stroke (Ruff et al., 2001). Increasing the length of the lever arm resulted in predictable increases in actin filament speed in an in vitro motility assay and in larger steps compared with the values for wild-type myosin. During contraction, individual myosin cross-bridges attach to actin, cycle through chemical and mechanical states that are not yet fully defined, detach, and then reattach (Fig. 1.3). This so-called cross-bridge cycle is driven in one direction by coupling the transitions between cross-bridge states to the steps of the hydrolysis of ATP by myosin with a single molecule of ATP hydrolyzed for each power stroke of the myosin cross-bridge. Overall, the cross-bridge cycle can be summarized as (i) the binding of ATP to myosin, resulting in its dissociation from actin; (ii) the hydrolysis of ATP and subsequent reassociation of myosin to actin; and (iii) the release of hydrolysis products during the power stroke (Geeves and Holmes, 2003). Two steps in the ATP hydrolysis reaction in muscle, ATP binding to myosin and phosphate release from myosin, are highly favorable energetically and act to drive the cycle in a clockwise direction, as illustrated in Fig. 1.3. The phosphate release step has been hypothesized to be tightly coupled

6

S.V. BROOKS ET AL.

Fig. 1.3. Cross-bridge cycle. The cross-bridge cycle can be described by key steps in the hydrolysis of ATP myosin: 1 ! 2. ATP binding to myosin in the rigor state results in in the dissociation of actin and myosin, i.e., the detachment of the cross-bridge from the thin filament; 2 ! 3. ATP hydrolysis, i.e., splitting of ATP into ADP and inorganic phosphate (Pi), 3 ! 4. Subsequent re-association of myosin and actin, i.e., reattachment of the cross-bridge to the thin filament; and 4 ! 5. The release of the hydrolysis products, ADP and Pi, during the power stroke. The cross-bridge cycle can only proceed in the presence of calcium that allows the cross-bridges to access their binding sites on the think filament. Created with BioRender.com.

to the initiation of the power stroke based on the observation that addition of phosphate during contraction causes a decrease in force generation. In contrast, neither the addition of ADP nor the additions of ADP and phosphate to fibers in rigor, with the rigor state corresponding to the postpower stroke actomyosin state with no nucleotide bound (state ⑤/① in Fig. 1.3), reduces tension. Consequently, while the phosphate release step appears to be reversible and strongly associated with force generation, once both ADP and phosphate have been released, driving the cycle counterclockwise from the rigor state is difficult if not impossible. However, whether the power stroke is coupled directly to the release of phosphate from AM
ADP
Pi (where A is actin and M is myosin), to an isomerization of AM
ADP
Pi or AM
ADP, to the dissociation of ADP from AM
ADP, or to some combination of these steps is not as yet definitively known (Månsson, 2016).

MUSCLE FIBER TYPES The coupling of the hydrolysis of a single molecule of ATP to each cross-bridge power stroke necessitates that the rate of cross-bridge cycling is determined by the

ATPase activity. The rate of cycling establishes how rapidly the thick and thin filaments can slide past one another and therefore how rapidly a sarcomere and ultimately a muscle fiber can shorten. Thus, velocity of shortening is determined by ATPase activity, which in turn is determined primarily by which myosin heavychain (MHC) isoform is present in the fiber. Each myosin heavy-chain isoform has its own characteristic ATPase activity, and the different myosins all derive from separate genes. There are at least 11 separate sarcomeric MHC genes (Schiaffino and Reggiani, 2011). In humans, these include both an embryonic and a neonatal skeletal muscle MHC gene in addition to the adult isoforms that are arranged in series on chromosome 17. Two cardiac MHC genes are located in tandem on chromosome 14, with the b-cardiac MHC being the predominant isoform expressed in slow skeletal muscle fibers, known as Type I fibers. The fast MHC isoforms expressed in the skeletal muscles of humans are Types 2A and 2X (also known as 2D), and an additional, very fast myosin, Type 2B, is found in the muscles of rodents and other small mammals. Fibers expressing these isoforms are correspondingly known as Type IIa, Type IIx, and Type IIb fibers. Finally, there is a specialized extraocular isoform as well as additional tissue and species-specific isoforms

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION that are not expressed in humans. The b-cardiac MHC is a myosin with a long ATPase cycle time that, as a consequence, uses ATP slowly and results in slow shortening velocities for Type I muscle fibers. The fast Type 2 myosins have high ATPase activities and more rapid ATPase cycle times; therefore, Type II muscle fibers expressing these myosins can achieve much higher shortening velocities.

CALCIUM ACTIVATION OF CONTRACTION The intracellular signal that allows cross-bridge cycling and initiates contraction is calcium. Two additional proteins associated with the thin filament mediate this control mechanism. Tropomyosin is a rod-shaped protein that is arranged end-to-end and winds around the thin filament backbone, following its helical structure (Fig. 1.3). Situated at the point of overlap between two tropomyosin molecules is the second critical regulatory protein, troponin (Fig. 1.3). Troponin is a heterotrimeric protein complex consisting of the calcium-binding subunit (TnC), the tropomyosin-binding subunit (TnT), and the so-called inhibitory subunit (TnI). Through tropomyosin’s binding to TnT and the binding of TnI to actin, tropomyosin is tethered to the thin filament in a position that physically blocks the cross-bridge binding sites on actin in the absence of calcium. When calcium becomes elevated in the cytoplasm of the muscle cell, calcium binds to TnC, resulting in the detachment of TnI from actin (Gordon et al., 2000). Once it is no longer tethered to the surface of the thin filament, the position of tropomyosin becomes highly dynamic. Tropomyosin is a flexible molecule and either rolls or slides along the surface of the thin filament resulting in transient exposure of actin-binding sites to the myosin cross-bridges (Vibert et al., 1997; Xu et al., 1999). When a muscle fiber is relaxed, cytosolic calcium level is extremely low but physiologic ATP concentrations in the cell are quite high (3–5 mM) favoring the accumulation of cross-bridges in an “energized” state, i.e., having hydrolyzed ATP but retaining the hydrolysis products bound within the nucleotide-binding site. Thus, a large proportion of the cross-bridges are poised to very rapidly associate with actin upon the movement of tropomyosin from its position blocking the myosin-binding sites on the thin filament (“cocked” state ③ in Fig. 1.3). Once one myosin cross-bridge is strongly bound to actin, tropomyosin is locally stabilized in a position that enhances the likelihood of additional myosin binding on nearby actin molecules. In this way, the activation of the thin filament can quickly propagate along its length to achieve full disinhibition of cross-bridge binding and maximum levels of cross-bridge cycling. Cross-bridge cycling continues until the thin filament once again becomes inhibited by a

7

decrease in calcium and the resultant reattachment of TnI to actin, which once again tethers tropomyosin in its position to block cross-bridge binding. In skeletal muscle, therefore, calcium plays a permissive role in allowing cross-bridge binding, rather than acting directly on myosin to impact cross-bridge cycling. This regulatory mechanism requires the ability of the muscle to rapidly increase and decrease the calcium concentration surrounding the myofilaments. In skeletal muscle, this is achieved by the maintenance of a large releasable intracellular calcium store and an exquisitely designed mechanism for the release of the calcium in response to neural signals activating contraction. All of the activating calcium in a skeletal muscle fiber is stored and released from an intracellular membrane compartment called the sarcoplasmic reticulum (SR). The SR surrounds the myofibrils like a sleeve (Fig. 1.4) creating very short diffusion distances for calcium to reach its binding sites on TnC. The SR membrane contains a high density of calcium transporters, known as the sarco/ endoplasmic reticulum Ca2+-ATPase (SERCA), that are constantly active at physiologic ATP concentrations pumping calcium ions from the cytoplasm into the SR lumen. The activity of the SERCA pumps maintains the cytosolic [Ca2+] very low and the SR [Ca2+] very high. Thus, a large concentration gradient drives rapid calcium release from the SR upon the activation and opening of SR calcium release channels, the ryanodine receptors (RyR1) (Rebbeck et al., 2014), in response to muscle action potentials.

Fig. 1.4. Triad structure. Muscle action potentials are propagated along the surface membrane (sarcolemma) and communicated to intracellular calcium stores (sarcoplasmic reticulum, SR) via the transverse tubules (t-tubule) that lie proximal to the terminal ends of the SR (terminal cisternae) in the triad region. Components of calcium release machinery, including the voltages sensors and calcium release channels, are located in the triad region with the voltage sensors embedded in the t-tubule membrane and matched in both position and orientation with the calcium release channels in the SR membrane. OpenStax https://commons.wikimedia.org/wiki/ File:1023 T-tubule.jpg.

8

S.V. BROOKS ET AL.

The muscle action potential is propagated across the surface of the fiber via the sarcolemma and is communicated throughout the muscle cell by traveling down specialized invaginations in the surface membrane called transverse tubules (T tubules) (Fig. 1.4). The T tubules propagate the action potential to the center of the muscle fiber where the depolarization is detected by voltagegated L-type Ca2+ channels, CaV1.1, also known as the dihydropyridine receptor (DHPR), that are embedded in the T tubular membrane (Rios and Brum, 1987; Tanabe et al., 1988). In skeletal muscle, the terminal ends of the SR come in very close proximity to the T tubules (Fig. 1.4), forming a structure known as the triad junction (Flucher et al., 1992; Franzini-Armstrong and Jorgensen, 2003). Detailed structural studies, nicely reviewed in Protasi (2002) and Hernández-Ochoa et al. (2016), show that RyR1 is located on the SR surface facing the T-tubules, while the DHPRs within the T tubular membrane are precisely matched in both position and orientation with RyR1. Depolarization-induced calcium release does not occur if the alignment between DHPRs and RyR1s is disrupted suggesting that the communication between the voltage sensor (DHPR) and the calcium release channel (RyR1) involves a physical interaction. Although mechanical coupling between these two proteins is widely accepted (Block et al., 1988; Paolini et al., 2004), the precise nature of the interaction has not been elucidated. Current views of the molecular mechanisms by which DHPR regulates RyR1 are nicely summarized in a recent review (Shishmarev, 2020). DHPR was first identified as the skeletal muscle voltage sensor by Rios and Brum (1987). DHPR functions as a voltage-gated L-type Ca2+ channel but its function as a Ca2+ channel is entirely dispensable for skeletal muscle excitation-contraction (EC) coupling, i.e., depolarizationinduced SR calcium release and activation of the myofilaments, to proceed normally. Rather than the calcium current, it is the conformational change within DHRP elicited in response to the action potential that is responsible for transducing the signal to RyR1. The mechanism may involve a direct mechanical coupling between the two proteins or may be mediated by an as yet unidentified closely associated protein, and recent reports have identified two additional proteins that appear required to maintain EC coupling intact. In 2013, studies in both zebrafish (Horstick et al., 2013) and mice (Nelson et al., 2013) showed that deletion of STAC3 (SH3 and cysteine-rich domain containing protein 3) abolished membrane depolarization-induced SR calcium release, indicating that STAC3 is a required component of the EC coupling machinery (Polster et al., 2016). STAC3 is localized to the triad region and is now known to bind to DHPR (Wong King Yuen et al., 2017), although direct interaction between STAC3 and RyR1 has not yet been demonstrated. Finally, a fourth component of the EC

coupling machinery, junctophilin2, is not directly involved in transducing the depolarization signal to SR calcium release; rather, junctophilins are important for the formation and maintenance of the triad junctions (Takeshima et al., 2000). The triad structure is essential for the proper alignment of DHPR and RyR1 and thus critical for the maintaining EC coupling.

SKELETAL MUSCLE FUNCTION In vivo, whether a muscle shortens, remains at a fixed length, or is lengthened during contractions depends on the interaction between the magnitudes of the force developed by the muscle and the external load placed on the muscle (Faulkner, 2003). When the force developed by the muscle is greater than the load on the muscle, the fibers shorten during the contraction. When the force developed by the muscle is equal to the load or if the load is immovable, the overall length of the muscle remains the same, resulting in an isometric contraction. If the force developed by the muscle is less than the load placed on the muscle, the muscle is stretched during the contraction. In general, the initiation of a movement or the acceleration of a body part requires a shortening contraction and the generation of power (work/time or force  velocity), whereas stopping or slowing that involves braking actions result in lengthening contractions. Most activities require varying proportions of each type of contraction, but clearly the ability to generate power, rather than simply isometric force, is the most physiologically relevant marker of performance. Although there are conflicting reports in the literature regarding the force generating capacity of different myosin isoforms, single molecule data (Bottinelli and Reggiani, 2000) suggest that the step size and unitary force are not substantially different between muscle myosins. Thus, power output is determined largely by shortening velocity, which varies many-fold between slow and fast adult myosin isoforms within a given species.

NEUROMUSCULAR JUNCTION STRUCTURE AND DEVELOPMENT Each individual muscle fiber is innervated by a single axonal branch of a motor neuron. Action potentials are transmitted to the muscle fiber through a specialized synapse called the neuromuscular junction (NMJ) (Fig. 1.5). NMJs are the fundamental site where the nervous system transmits signals responsible for voluntary contractions to skeletal muscle. The NMJ is considered a “slave” synapse based on the expectation that action potentials will be initiated on the muscle fiber every time an action potential arrives at the nerve terminal (Ruff, 2011). Thus, defects in NMJ formation and development can lead to

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION

9

Fig. 1.5. Components of the neuromuscular junction (NMJ). Action potentials are propagated down the presynaptic motor neuron that is surrounded and supported by perisynaptic Schwann cells. Depolarization of the motor neuron results in the opening of voltage-sensitive calcium channels leading to an influx of calcium ions, which contributes to synaptic vesicle fusion and the release of the neurotransmitter, acetylcholine (ACh). ACh crosses the synaptic gap and binds to acetylcholine receptors (AChR) on the muscle membrane leading to Na+ ion influx, depolarization, and muscle contraction. Created with BioRender.com.

neuromuscular disorders. The intricate structure of the NMJ forms and develops early in life through a series of steps involving the nerve terminal, muscle fiber, and Schwann cells to produce a chemical synapse (Sanes and Lichtman, 2003). Our understanding of synapse formation and maturation comes largely from studies of rodents (Cetin et al., 2020) and amphibians (Heuser and Reese, 1973; Peng and Nakajima, 1978), so it should be recognized that this knowledge may not universally apply directly to human NMJs. Indeed, comparative analyses indicate a high degree of heterogeneity in mammalian NMJ morphology, with the NMJ morphology of sheep closely matching humans, while mice display very large NMJs, especially relative to the small muscle fiber diameters. Nevertheless, mice are a practical animal model for these studies, and the processes involved in NMJ formation and maintenance are undoubtedly comparable across species (Boehm et al., 2020). During myogenesis, myoblasts fuse to form myotubes, and prior to the appearance of the motor nerve,

acetylcholine receptor (AChR) genes are transcribed. The AChR gene products spontaneously form aneural clusters with a density of 1000/mm2 throughout the muscle fiber membrane during a phenomenon referred to as muscle prepatterning. Upon arrival of the motor nerve, the axon terminal, also known as a synaptic bouton, secretes agrin at the synaptic cleft (Kummer et al., 2006; Li et al., 2018). Agrin is a potent activator of AChR cluster formation and regulator of synapse development (Mittaud et al., 2004) through its controls of a core group of proteins including low-density lipoprotein receptorrelated protein 4 (LRP4), muscle-specific tyrosine kinase (MuSK), Docking Protein 7 (Dok7) (Wu et al., 2010; Tintignac et al., 2015) and rapsyn, which directly interacts with AChRs (Borges et al., 2008). NMJ development proceeds over the course of weeks as AChR clusters mature and structurally organize into a pretzel-like morphology (Marques et al., 2000). As AChRs organize during NMJ development, the postsynaptic membrane specializes by forming invaginations

10

S.V. BROOKS ET AL.

known as junctional folds (JFs). These folds are unique to the NMJ and contain high-density clusters of nicotinic AChRs. Once formed, the density of AChRs at the endplate is >10,000 AChR/mm2. Thus, the JFs are thought to increase the reliability of transmission and enhance depolarization. The mechanism of JF formation is unknown, but a leading theory suggests that JFs develop opposite to the active zone, the area where vesicles release acetylcholine across the synapse. While acetylcholine release likely plays a role in establishing the JF, the mechanisms by which the membrane is physically drawn in to form the actual invaginations remains unknown (Hong and Etherington, 2011; Zou and Pan, 2022). Nevertheless, the JFs appear to be highly relevant functionally, as defects in JFs are a consistent phenotype in disease states such as amyotrophic lateral sclerosis (ALS), Myasthenia gravis (MG), and muscular dystrophy (MD) (York and Zheng, 2017). Although AChRs have an expected half-life of about 14 days at fully functional mature synapses (Akaaboune et al., 1999), AChRs are also recycled throughout the lifespan of an animal, and recycled AChRs turnover more rapidly than pre-existing AChRs, although the mechanism is still unclear. Several regulatory signaling molecules converge to control the stability of AChRs and are well-reviewed (Valenzuela and Akaaboune, 2021).

NEUROTRANSMISSION The stability of the NMJ at mature synapses is highly robust enabling neurotransmission and supporting essentially lifelong motor performance (Sanes and Lichtman, 2001; Slater, 2017). The sequence of events responsible for neuromuscular transmission has been well characterized. In response to a motor neuron action potential, acetylcholine-carrying vesicles fuse with the nerve terminal membrane resulting in the release of the neurotransmitter into the synaptic cleft (Fig. 1.5). Acetylcholine diffuses across the synapse and binds to the postsynaptic AChRs. Each AChR contains two agonist-binding sites on the extracellular domain (Nayak et al., 2014, 2016), and the binding of two acetylcholine molecules activates the opening of the AChR channel allowing sodium influx and potassium efflux resulting in a net positive flow of charge and membrane depolarization known as the endplate potential (EPP). The density of the receptors and the amount of neurotransmitter released ensure that in a healthy muscle every nerve action potential results in an EPP with sufficient magnitude of depolarization to trigger a muscle action potential and muscle contraction (Aidley, 1998). The ligand-gated nicotinic AChR ion channels in muscle are heteropentamers composed of two-a, oneb, and one fetal-specific subunit that forms a central pore.

The E-subunit replaces the embryonic-specific g-subunit within the first 2 weeks of postnatal life in mice (Missias et al., 1996; Villarroel and Sakmann, 1996), whereas in humans the g-to-E subunit transition occurs prenatally during the second to the third trimester of development (Hesselmans et al., 1993). Complex molecular pathways regulate the g-to-E subunit change; however, motor neuron innervation appears critical for stimulating subsynaptic nuclei to initiate transcription of the epsilon subunit. The gamma subunit is active in all myonuclei in the initial formation of myotubes but becomes restricted to subsynaptic myonuclei upon the arrival of the nerve. mRNA expression of the E-subunit also seems to be exclusive to the adult subsynaptic nuclei (Sakmann and Brenner, 1978; Missias et al., 1996). Defects in this developmental milestone have been associated with null mutations to the CHRNE gene responsible for the production of the E-subunit leading to NMJ disorders such as congenital myasthenic syndrome (Croxen et al., 2001). The physiology of the neuromuscular junction is dictated by both the structure and function of the presynaptic motor neuron and the postsynaptic AChRs. The presynaptic active zone is the synaptic vesicle release site, while the number of active zones per presynaptic terminal is referred to as active zone density. The size of the active zone is significantly smaller in mice (Fukuoka et al., 1987) than in humans (Fukunaga et al., 1982) with a similar density observed between species (Fukunaga et al., 1982; Fukuoka et al., 1987; Chen et al., 2012). The mouse NMJ also is estimated to have more than double the amount of neurotransmitter released per nerve impulse, known as quantal content, compared with human NMJs (Slater, 2017), and mouse nerve terminal areas are substantially greater (Boehm et al., 2020). Finally, mice display larger endplates that are densely occupied by AChRs compared with humans (Albuquerque et al., 1974; Pestronk et al., 1985; Bollen et al., 1992). While the mouse NMJ possesses anatomic and physiologic variations from humans, mouse models continue to be essential to further understanding of the development of the NMJ and the mechanisms underlying pathology due to NMJ dysfunction. Novel proteins and molecules associated with the development and maintenance of the NMJ continue to be discovered. Podosomes are organelles capable of remodeling the extracellular matrix and have been identified in the postsynaptic apparatus (Proszynski et al., 2009; Bernadzki et al., 2014). While the role of podosomes at the NMJ remains to be fully understood, podosomes may remodel the endplate through interaction with dynamin. Dynamin is a well-studied mechanochemical enzyme that catalyzes membrane fission during endocytosis and synaptic vesicle recycling.

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION The Dynamin-2 isoform has been reported to be enriched in the synaptic podosome (Lin et al., 2020) and has been established to have a pivotal role at the postsynaptic membrane of NMJs by acting as an actin-remodeling GTPase, promoting AChR cluster perforation, and organizing the postsynaptic cytoskeleton to maintain the function of the NMJ (Lin et al., 2020). The extracellular matrix proteins Hevin and secreted protein acidic and rich in cysteine (SPARC) were initially identified for their role in the formation of brain synapses but have recently been shown to be involved in the development and reformation of endplates following motor neuron injury (Brayman et al., 2021). Finally, the Van Gogh-like protein 2 (Vangl2), critical for Wnt signaling, has recently been reported to reproduce NMJ disassembly and impaired neurotransmission with deficits in motor function when conditionally ablated in muscle. Furthermore, Vangl2 was shown to couple with MuSK, and loss of Vangl2 disrupted postsynaptic AChR clustering of NMJs in vivo (Boëx et al., 2022), likely adding to the list of core proteins required for NMJ development. As novel proteins and mechanisms at the NMJ continue to be discovered, our understanding and development of druggable targets may lead to improved prognosis and treatment of clinical conditions.

MOTOR UNIT PROPERTIES While each muscle fiber is innervated by a single axonal branch at a single NMJ, each motor neuron branches to innervate many muscle fibers. A motor neuron, its branches, and the muscle fibers innervated by the branches constitute a motor unit (Fig. 1.6). The motor unit is the smallest group of fibers within a muscle that can be activated volitionally. Activation of a motor unit occurs when action potentials emanating from the motor cortex depolarize the cell bodies of motor neurons in the ventral root of the spinal cord. The depolarization generates an action potential in the motor neuron that is transmitted to each muscle fiber in the motor unit, and each fiber then contracts more or less simultaneously. Motor units range in size from small units with perhaps only several dozen muscle fibers to large units containing many thousands of fibers and are classified functionally on the basis of the mechanical and metabolic properties of the fibers in the motor unit. Muscle fiber types are defined by speed of shortening and resistance to fatigue but are often simply referred to as slow twitch or fast twitch or sometimes red or white. The binning of fiber types into binary classifications has a historical basis but has also created the temptation to assume that the classifications map onto one another. This is simply not so, as the mechanistic basis for contractile speed vs. fatigability are entirely distinct. Moreover, the

11

Fig. 1.6. Anatomy of the motor unit. Axons from motor neurons located in the ventral root of the spinal cord extend to the muscle. Each axon divides into a number of axon branches which are further divided into axon terminals that innervate individual muscle fibers at the neuromuscular junctions. Muscle fibers innervated by the same motor neuron constitute a single motor unit. Created with BioRender.com.

expression of specific myosin isoforms as described previously in this chapter, which define contractile speed create discreet classifications of fiber type, whereas fatigability constitutes a continuum that is widely variable across muscles, species, habitual levels of activity and disease state. Fatigability, or more appropriately fatigue resistance is determined by the ability of the muscle fibers to establish and maintain energy balance. To maintain energy balance, the muscle must be able to generate ATP as rapidly as it is utilized. The ability to generate ATP is governed primarily by the concentration and activities of the enzymes of oxidative metabolism, or the oxidative capacity of the fibers. Skeletal muscle has an enormous capacity to adapt to the habitual level of demand for physical activity placed on it by enhancing its oxidative capacity and probably altering MHC expression. Because all muscle fibers in a motor unit contract simultaneously, the pressures for gene expression associated with activity, in terms of both frequency and loading, are identical in all of the fibers of the motor unit. Consequently, in normal healthy muscles, all fibers within a motor unit have similar biochemical and, therefore, functional properties. The details of how the effects of frequency and loading are transduced into fiber type determination at the molecular level are not fully understood (Goldspink, 1999). The mesodermal transcription factor T-box 15 (Tbx15) has specific and robust expression in glycolytic

12

S.V. BROOKS ET AL.

myofibers, while PGC-1 alpha is expressed preferentially in muscle enriched in Type I oxidative fibers. When PGC-1 alpha expression is increased in a muscle specific manner, muscles normally rich in Type II fibers display activation mitochondrial genes and also express proteins characteristic of Type I fibers. The muscles of the PGC-1 alpha overexpressing transgenic mice also show a much greater resistance to fatigue during repeated contractions (Lin et al., 2002). Upon ablation of Tbx15 in vivo, the number of glycolytic fibers decreases with an increase in the number of oxidative fibers and an overall reduction in muscle size (Lee et al., 2015). Recent single-fiber proteomic studies have established fiber type specific protein profiles that provide further insight into muscle fiber molecular heterogeneity (Murgia et al., 2021) and may lead to further understanding of the molecular transducers of physical activity that define fiber types.

MOTOR UNIT RECRUITMENT Slow (S) motor units generally have the fewest muscle fibers per motor unit, referred to as the innervation ratio, and the lowest velocity of shortening. Slow muscle fibers contain many mitochondria, giving the S motor units a high capacity to replenish ATP. As a consequence of their small size and slow shortening velocity, S motor units are recruited during tasks that require low force or power. Their small size allows for small increments in force generation with recruitment of additional motor units, thus also allow for highly precise actions. The low rate of ATP usage results in the economical maintenance of force during isometric contractions as well as efficient performance of repetitive, slow, shortening contractions (Walklate et al., 2016). Fast motor units display larger innervation ratios, and the muscle fibers within the motor units express fast myosins. Thus, fast motor units generate higher forces and are capable of high shortening velocities and are recruited under circumstances when high power output is needed or when isometric force produced by slow motor units is insufficient. The fast motor units are classified further as fast fatigable (FF) or fast fatigue resistant (FR). FF motor units tend to be the largest and have the highest velocities of shortening. The maximum normalized power (W/kg) developed by FF units is as much as fourfold greater than that of the S units due to the higher velocity of shortening for the fibers in the FF units; however, these motor units are typically recruited for only individual or very short-duration high-force contractions or high-power movements due to their low oxidative capacity and consequential high fatigability. The FR units are intermediate in terms of size and velocity but can have extremely high oxidative capacity (Kugelberg and Lindegren, 1979).

The ability to accomplish specific motor tasks is determined by the composition of one’s muscles, i.e., the proportion and properties of each type of motor unit, and the ability to optimally activate specific populations of motor units. Classically, recruitment of motor units is thought to be dictated by the size of the motor neurons, which defines their excitability. Smaller motor neurons that innervate small S units are the most highly excitable and are therefore recruited first followed by larger and faster motor units with the eventual recruitment of FF units. This phenomenon is known as the Heneman Size Principle (Cope and Pinter, 1995). The size principle is also observed in the opposite direction when reducing muscle contractile force, whereby larger motor units are deactivated before smaller motor units. Furthermore, in muscles that that are subdivided into anatomic compartments with distinct primary nerve branching and mechanical separation, the muscle fibers for a single motor unit also occupy a compartmentalized space within the CSA of the muscle (Monti et al., 2001). Based on the size principle and compartmentalization of motor unit fibers, recruitment of motor units would occur first in smaller oxidative muscle fibers, typically found in the deeper portions of the muscle, followed by gradual recruitment of larger glycolytic muscle fibers, typically located in the more superficial regions of the whole muscle (Delp and Duan, 1996). Although the size principle likely holds for many, if not most, activities, the motor control system is capable of adapting to optimize motor unit recruitment patterns for the complexity of movements that animals, including humans (Del Vecchio et al., 2019), can achieve. The progressive recruitment of motor units that leads to gradual increases in force production requires an increase in the frequency of the motor neuron action potentials or motor unit discharge rate. The control of motor unit force by changes in the frequency of the motor neuron action potentials is referred to as rate coding. Motor units that are recruited early during contraction reach a maximal discharge rate that plateaus and remains constant despite increases in the net muscle force production as additional larger motor units are recruited. All motor units have a saturation discharge rate where increases in motor neuron firing rate can no longer elicit an increase in muscle force output from the individual fibers of that motor unit. This limit in motor unit discharge rate is due to the intrinsic capacity of the motor neuron to produce action potentials despite further increases in excitatory inputs from excitatory neurons or exogenous stimulation (Fuglevand et al., 2015). Moreover, force generation by an individual fiber, muscle or motor unit only increases with frequency to the point where the thin filament regulatory system is maximally activated, and a maximum number of cross-bridges are cycling.

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION The force-frequency relationship for a motor unit can be determined relatively easily by stimulating motor unit axons over a range of frequencies and measuring the evoked forces. Experimentally, evoked forces in humans require stimulation frequencies well over 50 Hz; however, during voluntary contractions, motor unit discharge rates are typically less than 50 Hz (Heckman and Enoka, 2012). Ballistic contractions, i.e., those that involve very rapid force development, exhibit different firing rate patterns compared to standard muscle contractions, such that when test subjects are asked to rapidly contract a muscle, rapid increases in motor unit firing rates, exceeding 60–120 Hz, are seen at the beginning of the contractions followed by a decline in firing rate (Desmedt and Godaux, 1977; Bawa and Calancie, 1983; Klass et al., 2008). In addition to the contribution of rate coding to muscle force modulation during rapid compared with slower-developing contractions, differences in firing rate are also observed between lengthening and shortening contractions at submaximal levels (5%–30% MVC). When comparing submaximal shortening and lengthening contractions while controlling for torque and fascicle length in dorsiflexor muscles, discharge rates for lengthening contractions remained stable throughout the contractions, whereas motor unit firing rates progressively increased throughout the shortening phase of the task (Pasquet et al., 2006). The difference in discharge rate from between lengthening and shortening contractions is thought to be from an increase in neural drive in shortening contractions to compensate for the reduced force capacity at shorter fascicle lengths (Duchateau and Enoka, 2016). Motor unit firing rates are also modifiable by exercise training. Twelve weeks (five sessions per week) of training the ankle dorsiflexor muscles to lift a load (30–40 MVC) with rapid contractions enhanced the both the rate of force development and the average discharge rate of motor units in human tibialis anterior muscles (Van Cutsem et al., 1998). In addition, there were a greater incidence of double discharge motor units (>200 Hz) with training, an important yet uncommon motor unit discharge speed that is thought to be an important strategy for motor unit control during the start of their contraction (Mrówczy nski et al., 2015). Finally, motor unit discharge rates appear to be negatively impacted by aging such that discharge rate decline in older adults. With aging, both motor unit discharge rates and the rate of force development for motor units in the tibialis anterior are reduced in older adults (71–84 years of age) compared to young adults (18–22 years of age) during rapid dorsiflexion (Klass et al., 2008). Training effect size (increased discharge rate) is also reduced in older adults compared to young adults. Finally, the presence of double discharge motor units that discharge greater than

13

200 Hz in the tibialis anterior are fewer in proportion in older adults (4.6%) compared to young (8.4%) adults. Although rate coding is an important means to modulate force at the level of individual motor units, especially to allow rapid force development, it should be recognized that motor units are most often recruited at a frequency that results in near maximum activation of the fibers in the motor unit and controlling the number of active motor units is the most important physiologic means of governing muscle force generation and power output in vivo.

REFERENCES Aidley DJ (1998). The physiology of excitable cells, fourth edn. Cambridge University Press. Akaaboune M, Culican SM, Turney SG et al. (1999). Rapid and reversible effects of activity on acetylcholine receptor density at the neuromuscular junction in vivo. Science 286: 503–507. Albuquerque E, Barnard EA, Porter CW et al. (1974). The density of acetylcholine receptors and their sensitivity in the postsynaptic membrane of muscle endplates. Proc Natl Acad Sci 71: 2818–2822. Bawa P, Calancie B (1983). Repetitive doublets in human flexor carpi radialis muscle. J Physiol 339: 123–132. Bernadzki KM, Rojek KO, Pro´szy nski TJ (2014). Podosomes in muscle cells and their role in the remodeling of neuromuscular postsynaptic machinery. Eur J Cell Biol 93: 478–485. Block BA, Imagawa T, Campbell KP et al. (1988). Structural evidence for direct interaction between the molecular components of the transverse tubule/sarcoplasmic reticulum junction in skeletal muscle. J Cell Biol 107: 2587–2600. Boehm I, Alhindi A, Leite AS et al. (2020). Comparative anatomy of the mammalian neuromuscular junction. J Anat 237: 827–836. Boe¨x M, Cottin S, Halliez M et al. (2022). The cell polarity protein Vangl2 in the muscle shapes the neuromuscular synapse by binding to and regulating the tyrosine kinase MuSK. Sci Signal 15: 1–14. Bollen E, Den Heyer JC, Tolsma MHJ et al. (1992). Structure and function of neuromuscular junctions in the vastus lateralis of man. Brain 115: 451–478. Borges LS, Yechikhov S, Lee YI et al. (2008). Identification of a motif in the acetylcholine receptor b subunit whose phosphorylation regulates rapsyn association and postsynaptic receptor localization. J Neurosci 28: 11468–11476. Bottinelli R, Reggiani C (2000). Human skeletal muscle fibres: molecular and functional diversity. Prog Biophys Mol Biol 73: 195–262. Brayman VL, Taetzsch T, Miko M et al. (2021). Roles of the synaptic molecules Hevin and SPARC in mouse neuromuscular junction development and repair. Neurosci Lett 746. Cetin H, Beeson D, Vincent A et al. (2020). The structure, function, and physiology of the fetal and adult acetylcholine receptor in muscle. Front Mol Neurosci 13: 1–14.

14

S.V. BROOKS ET AL.

Chen J, Mizushige T, Nishimune H (2012). Active zone density is conserved during synaptic growth but impaired in aged mice. J Comp Neurol 520: 434–452. Cope TC, Pinter MJ (1995). The size principle: still working after all these years. Am J Phys 10: 280–286. https://doi. org/10.1152/physiologyonline. Croxen R, Young C, Slater C et al. (2001). End-plate g- and E-subunit mRNA levels in AChR deficiency syndrome due to E-subunit null mutations. Brain 124: 1362–1372. Del Vecchio A, Casolo A, Negro F et al. (2019). The increase in muscle force after 4 weeks of strength training is mediated by adaptations in motor unit recruitment and rate coding. J Physiol 597: 1873–1887. Delp MD, Duan C (1996). Composition and size of type I, IIA, IID/X, and IIB fibers and citrate synthase activity of rat muscle. J Appl Physiol (Bethesda, Md: 1985) 80: 261–270. Desmedt JE, Godaux E (1977). Ballistic contractions in man: characteristic recruitment pattern of single motor units of the tibialis anterior muscle. J Physiol 264: 673–693. Duchateau J, Enoka RM (2016). Neural control of lengthening contractions. J Exp Biol 219: 197–204. Faulkner JA (2003). Terminology for contractions of muscles during shortening, while isometric, and during lengthening. J Appl Physiol (1985) 95: 455–459. https://doi.org/10.1152/ japplphysiol.00280.200395. Flucher BE, Phillips JL, Powell JA et al. (1992). Coordinated development of myofibrils, sarcoplasmic reticulum and transverse tubules in normal and dysgenic mouse skeletal muscle, in vivo and in vitro. Dev Biol 150: 266–280. Franzini-Armstrong C, Jorgensen AO (2003). Structure and development of E-C coupling units in skeletal muscle. Annu Rev Physiol 56: 509–534. https://doi.org/10.1146/ annurev.ph.56.030194.002453. Fuglevand AJ, Lester RA, Johns RK (2015). Distinguishing intrinsic from extrinsic factors underlying firing rate saturation in human motor units. J Neurophysiol 113: 1310–1322. Fukunaga H, Engel AG, Osame M et al. (1982). Paucity and disorganization of presynaptic membrane active zones in the Lambert-Eaton myasthenic syndrome. Muscle Nerve 5: 686–697. Fukuoka T, Engel AG, Lang B et al. (1987). Lambert-Eaton myasthenic syndrome: I. early morphological effects of IgG on the presynaptic membrane active zones. Ann Neurol 22: 193–199. Geeves MA, Holmes KC (2003). Structural mechanism of muscle contraction. Annu Rev Biochem 68: 687–728. https://doi.org/10.1146/annurev.biochem.68.1.687. Goldspink G (1999). Changes in muscle mass and phenotype and the expression of autocrine and systemic growth factors by muscle in response to stretch and overload. J Anat 194: 323–334. Gordon AM, Homsher E, Regnier M (2000). Regulation of contraction in striated muscle. Physiol Rev 80: 853–924. Heckman CJ, Enoka RM (2012). Motor unit. Comprehens Phys 2: 2629–2682.

Herna´ndez-Ochoa EO, Pratt SJP, Lovering RM et al. (2016). Critical role of intracellular RyR1 calcium release channels in skeletal muscle function and disease. Front Physiol 6: 1–11. Hesselmans LFGM, Jennekens FGI, Van Den Oord CJM et al. (1993). Development of innervation of skeletal muscle fibers in man: relation to acetylcholine receptors. Anat Rec 236: 553–562. Heuser JE, Reese TS (1973). Evidence for recycling of synaptic vesicle membrane during transmitter release at the frog neuromuscular junction. J Cell Biol 57: 315–344. Hong IHK, Etherington SJ (2011). Neuromuscular junction. eLS Horstick EJ, Linsley JW, Dowling JJ et al. (2013). Stac3 is a component of the excitation–contraction coupling machinery and mutated in Native American myopathy. Nat Commun 2952: 4:1–4:1-11. Klass M, Baudry S, Duchateau J (2008). Age-related decline in rate of torque development is accompanied by lower maximal motor unit discharge frequency during fast contractions. J Appl Physiol 104: 739–746. Kugelberg E, Lindegren B (1979). Transmission and contraction fatigue of rat motor units in relation to succinate dehydrogenase activity of motor unit fibres. J Physiol 288: 285–300. Kummer TT, Misgeld T, Sanes JR (2006). Assembly of the postsynaptic membrane at the neuromuscular junction: paradigm lost. Curr Opin Neurobiol 16: 74–82. Lee KY, Singh MK, Ussar S et al. (2015). Tbx15 controls skeletal muscle fibre-type determination and muscle metabolism. Nat Commun 6: 1–12. Li L, Xiong WC, Mei L (2018). Neuromuscular junction formation, aging, and disorders. Annu Rev Physiol 80: 159–188. https://doi.org/10.1146/annurev-physiol-022516034255. Lin J, Wu H, Tarr PT et al. (2002). Transcriptional co-activator PGC-1 alpha drives the formation of slow-twitch muscle fibres. Nature 418: 797–801. Lin SS, Hsieh TL, Liou GG et al. (2020). Dynamin-2 regulates postsynaptic cytoskeleton organization and neuromuscular junction development. Cell Rep 33: 1–16. Ma˚nsson A (2016). Actomyosin based contraction: one mechanokinetic model from single molecules to muscle? J Muscle Res Cell Motil 37: 181–194. Marques MJ, Conchello JA, Lichtman JW (2000). From plaque to pretzel: fold formation and acetylcholine receptor loss at the developing neuromuscular junction. J Neurosci 20: 3663–3675. Missias AC, Chu GC, Klocke BJ et al. (1996). Maturation of the acetylcholine receptor in skeletal muscle: regulation of the AChR g-to-e switch. Dev Biol 179: 223–238. Mittaud P, Camilleri AA, Willmann R et al. (2004). A single pulse of agrin triggers a pathway that acts to cluster acetylcholine receptors. Mol Cell Biol 24: 7841–7854. Monti RJ, Roy RR, Reggie Edgerton V (2001). Role of motor unit structure in defining function. Muscle Nerve 24: 848–866.

SKELETAL MUSCLE STRUCTURE, PHYSIOLOGY, AND FUNCTION Mro´wczynski W, Celichowski J, Raikova R et al. (2015). Physiological consequences of doublet discharges on motoneuronal firing and motor unit force. Front Cell Neurosci 9: 1–6 Murgia M, Nogara L, Baraldo M et al. (2021). Protein profile of fiber types in human skeletal muscle: a single-fiber proteomics study. Skelet Muscle 11: 1–19. Nayak TK, Bruhova I, Chakraborty S et al. (2014). Functional differences between neurotransmitter binding sites of muscle acetylcholine receptors. Proc Natl Acad Sci U S A 111: 17660–17665. Nayak TK, Chakraborty S, Zheng W et al. (2016). Structural correlates of affinity in fetal versus adult endplate nicotinic receptors. Nat Commun 7: 1–14. Nelson BR, Wu F, Liu Y et al. (2013). Skeletal muscle-specific T-tubule protein STAC3 mediates voltage-induced Ca2+ release and contractility. Proc Natl Acad Sci U S A 110: 11881–11886. Paolini C, Protasi F, Franzini-Armstrong C (2004). The relative position of RyR feet and DHPR tetrads in skeletal muscle. J Mol Biol 342: 145–153. Pasquet B, Carpentier A, Duchateau J (2006). Specific modulation of motor unit discharge for a similar change in fascicle length during shortening and lengthening contractions in humans. J Physiol 577: 753–765. Peng HB, Nakajima Y (1978). Membrane particle aggregates in innervated and noninnervated cultures of Xenopus embryonic muscle cells. Proc Natl Acad Sci 75: 500–504. Pestronk A, Drachman DB, Self SG (1985). Measurement of junctional acetylcholine receptors in myasthenia gravis: clinical correlates. Muscle Nerve 8: 245–251. Polster A, Nelson BR, Olson EN et al. (2016). Stac3 has a direct role in skeletal muscle-type excitation-contraction coupling that is disrupted by a myopathy-causing mutation. Proc Natl Acad Sci U S A 113: 10986–10991. Proszynski TJ, Gingras J, Valdez G et al. (2009). Podosomes are present in a postsynaptic apparatus and participate in its maturation. Proc Natl Acad Sci U S A 106: 18373–18378. Protasi F (2002). Structural interaction between RYRs and DHPRs in calcium release units of cardiac and skeletal muscle cells. Front Biosci 7: 650–658. Rebbeck RT, Karunasekara Y, Board PG et al. (2014). Skeletal muscle excitation–contraction coupling: who are the dancing partners? Int J Biochem Cell Biol 48: 28–38. Rios E, Brum G (1987). Involvement of dihydropyridine receptors in excitation–contraction coupling in skeletal muscle. Nature 325: 717–720. Risal D, Gourinath S, Himmel DM et al. (2004). Myosin subfragment 1 structures reveal a partially bound nucleotide and a complex salt bridge that helps couple nucleotide and actin binding. Proc Natl Acad Sci U S A 101: 8930–8935. R€ uegg C, Veigel C, Molloy JE et al. (2002). Molecular motors: force and movement generated by single myosin II molecules. News Physiol Sci 17: 213–218.

15

Ruff RL (2011). Endplate contributions to the safety factor for neuromuscular transmission. Muscle Nerve 44: 854–861. Ruff C, Furch M, Brenner B et al. (2001). Single-molecule tracking of myosins with genetically engineered amplifier domains. Nat Struct Biol 8: 226–229. Sakmann B, Brenner HR (1978). Change in synaptic channel gating during neuromuscular development. Nature 276: 401–402. Sanes JR, Lichtman JW (2001). Induction, assembly, maturation and maintenance of a postsynaptic apparatus. Nat Rev Neurosci 2: 791–805. Sanes JR, Lichtman JW (2003). Development of the vertebrate neuromuscular junction. Annu Rev Neurosci 22: 389–442. https://doi.org/10.1146/annurev.neuro.22.1.389. Schiaffino S, Reggiani C (2011). Fiber types in mammalian skeletal muscles. Physiol Rev 91: 1447–1531. Shishmarev D (2020). Excitation-contraction coupling in skeletal muscle: recent progress and unanswered questions. Biophys Rev 12: 143–153. Slater CR (2017). The structure of human neuromuscular junctions: some unanswered molecular questions. Int J Mol Sci 18: 2183. Takeshima H, Komazaki S, Nishi M et al. (2000). Junctophilins: a novel family of junctional membrane complex proteins. Mol Cell 6: 11–22. Tanabe T, Beam KG, Powell JA et al. (1988). Restoration of excitation—contraction coupling and slow calcium current in dysgenic muscle by dihydropyridine receptor complementary DNA. Nature 336: 134–139. Tintignac LA, Brenner HR, R€ uegg MA (2015). Mechanisms regulating neuromuscular junction development and function and causes of muscle wasting. Physiol Rev 95: 809–852. Uyeda TQP, Abramson PD, Spudich JA (1996). The neck region of the myosin motor domain acts as a lever arm to generate movement. Proc Natl Acad Sci 93: 4459–4464. Vale RD, Milligan RA (2000). The way things move: looking under the hood of molecular motor proteins. Science 288: 88–95. Valenzuela IMPY, Akaaboune M (2021). The metabolic stability of the nicotinic acetylcholine receptor at the neuromuscular junction. Cell 10: 358. Van Cutsem M, Duchateau J, Hainaut K (1998). Changes in single motor unit behaviour contribute to the increase in contraction speed after dynamic training in humans. J Physiol 513: 295–305. Vibert P, Craig R, Lehman W (1997). Steric-model for activation of muscle thin filaments. J Mol Biol 266: 8–14. Villarroel A, Sakmann B (1996). Calcium permeability increase of endplate channels in rat muscle during postnatal development. J Physiol 496: 331–338. Walklate J, Vera C, Bloemink MJ et al. (2016). The most prevalent Freeman-Sheldon syndrome mutations in the

16

S.V. BROOKS ET AL.

embryonic myosin motor share functional defects. J Biol Chem 291: 10318–10331. Wong King Yuen SM, Campiglio M, Tung CC et al. (2017). Structural insights into binding of STAC proteins to voltage-gated calcium channels. Proc Natl Acad Sci U S A 114: E9520–E9528. Wu H, Xiong WC, Mei L (2010). To build a synapse: signaling pathways in neuromuscular junction assembly. Development 137: 1017–1033.

Xu C, Craig R, Tobacman L et al. (1999). Tropomyosin positions in regulated thin filaments revealed by cryoelectron microscopy. Biophys J 77: 985–992. York AL, Zheng JQ (2017). Super-resolution microscopy reveals a nanoscale organization of acetylcholine receptors for transsynaptic alignment at neuromuscular synapses. eNeuro 4. Zou S, Pan B-X (2022). Post-synaptic specialization of the neuromuscular junction: junctional folds formation, function, and disorders. Cell Biosci 12: 1–14.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00018-2 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 2

Upper and lower motor neuron neurophysiology and motor control MAMEDE DE CARVALHO1,2* AND MICHAEL SWASH2,3 1

Department of Neurosciences and Mental Health, Hospital de Santa Maria, Centro Hospitalar Universitário Lisboa-Norte, Lisbon, Portugal

2

Faculdade de Medicina-Instituto de Medicina Molecular-Centro de Estudos Egas Moniz, Universidade de Lisboa, Lisbon, Portugal 3

Department of Neurology, Barts and London School of Medicine, Queen Mary University of London and Royal London Hospital, London, United Kingdom

Abstract This chapter considers the principles that underlie neurophysiological studies of upper motor neuron or lower motor neuron lesions, based on an understanding of the normal structure and function of the motor system. Human motor neurophysiology consists of an evaluation of the active components of the motor system that are relevant to volitional movements. Relatively primitive motor skills include locomotion, much dependent on the spinal cord central pattern generator, reaching, involving proximal and distal muscles activation, and grasping. Humans are well prepared to perform complex movements like writing. The role of motor cortex is critical for the motor activity, very dependent on the continuous sensory feedback, and this is essential for adapting the force and speed control, which contributes to motor learning. Most corticospinal neurons in the brain project to brainstem and spinal cord, many with polysynaptic inhibitory rather than excitatory connections. The monosynaptic connections observed in humans and primates constitute a specialized pathway implicated in fractional finger movements. Spinal cord has a complex physiology, and local reflexes and sensory feedback are essential to control adapted muscular contraction during movement. The cerebellum has a major role in motor coordination, but also consistent roles in sensory activities, speech, and language, in motor and spatial memory, and in psychological activity. The motor unit is the final effector of the motor drive. The complex interplay between the lower motor neuron, its axon, motor end-plates, and muscle fibers allows a relevant plasticity in the movement output.

INTRODUCTION In this chapter, we consider the principles that underlie neurophysiological studies of upper motor neuron (UMN) or lower motor neuron (LMN) lesions, based on an understanding of the normal structure and function of the motor system. These two types of motor dysfunction commonly occur separately but they may occur in

combination, as in amyotrophic lateral sclerosis. Their physiological characteristics are best conceived in terms of ideas derived from clinical neurology, since these neurophysiological concepts derive in large measure from the seminal observations made in the late 19th century in patients with lesions in the motor pathways in the brain and spinal cord, when it was observed that cortical and subcortical motor system lesions caused different clinical

*Correspondence to: Mamede de Carvalho, MD, PhD, Instituto de Fisiologia, Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal. Tel: +351217805219, E-mail: [email protected]

18

M. DE CARVALHO AND M. SWASH

phenomena from more caudal disorders. At that time, neurophysiological assessments were necessarily restricted to study of volitional and reflex motor responses analyzed in electromyography (EMG) studies. Only in recent times has it become possible to image motor pathways in the brain and spinal cord using diffusionweighted magnetic resonance imaging (MRI). This has resulted in new approaches to studies of motor function in humans.

FUNCTIONAL MOTOR SYSTEMS IN BRAIN AND SPINAL CORD Human motor neurophysiology consists of an evaluation of the active components of the motor system that are relevant to volitional movements. Much more is known about motor neurophysiology command and control systems in nonhuman species than in the human (Brownstone et al., 2015). However, studies in apes and macaques have shown that the basic architecture of the motor system is comparable among mammalian species and much of our current understanding stems from these earlier studies (Sherrington, 1906; Kuypers, 1962). Motor functionality commences with a central pattern generator in the frontal and prefrontal regions of the brain. This central pattern generator integrates psychological processing with synthesis of motor programs, often requiring amendments to pre-existing programs. This corresponds to the experience of learning complex tasks, everyday activities such as running and moving among crowds, and new tasks such as cycling or writing (Wise, 1985). Little is known about this essential brain function, but its dissolution is evident in aging, stroke, and dementia, especially in frontotemporal dementia syndromes. This engram feeds back to the predicted sensorimotor coordination necessary for efference copy, the process by which the brain checks that a motor command is effectively and accurately completed during its smooth execution. An essential component of this command signal concerns the timing of sequences of activity in its coding, a function of the cerebellum and its wide intracerebral connections (Holmes, 1904; Guell and Schmahmann, 2020). The corticospinal (CST) tract looms large in every neurologist’s memory but its fast conducting component consists of only 2%–4% of the more than 1,000,000 axons within it. It is these large, fast-conducting fibers that are tested by transcranial magnetic stimulation of the motor cortex. Their function in life is probably directed to rapid coordinated movements of the extremities, and especially the fingers and hands. The remaining smaller diameter fibers consist of cortical and basal ganglia connections to the brainstem and to the spinal cord,

loosely classified as propriospinal pathways. The latter are concerned with postural maintenance to facilitate limb movement in upper limb reaching and manipulating tasks, and for lower limb rhythmic activities, especially locomotion. Propriospinal pathways were first defined as integrative pathways connecting spinal cord segments through interneurons but more recently the terminology has been used in a wider sense to incorporate its interplay with, for example, rubrospinal, tectospinal, olivospinal, and, most importantly, vestibulospinal pathways (Kuypers, 1982; Holstege, 1991). Afferents from limb and trunk muscles pass rostrally through the spinocerebellar tracts to Clarke’s column and Clarke’s nucleus at the C3 and C4 level to project to cerebellum and motor cortex as part of monitoring of the execution of motor commands by efference copy. There are also extensive connections through the vestibular system to cerebellum, basal ganglia and cortex providing automatic input to posture and balance. Sensory input from muscles in the neck is relevant to this system and to the control of rapid eye movements and head and neck postural responses (Pettorossi and Schieppati, 2014). For the neurologist the UMN syndrome is well defined, although it is often incomplete. It is important to recognize that its various components signify different aspects of the disturbed physiology (Swash, 2012; Swash et al., 2020). The description of weakness of varying extent, usually in a distribution especially involving antigravity muscles—a distribution that is reversed if the patient is inverted, illustrating the powerful influence of brainstem vestibular connections on limb and body posture—with spasticity, increased tendon reflexes, and an extensor plantar response form the basis of the clinical syndrome, but clumsy and impoverished finger movements and a similar reduced gait disorder are also important components. These clinical findings led to work that defined the tendon reflexes (Liddell and Sherrington, 1924), muscle tone (Tower, 1939), and muscle spindle physiology (Matthews, 1981) in UMN weakness as distinct from LMN weakness and, later, the higher motor system processes involved in normal motor function. Afferent input modulating these systems has also been studied as an integral factor in motor control (Swash and Fox, 1972). The UMN syndrome is not so simple as is often taught, however, since it varies according to site and type of lesion. The clinical features of a discrete internal capsular lesion are different from those associated with neurodegeneration of the CST pathways, which is often associated with degeneration of other cord pathways, or even with loss of anterior horn cells causing additional LMN features as in amyotrophic lateral sclerosis (Swash, 2012; Swash et al., 2020). The LMN syndrome is more distinct than the UMN syndrome, although also often partial.

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL

EVOLUTION OF THE MOTOR SYSTEM Relatively primitive motor skills include locomotion, much dependent on the spinal cord central pattern generator (MacKay-Lyons, 2002), reaching, involving proximal and distal muscles activation, and grasping (Whishaw and Pellis, 1990). Humans are well-prepared to perform complex movements like writing. Two key developments have determined the nature of the motor control system in humans: the increasingly dominant role of the motor cortex and CST at the expense of other descending pathways such as the rubrospinal tract; and the appearance of the direct corticomotoneuronal connection—originating from large pyramidal neurons in the deepest part of motor cortical layer V (Vb) (Lemon, 1993). Direct cortical motor neuron projections allowed the descending motor systems to bypass spinal segmental mechanisms and directly project to anterior horn cells, and bulbar motor neurons (Lassek, 1940).

MOTOR CORTEX Organization and function In the split-brain macaque Brinkman and Kuypers demonstrated that each hemisphere controls contralateral forearm distal movements and proximal movements bilaterally (Brinkman and Kuypers, 1972). Approximately 75% of the CST fibers decussate in the pyramids, 15% in the spinal cord, and 10% do not cross; the latter terminate in either the medial motor nuclei or intermediate zones of the spinal cord innervating axial and proximal limb muscles (Chouinard and Paus, 2006). The influence of nonprimary motor areas on the spinal cord reflects preparation and modulation of intrinsic spinal circuitry (Prut and Fetz, 1999). Motor cortex is distinct from somatosensory cortical areas due to the absence of granular layer L4, a sensory area that represents the principal thalamic recipient layer (Sherman and Guillery, 2009). Brodmann (1909) found differences between agranular cortex with large pyramidal cells in the anterior bank of the precentral sulcus (area 4) and the agranular cortex in the precentral gyrus and the posterior portion of the superior frontal gyrus on both lateral and medial surfaces of the brain (area 6) (Fulton, 1935). Telencephalic neurons located in layers 2–6 project to striatum bilaterally and to the contralateral cortex (corpus callosum and anterior commissure). Pyramidal neurons located in layer 5 (Betz cells in primary motor cortex) project to spinal cord and striatum, and corticothalamic neurons are in layer 6 (McColgan et al., 2020).

Local circuitries of the primary motor cortex are layer and cell-type specific Motor cortex organization consists of columnar modules (vertical cylinder of 0.5–1 mm radius) functionally

19

related to different aspects of a movement, but different columnar modules interact for the execution of complex movements involving multijoint activity (Keller, 1993). Neurons within each cortical cell column are functionally dependent on other cells within the adjoining few millimeters (Gatter and Powell, 1978). Each column activates spinal neurons involved in specific movements and muscles. In addition, their discharge is tuned to a particular movement direction (the cell’s preferred direction) (Amirikian and Georgopoulos, 2003). A small group of 3–5 neurons in a column can activate a single muscle group (Cheney and Fetz, 1985). Within a column, different neurons may be involved in different aspects of the movement, in particular regarding force and direction (Kalaska et al., 1989; Georgopoulos et al., 1992). In this way, some neurons are recruited during a controlled movement but not in ballistic contraction activating the same muscles (Cheney and Fetz, 1980). The motor columns consist of both input and output zones, with most of the neurons in the superficial layers projecting to other ipsilateral and contralateral cortical areas. Those in the deep layers send efferent connections to subcortical nuclei (Gatter and Powell, 1978). Afferents from L2–3 project to L5 (Weiler et al., 2008). The main afferents originate from other cortical areas and from the thalamic nuclei, which project monosynaptically and topographically to large pyramidal cells (Dormont and Massion, 1970). Thalamic afferents transmit peripheral sensory information and input also from the cerebellum (Brooks and Stoney, 1971). Cortical motor neurons have long horizontal collaterals (>2 mm) in layers 5–6 and 2–3, terminating in clusters (Feldman, 1984) that arborize in the vicinity of their parent neurons, in particular in layer 5 (Ghosh et al., 1988). Vertical axons are mainly those of the afferent and efferent pathways (Gatter and Powell, 1978). Axonal branches of nonpyramidal cells also contribute to the intrinsic synaptic circuitry of the motor cortex. Most of the nonpyramidal cells in the motor cortex are inhibitory GABAergic interneurons (Solberg et al., 1988). Cortical motor zones are recognized, consisting of low threshold neurons that converge on individual motor neuron pools, and others of high threshold neurons diverging in the spinal cord to contact several different motor neuron pools (Keller, 1993). A single muscle may be activated by several spatially segregated zones intermixed with zones related to different muscles (Asanuma and Sakata, 1967), and zones representing the same part of the body are strongly interconnected (Keller, 1993). This process of interaction between cortical neurons directed to different parts of the limb for a movement (convergence and overlap) facilitates control of the intended movement regarding speed, timing and amplitude (Lemon, 1993).

20

M. DE CARVALHO AND M. SWASH

Corticomotor connections are primarily excitatory. Inhibition is mainly exerted through disynaptic pathways via spinal interneurons. Indeed, some fast axons produce EPSPs that are immediately followed by large inhibitory postsynaptic potentials (IPSPs) mediated by spinal interneurons (Jankowska et al., 1976). This is important for inhibition of antagonists of the activated muscle (Keller, 1993). Some corticomotor connections are known to activate Ia inhibitory interneurons, which might be protective from dysfacilitation of the Ia cells leading to inappropriate co-contraction (Lemon, 1993).

Sensory feedback Cortical motoneurons receive input from peripheral sensory fields in the same anatomical region (Brooks and Stoney, 1971). Cutaneous input tends to provide a positive feedback action, i.e., contact and exploration (Asanuma et al., 1968). Cortical layers 2–3 receive thalamic (from the lower layer 4) and somatosensory inputs necessary for encoding motor representation. Motor learning will promote dendritic spine remodeling (Guo et al., 2015; McColgan et al., 2020). The somatosensory cortex directly innervates brainstem and spinal centers (Matyas et al., 2010), where it corresponds to cerebellar links, so refining motor action. But the somatosensory cortex can influence the motor activity more directly. Using optogenetics and pharmacogenetics in combination with in vivo and in vitro electrophysiology in mice, it has been show that the walking program can be generated in CST pathways from an origin in the primary somatosensory cortex through CST projections to cervical and lumbar locomotor network independently of the motor cortex activity (Karadimas et al., 2020).

Force and speed control The strength of muscle contraction depends on the number of activated motor units, their synchronicity and the activity of antagonist muscles. The motor cortex can control the degree of muscle contraction by regulating antagonist muscle activity through spinal interneurons and by coactivation of alpha and gamma motor neurons (Brooks and Stoney, 1971). Speed and force of movement are determined by the same cortical mechanisms, and supported by peripheral sensory afferents and by cerebellar function. Indeed, except in pure isometric contractions, they fully interact. Movement direction is controlled by specific cortical motor columns with connections to particular spinal motor neurons, influenced by the dentato-thalamic system and joint afferents (Brooks and Stoney, 1971).

Motor learning Learning from sensorimotor association (a stimulus– response paradigm) and perceptual training (distinguishing

between cues) are both important for cortical motor plasticity, influencing the excitability and size of a muscle’s cortical motor area. This will depend on modification of synaptic strength, axonal arborization, and the formation of new dendritic spines for long-term memory (Yang et al., 2009; Fu et al., 2012; Papale and Hooks, 2018). Functional magnetic resonance imaging studies have confirmed that after a few weeks of training in a complex motor task, the activated cortical area expanded, remaining expanded for several months, indicating a long-term, experience-dependent reorganization of the adult motor cortex (Kami et al., 1995). Increased somatosensory cortical excitability can precede motor cortex changes during human motor learning and, indeed, motor learning is associated with plasticity in both motor and somatosensory cortex (Ohashi et al., 2019). Explicit sequence learning (from an awareness or instruction set) originates in the prefrontal cortex, but during implicit sequence learning the initial encoding probably occurs in the primary motor cortex and reflects element-to-element associations within a sequence (Ashe et al., 2006). The practice effect derived from across-session learning implicates subcortical structures, in particular the striatum, in order to attain faster stereotyped movements (Van Der Meer et al., 2012). Motor learning is dependent on the basal forebrain cholinergic inputs, in addition to factors such as motivation, attention and fatigue (Conner et al., 2003). Motor cortex plasticity has been explored in protocols for rehabilitation following brain lesion, in particular stroke, for example during constraint-induced movement therapy, increasing the motor areas and the cortical excitability of the affected side (Liepert et al., 1998). Plasticity and learning depend on local persisting stimulation, as demonstrated in the monkey with implantable electronic circuits. Electrical brain stimulation caused synchronization of different populations of cortical neurons, inducing functional reorganization in vivo, supporting a role for a putative neural prosthesis to replace damaged cortical pathways after brain lesion (Jackson et al., 2006).

Motor program Premotor cortex is essential for movement preparation and motor planning (Fuster, 2000). However, preparatory activity has been described in primary motor cortex, frontal eye field, parietal cortex, thalamus, superior colliculus, and cerebellum. This preparatory activity is modular, redundant and controlled by feedback connections (Svoboda and Li, 2018). Patients with premotor area lesion have difficulty planning movements in the contralateral space, but remain able to activate the necessary muscles for the action (Mesulam, 1981). Planning activity predicts timing, speed, and adaptability of the motor action (Churchland et al., 2006).

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL Dorsomedial premotor cortex inhibits temporally inappropriate responses from the motor cortex, utilizing a complex circuit involving the thalamus, dorsal striatum, and the monoaminergic nuclei in the brainstem (Narayanan and Laubach, 2006). In addition, this region is critical to find associations between arbitrary cues and motor responses (Petrides, 1985). Motor responses based on spatial cues are supported by somatosensory and visual information from the parietal cortex (Wise, 1985). Ventral premotor cortex is essential to hand control, as in grasping, and in understanding actions since it contains mirror neurons (Chouinard and Paus, 2006). The supplementary motor area (SMA) has a major role in the planning and generation of simple and complex movements and is involved in motor learning and adaptation (Paz et al., 2005), including learning of new sensorimotor skills (Nakamura et al., 1998). Medial motor areas and SMA have a role in the temporal representation of sequences; in the manner that coding new sequence-learning is more likely to involve pre-SMA and repetition of old sequences is more evident in SMA (Ashe et al., 2006). The SMA is involved in the control of bimanual movements (Swinnen, 2002). In this preparatory phase, its activation occurs several hundred milliseconds before movement initiation. Initiation of movement occurs in response to a disinhibitory signal from basal ganglia or cerebellum, before muscle contraction can commence (Li et al., 2015). Recently two types of pyramidal tract neurons in layer 5 have been recognized, one projecting to thalamus forming a feedback loop involved in movement preparation, and another involved in movement execution (Economo et al., 2018). On the other hand, it is probable that motor cortex can suppress reflex motor responses from basal ganglia and brainstem nuclei (Moore et al., 2013). Functional imaging and transcranial magnetic stimulation studies have shown that ipsilateral dorsal premotor cortex is relevant in the adaptive response to the brain injury, in particular regarding recovering of hand movements after stroke (Johansen-Berg et al., 2002; MoreauDebord et al., 2021).

Mirror neuron system Mirror neurons are visuomotor neurons originally described in area F5 (bilaterally) of the monkey premotor cortex, which are activated when the animal performs a particular movement or when watching another subject performing a similar action. Interest, as in just observing the object, or watching someone making an intransitive (not object directed) gesture are ineffective for activating mirror neurons (Rizzolatti and Craighero, 2004). These neurons are essential for the coupling of understanding and learning by imitation (Rizzolatti et al., 2001). There is indirect evidence that mirror neurons exist in humans,

21

by electroencephalography (EEG) and magnetic electroencephalography (MEG) signal desynchronization, motor amplitude responses on TMS, H-reflex amplitude responses, and imaging findings (e.g., activation of the rostral part of the inferior parietal lobule, lower part of the precentral gyrus, and the posterior part of the inferior frontal gyrus—area 44 or pars opercularis). The cortical area activation in humans depends on the observed activity; area 44 for observing hand-mouth action; precentral gyrus for arm/neck action; and dorsal sector of the precentral gyrus for foot/leg action (Rizzolatti and Craighero, 2004). Observed actions not part of the motor repertoire of the observer do not excite the mirror system, although they are perceived by the visual system (Rizzolatti and Craighero, 2004). The greater the similarity between the observed action and the motor repertoire the stronger the priming (Prinz, 2002). A necessary step for speech evolution may have been the transfer of gestural meaning, intrinsic to gesture itself, to abstract sound meaning (Rizzolatti and Craighero, 2004). The complexity of Broca’s area in humans, in which phonology, semantics, echo-neurons, hand actions, and syntax are intermixed, is a likely consequence of this evolutionary process (Bookheimer, 2002).

SPINAL CORD The spinal cord is an intrinsic part of the brain: it is not a separate structure. Spinal cord neurophysiology is expressed by motor activity and by cutaneous and deep sensation, but complex functional mechanisms underlie these properties. The spinal cord has both segmentally restricted and long tract functions.

Spinal descending motor pathways—Corticospinal tract The CST at the level of the pyramids contains about one million axons in man, but only 2%–4% of the pyramidal fibers originate in the giant Betz neurons localized in layer 5 of the primary motor cortex (Lassek, 1940). Between 60% and 94% of all the fibers in the tract are myelinated, and the conduction velocity of these fibers varies between 50 and 80 m/s (Lassek, 1942). In humans, about 60% of the pyramidal fibers originate in area 4, the remaining 40% originating from other cortical areas, such as premotor cortex, supplementary motor area and the cingulate gyrus (Jane et al., 1967; Kombos and S€uss, 2009). This has been more extensively investigated in macaca rhesus, in which it was found that 31% of the fibers originated from area 4, 29% from area 6, and as many as 40% from the parietal cortex (Russell and DeMyer, 1961). The postcentral pyramidal tract neurons terminate in the dorsal horns, and are important for the feedback sensory control of movement (Lemon, 1993). In their subcortical diencephalic, mesencephalic, and

22

M. DE CARVALHO AND M. SWASH

medullary course, the pyramidal fibers are intermixed with cortical fibers terminating in various subcortical structures, forming 3 groups of fibers: 1.

2.

3.

those terminating on the cell of origin of descending brain stem pathways (including red nucleus and reticular formation); fibers terminating on cerebellar relay nuclei (including olive nucleus, pontine nuclei and reticular formation); fibers terminating on relay nuclei of sensory pathways (dorsal column nuclei receiving afferents from the limbs, and thalamic nucleus receiving lemniscal afferents) (Armand, 1982). Descending rubral fibers are important contributors to distal flexor movement of the upper limbs; the red nuclei are activated by ipsilateral corticorubral, inferior olivary, and monosynaptic contralateral cerebellar from the nucleus interpositus (Brooks and Stoney, 1971).

In primates a pure CST lesion does not impair potential functional recovery, including grasping, due to the major role of the brainstem reticular formation in movement control (Baker, 2011), but there will remain some residual motor limitation in the use of distal flexors, with decreased dexterity and a requirement for greater use of synergistic muscles (Beck and Chambers, 1970). The reticular spinal tracts originate from the pontine and medullary reticular formation and innervate proximal and axial muscles bilaterally, as part of the propriospinal motor system. Connections to hand muscles have also been described (Baker, 2011). Lesions of these tracts tend to cause marked motor deficits (Lawrence and Kuypers, 1968). Vestibular nuclei also project to bilateral proximal and axial muscles to control posture and balance. The basal ganglia are essential to motor output and automatic movements, receiving information from sensorimotor cortex and sending back information to thalamus and motor cortex, and links to the cerebellum are essential in connecting motor output to sensory feedback (as detailed below). Most CST neurons in the brain project to brainstem and spinal cord, many with polysynaptic inhibitory rather than excitatory connections. The monosynaptic connections observed in humans and primates constitute a specialized pathway implicated in fractional finger movements (Ebbesen and Brecht, 2017). A single cortical motor neuron projecting to the spinal cord connects with a large number of lower motor neurons. The resulting proliferation of collaterals with a prominent rostrocaudal orientation is consistent with the axon of the cortical motoneuron contacting a long column of spinal neurons belonging to a target muscle, representing a muscle field (Sherrington, 1889; Lemon, 1993).

Pierrot-Deseilligny (1996, 2002) has stressed that the propriospinal system is a disynaptic system with both excitatory and inhibitory effects on spinal cord motor pathways, after processing in a C3–C4 motor nucleus. This upper cervical motor nucleus was first recognized in studies of feline and monkey motor pathways (Alstermark et al., 2007). Pierrot-Deseilligny (1996, 2002) also noted that propriospinal neurons are potently inhibited by feedback from inhibitory neurons delivered to the corticospinal system, and that the strength of this indirect corticospinal projection may be indirectly estimated by the amount of suppression of the ongoing electromyogram of a suitable muscle, e.g., extensor carpi radialis, elicited by a cutaneous volley (the cutaneous silent period). The propriospinal motor system is important in the control of movements in humans, especially in stance and posture.

Motor unit activity Independent finger movements are well developed in the chimpanzee related to extensive cortical motor neuron connections to hand muscles (Lemon, 1993). This phenomenon is mirrored by the presence of large EPSPs generated in hand muscles by stimulation of the motor cortex (Clough et al., 1968). Cortical dominance over spinal motor neurons may be confirmed by their degree of discharge irregularity during voluntary contractions (Brooks and Stoney, 1971). The motor cells in the ventral gray matter of the cord segments are arranged in associated groups representing different muscles, usually extending across several adjacent spinal levels (Asanuma, 1975). Motor cells innervating distal muscles are located in anterolateral spinal gray matter. Those representing proximal muscles are located medially and motor cells innervating extensor muscles are ventral to those sending axons to flexors. Motor unit recruitment and firing is essential for movement and motor control (Adrian and Bronk, 1929). Motor units are recruited in patterns consistent with a proposed movement (Desmedt and Godaux, 1977), thus following John Hughlings Jackson’s concept that movements are represented in the brain and muscles are recruited in the spinal cord as required for performing these movements. Motor unit recruitment can be recorded by surface or needle electrodes, to evaluate the activity of anterior horn cells at rest and during movement (Duchateau and Enoka, 2011). Voluntary motor unit firing rates can be finely modulated resulting in attempts to classify them as tonic or phasic motor units (Grimby and Hannerz, 1968). Differences between the after-hyperpolarizing potential in spinal motor neurons of different sizes (large vs. small) and types (fast, type II, fast-resistant IIa and fast-fatigable IIb vs. slow, type I)

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL are associated with different firing features (De Carvalho et al., 2014). Histological muscle fiber types, as described by their ATPase and oxidative enzyme characteristics, have supported this classification, which is much used in muscle pathology work (where ATPase enzyme activity in extreme, nonphysiological pH preincubations are used to characterize fiber types), but the physiological separation of these fiber types is not discrete and there is very considerable overlap. The motor unit recruitment pattern is also of interest in assessing segmental motor activity. During stable constant and moderate muscle contraction, motor unit potential firing is about 5–6 Hz, although the interspike intervals show a physiological variability depending on the slightly variable spinal motor neuron membrane threshold (Kernell et al., 1999). This variability (“synaptic noise”) is dependent on inward membrane currents modulated by descending reticulospinal projections through monoaminergic pathways, including serotoninergic and noradrenergic fibers (Henneman et al., 1965; Jordan et al., 2008). Corticospinal tract lesions influence motor unit recruitment causing a more regular firing during mild muscular contraction (De Carvalho et al., 2012, 2017), this could derive from abnormal activation of persistent inward currents producing low-voltage stable plateau potentials (Floeter et al., 2005) leading to lower membrane thresholds in spinal motor neurons (Gorassini et al., 2002). Motor units in a spinal segment are recruited in a rotational pattern, allowing intermittent rest of units in a contracting muscle without disturbing the force output, but this characteristic has scarcely been evaluated in clinical studies (Fallentin et al., 1993). The size principle, that small motor neurons are first recruited, followed by motoneurons of increasing size, has greatly clarified understanding of recruitment patterns in movements, by introducing a simple orderly pattern of activity. In general, small motor neurons fire in slow tonic fashion and large neurons in phasic bursts, but both small and large neurons can fire across the whole range of firing frequency (see De Carvalho et al., 2014, for discussion).

Spinal sensory input Sensory feedback is essential for the controlled coordination of movement. Cutaneous sensation has obvious clinical relevance, especially in fractionated finger movement and finger and thumb opposition. Fastconducting afferents are important in fine sensation, such as touch and movement of sensory stimuli across the skin surface, and joint position sense, all relevant to motor control (Windhorst, 2007). Slower-conducting afferents represent nociceptive input to the cord, determining withdrawal responses and selective sensory experiences,

23

in combination with large fiber input. Discriminative sensory data enters the spinal cord for transmission cephalad to the brain via large diameter afferents through the posterior columns. Small fiber, predominantly nociceptive, input projects cephalad via the lateral spinothalamic pathways. There is evidence that the cord responds to the mix of small and large fiber input by selective cephalad transmission that determines sensory experience and motor responses (Mendell, 2014). Since Renshaw’s discovery of the inhibitory spinal interneuron, inhibitory circuits have been recognized as universally important (Jankowska and Lundberg, 1981). Motor axon collaterals provide excitatory drive to Renshaw cells in a recurrent inhibitory circuit that restricts the pool of activated spinal motor neurons (Moore et al., 2015). These interneurons not only inhibit spinal motor neurons but also Ia inhibitory interneurons mediating disynaptic reciprocal inhibition, and both alpha and gamma-motor neurons (Hultborn et al., 1979). Renshaw cells are influenced by cortical input, which may induce a decreasing effect in stronger contractions (Haase et al., 1975). Sharpening of muscular contraction is also dependent on Ib interneurons. In real life during selective voluntary contraction of isolated groups of muscles, there is a strong Ib inhibition of the synergistic motor neurons but reduction of Ib inhibition to motor neurons involved in the movement (Fournier et al., 1983).

Local functions of the spinal cord In addition to its motor and sensory functions the spinal cord has major autonomic functions, e.g., for temperature regulation through sweating reflexes, for blood pressure control through sympathetic responses, for control of micturition and defecation and, therefore, of urinary and fecal continence and for sexual activity (De Groat et al., 1996). Blood pressure regulation, thermoregulation, bladder continence and sexual responses are disturbed in progressive autonomic failure (Cohen et al., 1987), a degenerative disorder in which there is loss of autonomic neurons in the intermediolateral cell column, found predominantly in the thoracic cord. Bladder and bowel continence depend on a spinal pathway running cephalad in deep white matter adjacent to the medial gray matter that signals bladder fullness to the pontine micturition center (Griffiths, 2002). The latter is influenced by frontal commands determining sphincter relaxation or closure. Similar mechanisms modulate bowel sphincter control. These systems relate visceral sensory input of organ filling to a combined visceral and somatic innervation—contraction or relaxation of the smooth muscle and striated sphincters (Snooks et al., 1984; De Groat et al., 1996). Sexual activity consists of a complex of sensory, somatic and visceral motor responses,

24

M. DE CARVALHO AND M. SWASH

with stimuli reflexes involving the sacral segment of the spinal cord (Lundberg, 1992).

Motor learning The spinal cord is an important structure for motor learning. One essential mechanism is the use of efference copy by which the motor output is continuously forwarded cephalad from the spinal cord to the cerebellum, via the ventral spinocerebellar tract, permitting rapid corrections of errors in motor task execution, without the delay associated with somatosensory feedback (Brownstone et al., 2015). This is achieved by a comparator mechanism that integrates the motor output with the encoded motor program (Wolpert et al., 1998). Ataxia may result when there is a mismatch between predicted motor output and input (Bhanpuri et al., 2014). Spinal motor neurons could also act as a component of this comparator system since they receive Ia afferent input and send predictive input to Renshaw cells, thus modulating their activity (Brownstone et al., 2015). After spinal cord injury the improvement of locomotion with training provides an example of the spinal cord’s motor learning potential (Martinez et al., 2013). Inhibition is a fundamental property of the motor system as emphasized by Sherrington long ago, in the spinal cord and brain, including the output from the cerebellar Purkinje cells which is entirely inhibitory.

THE CEREBELLUM Cerebellar function was systematically studied by Holmes (1917) in soldiers with gunshot wounds involving the cerebellum, sustained in the First World War. These observations were accompanied by smoked drum tracings of rapid alternating movements illustrating the impairment of rhythmic alternating movement of the upper limb. These defects consisted of errors in movement direction, as well as of judgment of distance of the moving finger from the target object. The latter errors have been seen as underestimation and overestimation errors, requiring the interpolation of corrective movements into the task, causing secondary problems with movement timing and with the construction of a smoothly conducted movement. Holmes (1917) surmised that the cerebellum was concerned with the execution of motor tasks, and for many years this was regarded as its principal function. The four categories of cerebellar symptomatology recognized by Holmes after cerebellar damage, basing his description on earlier work, were hypotonia, impaired ability or stand or walk, ataxia and action (intention) tremor. An additional feature, also described by Holmes, was loss of the automatic nature of movement control and irregular delay in initiation of voluntary movement.

The complexity of cerebellar connections, however, suggests that the cerebellum has a more fundamental role in brain organization. There are three paired connecting links to the brainstem; the inferior cerebellar peduncle (restiform body), and the middle and superior cerebellar peduncles. The deep cerebellar nuclei all contain homotypical maps of body structure. These maps include kinetic and kinematic information about the limbs, implying that they are important in the integration and modulation of movement. Coordinated movement requires integration of balance mechanisms for stance with rhythmic gait and directional limb actions. Purkinje neurons and neurons in the deep cerebellar nuclei are active during voluntary movement and this involves both “feed-forward” and efference copy monitoring of motor activity. It is striking that the cerebellar cortical “wiring diagram” is uniform throughout although cerebellar connections to the brain are widespread, suggesting a uniform functional output to different brain regions. The Purkinje output, which is finely modulated in character, represents a synthesis of excitatory and inhibitory input processed by the connected neurons related to each Purkinje cell (D’Angelo, 2018). There are prominent input connections, but only one output system, that from the Purkinje cells to the deep cerebellar nuclei and thence through the superior and middle peduncles to cerebral hemispheres and spinal cord, and to the brainstem vestibular system and olivary nuclei through the inferior cerebellar peduncle. The majority of connecting axons pass through the superior cerebellar peduncle. There are three overall functional considerations in cerebellar physiology. The vestibulocerebellum consists of the medially located flocculonodular lobe, which is developmentally the oldest part of the cerebellum and is concerned with oculovestibular functions, important in posture and balance. The spinocerebellum includes the vermis and the intermediate cerebellar lobes; it is linked to somatosensory and proprioceptive spinal cord inputs and to the vermis, which itself receives spinal and oculovestibular inputs. It projects to the fastigial nucleus and onward to brainstem systems concerned with proximal muscle coordination, important in posture and locomotion. Vermis degeneration, for example, is associated with a specific syndrome of truncal unsteadiness. The cerebrocerebellum, consisting of the lateral cerebellar lobes, is linked through the dentate nucleus with the motor thalamus and motor and premotor areas of the cerebral cortex, and is important in planning, executing, and monitoring movement. In addition, the cerebellum probably plays a role in learning motor programs, mainly through ascending mossy fiber activity (Middleton, 1998; D’Angelo, 2018).

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL MR imaging has revealed cerebellar activity not only in classical motor tasks, with precise anatomical connections, but also consistent roles in sensory activities, speech, and language, in motor and spatial memory and in psychological activity. Schmahmann and others (Schmahmann, 1998; Schmahmann and Caplan, 2006; Mariën et al., 2014; Van Overwalle et al., 2020) have explored these aspects deriving the theory that the cerebellum is concerned principally with timing as a generic function in brain. This timing function is essential in planning, executing, and monitoring ongoing brain activities, including registering timing of sensory input to the brain. This suggests a major role for cerebellum beyond the older concepts of the cerebellum as a purely motor organ and even extending to neuropsychiatric aspects. Schmahmann’s group have postulated a “universal cerebellar transform” to describe this overriding cerebellar activity (Guell et al., 2018). This fundamental reappraisal of cerebellar function accounts nicely for its anatomical prominence in the posterior fossa and its widespread input and output connections. Timing of the execution and monitoring of brain programs, whether motor or involving other aspects of brain function is clearly of fundamental importance. It also underlies perceptual functions such as speech recognition and musical and rhythmic perception.

THE LOWER MOTOR NEURON SYSTEM Each spinal motor neuron consists of a large neural cell giving off a long, thickly myelinated axon which branches near the target muscle endplate in a variable number of sprouts, each innervating a single striated muscle fiber. This cluster of muscle fibers and their innervating LMN spinal cell forms the motor unit, meaning that under physiological conditions all muscle fibers within a motor unit should depolarize nearly synchronously when the spinal motor cell is activated by upstream inputs (Duchateau and Enoka, 2011). Based on contractile speed, motor units are classified as either slow-twitch (S) or fast-twitch (F). The F motor units are further subdivided into fast-twitch fatigue-resistant (FR), and fast-twitch fatigable (FF), although an intermediate type has been reported. Motor unit properties have been assessed by studying their neurophysiological features in needle electromyographic studies (De Carvalho and Swash, 2016), but this recording technique has recently been extended by the use of high-density surface EMG (Del Vecchio et al., 2020). The latest application of muscle magnetic resonance imaging to record individual motor unit responses after threshold motor nerve stimulation may facilitate investigations of motor unit sizes, distribution, and properties in various muscles (Heskamp et al., 2022).

25

Communication between the motor axon and the muscle fibers in the motor unit occurs at the motor end-plates, a specific muscle membrane specialized zone, where acetylcholine is released from the terminal motor boutons to contact acetylcholine (Ach) receptors, that initiate sodium ion entry into the muscle fiber, causing membrane depolarization and calcium release from the sarcoplasmic reticulum. Acetylcholine release from the presynaptic membrane depends on calcium entry inside the terminal bouton. Both pre- and postsynaptic dysfunction are frequently associated with immunological disorders partially blocking this delicate system, but drugs and toxins are other agents causing neuromuscular transmission impairment. Clinical neurophysiological neuromuscular junction studies are generally limited to repetitive nerve stimulation and investigating neuromuscular jitter (Oliveira Santos et al., 2018), the variability in firing rates between activation of adjacent muscle fibers belonging to the same motor unit. The role of calcium is critical at this site, as it links to troponin C that initiates the sliding movement between actin and thick myosin filaments that shorten muscle fiber length, the basis of muscular contraction. All muscle fibers innervated by the same spinal motor neuron have similar histochemical, metabolic, and physical properties, in particular regarding fatigability, and speed and force of twitch contraction. However, these features can be partially changed by physical training (Plotkin et al., 2021) and an orphan muscle fiber, following denervation can change its histochemical features if reinnervated by a different spinal motor neuron during the process of reinnervation (Gordon, 1987). This represents an incredible plasticity in the peripheral motor system. However, each muscle has a particular motor unit type predominance appropriate for its specific functional demands. For example, soleus muscle is particularly rich in slow, fatigue-resistant motor units, unlike gastrocnemius. Ocular muscle fibers and some craniofacial muscles, e.g., masseter, are highly specialized with muscle fibers of mixed histochemical type; ocular muscles are remarkable in that they are hyperneurotized, that is they receive innervation form more than one motor unit. The process whereby the muscle fiber can influence axonal and spinal motor neuron function is itself a matter of exciting investigation, very relevant for some neurodegenerative diseases (Comella et al., 1994).

CONCLUSION Motor control in humans and many other species is a highly refined system, involving timing of inputs and outputs, integration of motor programs, feedback checking and control of the motor output, and integration with a wide range of sensory and higher level, premotor brain

26

M. DE CARVALHO AND M. SWASH

activity. There is redundancy in the system, based on the complexity of its evolutionary development. This has allowed for compensatory processes that adapt to unusual environmental situations as well as providing the capacity for some adaptation to disability in the lesioned nervous system. Redundancy and plasticity are fundamental properties of the motor system, essential to its day to day function and adaptational capacity.

REFERENCES Adrian ED, Bronk DW (1929). The discharge of impulses in motor nerve fibres. J Physiol 67: 9–151. Alstermark B, Isa T, Pettersson LG et al. (2007). The C3?C4 propriospinal system in the cat and monkey: a spinal premotoneuronal centre for voluntary motor control. Acta Physiol 189: 123–140. Amirikian B, Georgopoulos AP (2003). Modular organization of directionally tuned cells in the motor cortex: is there a short-range order? Proc Natl Acad Sci U S A 100: 12474–12479. Armand J (1982). The origin, course and terminations of corticospinal fibers in various mammals. Prog Brain Res 57: 329–360. Asanuma H (1975). Recent developments in the study of the columnar arrangement of neurons within the motor cortex. Physiol Rev 55: 143–156. Asanuma H, Sakata H (1967). Functional Organization of a Cortical Efferent System Examined with focal depth stimulation in cats. J Neurophysiol 30: 35–54. Asanuma H, Stoney SD, Abzug C (1968). Relationship between afferent input and motor outflow in cat motorsensory cortex. J Neurophysiol 31: 670–681. Ashe J, Lungu OV, Basford AT et al. (2006). Cortical control of motor sequences. Curr Opin Neurobiol 16: 213–221. Baker SN (2011). The primate reticulospinal tract, hand function and functional recovery. J Physiol 589: 5603–5612. Beck CH, Chambers WW (1970). Speed, accuracy, and strength of forelimb movement after unilateral pyramidotomy in rhesus monkeys. J Comp Physiol Psychol 70: 1–22. Bhanpuri NH, Okamura AM, Bastian AJ (2014). Predicting and correcting ataxia using a model of cerebellar function. Brain J Neurol 137: 1931–1944. Bookheimer S (2002). Functional MRI of language: new approaches to understanding the cortical organization of semantic processing. Annu Rev Neurosci 25: 151–188. Brinkman J, Kuypers HGJM (1972). Splitbrain monkeys: cerebral control of ipsilateral and contralateral arm, hand, and finger movements. Science 176: 536–539. Brodmann K (1909). Vergleichende Lokalisationslehre der Grosshirnrinde in ihren Prinzipien dargestellt auf Grund des Zellenbaues, Barth, Leipzig. Brooks VB, Stoney SD (1971). Motor mechanisms: the role of the pyramidal system in motor control. Annu Rev Physiol 33: 337–388. Brownstone RM, Bui TV, Stifani N (2015). Spinal circuits for motor learning. Curr Opin Neurobiol 33: 166–173.

Cheney PD, Fetz EE (1980). Functional classes of primate corticomotoneuronal cells and their relation to active force. J Neurophysiol 44: 773–791. Cheney PD, Fetz EE (1985). Comparable patterns of muscle facilitation evoked by individual corticomotoneuronal (CM) cells and by single intracortical microstimuli in primates: evidence for functional groups of CM cells. J Neurophysiol 53: 786–804. Chouinard PA, Paus T (2006). The primary motor and premotor areas of the human cerebral cortex. Neuroscientist 12: 143–152. Churchland MM, Afshar A, Shenoy KV (2006). A central source of movement variability. Neuron 52: 1085–1096. Clough JF, Kernell D, Phillips CG (1968). The distribution of monosynaptic excitation from the pyramidal tract and from primary spindle afferents to motoneurones of the baboon’s hand and forearm. J Physiol 198: 145–166. Cohen J, Low P, Fealey R et al. (1987). Somatic and autonomic function in progressive autonomic failure and multiple system atrophy. Ann Neurol 22: 692–699. Comella JX, Sanz-Rodriguez C, Aldea M et al. (1994). Skeletal muscle-derived trophic factors prevent motoneurons from entering an active cell death program in vitro. J Neurosci Off J Soc Neurosci 14: 2674–2686. Conner JM, Culberson A, Packowski C et al. (2003). Lesions of the basal forebrain cholinergic system impair task acquisition and abolish cortical plasticity associated with motor skill learning. Neuron 38: 819–829. D’Angelo E (2018). Physiology of the cerebellum. In: The Cerebellum: From Embryology to Diagnostic Investigations, Elsevier, pp. 85–108. De Carvalho M, Swash M (2016). Lower motor neuron dysfunction in ALS. Clin Neurophysiol 127: 2670–2681. De Carvalho M, Turkman A, Swash M (2012). Motor unit firing in amyotrophic lateral sclerosis and other upper and lower motor neurone disorders. Clin Neurophysiol 123: 2312–2318. De Carvalho M, Eisen A, Krieger C et al. (2014). Motoneuron firing in amyotrophic lateral sclerosis (ALS). Front Hum Neurosci 8: 719. De Carvalho M, Poliakov A, Tavares C et al. (2017). Interplay of upper and lower motor neuron degeneration in amyotrophic lateral sclerosis. Clin Neurophysiol 128: 2200–2204. De Groat CW, Vizzard MA, Araki I et al. (1996). Spinal interneurons and preganglionic neurons in sacral autonomic reflex pathways. In: G Holstege, R Bandler, CB Saper (Eds.), The Emotional Motor System, Elsevier, Amsterdam, pp. 97–111. Del Vecchio A, Holobar A, Falla D et al. (2020). Tutorial: analysis of motor unit discharge characteristics from high-density surface EMG signals. J Electromyogr Kinesiol 53: 102426. Desmedt JE, Godaux E (1977). Ballistic contractions in man: characteristic recruitment pattern of single motor units of the tibialis anterior muscle. J Physiol 264: 673–693. Dormont JF, Massion J (1970). Duality of cortical control on ventrolateral thalamic activity. Exp Brain Res 10: 205–218.

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL Duchateau J, Enoka RM (2011). Human motor unit recordings: origins and insight into the integrated motor system. Brain Res 1409: 42–61. Ebbesen CL, Brecht M (2017). Motor cortex—to act or not to act? Nat Rev Neurosci 18: 694–705. Economo MN, Viswanathan S, Tasic B et al. (2018). Distinct descending motor cortex pathways and their roles in movement. Nature 563: 79–84. Fallentin N, Jørgensen K, Simonsen EB (1993). Motor unit recruitment during prolonged isometric contractions. Eur J Appl Physiol Occup Physiol 67: 335–341. Feldman ML (1984). Morphology of the neocortical pyramidal neuron. In: A Peters, EG Jones (Eds.), Cerebral Cortex. Cellular Components of the Cerebral Cortex. vol 1. Plenum Press, New York, pp. 123–200. Floeter MK, Zhai P, Saigal R et al. (2005). Motor neuron firing dysfunction in spastic patients with primary lateral sclerosis. J Neurophysiol 94: 919–927. Fournier E, Karz R, Pierrot-Deseilligny E (1983). Descending control of reflex pathways in the production of voluntary isolated movements in man. Brain Res 288: 375–377. Fu M, Yu X, Lu J et al. (2012). Repetitive motor learning induces coordinated formation of clustered dendritic spines in vivo. Nature 483: 92–95. Fulton JF (1935). A note on the definition of the “motor” and “premotor” areas. Brain 58: 311–316. Fuster JNM (2000). Prefrontal neurons in networks of executive memory. Brain Res Bull 52: 331–336. Gatter KC, Powell TPS (1978). The intrinsic connections of the cortex of area 4 of the monkey. Brain 101: 513–541. Georgopoulos AP, Ashe J, Smyrnis N et al. (1992). The motor cortex and the coding of force. Science 256: 1692–1695. Ghosh S, Fyffe REW, Porter R (1988). Morphology of neurons in area 4? Of the cat’s cortex studied with intracellular injection of HRP. J Comp Neurol 269: 290–312. Gorassini M, Yang JF, Siu M et al. (2002). Intrinsic activation of human motoneurons: possible contribution to motor unit excitation. J Neurophysiol 87: 1850–1858. Gordon T (1987). Muscle plasticity during sprouting and reinnervation. Am Zool 27: 1055–1066. Griffiths DJ (2002). The pontine micturition centres. Scand J Urol Nephrol 36: 21–26. Grimby L, Hannerz J (1968). Recruitment order of motor units on voluntary contraction: changes induced by proprioceptive afferent activity. J Neurol Neurosurg Psychiatry 31: 565–573. Guell X, Schmahmann J (2020). Cerebellar functional anatomy: a didactic summary based on human fMRI evidence. Cerebellum 19: 1–5. Guell X, Gabrieli JDE, Schmahmann JD (2018). Embodied cognition and the cerebellum: perspectives from the dysmetria of thought and the universal cerebellar transform theories. Cortex 100: 140–148. Guo L, Xiong H, Kim J-I et al. (2015). Dynamic rewiring of neural circuits in the motor cortex in mouse models of Parkinson’s disease. Nat Neurosci 18: 1299–1309. Haase J, Cleveland S, Ross HG (1975). Problems of postsynaptic autogenous and recurrent inhibition in the mammalian

27

spinal cord. In: Reviews of Physiology, Biochemistry and Pharmacology, Springer Berlin Heidelberg. Henneman E, Somjen G, Carpenter DO (1965). Functional significance of cell size in spinal motoneurons. J Neurophysiol 28: 560–580. Heskamp L, Miller AR, Birkbeck MG et al. (2022). In vivo 3D imaging of human motor units in upper and lower limb muscles. Clin Neurophysiol 141: 91–100. Holmes G (1904). On certain tremors in organic cerebral lesions. Brain 27: 327–375. Holmes G (1917). The symptoms of acute cerebellar injuries due to gunshot injuries. Brain 40: 461–535. Holstege G (1991). Descending motor pathways and the spinal motor system: limbic and non-limbic components. In: Prog. Brain Res, Elsevier, pp. 307–421. Hultborn H, Lindstr€ om S, Wigstr€ om H (1979). On the function of recurrent inhibition in the spinal cord. Exp Brain Res 37. Jackson A, Mavoori J, Fetz EE (2006). Long-term motor cortex plasticity induced by an electronic neural implant. Nature 444: 56–60. Jane JA, Yashon D, Demyer W et al. (1967). The contribution of the precentral gyrus to the pyramidal tract of man. J Neurosurg 26: 244–248. Jankowska E, Lundberg A (1981). Interneurones in the spinal cord. Trends Neurosci 4: 230–233. Jankowska E, Padel Y, Tanaka R (1976). Disynaptic inhibition of spinal motoneurones from the motor cortex in the monkey. J Physiol 258: 467–487. Johansen-Berg H, Rushworth MFS, Bogdanovic MD et al. (2002). The role of ipsilateral premotor cortex in hand movement after stroke. Proc Natl Acad Sci U S A 99: 14518–14523. Jordan LM, Liu J, Hedlund PB et al. (2008). Descending command systems for the initiation of locomotion in mammals. Brain Res Rev 57: 183–191. Kalaska JF, Cohen DA, Hyde ML et al. (1989). A comparison of movement direction-related versus load directionrelated activity in primate motor cortex, using a twodimensional reaching task. J Neurosci Off J Soc Neurosci 9: 2080–2102. Kami A, Meyer G, Jezzard P et al. (1995). Functional MRI evidence for adult motor cortex plasticity during motor skill learning. Nature 377: 155–158. Karadimas SK, Satkunendrarajah K, Laliberte AM et al. (2020). Sensory cortical control of movement. Nat Neurosci 23: 75–84. Keller A (1993). Intrinsic synaptic organization of the motor cortex. Cereb Cortex 3: 430–441. Kernell D, Bakels R, Copray JCVM (1999). Discharge properties of motoneurones: how are they matched to the properties and use of their muscle units? J Physiol Paris 93: 87–96. Kombos T, S€ uss O (2009). Neurophysiological basis of direct cortical stimulation and applied neuroanatomy of the motor cortex: a review. Neurosurg Focus 27: E3. Kuypers HGJM (1962). Corticospinal connections: postnatal development in the Rhesus monkey. Science 138: 678–680.

28

M. DE CARVALHO AND M. SWASH

Kuypers HGJM (1982). A new look at the organization of the motor system. In: Prog. Brain Res., Elsevier. Lassek AM (1940). The human pyramidal tract: II. A numerical investigation of the Betz cells of the motor area. Arch Neurol Psychiatr 44: 718–724. Lassek AM (1942). The pyramidal tract: the effect of pre-and postcentral cortical lesions on the fiber components of the pyramids in monkey. J Nerv Ment Dis 95: 721–729. Lawrence DG, Kuypers HGJM (1968). The functional organization of the motor system in the monkey. I. the effects of bilateral pyramidal lesions. Brain 91: 1–14. Lemon RN (1993). The G. L. Brown prize lecture. Cortical control of the primate hand. Exp Physiol 78: 263–301. Li N, Chen T-W, Guo ZV et al. (2015). A motor cortex circuit for motor planning and movement. Nature 519: 51–56. Liddell E, Sherrington C (1924). Reflexes in response to stretch (myotatic reflexes). Proc Roy Soc Lond B 96: 212–242. Containing Papers of a Biological Character. Liepert J, Miltner WHR, Bauder H et al. (1998). Motor cortex plasticity during constraint-induced movement therapy in stroke patients. Neurosci Lett 250: 5–8. Lundberg PO (1992). Sexual dysfunction in patients with neurological disorders. Annu Rev Sex Res 3: 121–150. Mackay-Lyons M (2002). Central pattern generation of locomotion: a review of the evidence. Phys Ther 82: 69–83. Marie¨n P, Ackermann H, Adamaszek M et al. (2014). Consensus paper: language and the cerebellum: an ongoing enigma. Cerebellum (London, England) 13: 386–410. Martinez M, Delivet-Mongrain H, Rossignol S (2013). Treadmill training promotes spinal changes leading to locomotor recovery after partial spinal cord injury in cats. J Neurophysiol 109: 2909–2922. Matthews PB (1981). Evolving views on the internal operation and functional role of the muscle spindle. J Physiol 320: 1–30. Matyas F, Sreenivasan V, Marbach F et al. (2010). Motor control by sensory cortex. Science 330: 1240–1243. Mccolgan P, Joubert J, Tabrizi SJ et al. (2020). The human motor cortex microcircuit: insights for neurodegenerative disease. Nat Rev Neurosci 21: 401–415. Mendell LM (2014). Constructing and deconstructing the gate theory of pain. Pain 155: 210–216. Mesulam MM (1981). A cortical network for directed attention and unilateral neglect. Ann Neurol 10: 309–325. Middleton F (1998). The cerebellum: an overview. Trends Neurosci 21: 367–369. Moore JD, Desch^enes M, Furuta T et al. (2013). Hierarchy of orofacial rhythms revealed through whisking and breathing. Nature 497: 205–210. Moore NJ, Bhumbra GS, Foster JD et al. (2015). Synaptic connectivity between Renshaw cells and motoneurons in the recurrent inhibitory circuit of the spinal cord. J Neurosci Off J Soc Neurosci 35: 13673–13686.  Quessy S et al. (2021). Rapid and Moreau-Debord I, Serrano E, bihemispheric reorganization of neuronal activity in premotor cortex after brain injury. J Neurosci Off J Soc Neurosci 41: 9112–9128.

Nakamura K, Sakai K, Hikosaka O (1998). Neuronal activity in medial frontal cortex during learning of sequential procedures. J Neurophysiol 80: 2671–2687. Narayanan NS, Laubach M (2006). Top-down control of motor cortex ensembles by dorsomedial prefrontal cortex. Neuron 52: 921–931. Ohashi H, Gribble PL, Ostry DJ (2019). Somatosensory cortical excitability changes precede those in motor cortex during human motor learning. J Neurophysiol 122: 1397–1405. Oliveira Santos M, Swash M, De Carvalho M (2018). Concentric or monopolar electrode for jitter determination in orbicularis oculi. Clin Neurophysiol 129: 2552–2556. Papale AE, Hooks BM (2018). Circuit changes in motor cortex during motor skill learning. Neuroscience 368: 283–297. Paz R, Natan C, Boraud T et al. (2005). Emerging patterns of neuronal responses in supplementary and primary motor areas during sensorimotor adaptation. J Neurosci Off J Soc Neurosci 25: 10941–10951. Petrides M (1985). Deficits in non-spatial conditional associative learning after periarcuate lesions in the monkey. Behav Brain Res 16: 95–101. Pettorossi VE, Schieppati M (2014). Neck proprioception shapes body orientation and perception of motion. Front Hum Neurosci 8: 895. Pierrot-Deseilligny E (1996). Transmission of the cortical command for human voluntary movement through cervical propriospinal premotoneurons. Prog Neurobiol 48: 489–517. Pierrot-Deseilligny E (2002). Propriospinal transmission of part of the corticospinal excitation in humans. Muscle Nerve 26: 155–172. Plotkin DL, Roberts MD, Haun CT et al. (2021). Muscle fiber type transitions with exercise training: shifting perspectives. Sports (Basel, Switzerland) 9: 127. Prinz W (2002). Experimental approaches to imitation. In: AN Meltzoff, W Prinz (Eds.), The Imitative Mind: Development, Evolution, and Brain Bases. Cambridge University Press, pp. 143–162. Prut Y, Fetz EE (1999). Primate spinal interneurons show premovement instructed delay activity. Nature 401: 590–594. Rizzolatti G, Craighero L (2004). The mirror-neuron system. Annu Rev Neurosci 27: 169–192. Rizzolatti G, Fogassi L, Gallese V (2001). Neurophysiological mechanisms underlying the understanding and imitation of action. Nat Rev Neurosci 2: 661–670. Russell JR, Demyer W (1961). The quantitative cortical origin of pyramidal axons of Macaca rhesus: with some remarks on the slow rate of axolysis. Neurology 11: 96–108. Schmahmann J (1998). The cerebellar cognitive affective syndrome. Brain 121: 561–579. Schmahmann JD, Caplan D (2006). Cognition, emotion and the cerebellum. Brain 129: 290–292. Sherman SM, Guillery RW (2009). Exploring the Thalamus and Its Role in Cortical Function, The MIT Press. Sherrington CS (1889). On nerve-tracts degenerating secondarily to lesions of the cortex cerebri. J Physiol 10: 429–432.

MOTOR NEURON NEUROPHYSIOLOGY AND MOTOR CONTROL Sherrington CS (1906). The Integrative Action of the Nervous System, Yale University Press. Snooks SJ, Barnes PR, Swash M (1984). Damage to the innervation of the voluntary anal and periurethral sphincter musculature in incontinence: an electrophysiological study. J Neurol Neurosurg Psychiatry 47: 1269–1273. Solberg Y, White EL, Keller A (1988). Types and distribution of glutamic acid decarboxylase (GAD)-immunoreactive neurons in mouse motor cortex. Brain Res 459: 168–172. Svoboda K, Li N (2018). Neural mechanisms of movement planning: motor cortex and beyond. Curr Opin Neurobiol 49: 33–41. Swash M (2012). Why are upper motor neuron signs difficult to elicit in amyotrophic lateral sclerosis? J Neurol Neurosurg Psychiatry 83: 659–662. Swash M, Fox KP (1972). Muscle spindle innervation in man. J Anat 112: 61–80. Swash M, Burke D, Turner MR et al. (2020). Occasional essay: upper motor neuron syndrome in amyotrophic lateral sclerosis. J Neurol Neurosurg Psychiatry 91: 227–234. Swinnen SP (2002). Intermanual coordination: from behavioural principles to neural-network interactions. Nat Rev Neurosci 3: 348–359.

29

Tower SS (1939). The reaction of muscle to denervation. Physiol Rev 19: 1–48. Van Der Meer M, Kurth-Nelson Z, Redish AD (2012). Information processing in decision-making systems. Neuroscientist 18: 342–359. Van Overwalle F, Manto M, Cattaneo Z et al. (2020). Consensus paper: cerebellum and social cognition. Cerebellum (London, England) 19: 833–868. Weiler N, Wood L, Yu J et al. (2008). Top-down laminar organization of the excitatory network in motor cortex. Nat Neurosci 11: 360–366. Whishaw IQ, Pellis SM (1990). The structure of skilled forelimb reaching in the rat: a proximally driven movement with a single distal rotatory component. Behav Brain Res 41: 49–59. Windhorst U (2007). Muscle proprioceptive feedback and spinal networks. Brain Res Bull 73: 155–202. Wise SP (1985). The primate premotor cortex: past, present, and preparatory. Annu Rev Neurosci 8: 1–19. Wolpert DM, Miall RC, Kawato M (1998). Internal models in the cerebellum. Trends Cogn Sci 2: 338–347. Yang G, Pan F, Gan W-B (2009). Stably maintained dendritic spines are associated with lifelong memories. Nature 462: 920–924.

This page intentionally left blank

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00022-4 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 3

Vestibular motor control KATHLEEN E. CULLEN* Departments of Biomedical Engineering, of Otolaryngology-Head and Neck Surgery, and of Neuroscience; Kavli Neuroscience Discovery Institute, Johns Hopkins University, Baltimore, MD, United States

Abstract The vestibular system is an essential sensory system that generates motor reflexes that are crucial for our daily activities, including stabilizing the visual axis of gaze and maintaining head and body posture. In addition, the vestibular system provides us with our sense of movement and orientation relative to space and serves a vital role in ensuring accurate voluntary behaviors. Neurophysiological studies have provided fundamental insights into the functional circuitry of vestibular motor pathways. A unique feature of the vestibular system compared to other sensory systems is that the same central neurons that receive direct input from the afferents of the vestibular component of the 8th nerve can also directly project to motor centers that control vital vestibular motor reflexes. In turn, these reflexes ensure stabilize gaze and the maintenance of posture during everyday activities. For instance, a direct three-neuron pathway mediates the vestibulo-ocular reflex (VOR) pathway to provide stable gaze. Furthermore, recent studies have advanced our understanding of the computations performed by the cerebellum and cortex required for motor learning, compensation, and voluntary movement and navigation. Together, these findings have provided new insights into how the brain ensures accurate self-movement during our everyday activities and have also advanced our knowledge of the neurobiological mechanisms underlying disorders of vestibular processing.

OVERVIEW The vestibular system senses the motion of the head relative to space using two types of sensory organs—the semicircular canals and the otoliths (Fig. 3.1A). There are three semicircular canals (i.e., horizontal, anterior, and posterior). The three canals are arranged such that they are mutually orthogonal to each other, thereby together providing the brain with sensory feedback about the head’s rotation in three dimensions. There are also two otolith organs (the saccule and utricle), which sense linear acceleration and thus detect both the direction and magnitude of gravity, as well as transient linear accelerations due to movement (i.e., gravity and translational movements). As a result of the structural organization

of the sensory neuroepithelium in each of the two otoliths, the utricle and saccule are sensitive to horizontal vs vertical linear head movement, respectively. Thus, together the otoliths provide the brain with sensory feedback about the head’s linear motion and orientation relative to gravity in three dimensions. During daily activities, the VIII cranial nerve (vestibulocochlear nerve) transmits information about our current rotational and linear head motion from the semicircular canals and otoliths to the vestibular nuclei within the brainstem. In turn, neurons within the vestibular nuclei comprise vestibular motor pathways underlying stabile gaze and posture, as well as accurate control of voluntary movements and navigation (Fig. 3.1B). Individual vestibular-nerve afferents innervate the hair cells of one of the five sensory organs (i.e., one of the three

*Correspondence to: Kathleen E. Cullen, Johns Hopkins University, 720 Rutland Ave. Traylor 504, Baltimore, MD 21205-2109, United States. Tel: +1-410-271-2713, Fax: +1-410-614-8796, E-mail: [email protected]

32

K.E. CULLEN Semicircular Canals Anterior Posterior Horizontal

Utricule Otoliths Saccule

II

II

I

I Irregular afferents

Ampullae

Middle Ear

Cochlea B

D

C

Regular afferents

Voluntary motor control & Naviga

VN

Gaze stabiliza (VOR)

VIII nerve Postural stability (VSR)

A

B

Fig. 3.1. The structure mammalian vestibular system comprises five sensory organs: the three semicircular canals and the two otoliths (utricle and the saccule). (A) Within each of these sensory organs is a neuroepithelia in which the receptor cells (termed hair cells) are located. (B) There are two classes of hair cells: cylindrical type II hair cells and the phylogenetically more recent vase shaped type I hair cells. Afferent fibers in the VIII cranial nerve innervate hair cells and transmit head movement information to both the vestibular nuclei and regions of the vestibular cerebellum. Afferent fibers originating in both the canals and otoliths are classified based on the regularity of their resting discharge. More irregularly firing afferents preferentially transmit information from type I hair cells to central pathways whereas more regular afferents preferentially transmit information from type II hair cells. Adapted from Cullen KE (2019). Vestibular processing during natural self-motion: implications for perception and action. Nat Rev Neurosci 20: 346–363.

semicircular canals, or saccule or utricle). Investigations in alert animals have well characterized how rotational and linear head motion is first encoded in the firing rates of individual vestibular afferents, then processed by target neurons in the vestibular nuclei (for a review, see Goldberg et al., 2012). A distinctive feature of central vestibular processing is that neurons at the first stage of central processing in the vestibular nuclei are already multimodal. Vestibular nuclei neurons integrate vestibular afferent input with extra-vestibular signals (i.e., proprioceptive, oculomotor, and predictive self-motion signals) arising from the brainstem, as well as the cerebellum and cortex. This integration is essential for the behaviorally dependent shaping processing that underlies the control of vestibular motor reflex behavior vs voluntary movements. Moreover, this integration ultimately provides the neural substrate that underlies the brain’s ability to compensate for the loss of peripheral vestibular function. Overall, our comprehensive knowledge of the functional circuitry of normal central vestibular pathways has provided fundamental insights into the deficits observed clinically in patients.

VESTIBULAR MOTOR CONTROL: NEURONAL CODING IN ALERT ANIMALS The predominant approach for understanding the signals coded by vestibular motor reflex pathways is based on linear systems analysis (reviewed in Cullen, 2012). To use this analytical approach, experimenters typically apply sinusoidal head motion and record the responses of individual afferents or neurons for different frequencies of stimulation (reviewed in Goldberg, 2000).

Changes in the depth of modulation (gain) and timing of the response relative to the head motion stimuli (phase) are then computed and plotted as a function of each tested frequency. Neuronal response gains and phases have been estimated for neurons at different stages along vestibular motor pathways and then compared to behavioral response gains and phases of the reflex motor output to understand the coding of vestibular information at each stage of processing in motor control. Fig. 3.2A shows example response gains computed from actual firing rate responses for populations of vestibular afferents. Both semicircular canal and otolith afferents demonstrate increasing neural response gain as a function of frequency and thus display what is termed high pass tuning (Fig. 3.2B and C). Notably in both systems, afferents with more irregular resting discharges—termed irregular afferents—are more sensitive to motion than their more regularly discharging counterparts—termed irregular afferents (Haque et al., 2004; Hullar et al., 2005, Ramachandran and Lisberger, 2006; Sadeghi et al., 2007a,b; Jamali et al., 2013, 2016, 2019). In the case of the otolith system, this difference reaches nearly an order of magnitude greater response gain relative to regular afferents (see arrow at 20 Hz, Jamali et al., 2013, 2019). Additionally, both semicircular canal and otolith afferents demonstrate increasing phase leads as a function of frequency. Neurons at the first central stage of processing in the vestibular nuclei typically integrate input from both regular & irregular afferents (Goldberg et al., 1987; Highstein et al., 1987; Sato and Sasaki, 1993). Furthermore, it is noteworthy that most vestibular nuclei neurons also

VESTIBULAR MOTOR CONTROL Head velocity (sinusoidal)

33

Canal Afferents

Afferents Firing rate (sp/s)

15 Hz

0.5 Hz

Head Velocity 50 deg/sec

B Otolith Afferents

Es mate 800 700

Firing rate Gain (sp/s/G)

600

Irregular

0 0.5 8 16 Frequency (Hz)

300 100

100 msec

A

400

50

200

50 sp/sec 1 sec

500

100

Gain (sp/s/G)

Regular

0

C

0.5 2

4

6 8 10 12 14 16 Frequency (Hz)

Fig. 3.2. The encoding of head motion by semicircular canal and otolith afferents. (A) The responses of an example regular and irregular semicircular canal afferent during sinusoidal head rotations at 0.5 and 15 Hz. (B and C) Schematics of the difference in the response gains of regular vs irregular semicircular canal (B) vs (C) otolith afferents recorded over a wide range of frequencies of head rotation in alert rhesus monkeys. Note, irregular afferents neurons have higher gains which increase more dramatically at higher frequencies. Side bands show 1 SEM. Panel (A) adapted from Cullen KE (2019). Vestibular processing during natural self-motion: implications for perception and action. Nat Rev Neurosci 20: 346–363; panel (B) adapted from Sadeghi SG, Minor LB, Cullen KE (2007b). Response of vestibular-nerve afferents to active and passive rotations under normal conditions and after unilateral labyrinthectomy. J Neurophysiol 97: 1503–1514; and panel (C) adapted from Jamali M, Carriot J, Chacron MJ, Cullen KE (2013). Strong correlations between sensitivity and variability give rise to constant discrimination thresholds across the otolith afferent population. J Neurosci 33: 11302–11313.

combined information from the semicircular canals and the otoliths (Carriot et al., 2013; Carriot et al., 2015a; Mackrous et al., 2019). This integration has important implications for the coding of vestibular information during everyday activities, since natural head movements are multidimensional and thus will simultaneously activate semicircular canal and otolith afferents (Carriot et al., 2014, 2017a,b). Indeed, as discussed in the section below entitled “The Vestibular Cerebellum: Internal Models of Externally-Applied and Active Self-Motion,” such integration is essential because it underlies the brain’s ability to distinguish the gravitoinertial forces that result from translational self-motion from those that result from changes in head orientation (also commonly referred to as tilts). Vestibular nuclei neurons can be characterized into two main subclasses, based on their functional role in vestibular motor control: (i) vestibulo-ocular reflex (VOR) neurons and (ii) “vestibular-only” neurons (reviewed in Cullen and Roy, 2004; Cullen, 2019). First, VOR neurons comprise position-vestibular-pause (PVP) neurons and

floccular target neurons (FTNs), which control and modulate the VOR, respectively, to ensure accurate gaze stability in everyday life. The name of PVP neurons reflects that these neurons encode eye position, are sensitive to vestibular stimulation, and cease firing or “pause” for saccades. Notably, PVP neurons comprise the main direct three-neuron-arc that mediates the VOR namely: afferents project to central neurons in the vestibular nuclei (i.e., PVP neurons), that in turn project to the extraocular motoneurons (Scudder and Fuchs, 1992; Cullen and McCrea, 1993; Cullen et al., 1993). In contrast, FTN neurons receive direct input from the cerebellar flocculus and play an essential role in VOR motor learning and calibration (reviewed in Cullen and Mitchell, 2016 & see the section below entitled “The Vestibulo-Ocular Reflex”). For example, calibration of the VOR circuitry is vital in the first years of life to compensate for significant changes in head circumference (
30% in the first year), as well as in later life to compensate for common conditions such as the need to wear corrective lenses for visual conditions such as myopia (i.e., nearsightedness).

34

K.E. CULLEN

Second, a distinct class of neurons in the vestibular nuclei termed vestibular-only or “VO” neurons do not project to the eye motor neurons to control the VOR, but instead control posture and balance. These neurons are termed vestibular-only since they respond to passive vestibular stimulation but do not respond to either saccadic or smooth pursuit eye movements (Scudder and Fuchs, 1992; Cullen and McCrea, 1993; Cullen et al., 1993; Massot et al., 2011, 2012). More specifically, the axons of VO neurons target the spinal cord and thus contribute to the pathways that produce vestibular spinal reflexes (see review Goldberg and Cullen, 2011). VO neurons also have reciprocal connections with the posterior vermis of the cerebellum—an area referred to as the nodulus/uvula (Reisine and Raphan, 1992). As reviewed in the section below entitled “The Vestibular Cerebellum: Internal Models of Externally-Applied and Active SelfMotion,” this recurrent feedback serves an important role in altering the dynamics and spatial orientation of vestibular signals. Finally, VO neurons also send direct projections to the Ventral posterolateral nucleus of the thalamus (Marlinski and McCrea, 2009) which in turn sends vestibular input to vestibular-sensitive regions of cortex (Lang et al., 1979; Gr€ usser et al., 1990). Thus, VO neurons and the thalamocortical pathways that underlie self-motion perception and spatial orientation, in addition to controlling vestibular spinal reflexes. Similar to vestibular afferents (Fig. 3.2B and C), PVP and VO neurons display high pass gain tuning, as well as phase leads that increase as a function of stimulation frequency. As discussed in the section below entitled “The Vestibulo-Ocular Reflex,” this increasing phase lead effectively offsets for the finite
5 ms delay of VOR pathway (i.e., synaptic, neural, and muscle activation times from afferents to extraocular muscles), thereby ensuring that the response dynamics of the VOR pathway remain compensatory across the frequency range of natural head motion (Huterer and Cullen, 2002). In general, the PVP and FTN neurons mediating the VOR receive stronger input from afferents with more regular resting discharges (i.e., regular afferents), whereas the VO neurons mediating vestibulospinal pathways instead receive stronger input from afferents with more irregular resting discharges (i.e., irregular afferents) (Boyle et al., 1992). In this context, the response dynamics of irregular vs regular afferents are best matched to the mechanical demands of the vestibulospinal reflex vs the VOR, respectively (Fernández and Goldberg, 1971; Bilotto et al., 1982; Minor and Goldberg, 1991). Specially, larger gains and phase leads of the irregular afferents are required to drive robust postural responses since vestibulospinal pathways must account for the higher inertia of the body relative to the eye (reviewed in Cullen, 2016).

NONLINEAR & SPIKE TIMING CODES: IMPLICATIONS FOR MOTOR CONTROL AND THE RESTORATION OF FUNCTION While the use of the linear system approach described above in the section entitled “Vestibular Motor Control: Neuronal Coding in Alert Animals” has been beneficial to furthering our understanding of the neuronal basis of vestibular motor control, its use is only valid over a relatively small head movement range (reviewed in Cullen, 2019). Indeed, we have only recently developed a clear understanding of the dynamic structure of the vestibular stimuli that we experience during everyday activities (Carriot et al., 2014, 2017a; Zobeiri et al., 2021a,b) as well those experienced by two popular animal models—monkeys and mice (Carriot et al., 2017b). Overall, both rotational and linear head movements can reach high intensities—up to 15,000 deg./s and 8G, respectively—and comprise significant power up to 20 Hz (Carriot et al., 2014). Fig. 3.3A contrasts the natural range of rotational vestibular stimuli experienced during everyday life, with those that are typically applied to test vestibular function. Comparison of these two distributions emphasizes the fact that natural stimuli routinely reach intensities 10 larger than those typically applied in single unit recording experiments. Because afferents and vestibular nuclei neurons display nonlinearities at such higher amplitudes, linear systems analysis is only valid for a limited range of this natural distribution. Thus, as detailed below, in everyday life the processing that takes place at early stages of vestibular pathways is strongly nonlinear for natural stimuli. More recently, the responses of afferents and central vestibular neurons have been recorded during natural vestibular stimulation. Importantly, neural responses demonstrate excitatory saturation and inhibitory rectification (i.e., cut-off ) in their preferred vs nonpreferred direction of stimulation, respectively (Sadeghi et al., 2007b; Massot et al., 2012; Schneider et al., 2015). These nonlinear responses cannot be explained by traditional linear models of vestibular processing. Instead, responses to more high amplitude naturalistic stimuli can be explained by linear–nonlinear (LN) cascade models (Chichilnisky, 2001; Massot et al., 2012; Schneider et al., 2015). There are two stages of the LN cascade model. In the first stage, the same linear systems approach described above is used to characterize the neuron to stimulation within its linear range (i.e., computing the “linear tuning” as shown in Fig. 3.2). In the second stage the output of this linear filter is then passed through a nonlinear function, which accounts for the neuron’s saturation and rectification. Notably, this second nonlinear stage is generally well represented by a static

VESTIBULAR MOTOR CONTROL A

35 B

Dual coding strategies (Natural range of moon)

1000

Linear system analysis (Typical Tesng Range)

1

Transmit more informaon via precise spike ming Higher gains and phase leads Beer opmized for natural vesbular smuli

MI (Spike ming)

Highly non-linear *

1

Frequeny (Hz)

0 Phase

0

Irregular Afferents

Gain

Irregular Regular

0.5

Angular Velocity (deg/s)

Beer detecon thresholds

MI (Firing rate)

*

Regular Afferents Transmit more informaon via firing rate

0.5

Frequeny (Hz)

0

-1000 101 103 Probability Density

C

Eye velocity

Prosthesis mapping Linear Transfer funcon

Smulaon rate

VOR Pathway Eye velocity

50 deg/s

Head velocity

Non linearity

Compensatory Latency of ~5ms

100 ms

Fig. 3.3. The dynamic range of natural vestibular stimulation and implications for the development of a vestibular prosthetic to restore sensorimotor function. (A) In everyday life, natural vestibular stimuli high intensities reach high intensities (green curve), while in contrast vestibular testing is done over a restricted region corresponding the afferent linear coding range for which minimal inhibitory cut-off or excitatory saturation is demonstrated (gray curve). Example motion is shown for horizontal head rotations. Testing over the linear range has established the high pass tuning of regular and irregular afferents (left panel). Testing over the full natural range has established that regular afferents transmit more information through firing rate, whereas irregular canal afferents demonstrate significant nonlinearities and transmit more information via a precise spike timing (right panel). (B) Actual firing rate of an example vestibular afferent plotted as a function of the linear prediction for the same afferent estimated over its linear range. The nonlinear relationship is well described by sigmoid (black line) and thus deviates from the unity linear line (dashed line). Inset, Goodness of nonlinear (blue) vs linear (black) fits quantified by R2. (C) Schematic of prosthetic stimulation of the VOR pathway. The head motion to pulse frequency mapping cascades a linear transfer function (mimicking the high pass tuning of vestibular afferents) with a sigmoidal nonlinearity (limiting the firing rate to be above zero and below a maximum rate of 400 Hz). Panel (A) adapted from Cullen KE (2019). Vestibular processing during natural self-motion: implications for perception and action. Nat Rev Neurosci 20: 346–363; panel (B) adapted from Schneider AD, Jamali M, Carriot J, Chacron MJ, Cullen KE (2015). The increased sensitivity of irregular peripheral canal and otolith vestibular afferents optimizes their encoding of natural stimuli. J Neurosci 35: 5522–36 (fig 4D); and panel (C) adapted from Wiboonsaksakul KP, Della Santina CC, Cullen KE (2022). A prosthesis utilizing natural vestibular encoding strategies improves sensorimotor performance in monkeys a prosthesis utilizing natural vestibular encoding strategies improves sensorimotor performance in monkeys. PLoS Biol 20: e3001798.

nonlinearity (sigmoid) which can be estimated by plotting the output of the linear filter vs the actual firing rate (Fig. 3.3B). The use of LN models of vestibular afferent responses has had important implications regarding the recent development of a vestibular sensory prosthetic for patients with profound bilateral vestibular loss (reviewed in van de Berg et al., 2020). Ongoing clinical trials aimed at restoring vestibular semicircular canal function use a prosthesis (reviewed in Lewis, 2016) in which a 3D gyroscope first senses 3D head motion. This motion information is projected into the three canal planes and then a processor converts the three aligned signals into a pulse train to stimulate three electrode arrays each implanted in

one of the three orthogonal canals (Fig. 3.3C). Notably, this prosthesis, which has been implanted in patients with nearly complete vestibular loss due to the lack of functioning receptor cells in the semicircular canals, improves motor performance during balance and gait (Chow et al., 2021). The prosthesis works by bypassing the nonfunctioning vestibular hair cells and directly stimulating the vestibular nerve, which then in turn carries head motion signal to the vestibular nuclei. Such prosthetic stimulation evokes significant motor responses including, compensatory eye movements with
5 ms latencies consistent with those of well characterized VOR pathways (see section entitled “The VestibuloOcular Reflex” below), as well compensatory head

36

K.E. CULLEN

movements with
35 to 40 ms latencies consistent with those of well characterized vestibulospinal pathways (see section entitled “Vestibulospinal Reflex Pathways” below). LN cascade models are now typically used in the mapping head motion to prosthetic stimulation since they can account for the nonlinearities naturally seen in vestibular afferents. Importantly, implementing the natural tuning dynamics of vestibular afferents in the linear stage of such LN models produces more temporally accurate VOR eye movement response (Wiboonsaksakul et al., 2022). To date, a current limitation of the prosthetic is that the pulsatile stimulation that is delivered to the implanted electrodes rapidly generates a reduction in the efficacy of the afferent-central neuron synapse—on the order of seconds (Mitchell et al., 2016, 2017). In turn, this resulting reduction in vestibular pathway efficacy decreases the gain of the prosthesis-evoked reflex motor behaviors (VOR: Mitchell et al., 2016, Vestibulospinal reflex: Mitchell et al., 2017). The observed reduction in pathway efficacy likely arises because the applied pulsatile stimulation induces synchrony across the vestibular afferent population. Notably, such sustained synchrony is not present across the afferent population during natural head motion. Future work is needed to improve the ability of the prosthesis to restore function—including stimulation approaches which do not reduces the efficacy of central pathway transmission. Finally, afferents and vestibular nuclei neurons can demonstrate another key nonlinearity in addition to the saturation and cut-off reviewed above. In particular, irregular semicircular canal and otolith afferents as well as VO vestibular nuclei neurons encode head motion information using “precise spike timing” in addition to their spike rates (Jamali et al., 2016, 2019). A given irregular afferent or VO vestibular nuclei neuron can reliably discriminate between different natural head motion stimulus waveforms via differential patterns of precise (
6 ms) spike timing. Furthermore, this “precise spike timing” code also manifests in the form of nonlinear phase locking for sinusoidal head motion stimulation (Jamali et al., 2019). Overall, at the level of the VO neurons this coding strategy likely gives rise to more compensatory behavior in vestibulospinal pathways in response to transient dynamic stimulation. Specifically, this coding property likely induces synchrony at the VO population level that, in turn, generates a pulse command to the neck muscles to compensate for the inertia of the head–neck system (reviewed in Cullen, 2016). In contrast, rate coding by more regularly firing VOR neurons is essential for the generation of compensatory eye movements to ensure stable gaze. Overall, the variability of vestibular pathways is linked to different coding strategies for motor control (Mackrous et al., 2020).

THE VESTIBULO-OCULAR REFLEX As reviewed above, PVP and FTN neurons in the vestibular nuclei control and modulate VOR eye movements. These eye movements play an essential role in ensuring gaze stability during our everyday activities. For example, consider, what would the world look like if you had to walk home without a vestibular system. The head motion generated by each foot strike, would result in the movement of the world relative to the retina and thus produce image blur. However, this is not the case. Instead, the compensatory eye movements generated by the VOR rapidly move the eye to rotate it an equal amount in the opposite direction so that the visual axis of gaze (gaze ¼ eye-in-head + head-in-space) is stable relative to the visual world. Indeed, the VOR is essential to our survival and is arguably the fastest sensorimotor behavior that we generate. The rotational VOR is mediated by a simple threeneuron arc pathway (Fig. 3.3A) comprising semicircular canal afferents that directly project to neurons in the vestibular nuclei, which then project to extraocular motoneurons (Lorente de Nó, 1933). Because this vestibulomotor pathway is so direct, the compensatory eye movements generated by the VOR have an extraordinarily short latency of 5 to 6 ms relative to head motion (Fig. 3.4A; Huterer and Cullen, 2002). Indeed, this short latency can be accounted by summing the known fixed synaptic delays and activation time of the extraocular muscles. Further, when we experience sustained head rotation in one direction, the VOR generates compensatory eye motion that rotates the eye eccentrically in the orbit toward its limit of excursion (approx. 50 deg. in humans). In such conditions, the brain reflexively commands the eye to move back to a new more centered starting position during the sustained head motion. The resulting pattern of alternating slow compensatory and rapid resetting eye movements (termed slow phases and quick phases, respectively) is referred to as vestibular nystagmus (Fig. 3.4B). The vestibular slow phase is driven by the VOR pathway, while the quick phase of vestibular nystagmus is driven by the of the same neural machinery involved in the generation saccadic eye movements. Fig. 3.4C illustrates the pathways mediating the rotational VOR in response to a leftward horizontal head rotation. Specifically, leftward head motion excites afferents innervating the left horizontal semicircular canal, which in turn excite PVP neurons in the left vestibular nuclei. The axons of the PVP neurons then cross the midline to directly excite the motoneurons that activate the lateral rectus muscle of the right eye. The lateral rectus contracts to rotate the right eye to the right thereby generating a compensatory rightward eye movement

VESTIBULAR MOTOR CONTROL

37

Left

Right

Response to transient head motion Left lateral rectus muscle

Head velocity

Eye velocity

Right lateral rectus muscle

Oculomotor nucleus

100 deg/s

0 deg/s

A

Left medial rectus muscle

Left medial longitudinal fasciculus

5 ms

Abducens nucleus

Quick Phases

Slow Phases

Lateral vestibular nucleus

Right Movement direction

Degrees Time

C

Movement direction Right horizontal canal

Left horizontal canal

Left

B

Medial vestibular nucleus

Excitatory pathway Inhibitory pathway

Fig. 3.4. (A) Because the direct rotational VOR is mediated by a robust three neuron pathway connecting afferents to extraocular motoneuron, it generates eye movement with a minimal latency (5 ms) in response to head movement. (B) In response to prolonged rotations, the rotational VOR generates a saw tooth pattern of slow compensatory/rapid resetting eye movements (slow phases and quick phases, respectively) referred to as vestibular nystagmus. (C) Schematic of the pathways mediating horizontal VOR in response to a leftward head rotation. Excitatory and inhibitory pathways are shown as filled and dashed blue lines, respectively. The thick filled line represents the direct VOR pathway. Leftward head motion drives an increase in right lateral and left medial rectus activation and decrease in the left lateral and right medial rectus activation. Panel (A) adapted from Huterer M, Cullen KE (2002). Vestibuloocular reflex dynamics during high-frequency and high-acceleration rotations of the head on body in rhesus monkey. J Neurophysiol 88: 13–28 and panel (C) adapted from Adlers physiology of the Eye, 11th edition).

(Fig. 3.4C; thick blue lines). In addition to this direct pathway, there are more indirect pathways innervating the right lateral rectus. Moreover, a subset of neurons in the abducens nucleus, called internuclear neurons, send projections to the medial rectus motoneurons of the contralateral eye, such that it also moves in a coordinated fashion to the right (Fig. 3.4C; standard blue lines). Correspondingly, in a push pull fashion, this same leftward horizontal head rotation correspondingly inhibits the afferents that innervate the right horizontal semicircular canal, which in turn inhibit PVP neurons in the right vestibular nuclei (Fig. 3.4A; dashed blue lines). The axons of these PVP neurons send an inhibitory drive across the midline to inactivate the motoneurons that activate the lateral rectus of the left eye. Thus, the left lateral rectus contracts to rotate the right eye rightward thereby generating a compensatory eye movement. Studies in humans and nonhuman primates have established that rotational VOR functions to effectively stabilize gaze over the wide range of head rotation frequencies and amplitudes generated during everyday activities, including frequencies up to 25 Hz (Grossman et al., 1988; Armand and Minor, 2001; Huterer and Cullen, 2002) and angular velocities up to 500deg./s (Carriot et al., 2014, 2017a,b). Remarkably, the high pass tuning of vestibular afferents and VOR neurons in the vestibular nuclei (i.e., VOR

neurons) effectively compensates for the fixed 5 ms latency of the VOR pathway to provide compensatory phase up to
25Hz. For example, a 5 ms latency would produce a
40° lag at 20 Hz if there was no compensatory phase lead in the system. However, the responses of VOR neurons lead head velocity by about 20 Hz at this frequency to generate near perfect phase compensation (Huterer and Cullen, 2002). It is noteworthy that while the rotational VOR effectively stabilizes gaze for head motion over a wide range of frequencies, it is not effective for head motion below 0.1 Hz due to the biomechanical properties of the semicircular canals. At this lower end of the frequency range, optokinetic eye movements make the main contribution to gaze stability. Optokinetic eye movements have a longer latency then VOR eye movements (>50 vs 5 ms) because they are driven by visual input (i.e., the motion of the world relative to the eye) rather than by vestibular input. In everyday life, the VOR and optokinetic eye movement pathways work together synergistically to ensure stable gaze over an extended range of head movements (see Goldberg et al., 2012). The limitations of the VOR at lower frequencies can be appreciated in the laboratory by continually rotating a subject at a constant velocity (equivalent to 0 Hz). In response to such stimulation, the evoked VOR eye velocity will initially be of

38

K.E. CULLEN

the similar magnitude and opposite direction to the applied head velocity but then decay with a time constant of
20 s. This is because (i) the responses encoded by canal afferents decay exponentially with a time constant of
5 s due to the biomechanics of the semicircular canals (reviewed in Goldberg et al., 2012), and (ii) this afferent time constant is then lengthened centrally so that the responses of PVP neurons decay exponentially with a time constant of
20 s. As described below in the section entitled “The Vestibular Cerebellum: Internal Models of Externally-Applied and Active Self-Motion,” a cerebellar-dependent mechanism termed velocity storage (Laurens and Angelaki, 2011; Karmali, 2019) underlies the central lengthening of the afferent time constant. The rotational VOR is nearly perfectly compensatory for horizontal (yaw) and vertical (pitch) rotations. The compensatory eye movements evoked for torsional (roll) head rotations are also well aligned with the phase of head motion but have lower gain (eye speed/head speed) (reviewed in Goldberg et al., 2012). The vestibular system also generates eye movements to compensate for translational head movements: for example, side-to-side (interaural), fore and aft (anterior–posterior), or up and down (vertical) motion. The geometry of maintaining stabilize gaze on the fovea of the two eyes during translation requires that the gain of the translational VOR varies as a function of both the eye’s current position in the orbit and the viewing distance (reviewed in Goldberg et al., 2012). For example, to compensate for the motion parallax typically experienced during translation, the translational VOR gain scales as a function of viewing distance. However, in contrast to the rotational VOR, the translational VOR is not fully effective in stabilizing gaze in response to translational motion experienced during everyday activities. The translational VOR has a longer response time latency compared to the aVOR (>10 rather than 5 ms) because it not predominately mediated by a direct three-neuron arc but rather by more complex polysynaptic pathways. While the VOR pathways are essential for stabilizing our gaze during voluntary behaviors such as walking, they would be counterproductive when the goal is to redirect the visual axis of gaze in space. Indeed, in interacting with our environment, we frequently make rapid coordinated eye and head movements—termed gaze shifts—to voluntarily redirect our gaze to a new target. If the VOR were fully compensatory during gaze shifts, it would produce an eye movement in the opposite direction to that of the intended shift in gaze (reviewed in Cullen and Roy, 2004). To prevent the generation of such a counterproductive eye movement command, the brain sends a copy of the voluntary eye movement command to the

vestibular nuclei to suppress the efficacy of the direct VOR pathway during gaze shifts (Fig. 3.5; Roy and Cullen, 1998, 2002). This suppression of the direct VOR pathway is maximal at the start of the gaze shift, with the VOR recovering to reach normal values by gaze shift end (Cullen et al., 2004). Thus, the VOR is actually not a hard-wired reflex, but rather it is suppressed by a voluntary eye movement command from the premotor saccadic pathway when the current gaze strategy is to redirect rather than stabilize gaze. Likewise, a similar mechanism underlies the suppression of the VOR when we make coordinated eye and head movements to pursue a moving target—termed gaze pursuit (reviewed in Cullen and Roy, 2004).

Adaptation and compensation of the VOR Finally, it is noteworthy that the VOR has the capacity for impressive adaptation (also called VOR motor learning) in response to environmental requirements across our life span. For example, the VOR pathways adapt to account for the magnification of corrective lenses worn for common visual conditions, such as convex lenses worn for myopia. A particularly dramatic illustration such adaptation was provided by the pioneering study in which participants wore dove prisms that reversed the world such that left was right and vice versa (Gonshor and Melvill Jones, 1976). The gain of the VOR substantively declined within minutes of wearing these prisms, and then ultimately reversed sign when the prisms were worn for extended periods (3–4 weeks). The VOR also shows adaptive compensation in response to peripheral vestibular loss due to aging, disease, and/or surgical intervention. For instance, initially following unilateral lesion of the vestibular nerve, VOR responses are asymmetric and subpar for rotations to the affected side. Fortunately, over the weeks that follow, VOR responses largely recover for lower frequencies and velocity head movements (Curthoys and Halmagyi, 1995). Compensation for more dynamic and challenging stimuli, however, is not complete (Sadeghi et al., 2006). Over the past several decades, numerous studies have focused on understanding the neural mechanisms that underlie VOR adaptation (reviewed in Cullen and Mitchell, 2016). As further considered below in the section entitled “The Vestibular Cerebellum: Internal Models of Externally-Applied and Active Self-Motion,” a region of the vestibular cerebellum termed the floccular complex plays an essential role in mediating the formation of the required long-term synaptic changes required for VOR adaptation and compensation.

VESTIBULAR MOTOR CONTROL

39

Gaze stabilization vs. redirection Premotor Saccadic drive

Afferents

FR prediction based on head motion sensitivity Observed firing rate

PVP

VOR suppressed during gaze redirection

Head Motion VO

Vestibular Nuclei

Fig. 3.5. The pathway mediating the VOR demonstrates context-dependent gating at the level of the vestibular nuclei. The activity of position-vestibular-pause (PVP) neurons that mediate the middle link in direct VOR pathways are strongly inhibited by the premotor saccadic pathway. As a result, these central VOR neurons are suppressed when the behavioral goal is to voluntarily redirect rather than stabilize gaze (for example, via a coordinated eye-head gaze shift).

VESTIBULOSPINAL REFLEX PATHWAYS The vestibular system is often described as our balance system. Indeed, the vestibular system serves an essential role in controlling balance and maintaining our posture. Specifically, during our everyday activities the head rotates and translates relative to space. This head motion is the encoded by vestibular canal and otolith afferents, respectively, which in turn activate neurons in the vestibular nuclei. The vestibular nuclei send direct and indirect projections (via the reticular formation, as well as other brainstem nuclei) to different levels of the spinal cord via the vestibulospinal reflex pathways. In turn, the vestibulospinal reflexes (VSR) function to stabilize the head and body relative to space in response. The vestibulocollic reflex (VCR) is a specific subclass of VSR that plays an important role in activating the neck musculature in response to head movement in order to maintain the head in an upright position (reviewed in Goldberg and Cullen, 2011). The VCR is mediated by the medial vestibulospinal tract (MVST), which provides the primary input to the cervical levels of the cord spinal cord. As reviewed above in the section entitled "Vestibular Motor Control: Neuronal Coding in Alert Animals," VO neurons in the vestibular nuclei send projections to the cervical spinal cord to mediate the vestibulocollic reflex (VCR). There is a direct three-neuron pathway linking the canal afferents to dorsal neck motoneurons (Fig. 3.6; Wilson and Maeda, 1974); however, the major pathway mediating the rotational VCR is more complex comprising indirect pathways that include spinal cord interneurons as well as reticulospinal pathways. The pathways mediating the translational VCR circuitry

are less well characterized, but again indirect pathways play a more important role compared to direct pathways. During our everyday activities, the rotational and translational VCRs serves a vital function in stabilizing the head during the pitch and vertical linear displacements generated during locomotion (e.g., Pozzo et al., 1990, 1991; Takahashi, 1990). Recent work has shown that neck muscles are increasingly sensitive to vestibular stimulation spanning a frequency range up to 70 to 80 Hz (Forbes et al., 2020). Such high-frequency signal transmission in VCR pathways is thought to underlie the generation of robust neck muscle reflex responses that stabilize the head in response to unexpected perturbations. Vestibulospinal reflexes (VSR) also send important motor commands to the lower levels of the spinal cord that activate the trunk and limb musculature. Similar to the VCR, these VSR pathways include spinal cord interneurons as well as reticulospinal pathways and thus are less direct than the VOR. In contrast to the VCR, which is mediated by the MVST, the lateral vestibulospinal tract (LVST) provides the main drive to the lower levels of the spinal cord to activate muscles in the limbs and torso to maintain balance. Recent studies in humans have demonstrated that an individual’s postural stability depends on their vestibular sensory precision (Karmali et al., 2021). To noninvasively drive the VSR, numerous studies have utilized galvanic vestibular stimulation (GVS), which activates vestibular afferents through current applied via surface electrodes on the mastoid process (Kwan et al., 2019; Forbes et al., 2020). In turn, this activation drives the VSR—evoking a rapid electromyographic response in both upper and lower limb muscles during standing

40

K.E. CULLEN

Postural stability vs. active movement Vestibular Nuclei Voluntary motor control & Navigation

PVP Afferents

VO

INC

Head Motion Reticular formation Neck MN

A

Posture Stabilization (VSR)

VSR central pathway Vestibular Nuclei Afferents

VO

VSR central pathway

Head Motion

B

Reafference Cancellation signal

Posture Stabilization (VSR) FR prediction based on: Total head motion Passive head motion only Observed firing rate

Fig. 3.6. (A) The pathways mediating the Vestibulospinal/ascending pathways. (B) The vestibular nuclei neurons mediating Vestibulospinal/ascending pathways demonstrate context dependent gating. The activity of vestibular-only (VO) neurons are strongly inhibited when the behavioral goal is to generate voluntary head motion. During voluntary head motion, VO neurons receive a strong inhibitory cancelation signal when there is a match between (i) the expected sensory consequence of neck motor command and (ii) the actual neck proprioceptive feedback. This cancelation signal serves to effectively cancel the actively generated component of head motion (vestibular reafference). The mechanism for cancelation is represented by a gate that closes when such a match occurs. Abbreviations: INC, interstitial nucleus of Cajal; MN, motoneuron.

balance, which depend on the alignment of the head relative to the body (Nashner and Wolfson, 1974; Lund and Broberg, 1983; Mian and Day, 2014). This latter feature of the VSR pathways is essential because the same vestibular afferent vestibular input requires a different postural response depending on where the head is facing. Furthermore, GVS evoked responses are modulated in a manner consistent with a given muscle’s current ability to stabilize the body along the direction of a vestibular-evoked disturbance (Forbes et al., 2016). The application of GVS has also become increasing popular for the noninvasive assessment/treatment of individuals with impaired balance. However, to date, the mechanisms underlying reported improvements are not currently understood (reviewed in Stefani et al., 2020). In everyday life our vestibular system is activated both by unexpected head motion stimuli (vestibular exafference) as well as stimuli that are the result of our own voluntary motor actions (vestibular reafference). As reviewed above, the VCR and VSR mediated by the LVST play a critical role in the maintenance of

posture and balance by coordinating activation of the neck, trunk, and limb musculature to stabilize the head in space in response to vestibular exafference. However, while these vestibular-reflexes are essential for providing robust postural response to unexpected stimuli, they would be counter-productive when the goal is to make voluntary head movements (reviewed in Cullen, 2004). Consider the situation where an individual actively moves their head to look between the far sides of a room. In this situation, an intact VCR would function to generate a compensatory activation of the neck musculature to stabilize the head relative to space and would thus prevent the desired movement. Indeed, during our everyday activities, much of our head motion is voluntary and as such vestibular stimuli are reafference rather than exafference. Recent work in rhesus monkeys has focused on understanding how vestibular pathways encode vestibular reafference during voluntary behaviors (reviewed in Cullen, 2012, 2019). During active head orienting, vestibular afferents encode head motion in a

VESTIBULAR MOTOR CONTROL context-independent manner; head motion is encoded in the same way regardless of whether head motion is actively generated or passively applied (Cullen and Minor, 2002; Sadeghi et al., 2007a,b; Jamali et al., 2009; Mackrous et al., 2022). In contrast, neurons at the next stage in the VCR—namely VO vestibular nuclei neurons—demonstrate markedly reduced responses to self-generated as compared to externally applied head motion (Roy and Cullen, 2001, 2004; Cullen and Roy, 2004; Brooks and Cullen, 2014). Overall, the head motion sensitivity of VO neuron population is markedly attenuated (
70%) in response to vestibular reafference vs exafference. Furthermore, when monkeys make active head movements while experiencing simultaneous whole body passive motion, VO neurons remarkably encode only the passive exafferent component of the total head-in-space motion (Roy and Cullen, 2001, 2004; Brooks and Cullen, 2014). A systematic series of studies has further established that the robust vestibular afferent reafferent input to VO neurons is only suppressed in active conditions where there is a match between the actual activation of neck proprioceptors and that expected based on the brain’s prediction of the sensory consequences of the voluntary movement—as would be the case during normal voluntary head movements (reviewed in Cullen and Zobeiri, 2021). The suppression of VO neurons responses during voluntary orienting movements is a common strategy across movements and species. VO responses are markedly attenuated for active rotational and translational orienting head movements, as well as head movements that reposition the head relative to gravity (Mackrous et al., 2019). Further, comparable vestibular reafference suppression is observed in the VCR pathway across species, including other mammalian species (mice and cats) as well as new both Old World monkeys and New World monkeys (reviewed in Brooks and Cullen, 2019). To date, however, how the descending motor commands produced by the VCR and VSR mediated by the LVST maintain posture during active locomotion vs rest remains an open question. As reviewed above, recent work in rhesus monkeys has established that vestibular afferent coding is unaltered during voluntary movements. Notably, this is the case for locomotion (Mackrous et al., 2022) as well as for voluntary orienting head movements (Cullen and Minor, 2002; Sadeghi et al., 2007b; Jamali et al., 2009). Thus, the vestibular afferents transmit a contextindependent representation of head motion to the central vestibulospinal reflex pathways during locomotion. Prior recording experiments in cat have shown that reflex pathways provide a phasic excitatory drive to the antigravity extensor muscles during locomotion (Matsuyama and Drew, 2000a,b) to maintain posture. Yet whether and

41

how the efficacy of the VCR and VSR pathway are specifically modulated during locomotion remains unknown. Activation of the human vestibular system using GVS suggests that the vestibular drive to leg muscles is increasingly suppressed with increased cadence and walking velocity (Dakin et al., 2013). Furthermore, this resultant EMG is modulated in a manner that depends on the stabilization demands required to maintain a stable gait pattern (i.e., walking on a narrow vs wide base) rather than gait pattern itself (Magnani et al., 2021). Importantly, however, GVS stimuli are externally applied unnatural stimuli and not self-generated stimuli. Single-unit experiments during voluntary walking will be required to directly determine whether and how VCR and VSR pathways transmit self-generated vestibular signals during active walking.

Adaptation and compensation of postural reflexes Finally, it is noteworthy that the pathways that mediate vestibular postural reflexes, like those that mediate the VOR, are capable of impressive plasticity to adapt to new environmental conditions and compensate for vestibular injuries. Peripheral vestibular loss due to aging, disease, and/or surgical intervention cause impairments in postural as well as gaze stability. Fortunately, these symptoms typically resolve on the order of a few weeks. For example, behaviorally relevant patterns of vestibular nerve activation generate a rapid and substantial decrease in the responses of VO vestibular nuclei neurons within minutes of initiation (Mitchell et al., 2017). Strikingly, complementary plasticity on the same rapid time scale within the inhibitory bilateral vestibular nuclei network effectively offsets this decrease to help ensure a relatively robust behavioral output (Mitchell et al., 2017). As discussed in the next section, single unit recording experiments in macaque monkeys have provides insight into the mechanism that underlie the adaptation and compensation of the vestibular motor reflexes that stabilize gaze and posture.

THE VESTIBULAR CEREBELLUM: INTERNAL MODELS OF EXTERNALLYAPPLIED AND ACTIVE SELF-MOTION The cerebellum plays a key role in the coordination and calibration of movements. Three main areas of the cerebellar cortex respond to vestibular stimulation: floccular lobe, anterior vermis and nodulus/uvula (Fig. 3.7, reviewed in Goldberg et al., 2012). These areas serve multiple functions in ensuring accurate motor control. The first of these three, the floccular lobe (flocculus and ventral paraflocculus) is essential for calibrating the VOR in response to changes across the life span (e.g., aging, optical lens corrections, and peripheral

42

K.E. CULLEN Vermis I

Anterior lobe Posterior lobe

II

Anterior vermis

III IV V VI VII

Oculomotor vermis

Nodulus/Uvula

VIII IX X

Fastigial

Floccular lobe

Deep cerebellar nuclei

Vestibular Nuclei

Fig. 3.7. Organization of the vestibular cerebellum. Schematic of the five main cerebellar regions that receive vestibular input from the vestibular nuclei and/or vestibular afferents (i) lobules I–V of the anterior lobe (green), (ii) the nodulus and ventral uvula (blue), (iii) the flocculus and ventral paraflocculus (floccular lobe; pink), and (iv) the fastigial deep cerebellar nucleus and vestibular nuclei (yellow). In addition, the oculomotor vermis of posterior lobe (gray) receives some vestibular nuclei input. Adapted from Brodal A (1969). Neurological anatomy in relation to clinical medicine, Oxford University Press, New York.

vestibular loss). The second, the anterior vermis (lobules I–V) is essential for regulating muscle tone and postural control. And finally, the third area, the nodulus/uvula of the posterior cerebellar vermis, plays a vital role in the brain’s computation of its representation of our orientation relative to gravity. The anatomical organization of the cerebellar cortex is highly structured, comprising repeating motifs of five primary cell types: Purkinje cells, granule cells, basket cells, stellate cells, and Golgi cells. Importantly, of these five cell types only Purkinje cells send projections outside of the cerebellar cortex. Purkinje cells exclusively send inhibitory projections to target structures in the vestibular nuclei and deep cerebellar nuclei. Furthermore, there are two distinct input pathways to the cerebellum: mossy fibers and climbing fibers. Mossy fibers arise from many regions of the brain stem and spinal cord, while climbing fiber inputs arise from the inferior olive. The cerebellum’s mossy fiber input is integrated via the interneurons of the cerebellum and then serves to drive the simple spike firing rate responses of Purkinje cells which commonly reach rates up to 50 to 150 spikes/s. In contrast, each Purkinje cell only receives input from a single climbing fiber input. Climbing fibers make especially powerful excitatory synaptic connections with Purkinje cells and drive their complex spike firing rate responses, which can last 2 to 5 ms. In contrast to simple spike firing rate responses, Purkinje cell complex spike rates are relatively low (1–10 spikes/s).

The different regions of the vestibular cerebellar areas (Fig. 3.7) integrate sensory information from structures that encode both vestibular and extravestibular sensory signals as well as motor commands. These regions of the vestibular cerebellum, in turn, send projections to premotor and motor areas of the cerebral cortex as well as to premotor brain stem nuclei and the spinal cord to ensure and maintain accurate motor performance. As a result, damage to areas of vestibular cerebellum disrupt gaze stability, posture and muscle tone, coordination of locomotion, as well as motor learning in the associated pathways. In this context, the vestibular system has two unique features as compared to other sensory systems. First, primary vestibular afferents send direct projections to the cerebellum. Specifically, as described in more detail below, primary afferent fibers terminate as mossy fibers in the lobules I–II of the anterior vermis, nodulus, and ventral uvula as well as send collaterals to the deep cerebellar nuclei. Second, the Purkinje cells of the cerebellar cortex project directly to the vestibular nuclei as well as to the deep cerebellar nuclei. Thus, vestibular motor control is unique in its close connectivity to cerebellar processing.

Flocculus and ventral paraflocculus The flocculus and adjoining ventral paraflocculus are generally referred to as the floccular complex (Fig. 3.7, pink region). This cerebellar region plays an essential

VESTIBULAR MOTOR CONTROL role in vestibular motor control by mediating the adaption required to keep the VOR calibrated across our life span, as well as the compensation required to compensate for peripheral vestibular loss. In addition, the floccular complex plays a key role in the generation of OKN and pursuit eye movements. Patients with cerebellar disease affecting the flocculus and ventral paraflocculus display corresponding in deficits in gaze stability during their everyday activities. Like all areas of the cerebellar cortex, the floccular complex receives two major sources of input: mossy fiber and climbing fiber inputs. How do signals transmitted by these two inputs shape the neuronal responses of Purkinje cells to ensure accurate vestibular motor control? First, mossy fiber projections to the floccular lobe originate in a variety of brainstem nuclei, including the medial vestibular nucleus and the nucleus prepositus hypoglossi. These inputs drive the simple spike responses of Purkinje cells via the parallel fibers. As noted above, Purkinje cells can generate high frequency simple spike responses, whereas their complex spike rates generally do not exceed 10 spikes/s. As a result, simple spikes play the predominant role in shaping the response profiles of the downstream neurons in the vestibular nuclei, which are targeted by floccular Purkinje cells. Multiple regression and systems identification techniques have been used to evaluate the relative roles of visual retinal slip inputs, vestibular head motion information, and eye movement efference copy signals in shaping the simple spike responses of floccular Purkinje cells. During vestibular stimulation, simple spike responses encode head velocity and acceleration. During optokinetic stimulation and head-restrained smooth pursuit eye movements, the simple spike responses chiefly encode eye position, eye velocity, and to a much smaller degree, eye acceleration (Leung et al., 2000; Suh et al., 2000). Interestingly, simple spike responses increase for eye and head movements in the same direction during head-fixed pursuit eye movements and the visual cancelation of the VOR (e.g., Lisberger et al., 1994). Thus, floccular Purkinje cells effectively encode a gaze velocity signal (where gaze ¼ eye-in-head + head-inspace) (reviewed in Cullen and Mitchell, 2016). Second, in contrast with mossy fiber projections, climbing fiber inputs to the floccular lobe originate in the inferior olive, and project to floccular Purkinje cells where they control complex spiking (Maekawa and Simpson, 1973). Climbing fibers predominately relay visual image motion information to the floccular complex. The climbing fiber input to the flocculus functions to modify the efficacy of the synapses between parallel fibers and Purkinje cells, thereby underlying the circuit plasticity required so that the VOR and optokinetic reflexes remain compensatory to ensure stabile gaze (reviewed in Cullen

43

and Mitchell, 2016). At the cellular level, the climbing fiber spike produces a profound depolarization of the entire Purkinje cell resulting in long-term depression (LTD) that induces plasticity (reviewed in De Zeeuw et al., 2021). Consistent with its origin, the complex spike activity of floccular Purkinje cells is modulated in response to retinal image motion during VOR adaption, optokinetic stimulation, as well as during smooth pursuit eye movements. In contrast, simple spike activity encodes minimal retinal velocity and acceleration error information during optokinetic stimulation (Hirata and Highstein, 2001) or smooth pursuit (Suh et al., 2000). The connectivity of the floccular lobe with the vestibular nuclei effectively provides a parallel inhibitory side loop that can modulate the gain of the direct VOR pathway. Initially, when there is mismatch between head movement and compensatory eye movement—for example, after putting on new pair of glasses with a different prescription or due to peripheral vestibular loss— the climbing fiber input will signal that there is retinal image motion. Importantly, image motion during head movements is effectively an error signaling that gaze is not stable (i.e., the visual world is not stable on the retina). In turn, this climbing fiber error signal alters the responses of Purkinje cells to their mossy fiber input, which then produces a “gain change” in the relationship between Purkinje cells simple spike firing rate and vestibular input. This short-term plastic change in Purkinje cell firing next drives the formation of long-term plastic changes at the level of the vestibular nuclei (Broussard and Kassardjian, 2004). Overtime the site of synaptic plasticity shifts from the cerebellum to the brainstem during VOR adaptation via a process referred to as “transfer” or “consolidation” (Galiana, 1986; Peterson et al., 1991; Raymond et al., 1996; Broussard and Kassardjian, 2004). Targeted lesions in monkeys indicate that relative contribution of ventral paraflocculus may be more significant than that of the flocculus in mediating such VOR plasticity (Rambold et al., 2002). Finally, it is also noteworthy that synaptic changes outside of cerebellar pathways also guide initial changes in motor performance. Studies in mice have shown that climbing fiber input and/or plasticity within cerebellar pathways is not required for VOR adaptation (Ke et al., 2009; Nguyen-Vu et al., 2013) and that motorlearning deficits are not complete in transgenic mice with impaired cerebellar LTD (Boyden et al., 2006; Beraneck et al., 2008; Schonewille et al., 2011). Recording studies in monkeys have further shown that behaviorally relevant rates of vestibular nerve activation produce a rapid reduction in the efficacy monosynaptic response of direct VOR neurons (i.e., PVP neurons; Fig. 3.5), that in turn produces a lasting reduction in evoked eye movements. Thus, multiple sites of plasticity—both

44

K.E. CULLEN

in the cerebellum and brainstem—ultimately contribute to shaping motor performance of the VOR pathway.

Anterior vermis (lobules I–V) Lobules I–V of the anterior vermis of the vestibular cerebellum (Fig. 3.7, green region) play essential role in the maintenance of posture and balance. This region of the cerebellum integrates vestibular signals with proprioceptive information to ensure that the motor responses produced by vestibulospinal reflexes are appropriately referenced to the body to maintain postural stability. Patients with damage to the anterior vermis show impaired posture and balance, as well as deficits in motor coordination (Sullivan et al., 2006; Ilg et al., 2008; Mitoma et al., 2020). Anterior vermis Purkinje cells send direct descending projections the rostral portion of the most medial of the deep cerebellar nuclei (the rostral portion of the fastigial nucleus) as well to the vestibular nuclei (Fig. 3.7, yellow region). Thus expected, ablation of the rostral fastigial nucleus likewise significantly impairs postural control and balance (Thach et al., 1992; Kurzan et al., 1993; Pelisson et al., 1998). Mossy fiber inputs to the anterior vermis encode both vestibular and proprioceptive information. Purkinje cells in this region, in turn, integrate these two streams of sensory information to maintain our posture in response to unexpected movements (Manzoni et al., 1998a,b, 1999, 2004; Zobeiri and Cullen, 2022). Such integration is required for the vestibular motor control of vestibulospinal reflexes because it is essential that the brain accounts for the position of the head relative to the body to generate the appropriate postural motor commands (Tokita et al., 1989; Kennedy and Inglis, 2002; Tokita et al., 2009). In this context, neck proprioceptors provide the head position information required to transform selfmotion information from the head-centered reference frame of the vestibular system into the body-centered reference frame required for accurate postural control (reviewed in Cullen and Zobeiri, 2021). Indeed, the anterior cerebellum transforms head-centered vestibular information into a body-centric reference frame via its projection to the rostral fastigial nucleus (Zobeiri and Cullen, 2022). Specifically, Purkinje cells combine vestibular and proprioceptive information to dynamically encode an intermediate representation of self-motion between head and body motion. In contrast, their target neurons in the rostral fastigial nucleus unambiguously encode either head or body motion (Brooks and Cullen, 2009, 2014). Consistent with the Purkinje cell to deep cerebellar nucleus neuron projection ratio that has been anatomically demonstrated (Palkovits et al., 1977; Person and Raman, 2012), combining neural responses from a population of
40 to 50 Purkinje cells

can explain the transformed responses of these deep cerebellar nuclei neurons. Moreover, the anterior vermis is involved in a second essential computation, namely the brain’s computation of an internal estimate of the expected sensory consequences of actively generated head movements (Fig. 3.8A). As reviewed above in the section entitled "Vestibulospinal Reflex Pathways," robust vestibulospinal reflexes are essential for maintaining posture in response to unexpected vestibular stimuli. Yet the same intact reflexes are counter-productive when the behavioral goal is to instead move through space. Thus, as shown above in Fig. 3.6, vestibulospinal reflexes are largely suppressed during active movements as evidenced by the suppressed responses of VO neurons in the vestibular nuclei. Notably, this suppression only occurs when the brain’s estimate (internal model) of the expected consequences of actively-generated movement matches the actual sensory feedback experienced (for review, see Cullen, 2019; Cullen and Zobeiri, 2021)—as is the case during normal voluntary selfmotion. Likewise, the responses of the rostral fastigial nucleus show marked attenuation for active movements (Brooks and Cullen, 2009; Brooks et al., 2015). Further, when the correspondence between the motor command and actual sensory response is systematically altered, these same anterior vermis output neurons demonstrate rapid updating indicative of an internal model that enables the motor system to learn to expect unexpected sensory inputs (Fig. 3.8B; Brooks et al., 2015). Thus, by computing an estimate of the expected consequences of actively generated movements, the anterior vermis allows a distinction between actively generated vs passively applied vestibular inputs. This computation further underlies the continuous calibration of vestibulospinal pathways to ensure the accurate control posture and maintenance of balance over time (see Brooks and Cullen, 2019, for a review).

Nodulus/uvula of the posterior cerebellar vermis The nodulus and ventral uvula (lobules IX–X) of the posterior vermis of the vestibular cerebellum (Fig. 3.7, blue region) integrate vestibular information from the semicircular canals and otoliths to compute a representation of our orientation relative to gravity (reviewed in Angelaki and Cullen, 2008; Cullen, 2016). Neurons in this cerebellar region predominately respond to rotational vestibular stimuli that reorient the head relative to gravity (i.e., pitch and roll rotations) as well as head translations (reviewed in Goldberg et al., 2012). Lesions of the nodulus and ventral uvula alter the dynamics as

VESTIBULAR MOTOR CONTROL

45

Theorecal framework for computaon of vesbular reafference cancellaon signal

/ SPE

Reafference Cancellaon signal

No cancellaon No Match (SPE≠0) Actual sensory feedback during self-moon

1

Torque Removed

Learning Normalized Behavioral Error

Controls Match (SPE=0)

Compare

Torque Applied

B

Forward Model (expected sensory feedback)

Normalized Neuronal Sensivity

A

Catch Trials

Head Motor Learning 1 0.5 0

0.5

0

Controls (Before learning)

Learning

Fig. 3.8. (A) Schematic of the cerebellar-dependent mechanism that suppresses the responses of VO neurons to vestibular input generated by voluntary head motion (i.e., vestibular reafference). During voluntary head motion, the brain computes an internal (forward) model of the expected sensory consequences of its motor command. This mechanism then compares the brain’s estimate with the actual sensory inflow that occurs to compute the sensory prediction error (SPE). Vestibular reafference is suppressed when there is match between expected and actual sensory inflow. (B) Top: The relationship between the head motor command and resultant movement is altered via the application of resistive load (torque). Bottom: Neuronal vestibular sensitivities to active head motion initially increase to levels measured during passive head motion. As the brain’s internal model is updated to accommodate the new relationship between the voluntary head motor command and resultant movement, there is a re-emergence of reafferent suppression. This re-emergence of reafferent suppression follows the same time course as the corresponding change in head movement error (see inset). Adapted from Cullen KE (2019). Vestibular processing during natural self-motion: implications for perception and action. Nat Rev Neurosci 20: 346–363.

well as three-dimensional orientation of vestibularinduced eye movements relative to gravity. For example, in normal monkeys, the offset of constant velocity head motion results in a postrotatory VOR eye movement response. This postrotatory eye movement normally realigns relative to gravity when the head is tilted. However, removal of the nodulus and ventral uvula in monkeys eliminate the realignment relative to gravity, such that the postrotatory nystagmus remains aligned with the head when it is tilted (Waespe et al., 1985; Angelaki and Hess, 1995; Wearne et al., 1996, 1998). The network underlying this computation—commonly termed velocity storage—is thought to be mediated by reciprocal projections between nodulus/ventral uvula Purkinje cells and neurons in the vestibular nuclei (i.e., vestibular-only (VO) neurons) (Shojaku et al., 1987; Reisine and Raphan, 1992). The nodulus and ventral uvula receive substantial primary vestibular input directly from the afferents, with mossy fiber inputs providing convergent input from the otoliths and canals (Korte and Mugnaini, 1979; Maklad and Fritzsch, 2003). The vestibular nuclei also send direct projections to the nodulus and ventral uvula, effectively providing a secondary source of vestibular

mossy fiber input (Shojaku et al., 1991; Barmack and Shojaku, 1992; Ono et al., 2000). In addition to receiving primary and secondary vestibular input via these mossy fibers, the nodulus and ventral uvula also receive mossy fiber inputs from muscle proprioceptors (Sheliga et al., 1999). As a result, most Purkinje cells in nodulus and ventral uvula respond to proprioceptive as well as otolith and canal stimulation. In contrast, the dorsal uvula does not receive substantial vestibular input. Instead, it receives significant visual input from the dorsal pontine nuclei, which are an integral part of the major pathway related to smooth pursuit. By comparison, the corresponding visual input to the ventral uvula and nodulus is considerably weaker. As noted above, the nodulus and ventral uvula play an important role in computing the brain’s representation of our orientation relative to gravity, which in turn is required to ensure accurate behavior. Notably, however, there is no individual sensory input to the brain that itself encodes gravity. For example, the otolith afferents encode the linear forces produced during tilts and translations, which are physically indistinguishable from each other as indicated by Einstein’s equivalence principle. Thus, considered in isolation the otoliths afferents

46

K.E. CULLEN

transmit ambiguous information to the brain (reviewed in Angelaki and Cullen, 2008; Cullen, 2019). However head tilts, unlike head translations, comprise head rotations which will activate semicircular canal as well as otolith afferents. As a result, the brain integrates otolith and canal signals in order to distinguish between head tilts and translation. Indeed, single unit recording experiments in monkeys have shown that some nodulus/ ventral uvula Purkinje cells selectively encode head tilts (Laurens et al., 2013), whereas others encode selectively encode head translations (Yakusheva et al., 2007). Thus, Purkinje cells in this region of the cerebellum combine otolith and semicircular canal inputs to compute a neural representation of our orientation relative to gravity.

COMPENSATION AND EXTRAVESTIBULAR SENSORY SUBSTITUTION IN CENTRAL PATHWAYS: IMPLICATIONS FOR MOTOR CONTROL Immediately following peripheral vestibular loss, patients experience debilitating dizziness, gaze and postural instability, and an impaired sense of direction during their daily activities. Fortunately, within a few weeks, initial symptoms including spontaneous nystagmus and head tilt toward the side of the lesion are overcome and in fact almost completely disappear. Furthermore, during this period, the VOR response recovers, particularly for less challenging ipsilesional head movements (Curthoys and Halmagyi, 1995; Sadeghi et al., 2006). Common clinical tests of impaired vestibular function measure the eye movement and postural responses produced by VOR and vestibulospinal pathways (Box 3.1). Neurophysiological studies have led to a comprehensive understanding of the central mechanisms that mediate compensation in vestibular motor pathways. First, some compensation likely occurs at the level of the vestibular afferents (via modulation of the efferent

vestibular system) in mammals (reviewed in Cullen and Wei, 2021). Notably, following unilateral vestibular nerve lesion, the mean resting rates and sensitivities of afferents recorded in the intact nerve on the other side (i.e., the contralesional nerve) are unchanged, but there is a small but significant decrease in the proportion of regular afferents (Sadeghi et al., 2007a,b). Importantly, however, most compensation occurs centrally. In particular, the commissural vestibular system, which interconnects the vestibular nuclei on each side of the midline via reciprocal inhibitory projections, plays an essential role in mediating compensation. This reciprocal pathway serves to facilitate compensation by effectively rebalancing the synaptic weighting of commissural inhibition (Bergquist et al., 2008; Malinvaud et al., 2010). Additionally, as reviewed below, the brain uses two other key strategies to adapt to the loss of vestibular nerve input: (i) the unmasking of extra-vestibular information in early vestibular pathways, and (ii) cerebellar-dependent mechanisms (Fig. 3.9A).

The unmasking of extra-vestibular information in early vestibular pathways Vestibular nuclei neurons in healthy rhesus monkeys are not sensitive to proprioceptive stimulation. Instead, as reviewed above, integration of vestibular and proprioception only occurs at the next stages of vestibular processing, for example, in the rostral fastigial nucleus of the cerebellum (Brooks and Cullen, 2009; Brooks et al., 2015) as well as vestibular thalamus (Marlinski and McCrea, 2008a,b; Dale and Cullen, 2017). Remarkably, however, within a day of experiencing unilateral peripheral vestibular loss, vestibular nuclei neurons display strong responses to stimulation of proprioceptors. This rapid unmasking of proprioceptive inputs occurs in both VOR and vestibulospinal pathways and indicates a form of homeostatic plasticity, which effectively compensates

BOX 3.1. VESTIBULO-MOTOR PATHWAYS: DIAGNOSIS AND TREATMENT The function of each of the semicircular canals can be tested by measuring the compensatory VOR eye movements (e.g., Fig. 3.4) evoked by head rotation in the corresponding planes. In particular, the application of brief, high-acceleration head “impulses” have become a common method for detecting the loss of function of a semicircular canal (reviewed in Leigh and Zee, 2015). More recently, vestibular evoked myogenic potentials (VEMPs) have become an increasingly popular vestibular motor test for the assessment of otolith function. VEMPs are short latency electromyographic responses that are typically recorded in response to a transient acoustic stimulus. Cervical VEMPs (termed cVEMPs) are measured by recording from the neck musculature— typically from the ipsilateral sternocleidomastoid muscle. cVEMPs are mediated by the vestibulo-collic reflex pathway (Fig. 3.6) and are generally thought to result from the stimulation of the sacculus. Ocular VEMPs (termed oVEMPs) are measured by recording extraocular muscle activity via surface EMG placed under the contralateral eye. oVEMPs are mediated by the vestibulo-ocular reflex pathway (Fig. 3.5) and are generally thought to predominately result from the stimulation of the utricle (reviewed in Rosengren and Kingma, 2013; Scarpa et al., 2019).

VESTIBULAR MOTOR CONTROL Neck proprioceptive signal

PVP

Gaze Stabilization (VOR)

VO

Posture & Balance (VSR)

47

Vestibular signal Efference copy signal

Self-motion Perception & Navigation

A

Normalized Contribution

Sensory & Motor Substitution Following Vestibular Loss

B

Proprioceptive signal Vestibular signal Efference copy signal

1

0.5

Control Week 1

2

3

Post Lesion

Fig. 3.9. In response to the loss of peripheral vestibular input, the vestibular system demonstrates dynamic regulation of multimodal integration at the level of single vestibular nuclei neurons. Neck proprioceptive inputs are unmasked within 24 h followed by the dynamic upregulation of motor efference copy signals. Further, these changes are associated with increased recovery of vestibular sensitivity by individual neurons, suggesting a role for homeostatic mechanisms in adaptation.

for the decreased reliability of vestibular input (Sadeghi et al., 2010, 2011, 2012). The unmasking of this extra-vestibular input is linked to compensation as evidenced by faster and more significant recovery of neurons that respond to proprioceptive stimulation following peripheral loss (Sadeghi et al., 2010). Additionally, motor signals also contribute to neuronal responses over a longer time course (Sadeghi et al., 2010, 2011, 2012) which further serves to improve the efficacy of the vestibular motor pathways. For example, the VOR is enhanced for actively generated vs passively applied head movements following vestibular loss. It is also noteworthy that the increased sensitivity of vestibular nuclei neurons mediated via central compensation is accompanied by a parallel increase in response variability (i.e., decreased signal-to-noise; Jamali et al., 2014). Fig. 3.9B illustrates the time course of vestibular and extra-vestibular compensation observed at the level of the vestibular nuclei following complete unilateral vestibular loss (reviewed in Carriot et al., 2015b). Interestingly, sensory substitution via extravestibular inputs such as proprioception serves to improve signal-to-noise during self-motion (Jamali et al., 2014), which in turn is likely to improve the accuracy of patient vestibular motor behavior during everyday activities.

Cerebellar-dependent mechanisms for vestibular motor compensation Cerebellar-dependent mechanisms are also essential for vestibular motor compensation following peripheral sensory loss. As noted above, vestibular pathways through the cerebellum effectively provide a parallel inhibitory side loop which can modulate the gain of vestibulomotor reflexes. While most studies have focused on the cerebellum’s role in visually-induced motor learning (reviewed in: Broussard and Kassardjian, 2004), the cerebellum also plays an essential role in the normal resolution of static symptoms and initiation of behavioral recovery that is observed following peripheral lesions (reviewed in Cullen et al., 2009). Accordingly, cerebellar lesion studies in cats (Haddad et al., 1977; Courjon et al., 1982) and rats (Kitahara et al., 1997, 1998, 2000; Kim et al., 1997a,b) impair the resolution of static symptoms and initiation of behavioral recovery following peripheral vestibular loss. Similarly, Lurcher mutant mice—a strain of mouse demonstrating degeneration of the cerebellar cortex within the first 3 postnatal weeks—demonstrate significantly impaired compensation following peripheral lesion. Notably however, compensation is not completely abolished in these mice consistent with the parallel role played by

48

K.E. CULLEN

cerebellar-independent mechanisms in vestibulo motor compensation (see also, Mitchell et al., 2016, 2017). Thus, similar to mechanisms reviewed above for vestibular motor learning (see section entitled "The Vestibular Cerebellum: Internal Models of Externally-Applied and Active Self-Motion"), the brain uses both cerebellardependent and cerebellar-independent mechanisms to adapt to the loss of vestibular nerve input.

VOLUNTARY BEHAVIOR: STEERING, REACHING AND NAVIGATION So far, this chapter has focused on the vestibular motor control of essential reflexes, namely the VOR, VCR, and VSR, as well as the mechanisms that underlie their plasticity to optimize performance. However, recent studies have also emphasized the vestibular system’s vital contribution to other voluntary motor behaviors that are not classically associated with the vestibular system. As considered below recent studies of voluntary behaviors including reaching, steering, and navigation have furthered our understanding of the vestibular system’s pervasive role in voluntary motor control.

Reaching In everyday life, we often simultaneously reach for objects as we move through our environment—consider for example turning to pick up a cup of coffee off a counter. In such example, it is essential to keep track of our self-motion relative to the environment, as well as the additional physical forces (e.g., Coriolis and centrifugal forces) that must be accounted for in order to plan and execute an accurate reaching movement. In this context, vestibular information is well suited to provide on-line feedback to ensure accurate motor control. Indeed, the motor pathways controlling reaching movements demonstrate on-line updating that compensates for displacements of the body and limb relative to the target produced by externally applied perturbations (Adamovich et al., 2001; Bresciani et al., 2002, 2005; Raptis et al., 2007). As noted above, GVS is now commonly used to selectively and noninvasively target and perturb the vestibular system of human subjects in order to assess its specific contribution to voluntary motor behaviors. Studies using this technique have confirmed that vestibular feedback makes a significant contribution to the control of reaching by compensating spatial displacement resulting from self-motion (Bresciani et al., 2002; MoreauDebord et al., 2014; Keyser et al., 2017). Furthermore, the motor pathways that generate reaches process vestibular signals in a manner that accounts for the kinematic consequences of the forces imposed on it by the body’s motion (Guillaud et al., 2011; Moreau-Debord et al., 2014; Martin et al., 2021). Moreover, in addition to compensating for body and limb displacement and their biomechanics during

reaching, the vestibular system also contributes to ensuring postural stability during the preparation phase of reaching (Kennefick et al., 2020). Thus, in everyday life the vestibular system provides essential on-line feedback to ensure both the planning and execution of accurate reaching movements. These findings raise the question of what neural pathways underlie vestibular system’s contribution to voluntary reaching. Vestibular signals that can influence reach planning and execution have been described in parietal cortex, as well as somatosensory cortex (Kawano and Sasaki, 1984; Bottini et al., 1994; Andersen et al., 1999). For example, the parietal reach region (PRR) of primate parietal cortex plays a central role the planning and execution of arm reaching movements. Experiments in monkeys have shown that parietal cortex receives ascending vestibular input (Ugolini et al., 2019). Importantly, this input specifically targets the PRR and is thought to contribute to the sensorimotor transformation of visual and other sensory information for the generation of arm-reach signals. Consistent with this role, neurons in this cortical region demonstrate robust responses to vestibular stimulation (Schlack and Hoffmann, 2002; Klam and Graf, 2006) and integrate vestibular information with extravestibular (i.e., proprioception, vision, tactile) signals and motorrelated information thereby demonstrating feedback control during the high-level planning of reaching movements required to achieve accurate performance.

Steering and navigation Recent studies have also focused on the vestibular system’s contribution to our ability to voluntarily steer and navigate through our environment. Indeed, online feedback about where we are heading is vital for accurate steering and navigation. To this end, the brain integrates vestibular, visual (optic flow), and proprioceptive information (reviewed in Cullen, 2011, 2019). Behavioral studies in humans have demonstrated that manual control precision during steering depends on vestibular sensory precision (Rosenberg et al., 2018). Vestibular information is sent via vestibular nuclei neurons in the brainstem (Fig. 3.6A) to higher level centers via two main ascending pathways: anterior and posterior thalamocortical pathways (reviewed in Cullen and Taube, 2017). The posterior pathway comprises direct projections from the VO neurons of the vestibular nuclei to the ventral posterior thalamus, which in turn projects to vestibular cortical regions including parietal-insular vestibular cortex and dorsal medial superior temporal (MSTd). In contrast, the anterior pathway comprises indirect projections from the vestibular nuclei to head direction network of the anterior thalamus, which in turn projects to retrosplenial cortex and sends multisynaptic projections to the entorhinal cortex.

VESTIBULAR MOTOR CONTROL Surprisingly, when monkeys perform a goal directed steering task in which they manually control a steering wheel to move themselves relative to the world, VO vestibular nuclei neurons respond as if head motion was externally applied rather than voluntarily generated (Roy and Cullen, 2004; Cullen, 2019). This contrasts with the cancelation of their vestibular responses described above that occurs during actively generated orienting head movements (Fig. 3.6). Likewise, neurons in cortical area such as MSTd neurons targeted by the posterior thalamocortical pathway also encode vestibular signals during voluntary steering tasks (Duffy, 1998; Gu et al., 2007; Page and Duffy, 2008) in addition to showing visual responses to optic flow (Duffy and Wurtz, 1995; Page and Duffy, 2008; Bremmer et al., 2010). Further research is required to fully understand how the brain integrates self-motion (i.e., vestibular, proprioceptive, and visual) cues and motor information to generate accurate steering behavior. Further it remains to be determined whether and how such integration is enhanced with motor training to improve performance accuracy during a steering task. Finally, the vestibular system has long been implicated in providing a vital input to the neural mechanisms underlying our ability to accurately navigate our environment. Notably, lesions studies have shown that ablation of the peripheral vestibular system disrupts the direction signal found in head direction (HD) cells of the anterior thalamocortical pathway (reviewed in Cullen and Taube, 2017). HD neurons discharge in relation to the animal’s directional heading in the horizontal plane, independent of the animal’s location and behavior (reviewed in Moser et al., 2008, Dumont and Taube, 2015), and in turn contribute to the grid cell system of the medial entorhinal cortex whose tuning is also disrupted following peripheral vestibular lesions. However, as noted above vestibular input that is a consequence of active behavior is suppressed at the level of the vestibular nuclei. This fact has important implications for understanding the nature of the information that is relayed to cortex via the anterior thalamocortical pathway. Additionally, the anterior thalamocortical pathway appears to integrate eye motor signals with vestibular information—which will also contribute to HD cell tuning and grid cell firing (reviewed in Cullen and Taube, 2017). Thus, to date, how the ascending vestibular pathway utilizes vestibular signals vs eye motor signals during navigation remains an open question for the field to address in the next generation of research on vestibular motor control.

REFERENCES Adamovich SV, Archambault PS, Ghafouri M et al. (2001). Hand trajectory invariance in reaching movements involving the trunk. Exp Brain Res 138: 288–303.

49

Andersen RA, Shenoy KV, Snyder LH et al. (1999). The contributions of vestibular signals to the representations of space in the posterior parietal cortex. Ann N Y Acad Sci 871: 282–292. Angelaki DE, Cullen KE (2008). Vestibular system: the many facets of a multimodal sense. Annu Rev Neurosci 31: 125–150. Angelaki DE, Hess BJ (1995). Inertial representation of angular motion in the vestibular system of rhesus monkeys. II. Otolith-controlled transformation that depends on an intact cerebellar nodulus. J Neurophysiol 73: 1729–1751. Armand M, Minor LB (2001). Relationship between time- and frequency-domain analyses of angular head movements in the squirrel monkey. J Comput Neurosci 11: 217–239. Barmack NH, Shojaku H (1992). Vestibularly induced slow oscillations in climbing fiber responses of Purkinje cells in the cerebellar nodulus of the rabbit. Neuroscience 50: 1–5. Beraneck M, McKee JL, Aleisa M et al. (2008). Asymmetric recovery in cerebellar-deficient mice following unilateral labyrinthectomy. J Neurophysiol 100: 945–958. Bergquist F, Ludwig M, Dutia MB (2008). Role of the commissural inhibitory system in vestibular compensation in the rat. J Physiol 586: 4441–4452. Bilotto G, Goldberg J, Peterson BW et al. (1982). Dynamic properties of vestibular reflexes in the decerebrate cat. Exp Brain Res 47: 343–352. Bottini G et al. (1994). Identification of the central vestibular projections in man: a positron emission tomography activation study. Exp Brain Res 99: 164–169. Boyden ES, Katoh A, Pyle JL et al. (2006). Selective engagement of plasticity mechanisms for motor memory storage. Neuron 51: 823–834. Boyle R, Goldberg JM, Highstein SM (1992). Inputs from regularly and irregularly discharging vestibular nerve afferents to secondary neurons in squirrel monkey vestibular nuclei. III. Correlation with vestibulospinal and vestibuloocular output pathways. J Neurophysiol 68: 471–484. Bremmer F, Kubischik M, Pekel M et al. (2010). Visual selectivity for heading in monkey area MST. Exp Brain Res 200: 51–60. Bresciani JP, Blouin J, Sarlegna F et al. (2002). On-line versus off-line vestibular-evoked control of goal-directed arm movements. Neuroreport 13: 1563–1566. Bresciani JP, Gauthier GM, Vercher JL et al. (2005). On the nature of the vestibular control of arm-reaching movements during whole-body rotations. Exp Brain Res 164: 431–441. Brooks JX, Cullen KE (2009). Multimodal integration in rostral fastigial nucleus provides an estimate of body movement. J Neurosci 29: 10499–10511. Brooks JX, Cullen KE (2014). Early vestibular processing does not discriminate active from passive self-motion if there is a discrepancy between predicted and actual proprioceptive feedback. J Neurophysiol 111: 2465–2478. Brooks JX, Cullen KE (2019). Predictive sensing: the role of motor signals in sensory processing. Biol Psychiatry Cogn Neurosci Neuroimaging 4: 842–850. Brooks JX, Carriot J, Cullen KE (2015). Learning to expect the unexpected: rapid updating in primate cerebellum during voluntary self-motion. Nat Neurosci 18: 1310–1317.

50

K.E. CULLEN

Broussard DM, Kassardjian CD (2004). Learning in a simple motor system. Learn Mem 11: 127–136. Carriot J, Brooks JX, Cullen KE (2013). Multimodal integration of self-motion cues in the vestibular system: active versus passive translations. J Neurosci 33: 19555–19566. Carriot J, Jamali M, Chacron MJ et al. (2014). Statistics of the vestibular input experienced during natural self-motion: implications for neural processing. J Neurosci 34: 8347–8357. Carriot J, Jamali M, Brooks JX et al. (2015a). Integration of canal and otolith inputs by central vestibular neurons is subadditive for both active and passive self-motion: implication for perception. J Neurosci 35: 3555–3565. Carriot J, Jamali M, Cullen KE (2015b). Rapid adaptation of multisensory integration in vestibular pathways. Front Syst Neurosci 9: 59. Carriot J, Jamali M, Chacron MJ et al. (2017a). The statistics of the vestibular input experienced during natural self-motion differ between rodents and primates. J Physiol (Lond) 595: 2751–2766. Carriot J, Jamali M, Cullen KE et al. (2017b). Envelope statistics of self-motion signals experienced by human subjects during everyday activities: implications for vestibular processing. PLoS One 12: e0178664. Chichilnisky EJ (2001). A simple white noise analysis of neuronal light responses. Network 12: 199–213. Chow MR, Ayiotis AI, Schoo DP et al. (2021). Posture, gait, quality of life, and hearing with a vestibular implant. N Engl J Med 384: 521–532. Courjon JH, Flandrin JM, Jeannerod M et al. (1982). The role of the flocculus in vestibular compensation after hemilabyrinthectomy. Brain Res 239: 251–257. Cullen KE (2004). Sensory signals during active versus passive movement. Curr Opin Neurobiol 14: 698–706. Cullen KE (2011). The neural encoding of self-motion. Curr Opin Neurobiol 21: 587–595. Cullen KE (2012). The vestibular system: multimodal integration and encoding of self-motion for motor control. Trends Neurosci 185–196. Cullen KE (2016). Physiology of central pathways. Handb Clin Neurol 137: 17–40. Cullen KE (2019). Vestibular processing during natural selfmotion: implications for perception and action. Nat Rev Neurosci 20: 346–363. Cullen KE, McCrea RA (1993). Firing behavior of brain stem neurons during voluntary cancellation of the horizontal vestibuloocular reflex. I Secondary vestibular neurons. J Neurophysiol 70: 828–843. Cullen KE, Minor LB (2002). Semicircular canal afferents similarly encode active and passive head-on-body rotations: implications for the role of vestibular efference. J Neurosci 22: RC226. Cullen KE, Mitchell DE (2016). Procedural learning: VOR. In: The curated reference collection in neuroscience and biobehavioral psychology, Elsevier Science357–374. Cullen KE, Roy JE (2004). Signal processing in the vestibular system during active versus passive head movements. J Neurophysiol 91: 1919–1933. Cullen KE, Taube JS (2017). Our sense of direction: progress, controversies and challenges. Nat Neurosci 20: 1465–1473.

Cullen KE, Wei RH (2021). Differences in the structure and function of the vestibular efferent system among vertebrates. Front Neurosci 15: 684800. Cullen KE, Zobeiri OA (2021). Proprioception and the predictive sensing of active self-motion. Curr Opin Physiol 20: 29–38. Cullen KE, Chen-Huang C, McCrea RA (1993). Firing behavior of brain stem neurons during voluntary cancellation of the horizontal vestibuloocular reflex. II. Eye movement related neurons. J Neurophysiol 70: 844–856. Cullen KE, Huterer M, Braidwood DA et al. (2004). Time course of vestibuloocular reflex suppression during gaze shifts. J Neurophysiol 92: 3408–3422. Cullen KE, Minor LB, Beraneck M et al. (2009). Neural substrates underlying vestibular compensation: contribution of peripheral versus central processing. J Vestib Res 19: 171–182. Curthoys IS, Halmagyi GM (1995). Vestibular compensation: a review of the oculomotor, neural, and clinical consequences of unilateral vestibular loss. J Vestib Res 5: 67–107. Dakin CJ, Inglis JT, Chua R et al. (2013). Muscle-specific modulation of vestibular reflexes with increased locomotor velocity and cadence. J Neurophysiol 110: 86–94. Dale A, Cullen KE (2017). The ventral posterior lateral thalamus preferentially encodes externally applied versus active movement: implications for self-motion perception. Cereb Cortex 1–14. De Zeeuw CI, Lisberger SG, Raymond JL (2021). Diversity and dynamism in the cerebellum. Nat Neurosci 24: 160–167. Duffy CJ (1998). MST neurons respond to optic flow and translational movement. J Neurophysiol 80: 1816–1827. Duffy CJ, Wurtz RH (1995). Response of monkey MST neurons to optic flow stimuli with shifted centers of motion. J Neurosci 15: 5192–5208. Dumont JR, Taube JS (2015). The neural correlates of navigation beyond the hippocampus. Prog Brain Res 219: 83–102. Ferna´ndez C, Goldberg JM (1971). Physiology of peripheral neurons innervating semicircular canals of the squirrel monkey. II. Response to sinusoidal stimulation and dynamics of peripheral vestibular system. J Neurophysiol 34: 661–675. Forbes PA, Luu BL, Van der Loos HF et al. (2016). Transformation of vestibular signals for the control of standing in humans. J Neurosci 36: 11510–11520. Forbes PA, Kwan A, Rasman BG et al. (2020). Neural mechanisms underlying high-frequency vestibulocollic reflexes in humans and monkeys. J Neurosci 40: 1874–1887. Galiana HL (1986). A new approach to understanding adaptive visual-vestibular interactions in the central nervous system. J Neurophysiol 55: 349–374. Goldberg JM (2000). Afferent diversity and the organization of central vestibular pathways. Exp Brain Res 130: 277–297. Goldberg JM, Cullen KE (2011). Vestibular control of the head: possible functions of the vestibulocollic reflex. Exp Brain Res 210: 331–345.

VESTIBULAR MOTOR CONTROL Goldberg JM, Highstein SM, Moschovakis AK et al. (1987). Inputs from regularly and irregularly discharging vestibular nerve afferents to secondary neurons in the vestibular nuclei of the squirrel monkey. I. An electrophysiological analysis. J Neurophysiol 58: 700–718. Goldberg JM, Wilson VJ, Cullen KE et al. (2012). The vestibular system: a sixth sense, Oxford University Press. Gonshor A, Melvill Jones G (1976). Extreme vestibulo-ocular adaptation induced by prolonged optical reversal of vision. J Physiol 256: 381–414. Grossman GE et al. (1988). Frequency and velocity of rotational head perturbations during locomotion. Exp Brain Res 70: 470–476. Gr€ usser OJ, Pause M, Schreiter U (1990). Localization and responses of neurones in the parieto-insular vestibular cortex of awake monkeys (Macaca fascicularis). J Physiol 430: 537–557. Gu Y, DeAngelis GC, Angelaki DE (2007). A functional link between area MSTd and heading perception based on vestibular signals. Nat Neurosci 10: 1038–1047. Guillaud E, Simoneau M, Blouin J (2011). Prediction of the body rotation-induced torques on the arm during reaching movements: evidence from a proprioceptively deafferented subject. Neuropsychologia 49: 2055–2059. Haddad GM, Friendlich AR, Robinson DA (1977). Compensation of nystagmus after VIIIth nerve lesions in vestibulo-cerebellectomized cats. Brain Res 135: 192–196. Haque A et al. (2004). Spatial tuning and dynamics of vestibular semicircular canal afferents in rhesus monkeys. Exp Brain Res 155: 81–90. Highstein SM, Goldberg JM, Moschovakis AK et al. (1987). Inputs from regularly and irregularly discharging vestibular nerve afferents to secondary neurons in the vestibular nuclei of the squirrel monkey. II. Correlation with output pathways of secondary neurons. J Neurophysiol 58: 719–738. Hirata Y, Highstein SM (2001). Acute adaptation of the vestibuloocular reflex: signal processing by floccular and ventral parafloccular Purkinje cells. J Neurophysiol 85: 2267–2288. Hullar TE et al. (2005). Responses of irregularly discharging chinchilla semicircular canal vestibular-nerve afferents during high-frequency head rotations. J Neurophysiol 93: 2777–2786. Huterer M, Cullen KE (2002). Vestibuloocular reflex dynamics during high-frequency and high-acceleration rotations of the head on body in rhesus monkey. J Neurophysiol 88: 13–28. Ilg W, Giese MA, Gizewski ER et al. (2008). The influence of focal cerebellar lesions on the control and adaptation of gait. Brain 131: 2913–2927. Jamali M, Sadeghi SG, Cullen KE (2009). Response of vestibular nerve afferents innervating utricle and saccule during passive and active translations. J Neurophysiol 101: 141–149. Jamali M, Carriot J, Chacron MJ et al. (2013). Strong correlations between sensitivity and variability give rise to constant discrimination thresholds across the otolith afferent population. J Neurosci 33: 11302–11313.

51

Jamali M, Mitchell DE, Dale A et al. (2014). Neuronal detection thresholds during vestibular compensation: contributions of response variability and sensory substitution. J Physiol 592: 1565–1580. Jamali M, Chacron MJ, Cullen KE (2016). Self-motion evokes precise spike timing in the primate vestibular system. Nat Commun 7: 13229. Jamali M, Chacron MJ, Cullen KE (2019). Coding strategies in the otolith system differ for translational head motion vs. static orientation relative to gravity. Elife 8: e45573. Karmali F (2019). The velocity storage time constant: balancing between accuracy and precision. Prog Brain Res 248: 269–276. Karmali F, Goodworth AD, Valko Y et al. (2021). The role of vestibular cues in postural sway. J Neurophysiol 125: 672–686. Kawano K, Sasaki M (1984). Response properties of neurons in posterior parietal cortex of monkey during visual-vestibular stimulation. II. Optokinetic neurons. J Neurophysiol 51: 352–360. Ke MC, Guo CC, Raymond JL (2009). Elimination of climbing fiber instructive signals during motor learning. Nat Neurosci 12: 1171–1179. Kennedy PM, Inglis JT (2002). Interaction effects of galvanic vestibular stimulation and head position on the soleus H reflex in humans. Clin Neurophysiol 113: 1709–1714. Kennefick M, McNeil CJ, Burma JS et al. (2020). Modulation of vestibular-evoked responses prior to simple and complex arm movements. Exp Brain Res 238: 869–881. Keyser J, Medendorp WP, Selen LP (2017). Task-dependent vestibular feedback responses in reaching. J Neurophysiol 118: 84–92. Kim MS, Jin BK, Chun SW et al. (1997a). Effect of MK801 on cFos-like protein expression in the medial vestibular nucleus at early stage of vestibular compensation in uvulonodullectomized rats. Neurosci Lett 231: 147–150. Kim MS, Jin BK, Chun SW et al. (1997b). Role of vestibulocerebellar N-methyl-D-aspartate receptors for behavioral recovery following unilateral labyrinthectomy in rats. Neurosci Lett 222: 171–174. Kitahara T, Takeda N, Saika T et al. (1997). Role of the flocculus in the development of vestibular compensation: immunohistochemical studies with retrograde tracing and flocculectomy using Fos expression as a marker in the rat brainstem. Neuroscience 76: 571–580. Kitahara T, Takeda N, Uno A et al. (1998). Unilateral labyrinthectomy downregulates glutamate receptor delta-2 expression in the rat vestibulocerebellum. Brain Res Mol Brain Res 61: 170–178. Kitahara T, Fukushima M, Takeda N et al. (2000). Effects of preflocculectomy on Fos expression and NMDA receptormediated neural circuits in the central vestibular system after unilateral labyrinthectomy. Acta Otolaryngol 120: 866–871. Klam F, Graf W (2006). Discrimination between active and passive head movements by macaque ventral and medial intraparietal cortex neurons. J Physiol (Lond) 574: 367–386. Korte GE, Mugnaini E (1979). The cerebellar projection of the vestibular nerve in the cat. J Comp Neurol 184: 265–277.

52

K.E. CULLEN

Kurzan R, Straube A, B€uttner U (1993). The effect of muscimol micro-injections into the fastigial nucleus on the optokinetic response and the vestibulo-ocular reflex in the alert monkey. Exp Brain Res 94: 252–260. Kwan A, Forbes PA, Mitchell DE et al. (2019). Neural substrates, dynamics and thresholds of galvanic vestibular stimulation in the behaving primate. Nat Commun 10: 1904. Lang W, Buttner-Ennever JA, Buttner U (1979). Vestibular projections to the monkey thalamus: an autoradiographic study. Brain Res 177: 3–17. Laurens J, Angelaki DE (2011). The functional significance of velocity storage and its dependence on gravity. Exp Brain Res 210: 407–422. Laurens J, Meng H, Angelaki DE (2013). Neural representation of orientation relative to gravity in the macaque cerebellum. Neuron 80: 1508–1518. Leigh RJ, Zee D (2015). The neurology of eye movements, fifth edn. Oxford University Press, Oxford. Leung HC, Suh M, Kettner RE (2000). Cerebellar flocculus and paraflocculus Purkinje cell activity during circular pursuit in monkey. J Neurophysiol 83: 13–30. Lewis RF (2016). Vestibular implants studied in animal models: clinical and scientific implications. J Neurophysiol 116: 2777–2788. Lisberger SG, Pavelko TA, Bronte-Stewart HM et al. (1994). Neural basis for motor learning in the vestibuloocular reflex of primates. II. Changes in the responses of horizontal gaze velocity Purkinje cells in the cerebellar flocculus and ventral paraflocculus. J Neurophysiol 72: 954–973. Lorente de No´ R (1933). Vestibulo-ocular reflex arc. Arch Neurol Psychiatry 30: 245–291. Lund S, Broberg C (1983). Effects of different head positions on postural sway in man induced by a reproducible vestibular error signal. Acta Physiol Scand 117: 307–309. Mackrous I, Carriot J, Jamali M et al. (2019). Cerebellar prediction of the dynamic sensory consequences of gravity. Curr Biol 29: 2698–2710. Mackrous I, Carriot J, Cullen KE et al. (2020). Neural variability determines coding strategies for natural self-motion in macaque monkeys. Elife 11: e57484. Mackrous I, Carriot J, Cullen KE (2022). Context-independent encoding of passive and active self-motion in vestibular afferent fibers during locomotion in primates. Nat Commun 13: 120. Maekawa K, Simpson J (1973). Climbing fiber responses evoked in vestibulocerebellum of rabbit from visual system. J Neurophysiol 36: 649–666. Magnani RM, Bruijn SM, van Diee¨n JH et al. (2021). Stabilization demands of walking modulate the vestibular contributions to gait. Sci Rep 11: 13736. Maklad A, Fritzsch B (2003). Partial segregation of posterior crista and saccular fibers to the nodulus and uvula of the cerebellum in mice, and its development. Brain Res Dev Brain Res 140: 223–236. Malinvaud D, Vassias I, Reichenberger I et al. (2010). Functional organization of vestibular commissural connections in frog. J Neurosci 30: 3310–3325.

Manzoni D, Pompeiano O, Andre P (1998a). Convergence of directional vestibular and neck signals on cerebellar purkinje cells. Pflugers Arch 435: 617–630. Manzoni D, Pompeiano O, Andre P (1998b). Neck influences on the spatial properties of vestibulospinal reflexes in decerebrate cats: role of the cerebellar anterior vermis. J Vestib Res 8: 283–297. Manzoni D, Pompeiano O, Bruschini L et al. (1999). Neck input modifies the reference frame for coding labyrinthine signals in the cerebellar vermis: a cellular analysis. Neuroscience 93: 1095–1107. Manzoni D, Andre P, Bruschini L (2004). Coupling sensory inputs to the appropriate motor responses: new aspects of cerebellar function. Arch Ital Biol 142: 199–215. Marlinski V, McCrea RA (2008a). Activity of ventroposterior thalamus neurons during rotation and translation in the horizontal plane in the alert squirrel monkey. J Neurophysiol 99: 2533–2545. Marlinski V, McCrea RA (2008b). Coding of self-motion signals in ventro-posterior thalamus neurons in the alert squirrel monkey. Exp Brain Res 189: 463–472. Marlinski V, McCrea RA (2009). Self-motion signals in vestibular nuclei neurons projecting to the thalamus in the alert squirrel monkey. J Neurophysiol 101: 1730–1741. Martin CZ, Lapierre P, Hache S et al. (2021). Vestibular contributions to online reach execution are processed via mechanisms with knowledge about limb biomechanics. J Neurophysiol 125: 1022–1045. Massot C, Chacron MJ, Cullen KE (2011). Information transmission and detection thresholds in the vestibular nuclei: single neurons vs. population encoding. J Neurophysiol 105: 1798–1814. Massot C, Schneider AD, Chacron MJ et al. (2012). The vestibular system implements a linear-nonlinear transformation in order to encode self-motion. PLoS Biol 10: e1001365. Matsuyama K, Drew T (2000a). Vestibulospinal and reticulospinal neuronal activity during locomotion in the intact cat. II. Walking on an inclined plane. J Neurophysiol 84: 2257–2276. Matsuyama K, Drew T (2000b). Vestibulospinal and reticulospinal neuronal activity during locomotion in the intact cat. I. Walking on a level surface. J Neurophysiol 84: 2237–2256. Mian OS, Day BL (2014). Violation of the craniocentricity principle for vestibularly evoked balance responses under conditions of anisotropic stability. J Neurosci 34: 7696–7703. Minor LB, Goldberg JM (1991). Vestibular-nerve inputs to the vestibulo-ocular reflex: a functional-ablation study in the squirrel monkey. J Neurosci 11: 1636–1648. Mitchell DE, Della Santina CC, Cullen KE (2016). Plasticity within non-cerebellar pathways rapidly shapes motor performance in vivo. Nat Commun 7: 11238. Mitchell DE, Della Santina CC, Cullen KE (2017). Plasticity within excitatory and inhibitory pathways of the vestibulospinal circuitry guides changes in motor performance. Sci Rep 7: 853.

VESTIBULAR MOTOR CONTROL Mitoma H, Buffo A, Gelfo F et al. (2020). Consensus paper. Cerebellar reserve: from cerebellar physiology to cerebellar disorders. Cerebellum (London, England) 19: 131–153. Moreau-Debord I, Martin CZ, Landry M et al. (2014). Evidence for a reference frame transformation of vestibular signal contributions to voluntary reaching. J Neurophysiol 111: 1903–1919. Moser EI, Kropff E, Moser M-BB (2008). Place cells, grid cells, and the brain’s spatial representation system. Annu Rev Neurosci 31: 69–89. Nashner LM, Wolfson P (1974). Influence of head position and proprioceptive cues on short latency postural reflexes evoked by galvanic stimulation of human labyrinth. Brain Res 67: 255–268. Nguyen-Vu TDB, Kimpo RR, Rinaldi JM et al. (2013). Cerebellar Purkinje cell activity drives motor learning. Nat Neurosci 16: 1734–1736. Ono S, Kushiro K, Zakir M et al. (2000). Properties of utricular and saccular nerve-activated vestibulocerebellar neurons in cats. Exp Brain Res 134: 1–8. Page WK, Duffy CJ (2008). Cortical neuronal responses to optic flow are shaped by visual strategies for steering. Cereb Cortex 18: 727–739. Palkovits M, Mezey E, Ha´mori J et al. (1977). Quantitative histological analysis of the cerebellar nuclei in the cat. I. Numerical data on cells and on synapses. Exp Brain Res 28: 189–209. Pelisson D, Goffart L, Guillaume A (1998). Contribution of the rostral fastigial nucleus to the control of orienting gaze shifts in the head-unrestrained cat. J Neurophysiol 80: 1180–1196. Person AL, Raman IM (2012). Synchrony and neural coding in cerebellar circuits. Front Neural Circuits 6: 97. Peterson BW, Baker JF, Houk JC (1991). A model of adaptive control of vestibuloocular reflex based on properties of cross-axis adaptation. Ann N Y Acad Sci 627: 319–337. Pozzo T, Berthoz A, Lefort L (1990). Head stabilization during various locomotor tasks in humans: I. Normal subjects. Exp Brain Res 82. Pozzo T, Berthoz A, Lefort L et al. (1991). Head stabilization during various locomotor tasks in humans. II. Patients with bilateral peripheral vestibular deficits. Exp Brain Res 85: 208–217. Ramachandran R, Lisberger SG (2006). Transformation of vestibular signals into motor commands in the vestibuloocular reflex pathways of monkeys. J Neurophysiol 96: 1061–1074. Rambold H, Churchland A, Selig Y et al. (2002). Partial ablations of the flocculus and ventral paraflocculus in monkeys cause linked deficits in smooth pursuit eye movements and adaptive modification of the VOR. J Neurophysiol 87: 912–924. Raptis HA, Dannenbaum E, Paquet N et al. (2007). Vestibular system may provide equivalent motor actions regardless of the number of body segments involved in the task. J Neurophysiol 97: 4069–4078.

53

Raymond JL, Lisberger SG, Mauk MD (1996). The cerebellum: a neuronal learning machine? Science 272: 1126–1131. Reisine H, Raphan T (1992). Neural basis for eye velocity generation in the vestibular nuclei of alert monkeys during offvertical axis rotation. Exp Brain Res 92: 209–226. Rosenberg MJ, Galvan-Garza RC, Clark TK et al. (2018). Human manual control precision depends on vestibular sensory precision and gravitational magnitude. J Neurophysiol 120: 3187–3197. Rosengren SM, Kingma H (2013). New perspectives on vestibular evoked myogenic potentials. Curr Opin Neurol 26: 74–80. Roy JE, Cullen KE (1998). A neural correlate for vestibuloocular reflex suppression during voluntary eye-head gaze shifts. Nat Neurosci 1: 404–410. Roy JE, Cullen KE (2001). Selective processing of vestibular reafference during self-generated head motion. J Neurosci 21: 2131–2142. Roy JE, Cullen KE (2002). Vestibuloocular reflex signal modulation during voluntary and passive head movements. J Neurophysiol 87: 2337–2357. Roy JE, Cullen KE (2004). Dissociating self-generated from passively applied head motion: neural mechanisms in the vestibular nuclei. J Neurosci 24: 2102–2111. Sadeghi SG, Minor LB, Cullen KE (2006). Dynamics of the horizontal vestibuloocular reflex after unilateral labyrinthectomy: response to high frequency, high acceleration, and high velocity rotations. Exp Brain Res 175: 471–484. Sadeghi SG, Chacron MJ, Taylor MC et al. (2007a). Neural variability, detection thresholds, and information transmission in the vestibular system. J Neurosci 27: 771–781. Sadeghi SG, Minor LB, Cullen KE (2007b). Response of vestibular-nerve afferents to active and passive rotations under normal conditions and after unilateral labyrinthectomy. J Neurophysiol 97: 1503–1514. Sadeghi SG, Minor LB, Cullen KE (2010). Neural correlates of motor learning in the vestibulo-ocular reflex: dynamic regulation of multimodal integration in the macaque vestibular system. J Neurosci 30: 10158–10168. Sadeghi SG, Minor LB, Cullen KE (2011). Multimodal integration after unilateral labyrinthine lesion: single vestibular nuclei neuron responses and implications for postural compensation. J Neurophysiol 105: 661–673. Sadeghi SG, Minor LB, Cullen KE (2012). Neural correlates of sensory substitution in vestibular pathways following complete vestibular loss. J Neurosci 32: 14685–14695. Sato F, Sasaki H (1993). Morphological correlations between spontaneously discharging primary vestibular afferents and vestibular nucleus neurons in the cat. J Comp Neurol 333: 554–566. Scarpa A, Gioacchini FM, Cassandro E et al. (2019). Clinical application of cVEMPs and oVEMPs in patients affected by Menie`re’s disease, vestibular neuritis and benign paroxysmal positional vertigo: a systematic review. Acta Otorhinolaryngol Ital 39: 298–307.

54

K.E. CULLEN

Schlack A, Hoffmann KP, Bremmer F (2002). Interaction of linear vestibular and visual stimulation in the macaque ventral intraparietal area (VIP). Eur J Neurosci 16: 1877–1886. Schneider AD, Jamali M, Carriot J et al. (2015). The increased sensitivity of irregular peripheral canal and otolith vestibular afferents optimizes their encoding of natural stimuli. J Neurosci 35: 5522–5536. Schonewille M, Gao Z, Boele H-J et al. (2011). Reevaluating the role of LTD in cerebellar motor learning. Neuron 70: 43–50. Scudder CA, Fuchs AF (1992). Physiological and behavioral identification of vestibular nucleus neurons mediating the horizontal vestibuloocular reflex in trained rhesus monkeys. J Neurophysiol 68: 244–264. Sheliga BM, Yakushin SB, Silvers A et al. (1999). Control of spatial orientation of the angular vestibulo-ocular reflex by the nodulus and uvula of the vestibulocerebellum. Ann N Y Acad Sci 871: 94–122. Shojaku H, Sato Y, Ikarashi K et al. (1987). Topographical distribution of Purkinje cells in the uvula and the nodulus projecting to the vestibular nuclei in cats. Brain Res 416: 100–112. Shojaku H, Barmack NH, Mizukoshi K (1991). Influence of vestibular and visual climbing fiber signals on Purkinje cell discharge in the cerebellar nodulus of the rabbit. Acta Otolaryngol Suppl 481: 242–246. Stefani SP, Serrador JM, Breen PP et al. (2020). Impact of galvanic vestibular stimulation-induced stochastic resonance on the output of the vestibular system: a systematic review. Brain Stimul 13: 533–535. Suh M, Leung HC, Kettner RE (2000). Cerebellar flocculus and ventral paraflocculus Purkinje cell activity during predictive and visually driven pursuit in monkey. J Neurophysiol 84: 1835–1850. Sullivan EV, Rose J, Pfefferbaum A (2006). Effect of vision, touch and stance on cerebellar vermian-related sway and tremor: a quantitative physiological and MRI study. Cereb Cortex 16: 1077–1086. Takahashi M (1990). Head stability and gaze during vertical whole body rotations. Ann Otol Rhinol Laryngol 99: 883–887. Thach WT, Goodkin HP, Keating JG (1992). The cerebellum and the adaptive coordination of movement. Annu Rev Neurosci 15: 403–442. Tokita T, Ito Y, Takagi K (1989). Modulation by head and trunk positions of the vestibulo-spinal reflexes evoked by galvanic stimulation of the labyrinth. Observations by labyrinthine evoked EMG. Acta Otolaryngol 107: 327–332.

Tokita T, Miyata H, Takagi K et al. (2009). Studies on vestibulo-spinal reflexes by examination of labyrinthineevoked EMGs of lower limbs. Acta Otolaryngol 111: 328–332. Ugolini G, Prevosto V, Graf W (2019). Ascending vestibular pathways to parietal areas MIP and LIPv and efference copy inputs from the medial reticular formation: functional frameworks for body representations updating and online movement guidance. Eur J Neurosci 50: 2988–3013. van de Berg R, Ramos A, van Rompaey V et al. (2020). The vestibular implant: opinion statement on implantation criteria for research. J Vestib Res 30: 213–223. Waespe W, Cohen B, Raphan T (1985). Dynamic modification of the vestibulo-ocular reflex by the nodulus and uvula. Science 228: 199–202. Wearne S, Raphan T, Cohen B (1996). Nodulo-uvular control of central vestibular dynamics determines spatial orientation of the angular vestibulo-ocular reflex. Ann N Y Acad Sci 781: 364–384. Wearne S, Raphan T, Cohen B (1998). Control of spatial orientation of the angular vestibuloocular reflex by the nodulus and uvula. J Neurophysiol 79: 2690–2715. Wiboonsaksakul KP, Della Santina CC, Cullen KE (2022). A prosthesis utilizing natural vestibular encoding strategies improves sensorimotor performance in monkeys a prosthesis utilizing natural vestibular encoding strategies improves sensorimotor performance in monkeys. PLoS Biol 20: e3001798. Wilson VJ, Maeda M (1974). Connections between semicircular canals and neck motoneurons in the cat. J Neurophysiol 37: 346–357. Yakusheva TA, Shaikh AG, Green AM et al. (2007). Purkinje cells in posterior cerebellar vermis encode motion in an inertial reference frame. Neuron 54: 973–985. Zobeiri OA, Cullen KE (2022). Distinct representations of body and head motion are dynamically encoded by Purkinje cell populations in the macaque cerebellum. Elife 11: e75018. Zobeiri OA, Mischler GM, King SA et al. (2021a). Effects of vestibular neurectomy and neural compensation on head movements in patients undergoing vestibular schwannoma resection. Sci Rep 11: 517. https://doi.org/10.1038/s41598020-79756-3. Zobeiri OA, Ostrander B, Roat J et al. (2021b). Loss of peripheral vestibular input alters the statistics of head movement experienced during natural self-motion. J Physiol 599: 2239–2254.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00020-0 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 4

Autonomic failure: Clinicopathologic, physiologic, and genetic aspects DAVID S. YOUNGER1,2* 1

Department of Clinical Medicine and Neuroscience, CUNY School of Medicine, New York, NY, United States

2

Department of Medicine, Section of Internal Medicine and Neurology, White Plains Hospital, White Plains, NY, United States

Abstract Over the past century, generations of neuroscientists, pathologists, and clinicians have elucidated the underlying causes of autonomic failure found in neurodegenerative, inherited, and antibody-mediated autoimmune disorders, each with pathognomonic clinicopathologic features. Autonomic failure affects central autonomic nervous system components in the a-synucleinopathy, multiple system atrophy, characterized clinically by levodopa-unresponsive parkinsonism or cerebellar ataxia, and pathologically by argyrophilic glial cytoplasmic inclusions (GCIs). Two other central neurodegenerative disorders, pure autonomic failure characterized clinically by deficits in norepinephrine synthesis and release from peripheral sympathetic nerve terminals; and Parkinson’s disease, with early and widespread autonomic deficits independent of the loss of striatal dopamine terminals, both express Lewy pathology. The rare congenital disorder, hereditary sensory, and autonomic neuropathy type III (or Riley–Day, familial dysautonomia) causes life-threatening autonomic failure due to a genetic mutation that results in loss of functioning baroreceptors, effectively separating afferent mechanosensing neurons from the brain. Autoimmune autonomic ganglionopathy caused by autoantibodies targeting ganglionic a3-acetylcholine receptors instead presents with subacute isolated autonomic failure affecting sympathetic, parasympathetic, and enteric nervous system function in various combinations. This chapter is an overview of these major autonomic disorders with an emphasis on their historical background, neuropathological features, etiopathogenesis, diagnosis, and treatment.

OVERVIEW Historical background The notion that the autonomic nervous system (ANS) plays a key role in regulation of the body’s inner world is not only essentially correct but is a founding concept of modern physiology. In 1865, in his masterpiece, Introduction to the Study of Experimental Medicine, Claude Bernard described his earlier observations on the role of the liver in secreting glucose formed from glycogen stores and how his studies of heat regulation led to the discovery of vascular blood flow regulation by sympathetic nerves (Bernard, 1865). In 1921, Otto Loewi noted

that stimulation of the vagus branch slowed down the heartbeat by means of the release of a vagusstoff or vagal substance. In the same year, the English physiologist John Langley coined the terms autonomic nervous system and parasympathetic nervous system (PaNS) (Langley, 1921). However, the notion that automatic, unconscious, involuntary mechanisms orchestrate the functions of body organs is ancient. The 2nd-century Greek physician Galen, whose teachings dominated medical thought and practice for 14 centuries, viewed the chains of ganglia on each side of the spinal column and nerves emanating from them as tubes distributing the animal spirit in the body, producing consent, or

*Correspondence to: David S. Younger, MD, DrPH, MPH, MS, 333 East 34th Street, Suite 1J, New York, NY 10016, United States. Tel: +1-212-213-3778, Fax: +1-212-213-3779, E-mail: [email protected]

56

D.S. YOUNGER

sympathy, among the organs. This nerve network came to be called the sympathetic nervous system (SNS). Dale and Dudley (1929) found that the active substance in this process was found to be acetylcholine. In the same year, the American physiologist Walter Cannon expanded on the theory of the milieu int
erieur in a series of experiments over a quarter century demonstrating the critical role of adrenaline, the effector compound of the sympathetic adrenergic system maintaining the constancy of the inner world of the body (Cannon, 1929). He introduced the concepts of homeostasis, referring to the stability of the inner world, a direct extension from Bernard’s notion of the milieu int
erieur, which according to Cannon (1929) was coordinated by the brain. He also described fight-or-flight responses asserting that blood loss from trauma and even psychological emergencies evoked release of adrenaline into the bloodstream. Adrenaline relaxes blood vessels in skeletal muscle increasing local blood flow important in providing metabolic fuels to exercising muscle and in removing metabolic waste products, while constricting cutaneous blood vessels and promoting clotting to minimize blood loss from lacerations. It releases glucose from the liver into the bloodstream from the breakdown of glycogen and stimulates respiration, maximizing delivery of oxygen from the lungs into the bloodstream. From a psychological point of view, adrenaline intensifies emotional experiences (Schachter and Singer, 1962) by its antifatigue and energizing effects. Finally, Cannon theorized the sympathoadrenal system and the release of a substance different from adrenaline, which was later found to be the adrenaline precursor, norepinephrine (noradrenaline) (von Euler, 1946).

Components of the autonomic nervous system The ANS can be conceptualized as having five components: the sympathetic noradrenergic system, the sympathetic cholinergic system, the parasympathetic cholinergic system, the sympathetic adrenergic system, and the enteric nervous system (ENS) (Goldstein, 2006). Evidence has accumulated for differential noradrenergic and adrenergic responses in various situations. The largest sympathetic adrenergic system responses are seen when the organism encounters stressors that pose a global or metabolic threat.

SYMPATHETIC NORADRENERGIC SYSTEM The sympathetic noradrenergic system plays a dominant role in regulation of the circulation by the brain, not only during emergencies but in continual activities of daily living (ADLs) such as standing erect, in exercise, adjusting ingestion of meals and thermoregulation.

Sympathetic noradrenergic nerves enmesh arterioles throughout the body, and since their caliber determines total peripheral resistance to blood flow, sympathetic noradrenergic innervation of the smooth muscle cells in arteriolar walls represents a focal point in neural regulation of blood pressure. In the heart, sympathetic noradrenergic nerves form lattice-like networks around myocardial cells and also supply coronary arterial vessels. Because of the close architectural association between the sympathetic nerves and myocardial and arteriolar smooth muscle cells, lesions of the sympathetic noradrenergic system manifest clinically with dysregulation of cardiovascular performance. Indeed, orthostatic hypotension (OH) seen as the fall in blood pressure upon the erect stance is a cardinal manifestation of sympathetic noradrenergic failure. Through mechanisms that are poorly understood, neurogenic OH may be accompanied by supine hypertension (Biaggioni and Robertson, 2002). Failure of sympathetic noradrenergic outflows cephalad is accompanied by ptosis and miosis, as in the Horner syndrome, while failure of sympathetic noradrenergic cardiovascular outflows manifests as postprandial hypotension, inability to tolerate extremes of temperature, and exercise intolerance.

SYMPATHETIC CHOLINERGIC SYSTEM The sympathetic cholinergic system mediates sweating and failure of sympathetic cholinergic outflow results in anhidrosis, which in association with ptosis and miosis forms the Horner syndrome triad. In autonomic failure from inability to synthesize norepinephrine, the patients have severe neurogenic OH from sympathetic noradrenergic failure, while sweating remains normal due to intact sympathetic cholinergic function. Analogous neurotransmitter specificity occurs in Parkinson’s disease (PD) associated with OH, in which sympathetic noradrenergic denervation occurs without evidence of sympathetic cholinergic denervation (Sharabi et al., 2003). Small fiber neuropathy and postural tachycardia syndrome, often involve locally decreased sweating (Peltier et al., 2010), generally without evidence of sympathetic noradrenergic dysfunction. In pandysautonomia associated with autoimmune autonomic ganglionopathy (AAG), there is both OH as a result of sympathetic noradrenergic failure and anhidrosis due to sympathetic cholinergic failure.

PARASYMPATHETIC CHOLINERGIC SYSTEM The main nerve of the parasympathetic cholinergic system or PaNS, the vagus nerve, innervates the myocardium, splanchnic organs, and most of the gastrointestinal tract. Stimulation of parasympathetic cholinergic outflows increases salivation and lacrimation and evokes pupillary constriction. Vagal tone stimulates gastric acid secretion

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS and gut smooth muscle contraction, augments urinary bladder tone, and contributes to penile erection with associated slowing of heart rate and decreased atrioventricular node electrical conduction. Parasympathetic cholinergic system failure manifests as dry mouth and eyes (sicca syndrome), urinary retention, and constipation. Since the vagus nerve innervates the ENS, which is a local nerve network in walls of the gastrointestinal tract, it is difficult to clinically distinguish parasympathetic cholinergic denervation and enteric denervation especially as determinants of constipation, abdominal bloating, and esophageal reflux.

SYMPATHETIC ADRENERGIC SYSTEM Through the circulatory secretion of adrenaline, the sympathetic adrenergic system constitutes the main hormonal component of the ANS. Because of its potency however, the plasma of healthy humans at rest contains remarkably low levels of adrenaline, as little as a picograms per milliliter (picomolar). The technical problem of measuring levels of adrenaline directly in the bloodstream probably impeded accumulation of scientific understanding about involvement of the sympathetic adrenergic system in pathophysiological states. It is one of the three main hormones responsible for regulation of blood glucose levels, the other two being insulin and glucagon. Sympathetic adrenergic system failure is associated with a tendency toward hypoglycemia; however, the redundancy of effector systems results in few if any clinical manifestations, unless the other effector systems are inactivated. Soon after adequately sensitive assay methods of plasma levels of norepinephrine and adrenaline became available, evidence rapidly accumulated for the differential noradrenergic and adrenergic responses in various situations (Robertson et al., 1979). The largest sympathetic adrenergic system responses are seen in hemorrhagic shock, insulin-induced hypoglycemia and or cardiac arrest (Pacak et al., 1998) in which there are proportionate increments in adrenomedullary secretion that generally exceed increments in sympathetic noradrenergic outflow. The importance of sympathetic adrenergic and the hypothalamic–pituitary–adrenocortical (HPA) system has emerged with increased understanding of its components. Steroid-producing cells of the adrenal cortex and catecholamine-producing chromaffin cells of the adrenal medulla have until recently been regarded as independent. The cortex and medulla actually have multiple contact areas, and because of the cortical-medullary direction of blood flow in the adrenal gland, adrenomedullary chromaffin cells are bathed continuously in blood containing high concentrations of adrenocortical steroids. Adrenal glucocorticoids and corticosteroids (CS)

57

are trophic for phenylethanolamine-N-methyltransferase, the enzyme catalyzing synthesis of adrenaline from norepinephrine (Wurtman and Axelrod, 1965) while catecholamine biosynthesis seems trophic for adrenocortical cells (Bornstein et al., 2000). Corticotropin-releasing hormone (CRH) or corticotropin-releasing factor plays key roles in activity of the HPA system and the sympathetic adrenergic system. Experimental treatment with a CRH receptor antagonist attenuates stress-induced adrenaline responses (Brown et al., 1985), while deficiency of CRH reduces decreased basal and restraint-induced adrenaline levels (Jeong et al., 2000). Moreover, adrenalectomy activates the sympathetic noradrenergic system and adrenalectomized monkeys show markedly decreased plasma adrenaline concentrations but accentuated plasma norepinephrine responses to subsequent cholecystectomy, the magnitude of which is inversely related to the dose of cortisol administered preceding the second surgery (Udelsman et al., 1987). Norepinephrine fails to normalize completely even at the high dose of corticosteroid replacement, calculated to be similar to the cortisol production rate during maximal stress. Finally, prednisone administered daily over 2 weeks decreases directly recorded skeletal muscle sympathetic nerve activity and plasma norepinephrine levels, decreasing sympathetic noradrenergic outflows in humans (Golczynska et al., 1995). A meta-analysis of publications describing original data regarding plasma adrenaline, norepinephrine, and corticotrophin (ACTH) levels measured before and during or after exposure to stressors, in humans and in laboratory animals (Goldstein and Kopin, 2008) found justification for the concept of coordinated adrenocortical– adrenomedullary responses in stress, involving CRH driving adrenocortical and adrenomedullary outflows, as well as interactions between adrenocortical and adrenomedullary chromaffin cells.

Parasympathetic and sympathetic outflow While the terms autonomic failure, dysautonomia, and autonomic dysfunction imply the existence of a single disease entity, in reality, disorders of the ANS instead relate to disturbances of functionally and neurochemically distinct components, as reflected in differential responses to stressors resulting in finite pathophysiological different states. The ANS should not be regarded as a system merely carrying out the commands of the brain or a reflex circuit. It is infinitely more sophisticated employing the feedback of other organs to change, and precisely adapt its output by adjusting to a given physiological state of the body. These outputs are modulated by input from higher neural centers located in many different areas in the brainstem, hypothalamus, and even prefrontal cortex, which contain preautonomic neurons that

58

D.S. YOUNGER

provide input to autonomic motor neurons (MNs). By following the physiology and pathophysiology, one can appreciate the specialization within the PaNS and SNS, not so much in its targets but in the function, it supports as driven by the brain. Consequently, it is proposed that the ANS with a diverse output, must be controlled by multiple areas within the brain directing the body in separate compartments and different functions rather than in a singular vertical hierarchical fashion.

SYMPATHETIC ORGANIZATION AND OUTPUT The central control of the outflow of SNS is organized as a long chain of MNs in the intermediolateral (IML) column of the spinal cord. As a consequence of the segmental organization, the ACh-producing motor neurons reach different and multiple ganglions along the spinal cord. Neurons in these ganglions have the capacity to elaborate different neurotransmitters, of which noradrenaline/norepinephrine is the main one, often together with cotransmitters such as neuropeptide Y (Lundberg et al., 1983). Interestingly, the target structures of the ANS, although as diverse as the brain and the kidney, are very often the smooth muscles around the blood vessels within these organs. This allows the ANS, by vasoconstriction and vasodilatation, to control the blood supply. The segmental organization of the SNS allows topographical signal distribution such that the superior cervical ganglion (SCG) serves not only to provide the cranium with sympathetic innervation, it also supplies part of the sympathetic innervation to the heart. Despite its segmental distribution, it cannot be concluded that a

single segment or ganglion provides a unique outgoing signal. Rather, on the basis of the differences in activity and control in the hierarchy of segmentation, an examination of branches of the SNS suggests that it may be controlled by separate autonomic MNs and separate brain networks. This is illustrated in the projections of hypothalamus from the suprachiasmatic nucleus (SCN) and the paraventricular nucleus (PVN) that maintain sympathetic–parasympathetic separation whether to preganglionic sympathetic neurons in the IML column of the spinal column or to preganglionic neurons of the dorsal motor nucleus of the vagus (DMV), or via axonal collaterals from presympathetic PVN neurons to preparasympathetic neurons (Fig. 4.1). The involvement of similar hypothalamic areas in the control of different organs provokes the question of whether they are influenced by a general drive from the hypothalamus, or via dissimilar neurons projecting exclusively on motor neurons involved in sending signals to a particular organ.

PARASYMPATHETIC ORGANIZATION AND OUTPUT In contrast to the SNS, which mediates stimulus-specific patterns of responses affecting multiple effectors, the PaNS mediates reflexes activated in an organ-specific fashion via the release of ACh from its nerve terminals, which is especially accurate when intermediate ganglions are involved in the output of parasympathetic motor nuclei. For the transmission of information from parasympathetic ganglia, it is the case that, unlike noradrenaline, ACh exhibits a very short half-life of 1–2 ms, due to the presence of acetylcholinesterase,

Fig. 4.1. Scheme of interaction between the hypothalamic suprachiasmatic nucleus (SCN) and the paraventricular nucleus (PVN). Separate sympathetic (red) or parasympathetic (blue) neurons of the SCN project to preautonomic neurons of the PVN, where a similar sympathetic–parasympathetic separation can be observed. Preautonomic neurons of the PVN project either to the preganglionic sympathetic neurons in the intermediolateral (IML) column of the spinal cord, or to the preganglionic neurons of the dorsal motor nucleus of the vagus (DMV). The presympathetic PVN neurons have axon collaterals to preparasympathetic neurons, either in the PVN itself, or in the nucleus tractus solitarius. Reproduced from Buijs RM (2013). The autonomic nervous system: a balancing act. Handb Clin Neurol 117: 1–11 with permission.

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS which hydrolyzes ACh at a high rate. Close contact between cholinergic nerve terminals and cells expressing ACh receptors is required for the cholinergic control of these cells. The DMV and the NA are the two nuclei in the brainstem from which the parasympathetic output to most organs arises, either directly or via an intermediate ganglion. Considering the direct input from these two parasympathetic motor nuclei, it is important to keep in mind that ACh is the neurotransmitter in just a part of these neurons, since some of these colocalize with dopamine. The differentiation of function within the DMV has been more convincingly demonstrated than a topographic specialization as concerns the distribution of gastric MNs that innervated different parts of the stomach (Hayakawa et al., 2003).

AUTONOMIC CONTROL OF BLADDER, BOWEL AND SEXUAL FUNCTION The storage and voiding of urine are performed by the coordinated function of the bladder, smooth muscle urethral sphincter, and striated muscle urethral sphincter. The bladder wall, formed by three layers of interdigitating detrusor smooth muscle is comprised of an internal sphincter that is not anatomically isolated, but functions as a physiological sphincter. Reflex bladder contractions are activated by sympathetic, parasympathetic, and somatic nerves from the spinal cord. Three sets of peripheral nerves, each with efferent and afferent components, innervate the bladder, urethra, and associated striated muscle of the urethra and pelvic floor. Sympathetic stimulation typically includes contraction of sphincter muscle, relaxation of smooth muscle in the wall of hollow viscera, and constriction of blood vessels, while parasympathetic stimulation results instead in relaxation of sphincter muscles and contraction of smooth muscles of hollow viscera. Parasympathetic innervation of blood vessels in erectile tissue of the genitals via pelvic splanchnic nerves cause vasodilatation of vessels in erectile tissue upon stimulation. Visceral afferent fibers supplying the visceral peritoneum, pelvic organs, and vasculature transmit dull, aching, poorly localized pain via sympathetic afferent fibers, whereas bladder distention, rectal fullness, urge to void or defecate, and sexual sensations conveyed via parasympathetic afferent fibers. Parasympathetic efferent parasympathetic nerves originating in the intermediolateral cell column of the sacral spinal cord from the S2 to S4 segments, exit the spinal cord through the anterior roots, intermingle with somatic efferent fibers forming the spinal nerves, and continue in anterior primary rami reaching the pelvic splanchnic nerves to ganglion cells in the pelvic plexus, subsidiary vesical plexus, cavernous nerves, bladder wall and urethra. Sympathetic nerves supplying the bladder originate

59

in the intermediolateral nuclei of the thoracic spinal cord from the T11 to L2 segments, and exit the spinal cord through the anterior roots, intermingled with somatic efferent fibers forming spinal nerves and synapse in a nearby paravertebral ganglia in the sympathetic chain. Those destined for the bladder and urethra pass directly through paravertebral ganglia and exit as preganglionic visceral or splanchnic nerves synapsing on one of the prevertebral or collateral ganglia on the anterior aspect of the aorta or internal iliac vessels such as the inferior mesenteric ganglia before continuing inferiorly as the right and left hypogastric nerves to the pelvic plexus. The latter give rise to subsidiary plexi and nerves including the vesical plexus to the bladder and urethra, and cavernous nerves to the urethral sphincter complex. Activation of postganglionic sympathetic fibers lead to excitation of the bladder base and urethra and inhibit detrusor muscle located in the bladder dome, while and bladder parasympathetic ganglia neurotransmission facilitates urethral smooth muscle contraction. Striated muscle of the external urethral and external anal sphincters is supplied by pudendal nerve branches that originate in a motor neurons of Onuf’s nucleus in the anterior horn of sacral spinal cord segments from S1 to S3. The pudendal nerve exits the pelvic cavity below the piriformis via the greater sciatic foramen and reenters the pelvic cavity running along the lateral wall of the ischiorectal fossa in the pudendal canal supplying three major branches, one each to the dorsal nerve of the penis or clitoris for cutaneous supply; the inferior rectal nerve which supplies the lower anal canal, external anal sphincter and skin around the anus; and the perineal nerve which divides into a cutaneous branch to scrotum and ventral penis in men, or posterior labia and lower vagina in women, and distal urethra, with muscular branches to the superficial and deep perineal muscles including the external urethral sphincter and levator ani. Penile erection is mediated via parasympathetic impulses that initiate the necessary penile vascular changes. In women, parasympathetic activity increases vaginal secretions in association with clitoral swelling. Sympathetic nerves supply the vas deferens, seminal vesicle, prostate, and bladder vesicle neck. The sympathetic efferent fibers in men constrict the proximal bladder neck to retrograde flow and induce the ejaculation of semen through rhythmic smooth muscle contraction, while in women it induces contraction of genital smooth muscle during orgasm.

NEUROPATHOLOGIC CHARACTERIZATION The neuropathologic characterization of normal sympathetic ganglia has been complicated by the necessity of obtaining specimens from autopsy studies and apparently

60

D.S. YOUNGER

normal ganglia surgically to improve limb ischemia, resulting in inherent difficulty in defining normalcy.

Sympathetic ganglia Sympathetic ganglia are enclosed in a thin collagenous capsule continuous with the epineurium of afferent and efferent nerve trunks. Principal sympathetic neurons in the human SCG are distributed in large groups accompanied by large nerve fascicles. The SCG, the most rostral and largest of the paravertebral chain ganglia, is distributed to various cephalic structures. The prevertebral plexiform ganglia including the celiac, superior and inferior mesenteric (IMG and SMG) are represented by clumps of neurons intermixed with bundles of axons. Neurons in both prevertebral and paravertebral sympathetic ganglia range from 20 to 60 mm in diameter and

typically contain copious amounts of cytoplasm, poorly differentiated Nissl bodies, vesicular nuclei and prominent nucleoli (Fig. 4.2). The sympathetic neurons are typically surrounded by a thin capsule of flattened satellite cells which extend along proximal dendrites and enclose synapses. Approximately half of the SCG and SMG in one large autopsy series (Schmidt, 1996) demonstrated perivascular and parenchymal lymphocytic infiltrates (Fig. 4.3) that neither invaded the satellite cell capsule nor accompanied neuronal degeneration or any of the 15 examined disease states, and were thus correlated instead with increasing age, and of uncertain significance. Pathologic studies of autonomic dysfunction that show lymphocytic ganglionic infiltrates related to a given primary disease process should be individualized and always related to age-matched controls.

Fig. 4.2. Normal sympathetic neurons. Principal sympathetic neurons are separated by intraganglionic neuropil (Toluidine blue,
300). Reproduced from Bruch LA, Schmidt RE (2000). Pathology of the autonomic nervous system. Handb Clin Neurol 75: 1–52 with permission.

Fig. 4.3. Perivascular cuffing of sympathetic ganglia. Perivascular lymphocytes are seen cuffing (arrows) at postmortem in the vicinity of sympathetic ganglia of a normal individual (Hematoxylin and eosin,
150). Reproduced from Bruch LA, Schmidt RE (2000). Pathology of the autonomic nervous system. Handb Clin Neurol 75: 1–52 with permission.

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

Parasympathetic ganglia Brainstem projections and peripheral ganglia form the cranial portion of the PaNS. A population of axons travel in cranial nerve (CN) III from the brainstem Edinger– Westphal (EW) nucleus to synapse in the ciliary ganglion, which contains predominantly cholinergic neurons, some of which also contain VIP and NO. The parasympathetic portion of the CN VII originates in the superior salivatory nucleus of the brainstem and supplies the submandibular and sphenopalatine ganglia whose neurons contain ACh, NO synthase, as well as neuropeptides VIP and peptide histidine methionine (PHI). The neurons of the otic ganglion, innervated by the CN IX are also cholinergic with few colocalized neuropeptides. The parasympathetic portion of CN X, and its outflow in the vagus nerve, that arises from the DMV, nucleus tractus solitarius (NTS), spinal trigeminal nucleus and the NA, provides a widely distributed innervation of the viscera with axons immunoreactive for a variety of neurotransmitters and neuropeptides, as do parasympathetic vasodilator axons. The vagus nerve with both afferent and efferent components, is critical in triggering autonomic reflexes. The afferent vagus provides inputs to the NTS and convey information from cardiovascular, gastrointestinal, and respiratory receptors. The DMV receives inputs from the NTS and mediates all vagovagal reflexes controlling gastrointestinal motility and secretion via activation of neurons of the myenteric and the submucosal plexus of the ENS. Different subgroups of DMV neurons provide parallel excitatory and inhibitory pathways controlling gastrointestinal motility (Chang et al., 2003). The excitatory vagal motor pathway innervates myenteric neurons that utilize ACh and stimulate contraction of smooth muscle; the inhibitory pathway is mediated by postganglionic myenteric neurons that utilize nitric oxide (NO), vasoactive intestinal polypeptide, or adenosine triphosphate (Grundy and Schemann, 2007). Vagal influence is most prominent in the esophagus and stomach but is less important in the small bowel and colon, where motility and secretion are primarily controlled by local ENS reflexes. The sacral preganglionic output originates in neurons located in the lateral gray matter at the S2–S4 segments of the sacral spinal cord. These neurons are critical for normal micturition, defecation, and sexual organs; this involves their coordinated interactions with both lumbar sympathetic neurons located at T12–L2 levels and MNs of Onuf’s nucleus, located in the ventral horns of the sacral spinal cord. The MNs of Onuf’s nucleus innervate striated voluntary muscles of the pelvic floor and are histologically and biochemically comparable to the other somatic spinal MNs at the S2–S4 levels innervating the external urinary sphincter and

61

pelvic floor. The best characterized example of this coordinated interaction is the control of urine storage and micturition. Urine storage depends on guarding reflexes mediated by lumbosacral sympathetic neurons (which inhibit contraction of the detrusor and promote contraction of the bladder neck) and Onuf’s nucleus motoneurons that elicit contraction of the external urethral sphincter. Normal micturition involves a spinobulbospinal reflex that is triggered by bladder afferent inputs that reach the periaqueductal gray (PAG) and are then conveyed to the pontine micturition center (PMC) (Holstege, 2005). During voluntary voiding, there is withdrawal of prefrontal inhibition on the PAG, which activates the PMC or Barrington’s nucleus; descending projections directly activate the sacral preganglionic neurons (promoting detrusor contraction) and, via interneurons, inhibit the sphincter motoneurons of Onuf’s nucleus (allowing relaxation of the external sphincter).

The aging autonomic nervous system Dysfunction of the aging ANS is thought to underlie agerelated abnormalities in maintenance of blood pressure and cardiovascular reflexes, gastrointestinal function, pupillary kinetics, thermoregulation, and sudomotor responses (Schmidt, 1991; Low, 1997). Age-related autonomic dysfunction may result in clinical symptoms directly, subclinical disease, or a decrease in the safety margin upon which additional insults may be superimposed. A 5%–8% loss of IML neurons per decade was suggested by Low et al. (1977) in studies of myelinated fiber loss in the greater splanchnic nerve, (which provides preganglionic input to the celiac and SMG) in aged humans. Although clinical autonomic studies suggest substantial abnormalities in PaNS function with aging, particularly altered control of heart rate, systematic studies of the vagus nerve and cranial parasympathetic nuclei have not been performed in aged human subjects. Rather, the impact of aging on the ANS is confined mainly to systematic neuropathological studies in the SNS. Early studies (Botar, 1956) of aging human sympathetic ganglia found that sympathetic neuron numbers and the size of their perikaryal decreased with age and that the majority of residual aged celiac ganglia neurons were actively degenerated. However, more recent studies have not found compelling evidence for a decrease in neuronal density as measured by neuron number/mm2 or ganglionic surface area or cell body size with age (J€arvi et al., 1988; Schmidt, 1996). Both sympathetic perikaryal catecholamines and neuromelanin content, representing the normal accumulation of a by-product of catecholamine metabolism, were reported to fall with increasing age. However, the hallmark pathological alternation in aging prevertebral sympathetic celiac

62

D.S. YOUNGER

ganglia and SMG, but not paravertebral SCG, was the accumulation of markedly enlarged dystrophic axons typically located adjacent to the perikarya of principal sympathetic neurons or their primary dendrites (Schmidt et al., 1993). A pathological process selectively targeting synapses in prevertebral sympathetic ganglia likely contributes to the loss of integrated reflexes (Bruch and Schmidt, 2000). Although studies of unmyelinated axons in sural nerve of aged human subjects typically have not separated sympathetic axons from nonautonomic axons, there is evidence of unmyelinated axon loss and increase in the numbers of small diameter, unmyelinated axons (Ochoa and Mair, 1969), suggesting a role for intact regenerating autonomic small fibers.

Baroreflexes, autonomic, and baroreceptor failure Kaufmann and colleagues have described the functional anatomy underlying baroreceptor dysfunction (Kaufmann et al., 2020). Baroreflexes enable the circulatory system to adapt to varying conditions in daily life while maintaining blood pressure, heart rate, and blood volume within a narrow physiologic range. Baroreceptors embedded in the walls of major arteries and veins and the heart elicit distinct reflexes (Hainsworth, 2014). They continuously signal to the NTS through the vagus and glossopharyngeal nerves which are activated by stretch when blood pressure, blood volume, or both rises. To counter the rise, baroreceptors evoke reflex inhibition of efferent sympathetic signals to splanchnic, skeletal muscle, and renal blood vessels, causing vasodilatation. A concomitant increase in parasympathetic nerve traffic to the sinoatrial node slows the heart rate. Conversely, when a change to a standing position is made, baroreceptors are unloaded, allowing vasoconstriction and tachycardia to buffer the fall in blood pressure that would otherwise occur. Arterial baroreceptors in the carotid sinuses and aortic arch sense pressure changes, and cardiopulmonary baroreceptors in thoracic veins and the heart sense changes in blood volume. Both arterial and cardiopulmonary baroreceptors inhibit efferent sympathetic neurons, leading to vasodilatation, but only arterial baroreceptors influence the heart rate. Arterial baroreceptors preferentially target the splanchnic circulation, and cardiopulmonary baroreceptors inhibit sympathetic renal outflow, reducing renin release and proximal tubular sodium reabsorption. Baroreceptor activation also suppresses vasopressin release and sodium appetite, increasing urine output. Mechanosensing by arterial baroreceptors is mediated by the Piezo type mechanosensitive ion channel component 1 and 2 (PIEZO1, PIEZO2) mechanically activated excitatory ion channels (Zeng et al., 2018).

At rest, afferent baroreceptor discharge on the nucleus of the solitary tract maintains a tonic level of peripheral sympathetic inhibition and cardiovagal activation. In synchronicity with the pulse wave, afferent baroreceptor discharge increases with each systole and decreases during diastole, causing reciprocal changes in sympathetic and vagal efferent activity. Vagal efferent neurons also entrain with respiratory neurons and are inhibited during inspiration, giving rise to respiratory sinus arrhythmia. When the baroreflex pathways are damaged, these physiologic rhythms are lost or blunted. The nucleus of the solitary tract also receives and integrates information from other sources, including peripheral and central chemoreceptors, renal mechanoreceptors and chemoreceptors through renal afferent nerves, muscle afferents that are stimulated by muscle work and respiratory neurons, as well as cortical and hypothalamic neurons. Diseases affecting baroreflex neurons cause unstable blood pressure with acute symptoms of hypoperfusion or hyperperfusion. Lesions of afferent, central, or efferent baroreflex neurons result in distinct but overlapping cardiovascular phenotypes. Clinicians use the term autonomic failure (specifying neurologic or neurogenic), when referring to diseased efferent baroreflex neurons because other autonomic fibers are also frequently affected, impairing bladder, gastrointestinal, and sexual function. Baroreflex failure commonly refers to compromised afferent neurons, which affect cranial nerves IX and X but cause no additional autonomic deficits. Diseases affecting baroreflex sympathetic efferent neurons impair the release of norepinephrine at the neurovascular junction. Insufficient vasoconstriction on standing or exertion leads to OH and symptoms of organ hypoperfusion, including lightheadedness or dizziness, visual blurring, and syncope. Dyspnea, subtle cognitive slowing, and fatigue are common and disappear in the supine position. Supine hypertension develops in 50% of patients with efferent baroreflex failure, probably as a result of the activation of residual sympathetic fibers and denervation supersensitivity. Impaired release of norepinephrine may be the result of disorders affecting peripheral postganglionic sympathetic neurons or the preganglionic and premotor neurons in the spinal cord and brainstem that activate them. Although the lesions are different, the severity of OH is similar. The supine plasma norepinephrine levels tend to be low in patients with postganglionic lesions but normal in patients with preganglionic or premotor sympathetic lesions. In patients with postganglionic lesions, the pressor response to adrenergic agents is exaggerated because of adrenergic denervation supersensitivity, which is probably caused by an increased number of receptors. Conversely, in preganglionic or premotor lesions, sympathetic postganglionic neurons are spared but are

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS disconnected from central influences. In preganglionic but not postganglionic lesions, norepinephrine-reuptake inhibitors significantly increase norepinephrine levels at the neurovascular junction and raise blood pressure. The two most common synucleinopathies, Parkinson’s disease and Lewy body dementia (LBD), have a predominantly postganglionic phenotype. Multiple system atrophy (MSA) is rare but much more aggressive, and it has a preganglionic phenotype, with postganglionic sympathetic neurons largely spared but compromised connections between the nucleus of the solitary tract and the hypothalamic nuclei regulating vasopressin. Baroreflex dysfunction can be the initial presentation of all synucleinopathies and may allow for early diagnosis, before the appearance of typical motor or cognitive deficits. Prospective studies (Kaufmann et al., 2017) show that for patients with isolated efferent baroreflex failure (i.e., pure autonomic failure, PAF), the cumulative risk of a future diagnosis of PD, LBD, or MSA is 10% per year. Moreover, prospective, population-based studies of healthy persons have shown that a decrease in heart rate variability or peak exercise heart rate carries an increased risk of a later diagnosis of PD (Palma et al., 2013; Alonso et al., 2015). It is suspected that these persons already had a synucleinopathy, solely affecting autonomic neurons at the time.

NEURODEGENERATIVE AUTONOMIC FAILURE Involvement of central and peripheral autonomic components The central autonomic network includes the insular cortex, anterior cingulate cortex, amygdala, hypothalamus, PAG, parabrachial nucleus (PBN), nucleus of the solitary tract, ventrolateral reticular formation of the medulla, and medullary raphe. These areas are reciprocally interconnected and receive converging visceral and somatosensory information, and generate stimulus-specific patterns of autonomic, endocrine, and motor responses (Fig. 4.4). These areas not only control the sympathetic and parasympathetic outflows but also affect endocrine and motor outputs as components of integrated physiological responses. Although the functional anatomy of this central autonomic network was first characterized in experimental animals, many of these areas are clearly activated during autonomic responses in humans. The forebrain regions that are directly or indirectly involved in high-level control of autonomic functions include the insular cortex, anterior cingulate cortex, amygdala, and several areas of the hypothalamus. The brainstem areas controlling the sympathetic and parasympathetic outputs include the PAG matter of the midbrain, the PBN and adjacent areas of the pons, several medullary regions,

63

including the nucleus of the solitary tract (NTS), DMV, ventrolateral reticular formation of the medulla, and medullary raphe. Spinal cord elements involved in sympathetic outflow specifically include the IML cell column at vertebral levels T1–L3 with cell bodies residing in lamina VII in several well-defined nuclei including the nucleus dorsalis (Clarke’s column) and IML nuclei. Autonomic neurons of the spinal cord give rise to preganglionic sympathetic general visceral nerves that synapse along bilaterally symmetric sympathetic chain ganglia, also called the paravertebral ganglia, located ventral and lateral to the spinal cord extending from the upper neck to the coccyx. Each ganglion within the paravertebral this chain is either cervical, thoracic, lumbar, or sacral. Postganglionic fibers extend to an effector including the skin, along blood vessels, and to visceral organ in the thoracic cavity, abdominal cavity, and pelvic cavities. Although postganglionic neurons in both divisions of the ANS express nicotinic AChRs to receive signals from preganglionic neurons, postganglionic neurons in the PaNS division are cholinergic. By contrast, postganglionic sympathetic fibers are mostly adrenergic utilizing epinephrine and norepinephrine function as the primary neurotransmitters. Notable exceptions to this rule include the sympathetic innervation of sweat glands where the neurotransmitter at both pre- and postganglionic synapses is acetylcholine; and adrenal medulla, where chromaffin cells function as modified postganglionic nerves that release catecholamines into the blood stream as hormones instead of a synaptic cleft. Like other components of the SNS, both of these exceptions are still stimulated by cholinergic preganglionic fibers. Several components of the central autonomic networks are affected in neurodegenerative disorders and characterized by the presence of intracellular inclusions. They include Lewy bodies and Lewy neurites composed of aggregated and misfolded proteins which are a feature of a-synucleinopathies and the associated degenerative nigrostriatal areas in PD, cortical regions of DLB, and diffusely in the neuraxis and peripheral sympathetic tissues in PAF. Such disorders affected by Lewy body and neuronal loss in different components of the central autonomic network including preganglionic neurons and their target autonomic ganglia are associated with autonomic failure manifested by OH, neurogenic bladder, erectile failure, gastrointestinal dysmotility, and impaired thermoregulation. While there are clinical features that can help in the differentiation among them, the clinical picture does not always predict the underlying neuropathology. By comparison, MSA is characterized by the presence of cytoplasmic inclusion in the oligodendrocytes (GCIs) and to a lesser extent, neurons. The clinical, pathophysiological, and biochemical features of PAF, MSA and PD have been tabulated by Garland and Robertson (2009) (Table 4.1).

64

D.S. YOUNGER

Fig. 4.4. Central autonomic control areas. Reproduced from Cersosimo MG, Benarroch EE (2013). Central control of autonomic function and involvement in neurodegenerative disorders. Handb Clin Neurol 117: 45–57 with permission.

Table 4.1 Best clinical criteria for the diagnosis of multiple system atrophya Sporadic adult onset Dysautonomia Parkinsonism Corticospinal tract signs Cerebellar signs Absent levodopa response Absent cognitive dysfunction Absent supranuclear gaze palsy Adapted from Litvan I, Booth V, Wenning G et al. (1998). Retrospective application of a set of clinical diagnostic criteria for the diagnosis of multiple system atrophy. J Neural Transm 105: 217–227. a Different clinical criteria.

Consensus statement and definitions A 1995 meeting of the Consensus Committee of the American Autonomic Society (AAS) and the American Academy of Neurology (AAN) published a statement on the definition of OH, PAF, and MSA.

ORTHOSTATIC HYPOTENSION Orthostatic hypotension was defined as a reduction of systolic blood pressure (SBP) of at least 20 mm Hg or diastolic blood pressure of at least 10 mm Hg within 3 min of standing. It is a physical sign and not a disease. An acceptable alternative to standing is the demonstration of a similar drop in blood pressure within 3 min, with head-up tilting (HUT) at an angle of 60 degrees. The test

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS should be conducted before food ingestion, early in the day adequately hydrated at a normal ambient temperature apart from recent recumbency, postural deconditioning, uncontrolled hypertension, holding potentially disruptive medications. Symptoms evoked during HUT should resolve in resuming the recumbent position and may include lightheadedness, dizziness, blurred vision, weakness, fatigue, cognitive impairment, nausea, palpitations, tremulousness, headache, and neck ache. A patient with suggestive symptoms of, but undocumented OH should have repeated measurements of blood pressure and extend the period of observation for up to 10 min.

PURE AUTONOMIC FAILURE The Consensus Committee defined PAF as an idiopathic and sporadic disorder characterized by OH and reduced supine plasma norepinephrine levels, usually with evidence of more widespread autonomic failure in the absence of other neurological features. Some patients with manifestations of PAF can later prove to have other disorders such as MSA.

PARKINSON’S DISEASE WITH AUTONOMIC FAILURE The Consensus Committee noted that a minority of patients with PD as defined by United Kingdom Parkinson’s Disease Brain Bank criteria (Hughes et al., 1992b) may develop autonomic failure, including OH. It was not known if these patients have a more serious prognosis than PD without autonomic failure.

MULTIPLE SYSTEM ATROPHY The Consensus Committee defined MSA as a sporadic, progressive, adult-onset disorder characterized by autonomic dysfunction, parkinsonism (bradykinesia with rigidity or tremor or both often with a poorly sustained motor response to chronic levodopa therapy) and cerebellar signs often in combination with corticospinal tract signs; and OH, with frequent impotence, urinary incontinence or retention. Autonomic involvement typically precedes or occurs within 2 years of the onset of CNS motor symptoms. Characteristically, these features cannot be explained by medications or other disorders. A number of terms have been related to differing clinical phenotypes of MSA including striatonigral degeneration (SND) for patients with parkinsonian features; sporadic olivopontocerebellar atrophy (OPCA) for predominant cerebellar features, and Shy–Drager syndrome (SDS) for predominant autonomic failure. Such clinical manifestations may occur in various combinations and evolve over time.

65

Clinical and laboratory autonomic assessment AUTONOMIC SYMPTOM ASSESSMENT AND DISABILITY The severity of symptoms of OH and its impact on ADLs can be assessed using the Orthostatic Hypotension Questionnaire, a 10-item patient-reported outcome (Kaufmann et al., 2012) and composite autonomic symptom score 31 (COMPASS-31) (Sletten et al., 2012) with higher values representing more severe symptoms. It was distilled from the well-established Autonomic Symptom Profile questionnaire. The degree of disability can be rated using the Unified Multiple System Atrophy Rating Scale (UMSARS) disability domain (part IV) (Wenning et al., 2004). Several tests can help characterize peripheral and central autonomic disorders respectively seen in PAF and MSA and PD including, the quantitative sudomotor axon reflex test (QSART), thermoregulatory sweat test (TST), cardiovagal tests (including heart rate response to deep breathing [HRRDB]), Valsalva ratio (VR) and vascular response to HUT test, supine and standing catecholamines, cardiac sympathetic imaging including 123I-metaiodobenzylguanidine (123I-MIBG) scan, magnetic resonance imaging, biopsy findings, and electromyography and nerve conduction studies (NCSs) (Low and Sletten, 2008).

QUANTITATIVE SUDOMOTOR AXONAL REFLEX TEST QSART assesses the integrity of peripheral sympathetic cholinergic (sudomotor) function and is performed with the installation of an electric current for 5 min to iontophoresis ACh into the skin stimulating nicotinic receptors that activate sudomotor axons to stimulate neurosecretory synapses leading to sweat secretion. Humidity is measured during the iontophoresis and for 5 min afterward (10 min total). In healthy subjects, sweating occurs within 2 min after stimulation begins and returns toward baseline once current is turned off. Multiple sites are tested to map areas of possible dysfunction, including the lateral dorsum foot, distal leg, proximal leg, and medial forearm. Abnormalities are seen with disruption of the postganglionic limb of the sudomotor pathway and are therefore seen in PAF, any cause of autonomic neuropathy such as diabetic neuropathy, and PD when Lewy bodies disrupt the postganglionic axon. The QSART response is typically preserved in MSA, although it may become abnormal late in the disease, presumably due to secondary degeneration of axons. A normal QSART would be unexpected in wellestablished PAF, although variable organ involvement has been seen in some patients.

66

D.S. YOUNGER

THERMOREGULATORY SWEAT TEST The TST evaluates the body ability to dissipate heat by sweating in a patient lying supine in a chamber that maintains temperature at a high level (typically around 115–120°F). A dye applied to the skin, typically alizarin red mixed with cornstarch and sodium carbonate which changes from a light tan to a deep purple color where sweating occurs. A normal sweat pattern should be symmetric and extend to all distal extremities, with some variability in proximal limb segments, where some people may naturally sweat lightly, especially women. Distal, focal, segmental, or global anhidrosis patterns are abnormal, and may reflect neuropathic or central abnormalities. These abnormal patterns may be seen in MSA, PAF, and other causes of distal small fiber neuropathy. The test is most useful in conjunction with the QSART to determine whether a lesion is preganglionic or postganglionic such as when the QSART and TST are both abnormal at the same site suggesting a postganglionic localization; or when the QSART is preserved but the TST is abnormal, as in a likely preganglionic localization.

HEAD-UP TILT TABLE TESTING Head-up tilting table testing assesses the integrity of the cardiovascular functions of the ANS such that upon changing from a recumbent to an upright position on a tilt table, there is a 25%–40% shift of venous blood from central to peripheral compartments, leading to reduction in cardiac filling and stroke volume. A recent consensus statement of the AAN and AAS defined OH as a sustained drop in blood pressure of >20 mm Hg systolic or >10 mm Hg diastolic in the first 3 min of the tilt study (Freeman et al., 2011). The upright position decreases afferent baroreceptor vagal signaling to the NTS, resulting in reflex increased sympathetic vasomotor and cardiac adrenergic sympathetic output through the sympathetic chain. In a normal subject, venoconstriction increases venous return to mitigate cardiac output reduction, arterial constriction increases vascular resistance and maintains pressure to brain and other organs, and cardiac signals increase both heart rate and cardiac stroke volume. The occurrence of OH typically results from impaired efferent sympathetic vasomotor signaling, which may have either a central (MSA), ganglionic (PAF), or peripheral (diabetic neuropathy) origin. Involvement of cardiac sympathetic function may blunt any compensatory tachycardia. A simple, bedside diagnostic test to distinguish neurogenic from nonneurogenic causes of orthostatic

hypertension is the ratio of the increase in heart rate (in beats per minute) to the decrease in SBP (in millimeters of mercury). Since the heart rate response to hypotension is pronounced in patients with nonneurogenic orthostatic hypotension but is blunted in those with efferent baroreflex failure, a ratio below 0.5 indicates baroreflex failure and provides a sensitive and specific cutoff value during passive tilt and active standing (Norcliffe-Kaufmann et al., 2018).

HEART RATE RESPONSE TO DEEP BREATHING In HRRDB, the subject takes 6 deep breaths per minute for 1 min lying supine while recording beat-to-beat heart rate and blood pressures. The normal heart rate variation with respiration helps to pump additional blood into the lung at the end of inspiration when heart rate is maximal. In expiration when heart rate slows, pulmonary pressures aid the cardiac pump by pushing blood into the left heart. Pulmonary stretch receptors pass information through afferent and efferent pathways of the vagus nerve to the NTS, which appropriately increases or reduces cardiac parasympathetic outflow to the heart. Results are typically calculated by subtracting the expiratory from inspiratory heart rates across the five best cycles. The difference is reduced in patients with diabetic and other neuropathies, MSA, and PAF (Ewing et al., 1985).

VALSALVA MANEUVER While performing the Valsalva maneuver (VM), the patient is either supine or at 30 degrees upright and blows against a mouthpiece with an air leak to reach 40 mm Hg for 15 s. The maneuver involves four phases (I–IV): (I) initial pressure generation (1–3 s), (II) continued pressure maintenance (12–14 s), (III) pressure release (1–3 s), and (IV) continued maintenance of normal pressure (30 s). During phase I, there is a transient mechanical blood pressure increase due to increased intrathoracic pressure. Heart rate increases throughout phase II as blood pressure drops (phase II early), recovers to baseline, and typically exceeds baseline due to increased peripheral resistance (phase II late). Tachycardia results from vagal withdrawal initially followed by sympathetic stimulation in phase II late, when the heart rate increase becomes a good measure of sympathetic function. With strain release in phase III, blood pressure drops transiently as a mechanical event, mirroring the phase I increase, without heart rate changes. Phase IV results from venous return exceeding baseline, as vasoconstriction persists momentarily, despite release of chest

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS pressure, and heart rate typically dips below baseline due to activation of the baroreflex by the high blood pressure. The Valsalva ratio compares the maximum heart rate in phase II, that occurs about 1 s after the strain of blowing is released to the lowest heart rate in phase IV, that occurs about 15–20 s after releasing the strain. A ratio greater than 1.45 is considered normal but the ratio varies by age as well (a lower normal ratio down to 1.35 is considered normal in patients over 70 years of age). This test shows an abnormally low ratio in autonomic failure patients with MSA, PAF, and polyneuropathies.

CARDIAC RADIOISOTOPIC DENERVATION IMAGING 123

I-MIBG single-photon emission computed tomography (SPECT) and 6-[18]fluorodopamine (18FDA) positron emission tomography (PET) can show decreased cardiac sympathetic postganglionic innervation. Characteristic patterns of denervation occur in patients with PAF and PD associated with generalized autonomic failure while lack of innervation is not seen in MSA (Fig. 4.5).

SUPINE AND STANDING CATECHOLAMINE LEVELS A postganglionic peripheral lesion such as PAF will show low supine catecholamines compared with a central lesion associated with MSA. In practice, there is no specific level below which the distinction can be made however normally, norepinephrine levels will double from the supine to the upright position. In a disorder that affects postganglionic neurons, such as PAF, there is decreased spillover of norepinephrine into the bloodstream at baseline, because of the degeneration of the distal sympathetic nerves. However, in the upright position, the functioning neurons will double their output, so that one still finds an increase in the norepinephrine level

67

with standing. In contrast, an individual with a central autonomic disorder like MSAwill have a normal baseline supine level of norepinephrine as the distal sympathetic nerves are expected to be intact. With standing, central signals are impaired, leading to a much smaller increase in the norepinephrine level; however, the increase may be larger than expected due to decreased norepinephrine clearance while upright.

SKIN BIOPSY Skin biopsy tissue can be immunohistochemically stained for a-synuclein positivity in suspected patients with PAF who lack other neurological stigmata. Skin biopsy samples containing a-synuclein may also be seen in PD with autonomic involvement (Donadio et al., 2016).

ELECTRODIAGNOSTIC STUDIES Electrodiagnostic studies are useful in the evaluation peripheral neuropathy and in assessing anal sphincter function in MSA associated with denervation due to degeneration in Onuf’s nucleus of the sacral spinal region. Affected individuals show fibrillation potentials, complex repetitive discharges, and chronic motor axon loss motor unit potentials at rest and with graded volitional effort (Ravits et al., 1996).

AUTONOMIC REFLEX SCREEN AND THE COMPOSITE AUTONOMIC SCORE

The severity of autonomic dysfunction can be graded using a composite autonomic severity score, with higher scores indicating more severe autonomic failure (Low, 1993).

Fig. 4.5. Cardiac positron emission tomography (PET) in patients with neurodegenerative autonomic failure. Intravenous 6-[18] fluorodopamine (18FDA) in a normal control individual and in patients with pure autonomic failure (PAF), multiple system atrophy (MSA), and Parkinson’s disease (PD). Radioactivity was absent in the patients with PAF and PD, indicating loss of sympathetic nerve terminals. 18FDA-derived radioactivity in the patient with MSA indicates intact sympathetic terminals. Reproduced from Kabir MA, Chelimsky TC (2019). Pure autonomic failure. Handb Clin Neurol 161: 413–422 with permission.

68

D.S. YOUNGER

Multiple system atrophy OVERVIEW MSA is an adult-onset, sporadic, progressive neurodegenerative disease characterized by varying severity of parkinsonian features, cerebellar ataxia, autonomic failure, urogenital dysfunction, and corticospinal signs and symptoms (Geser et al., 2006). The disease frequently begins with bladder dysfunction, and in males, erectile dysfunction usually precedes this complaint (Kirchhof et al., 2003). The presenting motor disorder most commonly consists of parkinsonism with bradykinesia, rigidity, gait instability, and at times tremor, but cerebellar ataxia is the initial motor disorder in a substantial percentage of patients (Wenning et al., 1997). There can be prototypical contractures of one or both hands and feet commensurate with loss of volitional movement and increased rigidity and weakness (Fig. 4.6).

Fig. 4.6. (A, B) Joint deformities in a patient with MSA-P. (A) Photographs showing hand deformity with flexion at the metacarpophalangeal and proximal interphalangeal joints with difficulty in passively straightening giving rise to a fixed contracture and ulnar deviation of the fingers. (B) Photograph demonstrates inversion contracture of the foot. Reproduced from Chaudhuri K, Hu M (2000). Central autonomic dysfunction. Handb Clin Neurol 75: 161–202 with permission.

The motor manifestations span three distinct disease phenotypes including SND, OPCA, and SDS; Graham and Oppenheimer (1969) coined the term MSA to encompass the three entities. Depending upon the predominant motor feature, MSA is further divided into cerebellar (MSA-C) or parkinsonism (MSA-P) disease phenotypes. The discovery of GCI in all three supports the unified concept of MSA (Papp et al., 1989). Two consensus statements reaffirm the criteria MSA with varying degrees of diagnostic certainty as definite, probable and possible (Gilman et al., 1998, 2008). A diagnosis of definite MSA requires pathological confirmation. Autonomic failure, as exemplified by urinary incontinence or orthostatic hypotension, is a sine qua non for clinical diagnosing MSA. Other than autonomic failure and motor symptoms, nonmotor symptoms, including stridor, rapid eye movement (REM) sleep behavior disorder, pseudobulbar affect, and severe dysphonia and dysarthria, are often seen in MSA. At present, there are no curative treatments for MSA; symptomatic treatment is, therefore, important. There is a recent review (Koga and Dickson, 2018). Ataxia of gait, the most common cerebellar feature of MSA-C, is often accompanied by ataxia of speech (cerebellar dysarthria) and cerebellar oculomotor dysfunction. Limb ataxia may be seen but is generally less prominent than gait or speech disturbances. Although gaze-evoked nystagmus occurs in the majority of laterstage MSA-C patients, earlier oculomotor abnormalities may not involve nystagmus, but include square wave jerks, jerky pursuit, and dysmetric saccades. Limitations of supranuclear gaze and severe slowing of saccadic velocities are not features of MSA. Most MSA patients develop parkinsonism (bradykinesia with rigidity, tremor, or postural instability) at some stage. The tremor is usually irregular and postural/action, often incorporating myoclonus, but a classic pill-rolling rest tremor is uncommon. The parkinsonism can be asymmetric. Postural instability, as defined by item 30 of the Unified Parkinson’s Disease Rating Scale (UPDRS) part III (motor examination) (Fahn et al., 1987) occurs earlier and progresses more rapidly than in PD. Moreover, the UPDRS part III score typically worsens by less than 10% per annum in PD but by more than 20% in MSA (Seppi et al., 2005b). Parkinsonism usually responds poorly to chronic levodopa therapy; however, up to 30% of patients show a clinically significant, but usually waning, response (Hughes et al., 1992a). Responsiveness should be tested with escalating doses of levodopa with a peripheral decarboxylase inhibitor over 3 months up to at least 1 g/day (if necessary and tolerated). A positive response is defined as clinically significant motor improvement. This should be demonstrated by objective evidence such as an improvement of 30% or more on part III of the UPDRS or on part II of the UMSARS (Wenning et al., 2004).

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

PATHOPHYSIOLOGY The defining neuropathology of MSA consists of degeneration of striatonigral and olivopontocerebellar structures accompanied by profuse numbers of distinctive GCIs (Fig. 4.7) formed by fibrillized a-synuclein proteins (Papp et al., 1989; Spillantini et al., 1998; Ozawa et al., 2006). a-Synuclein is a highly conserved protein that is abundant in neurons, especially presynaptic terminals. Aggregated a-synuclein proteins form brain lesions that are hallmarks of neurodegenerative synucleinopathies. Oxidative stress has been implicated in the pathogenesis of some of these disorders. Using antibodies to specific nitrated tyrosine residues in a-synuclein, Giasson et al. (2000) demonstrate extensive and widespread accumulations of nitrated a-synuclein in the signature inclusions of PD, DLB, the Lewy body variant of Alzheimer disease (AD) and MSA brains. The a-synuclein (SNCA) gene maps to chromosome 4q22.1, and while cerebrospinal fluid (CSF) studies have not been part of the routine laboratory assessment, an a-synuclein-protein misfolding cyclic amplification assay discriminates between samples of patients diagnosed with PD and MSA with an overall sensitivity of 95.4% (Shahnawaz et al., 2020). Notwithstanding, MSA can be clinically difficult to differentiate from DLB, PD, and PSP not only in early stages, but also in late stages of the disease process. Although MSA is generally considered to be a sporadic disorder, Scholz et al. (2009) performed a candidate single nucleotide polymorphism association study in a genome-wide study of PD, MSA cases and control subjects noting significant associations for increased risk for

69

developing MSA (odds ratio [OR] 6.2) suggesting the influence of heritable factors in the pathogenesis and development of the disease. Moreover, rare familial manifestations were reported suggesting a familial monogenic form of MSA with autosomal recessive (AR) inheritance (Hara et al., 2007) in 8 patients of 4 unrelated Japanese families, manifesting definite, probable, and possible MSA based upon contemporaneous consensus criteria (Gilman et al., 1998). However, no mutations were found in several genes for hereditary ataxia or in the a-synuclein (SNCA) gene.

CLASSIFICATION A consensus conference on diagnosis held in 1998 defined two categories, MSA with predominant parkinsonism (MSA-P) and MSA with predominant cerebellar ataxia (MSA-C) (Gilman et al., 1998). Three levels of certainty were established, possible, probable, and definite MSA, with the diagnosis of definite MSA requiring autopsy confirmation. These guidelines emphasized the importance of autonomic features by requiring them for the diagnosis of probable MSA. Validation studies of the consensus criteria demonstrated high positive predictive value of 86% for the clinical diagnostic accuracy in a series of postmortem confirmed cases from the Queen Square Brain Bank for Neurological Disorders (Osaki et al., 2002) with clinical misdiagnoses predominantly due to confusion with PD, that could be ascribed to both autonomic and urinary dysfunction, and levodopa poorly responsive parkinsonism occurring in PD or PD with dementia cases. Two intriguing results came from a retrospective review of 134 consecutive patients

Fig. 4.7. Multiple system atrophy: glial cytoplasmic inclusions (arrows).
700. Reproduced from Bruch LA, Schmidt RE (2000). Pathology of the autonomic nervous system. Handb Clin Neurol 75: 1–52 with permission.

70

D.S. YOUNGER

(Koga et al., 2015) in whom the antemortem clinical diagnosis of probable or possible MSA was based upon available clinical information according to the second consensus statement for MSA (Gilman et al., 2008) and later studied at postmortem neuropathologic evaluation of the brain. The first was a diagnostic accuracy of 62%, 71%, and 60% for definite, probable, and possible, respectively, that did not differ generally between general neurologists and movement disorder specialists. The second was the misdiagnosis of MSA in life typically for cases with an older age at onset and age at death than comparable cases of PD or PSP; and misdiagnoses among those with atypical presentations of ataxia (as in PSP), and uncommon clinical features such as dysautonomia (in DLB). Two contemporaneous studies by Litvan et al. (1997, 1998) that were predicated on the difficulty of diagnosis at an early stage noted respectively a sensitivity for clinical diagnosis of 56% and 69%, at first clinical visit and at last visit 74 months postsymptom onset; was enhanced when six of eight clinical features mentioned were present (Table 4.2). The availability of additional clinical, laboratory, neuropathological, biochemical and neuroimaging studies that assist in the diagnosis of MSA (Ozawa et al., 2004, 2006; Seppi et al., 2004, 2006; Gilman, 2005) prompted a second consensus conference in 2007 producing new guidelines (Gilman et al., 2008). While keeping the clinical designations of the first consensus (Gilman et al., 1998), MSA-C and MSA-P was used more appropriately in referring to the predominant motor feature of cerebellar ataxia or parkinsonism respectively, at the time of first evaluation, recognizing that a patient might present with predominant cerebellar signs and but develop increasingly severe parkinsonian features until Table 4.2 Criteria of the diagnosis of probable multiple system atrophy* In adults (age >30 years) with sporadic and progressive disease onset characterized by: Autonomic failure involving urinary incontinence defined as the inability to control the release of urine from the bladder, with erectile dysfunction in males; or an orthostatic decrease in blood pressure within 3 min of standing by at least 30 mm Hg systolic or 15 mm Hg diastolic, and Poorly levodopa-responsive parkinsonism defined as bradykinesia with rigidity, tremor, or postural instability or A cerebellar syndrome defined as gait ataxia with cerebellar dysarthria, limb ataxia, or cerebellar oculomotor dysfunction Adapted from Gilman S, Wenning G, Low PA et al. (2008). Second consensus statement on the diagnosis of multiple system atrophy. Neurology 71: 670–676. Hg, mercury.

the latter dominated the presentation. Similar to the first consensus conference, the diagnosis of MSAwas divided into three groups definite, probable, and possible. In keeping with distinctions in the levels of diagnostic certainty, the current criteria used the term definite MSA for subjects with autopsy demonstration of typical histologic features of widespread and abundant CNS a-synucleinpositive GCIs (Papp–Lantos inclusions) in association with neurodegenerative changes in striatonigral or olivopontocerebellar structures (Trojanowski et al., 2007), which were not previously required in the first consensus statement (Gilman et al., 1998). In regards to the categories of possible and probable MSA of the first consensus statement (Gilman et al., 1998) that used “features” to described the clinical findings, and “criteria” to indicate the features that could be used for diagnosis proved to be confusing and difficult to retain, so those were discarded, and replaced with straightforward descriptions of the clinical findings required for diagnoses. The diagnosis of probable MSA required at least one feature suggesting autonomic dysfunction in addition to parkinsonism or a cerebellar syndrome (Table 4.3), while an additional clinical feature was required for the diagnosis of possible Table 4.3 Additional features for the diagnosis of possible multiple system atrophy* Possible MSA-P or MSA-C Babinski signs with hyperreflexia Stridor Possible MSA-P Rapidly progressive parkinsonism Poor response to levodopa Postural instability within 3 years of motor onset Gait ataxia, cerebellar dysarthria, limb ataxia, or cerebellar oculomotor dysfunction Dysphagia within 5 years of motor onset Atrophy of MRI of putamen, middle cerebellar peduncle, pons or cerebellum Hypometabolism on FDG-PET in putamen, brainstem, or cerebellum Possible MSA-C Parkinsonism as defined by bradykinesia and rigidity Atrophy on MRI of putamen, middle cerebellar peduncle, or pons Hypometabolism on FDG-PET in putamen Presynaptic nigrostriatal dopaminergic denervation on SPECT or PET Adapted from Gilman S, Wenning G, Low PA et al. (2008). Second consensus statement on the diagnosis of multiple system atrophy. Neurology 71: 670–676. FDG-PET, fluorodeoxyglucose positron emission tomography; MRI, magnetic resonance imaging; MSA, multiple system atrophy; MSAC, MSA with predominant cerebellar ataxia; MSA-P, MSA with predominant parkinsonism.

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

71

Table 4.4 Comparison of multiple system atrophy, primary autonomic failure, and Parkinson’s disease Characteristic

MSA

PAF

PD

Site (pre/postganglionic) Site (CNS/PNS) Sxs (Parkinson/cerebellar) Site (Inclusion) OH, present Sxs (sleep/SBD/apnea) Sxs urinary tract Supine NE Orthostatic rise NE Sens. postsynaptic. AdrRec

Preganglionic CNS Common Oligo (GCI) Common Common Early in disease Normal Subnormal Mildly increased

Postganglionic PNS Rare Neuron (Lewy body) Common Uncommon Late in disease Very low Subnormal Increased

Both CNS Always Neuron (Lewy body) Less common Common After extrapyramidal Low Normal Mildly increased

AdrRec, adrenal receptor; CNS, central nervous system; GCI, glial cytoplasmic inclusions; MSA, multiple system atrophy; NE, norepinephrine; OH, orthostatic hypotension; oligo, oligodendrites; PAF, primary autonomic failure; PD, Parkinson’s disease; PNS, peripheral nervous system; SBD, sleep behavior disorder; Sxs, symptoms.

MSA ascertained from the history, clinical examination, or results from either structural or functional imaging (Table 4.4).

AUTONOMIC FAILURE In MSA, autonomic failure is associated with abnormalities in control of automatic respiration and endocrine function. Orthostatic hypotension may indicate autonomic failure and can be asymptomatic or symptomatic. When symptomatic, it frequently occurs after the onset of ED and urinary symptoms (Kirchhof et al., 2003). Symptoms of OH result from hypoperfusion, and syncope may occur (Allcock et al., 2004). The second consensus statement (Gilman et al., 2008) required a reduction of SBP by at least 30 mm Hg or of diastolic blood pressure by at least 15 mm Hg after 3 min of standing from a previous 3-min interval in the recumbent position. This orthostatic decline is usually accompanied by a compensatory increase in heart rate that is inadequately low for the level of blood pressure decline. This is a more pronounced decrease of blood pressure than recommended previously in the AAS–AAN consensus statement on the definition of orthostatic hypotension (Consensus Committee of the American Autonomic Society the American Academy of Neurology, 1996). The mechanisms for these manifestations are probably multifactorial and reflect involvement of preganglionic efferent neurons, ventrolateral medullary, pontine and PAG regulatory circuits, and hypothalamic cell groups (Benarroch, 2002; Ozawa, 2007). The loss of C1 epinephrine-synthesizing neurons in the ventrolateral contributes to OH in MSA much more consistently than in Lewy body disorders, however there can also be involvement of sympathetic nerves at the late stages of MSA consistent with evidence of cardiac sympathetic

denervation (Nagayama et al., 2008) seen as reduced myocardia MIBG uptake on SPECT. There can be loss of cholinergic neurons in the DMV (Benarroch et al., 2006) and a primary contributor to gastrointestinal symptoms in this disorder. The involvement of vagal neurons of the NA controlling laryngeal function differs among cases with some studies showing loss (Isozaki et al., 2000) and others showing relative preservation of these neurons (Benarroch et al., 2003), thus, the contribution of loss of NA neurons to laryngeal stridor in MSA is yet to be defined. A loss of neurons on the ventrolateral portion of the NA implicated in cardiovagal control can affected in MSA and relatively spared in Lewy body disorders may contribute to impaired baroreflex cardioinhibition and respiratory sinus arrhythmia in MSA (Benarroch et al., 2006). Involvement of medullary respiratory neurons can contribute to central sleep apnea, alveolar hypoventilation, respiratory dysrhythmia and laryngeal stridor and a prominent cause of sudden death in MSA patients (Ghorayeb et al., 2005). Urinary incontinence in MSA commonly associated with incomplete bladder emptying, reflecting detrusor hypocontractility and with denervation of the external urinary sphincter, as detected in sphincter electromyogram (EMG) may result from involvement of the pontine micturition areas and the loss of neurons in the sacral preganglionic nucleus. Involvement of ventral PAG in MSA leads to loss of tyrosine hydroxylase immunoreactive neurons in MSA and cases with cortical stage Lewy body disease (Benarroch et al., 2009). a-Synuclein pathology affects all columns of the PAG (Benarroch et al., 2010), which may contribute to disturbances in cardiovascular, bladder, and respiratory control in this disorder. Autonomic failure should be evaluated not only by a history of urinary incontinence and orthostatic blood pressure measurements in the clinic, but also by a

72

D.S. YOUNGER

Fig. 4.8. T2-weighted MRI image (A) unaltered in PD, and (B) putaminal atrophy (arrow), putaminal hypointensity (dotted line) and hyperintense margin (dashed line) in a patient with MSA-P. Reproduced from Wenning GK, Krismer F (2013). Multiple system atrophy. Handb Clin Neurol 117: 229–241 with permission.

comprehensive battery that examines the distribution and severity of cardiovascular, sudomotor, and urinary bladder deficits (Suarez et al., 1999). Cardiovascular and sudomotor autonomic function tests may help to separate MSA from other sporadic cerebellar ataxias and from PD (Sandroni et al., 1991). Measurement of urine residual volume (RV) by ultrasound can reveal incomplete bladder emptying of >100 mL, and tends to increase as MSA progresses. Imaging of cardiac innervation with PET and [18F]fluorodopa have shown preserved sympathetic postganglionic neurons in MSA, in contrast to PD (Courbon et al., 2003). Others with SPECT and 123 I-MIBG (Nagayama et al., 2005) and PET and [11C] hydroxyephedrine revealed cardiac denervation in MSA (Raffel et al., 2006).

ADDITIONAL LABORATORY ASSESSMENT The MRI demonstration of T2-signal changes in the basal ganglia and brainstem, including posterior putaminal hypointensity and hyperintense lateral putaminal rim (Fig. 4.8), “hot cross bun” sign (Fig. 4.9) and middle cerebellar peduncle (MCP) hyperintensities (Seppi et al., 2005a) may be helpful in diagnosing MSA. Striatal or brainstem hypometabolism by PET with [18F]fluorodeoxyglucose can also be helpful (Gilman, 2005) especially in the absence of clinically evident ataxia in patients with parkinsonian features to demonstrate cerebellar hypometabolism pointing to the diagnosis of MSA-P rather than PD. Conversely, in the absence of parkinsonian features in a patient with cerebellar ataxia, evidence of nigrostriatal dopaminergic denervation from functional imaging (SPECT and PET) may point to the

Fig. 4.9. “Hot cross bun” sign in a T2-weighted MRI image of a patient with MSA-P. Reproduced from Wenning GK, Krismer F (2013). Multiple system atrophy. Handb Clin Neurol 117: 229–241 with permission.

diagnosis of MSA-C (Gilman et al., 1999). MCP regional apparent diffusion coefficient values in diffusionweighted MRI can discriminate MSA-P, progressive supranuclear palsy and PD from MSA-P with 100% sensitivity and specificity (Nicoletti et al., 2006). Progressive hypohidrosis due to postganglionic sudomotor failure may be associated with hyperthermia or heat stroke. TST may differentiate MSA from PD, but not from PAF.

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

Pure autonomic failure HISTORICAL TURNING POINTS Neurogenic idiopathic orthostatic hypotension (IOH) as we now recognize it was first described in the English literature by Bradbury and Eggleston in 1925 in 3 living patients who showed a total absence of normal vasomotor control with precipitous drop of 50% or more in blood pressure in the face of postural changes. Each patient complained of syncopal attacks after exertion or standing erect for a few minutes, often in association with weakness (Case 1), exhaustion and insomnia (Case 2), or dyspnea, chest pain, numbness in the limbs (Case 3), although without documentation of an associated underlying process aside from slight anemia and a lowered basal metabolism. The 3 patients were surmised to have a disorder of the “vegetative” or ANS in association with a uniformly slow heart rate unaltered by changes in blood pressure, posture or injection of atropine; impaired cardiac acceleration and augmentation of sympathetic function by epinephrine stimulation to offset postural hypotension; and decreased vagal response to elevated blood with the head-down or after epinephrine injection and absent sweating despite pharmacologic stimulation. The explanation for the observed blood pressure responses to postural and provocative maneuvers was a paralysis of sympathetic vasoconstrictor nerve endings. The later report of the postmortem examination of Bradbury and Eggleston’s Case 2 (Bradbury and Eggleston, 1925), a 39-year-old man who died 6 years after onset of symptoms who did not regain consciousness after a spontaneous fainting episode, excluded the brain and spinal cord (Bradbury and Eggleston, 1925) and showed myocarditis, cardiac dilatation, and chronic tuberculous lymphadenopathy. Nylin and Levander (1948) reviewed the world’s literature of 24 similar cases, noting that one such case cited in 1891, was later published by Laubry and Doumer (Ghrist and Brown, 1928; Riecker and Upjohn, 1930; Sanders, 1931; Laubary, 1932; Lian and Blondel, 1933; Ganshorn, 1934; Alvarez and Roth, 1935; Croll and Duthie, 1935; Korns and Randall, 1937, 1938; Capaccio and Donald, 1938; Ewert, 1938; Dietrich and Schimert, 1940; Jeffers et al., 1941; Nordenfelt, 1941; Stead and Ebert, 1941; Hammarstr€ om and Lindgren, 1942), and adding one of their own, referring to it as asympathicotonic hypotension. A decade later, Barnett and colleagues described widespread loss of sympathetic nervous activity and limited loss of parasympathetic activity in a young man (Barnett and Wagner, 1958) postulating a causative peripheral sympathetic lesion. A number of turning points ushered in modern understanding of IOH associated with PAF. The first came with clinicopathologic studies differentiating IOH from

73

PD-like (SDS) (Shy and Drager, 1960) and multiple system degeneration (MSA) (Graham and Oppenheimer, 1969), such that for the next several decades, investigators meticulously separated cases of IOH alone from others clinically (Thomas and Schirger, 1963; Hughes et al., 1970) and histopathologically at postmortem examination (Johnson et al., 1966; Goodall et al., 1968; Martin et al., 1968; Vanderhaeghen et al., 1970; Roessmann et al., 1971; Oppenheimer, 1980; Van Ingelghem et al., 1994; Hague et al., 1997). Moreover, there was good corroboration between living and postmortem-studied cases, the latter manifesting absent or minimal neuronal loss in the substantia nigra, DMV, locus coeruleus, inferior olives, and more prominent cell loss in the IML columns of the spinal cord and sympathetic ganglia, and generally prominent Lewy bodies along the neuraxis (Table 4.5). The second turning point was the recognition of disrupted peripheral autonomic pathways, first suggested by Goodall et al. (1968) who showed a marked decrease in the synthesis of noradrenaline as reflected by a decline in the urinary recovery of radioactive noradrenaline following an infusion of the immediate precursor to noradrenaline, 3-hydroxytyramine-2-14C, with a marked decrease in the formation of 3-methoxy-4-hydroxymandelic acid, the principal metabolite of noradrenaline. The localization to the peripheral SNS was supported by degenerative changes in the sympathetic ganglia and neurons with disruption and degeneration of peripheral sympathetic nerve fibers at postmortem in 1 patient (case MQ). Kontos et al. (1975) and Ziegler and colleagues in 1977 later separated the defects in OH physiologically. The latter (Ziegler et al., 1977) studied 10 patients with IOH, 6 of who were deemed to have a central basis because of coexistent parkinsonism and cerebellar dysfunction, with normal plasma levels of norepinephrine while recumbent, compared to 4 with presumed peripheral manifestations based upon low supine levels of plasma norepinephrine. Both central and peripheral groups failed to increase plasma norepinephrine levels normally after standing and exercising with low levels of plasma dopamine-b-hydroxylase, presumably due to prolonged deficits in sympathetic neuronal function. The authors (Ziegler et al., 1977) concluded that the defect in patients without CNS signs affected peripheral sympathetic nerves. The third turning point followed a series of detailed autonomic neurochemical, morphological and physiological studies. Bannister (1979) compared the results of cardiac reflex testing and responses to pressor agents in 10 patients with chronic autonomic failure and postural hypotension, designating autonomic failure in isolation as “pure” (PAF), autonomic failure associated with MSA, or autonomic failure associated with parkinsonism (MSA), noting abnormalities in Valsalva maneuvers and sweat tests in all groups. Polinsky assessed the pressor

Table 4.5 Postmortem clinicopathological features of pure autonomic failure

Caseno. Sex (M/F) Age onset/death Duration of illness (y, yrs.) Clinical OH Present: Y/N; ns, not stated Sphincter disturbance Loss of sweating Sexual impotence Ataxia/dysarthria Parkinsonian features Postmortem neuropathology Neuronal or fiber loss (Y/N)/Lewy bodies, LB; (+/) or ns, not stated Substantia nigra Dorsal motor nucleus vagus Locus coeruleus Inferior olives Intermediolateral column Sympathetic ganglia Lumbar parasympathetic ganglia Sympathetic nerve Vagus nerve Somatic nerve a-Synuclein deposition

Johnson et al. (1966)

Martin et al. Goodall et al. Vanderhaeghen et al. Roessmann et al. (1968) (1968) (1970) (1971)

Oppenheimer (1980)

Oppenheimer (1980)

Van Ingelghem et al. (1994)

Hague et al. (1997)

Isonaka et al. (2017)

1 M 62/66 4y Y

2 M 54/59 5y Y

3 M 46/49 3y Y

4 M 73/74 1y Y

5 M 56/67 11 y Y

6 M –/83 ns Y

7 M –/70 ns Y

8 M 57/64 7y Y

9 M 48/63 15 y Y

10 M 50/60 10 y Y

N Y – N N

Y – Y N N

Y Y Y ns ns

ns ns ns N N

Y Y N N N

ns ns ns Y N

ns ns ns N N

N Y Y N N

Y Y Y N N

Y Y ns N N

N/+ N/+ /+

N N N N Y/ns N/ns ns ns ns ns ns

ns ns ns ns ns Y/ns ns Y/ns ns ns ns

N/+ Y/+ N/+ N/+ ns/+ ns/+ ns ns ns ns ns

Y/+ Y/+ Y/+ Y/ Y/ N/LB N/ns N/ns N/ns Y/ns ns

N/ N/ Y/

N/ N/ N/

Y/+ ns ns Y/ns ns ns ns

Y/+ ns ns ns ns N/ns ns

N/ N/ N/ N/ Y/ Y/+ Y/+ ns ns ns ns

Y/+ ns/ Y/+ ns Y/ Y/+ ns/ns ns/+ ns ns ns

N/ N/ N/ N/ ns ns/ ns ns ns ns Negative

Y/+ N/+ ns Y ns N/ns ns

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS responses to vasoactive agents suggesting denervation in PAF (Polinsky et al., 1981). Dotson et al. (1990) applied microneurographic and histopathological techniques in a patient with PAF showing marked reductions in sympathetic efferent nerve impulses and supine norepinephrine that did not rise upon standing, with morphometric evidence of past degeneration in biopsied sural nerve tissue. Bannister et al. (1981) next provided evidence for the loss of sympathetic postganglionic neurons in PAF employing catecholamine fluorescence and electron microscopic studies of sympathetic perivascular nerve plexuses in quadriceps muscle of 2 patients with PAF, showing complete absence of catecholamines and reduced granular noradrenergic vesicles. Two subsequent postmortem cases of PAF (Table 4.5) (Van Ingelghem et al., 1994; Hague et al., 1997) described prototypical loss of IML column neurons with numerous Lewy bodies in lumbar paravertebral sympathetic ganglia and autonomic nerves, correlative in one (Hague et al., 1997) with the loss of end-organ sympathetic innervation. A fourth turning point was the recognition of a-synuclein as a major component of Lewy bodies and Lewy neuritis (Kaufmann et al., 2001) and the conceptualization of Lewy body synucleinopathy (Kaufmann and Goldstein, 2010) that accounted for the distinctive clinical phenotypes and predominant central and peripheral sites of Lewy body formation and neuronal loss: nigrostriatal in PD, cortical in DLB, and peripheral sympathetic in PAF. One patient with PAF, so studied (Shishido et al., 2010) had absent tyrosine hydroxylase immunoreactivity in the dermis and subcutaneous tissue and around blood vessels supporting an association between a-synuclein deposits and sympathetic noradrenergic denervation. Intraneural misfolded phosphorylated (psyn) a-synuclein was found in 14 patients with PAF with length-dependent somatic and autonomic small nerve fiber loss (Donadio et al., 2016). Native (n-syn) a-synuclein staining in dermal nerve tissue was found in patients with PAF and PD, while p-syn was conspicuously absent in the latter; cutaneous a-synuclein immunoreactivity was also absent in cases of MSA and other disorders without Lewy bodies (Ikemura et al., 2008). While autopsies and biopsies of patients suspected to have PAF have overwhelmingly shown a-synuclein in Lewy bodies or in nerve fibers of dermal tissues, a recent clinically compatible case with the characteristic triad of chronic neurogenic OH, absent clinical signs of central neurodegeneration and generalized sympathetic noradrenergic denervation, but without Lewy bodies or a-synuclein deposition in the brain, brainstem, sympathetic ganglion tissue, myocardium, or skin was reported (Isonaka et al., 2017). While considering the division of PAF into pathophysiologically distinctive subtypes based upon that case and other potential outliers, it’s

75

worth noting that the patient was atypical due to three potential considerations. First, in the presence of normal sweating and supported by QSART. Second, due to dream-enactment behavior suggestive of REM behavior disorder in keeping with PAF or evolving synucleinopathy (Boeve et al., 2001), yet not seen on polysomnography. Third, other diagnoses that mimic PAF needed to be excluded including seronegative AAG (Suarez et al., 1994) and selective sympathetic ganglioneuropathy associated with the history of exposure to potential neurotoxins.

DIAGNOSIS Selection for prospective cohort studies The Autonomic Disorders Consortium (Kaufmann et al., 2017) has recruited subjects for a prospective longitudinal observational natural history study of patients with PAF from September 2011 to September 2015, defining neurogenic OH as a fall in blood pressure (BP) of 20 mm Hg systolic or 10 mm Hg diastolic within the first 3 min of upright tilt (Freeman et al., 2011) (Fig. 4.10), with absence of phase IV BP overshoot after release of the Valsalva strain, consistent with sympathetic (autonomic) failure and a neurogenic cause (Goldstein and Low, 2007) (Fig. 4.11); excluding those meeting criteria for PD (Hughes et al., 1992b), MSA (Gilman et al., 2008), DLB (McKeith et al., 2005), or other neurodegenerative disorder, peripheral neuropathy, amyloidosis, diabetes, or autoimmune disease, ganglionic AChR antibodies, and secondary causes of OH such as medications, dehydration or severe anemia. Laboratory studies According to Coon et al. (2019) the diagnosis of PAF was supported by a low supine norepinephrine levels with minimal to no increase upon standing. However, the combination of autonomic function testing, functional imaging, and orthostatic catecholamines differentiated PAF from other synucleinopathies (Merola et al., 2017). Biopsy of skin was available to search for a-synuclein deposition.

NATURAL HISTORY AND PROGNOSIS Among 100 consecutive patients with PAF who met inclusion criteria and completed the clinical evaluations at entry into the Autonomic Disorders Consortium natural history study (Kaufmann et al., 2017), a significant majority of subjects were men (70% vs 30%) with symptoms of neurogenic OH beginning at a mean of 63 years. At the time of enrollment, the median duration of illness was 5  7 years with predominant symptoms of OH, however additional symptoms of autonomic

76

D.S. YOUNGER

Fig. 4.10. Tilt table testing in a patient with pure autonomic failure. The upper panel shows orthostatic hypotension as frequently seen in pure autonomic failure. X-axis shows the following curves in order: systolic blood pressure (SBP), mean arterial pressure (MAP), diastolic blood pressure (DBP), and heart rate (HR). Note the gradual descent of blood pressure from a baseline supine hypertension of 210/110 to 90/60, with minimal compensatory heart rate response from 75 to 90 bpm. The lower panel shows the normal response to a head-up tilt table study. Note the absence of a fall in blood pressure and any rise in heart rate. Reproduced from Kabir MA, Chelimsky TC (2019). Pure autonomic failure. Handb Clin Neurol 161: 413–422 with permission.

impairment included constipation (58%), bladder disturbances (50%), sweating abnormalities (44%), and erectile dysfunction in men (65%). The prevalence of supine hypertension was 47% with a mean resting supine blood pressure of 152/84 mm Hg, a mean heart rate of 65 bpm, which after 3-min of upright tilt, fell by 52  27 mm Hg systolic and 23  23 mm Hg diastolic. On entry, none of the patients had signs of

cognitive impairment, and none of the patients met clinical diagnostic criteria for PD, MSA, DLB or resting tremor and rigidity. Among 74 patients followed longitudinally, 34% of those with PAF phenoconverted to a manifest CNS synucleinopathy, either DLB (18%), PD (8%), or MSA (8%), presumably associated with the anatomical spread of a pathology already present at study entry, with a risk of phenoconversion from PAF to a

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

77

Fig. 4.11. Valsalva maneuver in a patient with pure autonomic failure. The upper panel shows the Valsalva maneuver from the same patient (with PAF) as the tilt study in Fig. 4.10. The X-axis again represents SBP, MAP, DBP, and HR in order, and the Y-axis of both represents time in minutes. Note the inexorable drop in blood pressure that never recovers during phase II late, the highly prolonged pressure recovery time, measured from the trough of phase III to the time blood pressure is at baseline, of about 20 s and absence of phase IV BP overshoot after release of the Valsalva strain, consistent with sympathetic (autonomic) failure and a neurogenic cause. The lower panel shows normal response to a Valsalva maneuver, with pressure recovery time of about 3 s. Reproduced from Kabir MA, Chelimsky TC (2019). Pure autonomic failure. Handb Clin Neurol 161: 413–422 with permission.

manifest CNS synucleinopathy of 14% per year. Almost all patients who phenoconverted had RBD at the time of enrollment, 88% of whom had subtle motor deficits, and 53% had olfactory loss. Such symptoms were previously

reported to predict phenoconversion in premotor PD cohorts (Mahlknecht et al., 2015; Postuma et al., 2015a,b). The presence of RBD, olfactory loss, or subtle motor deficits was considered as nonsupportive for PAF

78

D.S. YOUNGER

because they indicated that CNS neurons were already affected, and thus the disease was not exclusively autonomic and likely to progress. Of 42 patients who remained as PAF clinically at the last clinical visit, 30 had RBD, impaired olfaction, or subtle motor signs suggesting that synuclein-driven neurodegeneration was already present in their CNS and would likely spread further, resulting in phenoconversion. In contrast, 12 patients with neither RBD, olfactory loss, nor subtle motor deficits, implied they had a restricted synucleinopathy affecting only autonomic neurons. These patients were 57 12 years old when their symptoms of neurogenic orthostatic hypotension first began and had a disease duration of 6  5 years, and their plasma norepinephrine levels were very low (median ¼ 63 37 pg/mL; in all cases, C) in the ELP1 gene located on the long arm of chromosome 9q. Haplotype analysis has traced the origin of the mutation to Ashkenazi Jews in the 16th century living in the area of the Pale of Settlement between the Black sea and the Baltic sea (Norcliffe-Kaufmann et al., 2017). Transmission is autosomal recessive with 99.9% of the cases homozygous for this mutation. The carrier rate in the entire Ashkenazi Jewish population is around 1:30, but increases among those descending from regions of Poland from which waves of emigration carried the founder mutation to other continents including Israel and the United States where the largest number of cases is found. With the widespread use of whole-genome sequencing, both newborns homozygous for the founder mutation have described in families unaware of Jewish heritage (Leyne et al., 2003), implying that the causative mutation may have been introduced into other population clusters creating new founding populations; as well as, compatible cases heterozygous for the ELP1 mutations have been identified. The latter case was from mixed Ashkenazi Jewish parents who manifested a major FD haplotype from an Ashkenazi Jewish the father, and a 3051C-T transition in exon 26 resulting in a pro914-to-leu (P914L) substitution from the mother of Irish-German/Sicilian heritage (Leyne et al., 2003). Formerly described as IKBKAP (inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein), the elongator acetyltransferase complex, subunit 1 (ELP1) gene encodes a highly conserved protein and scaffolding subunit of the sixsubunit elongator complex. The common founder mutation in the ELP1 gene leads to a splicing defect in the pre-

85

mRNA-producing mutant mRNA transcripts and a truncated nonfunctional ELP1 protein. The splicing abnormality in FD is expressed in a tissue-specific manner with patients producing variable amounts of mutant or wild-type ELP1 transcripts in different nervous system tissues. How the ELP1 gene mutation results in the human clinical FP phenotype of widespread sensory deafferentation due to underdeveloped afferent somatic and autonomic sensory neurons, with their cell bodies in DRG and cranial nerves is not well known. This contrasts with functionally spared yet mildly reduced number of efferent motor somatic and sympathetic neurons, likely due to adaptive mechanisms at the target end organs junction that gives rise to denervation supersensitivity. It is noteworthy that in experimental mice, Elp1 is essential for the normal development of neural crest-derived dorsal root ganglia (DRG) sensory neurons (Tolman et al., 2022) and the circuitry underlying baroreception and chemoreception in visceral sensory neuron ganglia, as well as its peripheral target innervation and CNS synaptic partners in the medulla. The fundamental importance of Elp1 in their development was apparent by the fact that mice null for Elp1 die by embryonic day 10, owing to failures in neurulation and vasculature formation (Dietrich et al., 2011). Yet why dependence on Elp1 varies temporally and by cell type is also not well understood. Not all neuronal populations depend on Elp1 for their development and survival. In spite of robust expression, retinal neurons do not require Elp1 for their development, however, within 2 weeks of birth, Elp1 conditional knockout retinal ganglion cells progressively die, similar to humans in whom histopathological analysis of postmortem tissues of patients with FD and visual decline showed thinning of the retinal ganglion cell layer with widespread axonal loss due to depletion of macular ganglion cells (Mendoza-Santiesteban et al., 2017).

POSTMORTEM STUDIES Early pathological studies indicate that within the peripheral sensory and autonomic systems, individuals affected with FD suffer from incomplete neuronal development as well as progressive neuronal degeneration as indicated below. Sensory nervous system DRG are grossly reduced at postmortem examination due to decreased neuronal population (Pearson et al., 1974, 1978). Within the spinal cord, lateral root entry zones and Lissauer tracts are severely depleted of axons. With evidence of slow progressive deterioration, there was a definite trend with increasing age for further

86

D.S. YOUNGER

depletion of the number of neurons in DRG and an increase in the abnormal numbers of residual nodules of Nageotte in DRGs (Filler et al., 1965; Pearson et al., 1978). In addition, there was loss of dorsal column myelinated axons that was more evident in older patients. Neuronal depletion correlated well with the clinical observations of worsening pain and vibration sense with increasing age (Axelrod et al., 1981). Sural nerves contained markedly diminished number of nonmyelinated and small diameter myelinated axons (Pearson et al., 1974), with characteristic findings to distinguish it from other sensory neuropathies. There was a diminution of immunohistochemical staining for substance P-containing axons relevant for synaptic transmission, in the substantia gelatinosa of spinal cord and medullary tissue of FD patients with diminished pain sensitivity (Pearson et al., 1982). Autonomic nervous system Consistent with an actual decrease in neuronal numbers, the mean volume of superior cervical ganglia was reduced by 34% of normal size, with decreased numbers of neurons in the IML gray columns of the spinal cord suggesting involvement of preganglionic neurons (Pearson and Pytel, 1978b). Autonomic nerve terminals could not be demonstrated on peripheral blood vessels (Grover-Johnson and Pearson, 1976). This lack of innervation was consistent with postural hypotension, as well as the exaggerated response to sympathomimetic and parasympathetic agents (Dancis, 1968). Other than the sphenopalatine ganglia, which were consistently reduced in size with low total neuronal counts, parasympathetic ganglia, such as the ciliary ganglia, seemed not to be affected (Pearson and Pytel, 1978a).

NEUROPHYSIOLOGY OF AUTONOMIC FAILURE Denervation extending to chemoreceptors and baroreceptor functioning in FD is strongly suggested by physiological studies. Baroreflex failure autonomic storms Kaufmann and Norcliffe-Kaufmann et al. (NorcliffeKaufmann and Kaufmann, 2012; Kaufmann et al., 2020) have reviewed the baroreflex neuroanatomy and its dysfunction in FD. Afferent baroreflex mechanosensing neurons with axonal receptors in thoracic arteries of the aortic arch and carotid sinuses, and the heart have their cell bodies in the nodose and petrosal ganglia of the glossopharyngeal and vagal nerves (nerves IX and X respectively), synapsing with neurons in the medulla oblongata in the nucleus of the NTS. From the NTS, there are three important baroreflex pathways as described by Kaufmann et al. (2020).

One inhibitory pathway restrains sympathetic outflow to the vasculature through interneurons in the caudal ventrolateral medulla via neurons in the NTS that inhibit sympathetic (premotor) pacemaker neurons in the rostral ventrolateral medulla that are the source of sympathetic activity to the vasculature, and barosensitive axons that descend through the spinal cord and activate the twoneuron (preganglionic and postganglionic) sympathetic efferent pathway to skeletal muscle and mesenteric and renal vessels. A second pathway activates vagal efferents to slow the heart rate through a direct projection to preganglionic parasympathetic neurons in the nucleus ambiguus of the medulla, which activate postganglionic parasympathetic neurons to the sinoatrial node. A third pathway connects the NTS with the supraoptic nucleus and the PVN in the hypothalamus, controlling arginine vasopressin release from the pituitary. In addition, there are renal afferents and muscle ergoreceptors (thinly myelinated group III and IV afferents) that reach the spinal cord through DRG with important modulatory effects on the baroreflex. FD is characterized by an absence of afferent baroreceptor input due to depletion of primary peripheral afferent neurons (Fogelson et al., 1967). The genetic deficiency in FD specifically affects the afferent baroreflex pathways leaving the efferent sympathetic nerve reduced in number but functionally active (NorcliffeKaufmann et al., 2010). From a pathophysiological point of view, marked blood pressure instability is attributed to impaired baroreceptive inputs. Without mechanosensing afferent inputs, neurons in the nucleus of the solitary tract fail to inhibit brainstem premotor sympathetic neurons responsible for basal and reflex control of sympathetic activity (Mischel et al., 2015) that project efferent fibers upon sympathetic preganglionic neurons in the IML column of the spinal cord (Guyenet, 2006). As in acquired afferent baroreflex failure, direct recordings show that sympathetic efferent activity is no longer coupled to the cardiac cycle (Norcliffe-Kaufmann et al., 2017). Complete failure of the baroreceptor afferents results in orthostatic hypotension with a paradoxical slowing of the heart rate that is not blocked by atropine, indicating that it is not mediated by activation of vagal efferents (Norcliffe-Kaufmann et al., 2010). Slowing of the heart rate in patients with familial dysautonomia when they are in the standing position may be the result of decreased right atrial filling, as observed in denervated heart preparations. Increasing right atrial pressure increases the heart rate in the denervated heart, and increasing right atrial filling in patients with familial dysautonomia by placing them in the head-down position also raises their heart rate, perhaps revealing the intrinsic responses of a deafferented heart (NorcliffeKaufmann et al., 2010).

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS Historically, human physiological studies in FD patients showed absent corrective responses in heart rate responses, cardiac tone, heart rate variability, and rate of norepinephrine release to postural changes implicating extensive sympathetic as well as parasympathetic responses (Ziegler et al., 1976; Axelrod et al., 1997). In addition, there can be impaired cerebral autoregulation and paradoxical cerebral vasoconstriction during HUT (Hilz et al., 1996, 1997). Hilz et al. (2002) postulate reactive myogenic cerebral arterial wall thickening that compensates for decreased sympathetic vascular innervation. Supine plasma levels of norepinephrine are normal or elevated like most other patients with neurogenic OH, however FD patients do not show an appropriate increase in plasma levels of norepinephrine and dopamine betahydroxylase (DbH) with standing (Ziegler et al., 1976; Axelrod et al., 1996). In addition, they have a distinctive pattern of plasma levels of catechols such that regardless of posture, plasma levels of dihydroxyphenylalanine (DOPA) are disproportionately high and plasma levels of dihydroxyphenylglycol (DHPG) are low, resulting in elevated plasma DOPA:DHPG ratios atypical for other cases of neurogenic OH (Axelrod et al., 1996). Increased plasma DOPA levels in FD patients is consistent with an increased proportion of tyrosine hydroxylase in superior cervical ganglia (Pearson et al., 1979) that partially compensates for the decreased numbers of sympathetic neurons. When FD subjects are supine, there is a strong correlation between mean blood pressure and plasma levels of norepinephrine, but when they are upright, the correlation is seen only with plasma dopamine levels, suggesting that the increased dopamine may serve to maintain upright blood pressure. A characteristic feature of FD is recurrent episodes of vomiting accompanied by hypertension, tachycardia, red blotching of the skin and profuse diaphoresis triggered by emotional arousal or other stresses (Riley et al., 1949; Norcliffe-Kaufmann et al., 2010). Similar episodes occur in patients with acquired lesions in the afferent baroreflex pathways, but without the characteristic vomiting that occurs in patients with FD (Robertson et al., 1993; Biaggioni et al., 1994). During such autonomic storms, surges in sympathetic activity are not restrained by the normal afferent baroreceptor feedback. At times of emotional arousal increases in blood pressure and heart rate are amplified and prolonged (Fagius et al., 1985). Hypertensive vomiting attacks in patients with FD, are accompanied by unrestrained release of catecholamines (Norcliffe-Kaufmann et al., 2010). Plasma norepinephrine and dopamine levels rise markedly, the former due to the peripheral conversion of dopamine by DbH and vomiting that later ensues is coincident with high dopamine levels in contact with area postrema of the medulla.

87

Chemoreflex failure Inherent chemosensory deficits in FD result in ventilatory insufficiency associated with hypoxia, often with sleep-disordered breathing and daytime hypoventilation (Kazachkov et al., 2018). The lack of information from chemosensory receptor neurons in the carotid and aortic bodies underlies chemoreflex failure in FD. These receptors that sense the partial pressure of oxygen in arterial blood relay their signals through cranial nerve IX to the nucleus of the solitary tract in the medulla. While ventilatory responses to hypercapnia and to metabolic acidosis mediated by central chemoreceptors are partially preserved, those to hypoxia mediated by peripheral chemoreceptors are almost absent in all cases of FD (Edelman et al., 1970; Maayan et al., 1992; Bernardi et al., 2003). In response to hypoxia, patients may develop paradoxical hypotension, hypoventilation, bradycardia, and, potentially, death (Filler et al., 1965; McNicholas et al., 1983). Rapid correction of hypoxemia with 100% inspired O2 leads to prolonged apnea (Bernardi et al., 2003) whereas breath-holding spells potentially lead to hypotension and syncope occur in more than one-half of affected patients with FD (Maayan et al., 2015). Sleep-disordered breathing occurs overall in 85% of adults and 91% of pediatric patients with FD with signs of obstructive and central sleep apnea in up to one-half of cases (Singh et al., 2018). Sleep disorder breathing is a consequence of chemoreflex failure causing impaired ventilatory drive, neuromuscular dysfunction causing or aggravating upper airway obstruction, and chronic lung disease (Filler et al., 1965; Edelman et al., 1970; McNicholas et al., 1983; Bernardi et al., 2003; WeeseMayer et al., 2008). Untreated sleep apnea is a risk factor for sudden unexpected death during sleep (SUDS) in FD (Palma et al., 2017). Daytime hypoventilation in FD occurs due to chemoreflex failure, which causes impaired ventilatory drive, and restrictive lung disease largely due to kyphoscoliosis and abnormal proprioception. In clinical practice, a lower threshold of pCO2 45 mm Hg is used to define daytime hypercapnia in patients with FD. Hypoventilation is exaggerated by the use of sedative medications or drugs that induce metabolic alkalosis such as fludrocortisone, frequently used in this population for the treatment of orthostatic hypotension

PROGNOSIS Survival has been improving for patients with familial dysautonomia with mortality rates of 50% prior to 1960 typically before 5 years of age (Brunt and McKusick, 1970). By the turn of the century, a newborn

88

D.S. YOUNGER

with FD had a 50% probability of reaching 40 years of age (Axelrod et al., 2002) and QoL had also improved with many FD adults able to achieve independent function. Both men and women with FD married and reproduced with normal phenotype prodigy despite their obligatory heterozygote state. With an aging FD population there has been evidence for the progressive nature of the disorder with many adult FD patients appreciating a decline in sensory abilities with commensurately poor balance and unsteady gait; and worsening OH, supine hypertension, and cardiac bradyarrhythmia (Axelrod et al., 2002). Mortality is less often related to pulmonary complications indicating that aggressive treatment of aspirations has been beneficial (Axelrod and Abularrage, 1982; Axelrod et al., 2002).

TREATMENT OF ORTHOSTATIC HYPOTENSION Management of OH is similar to other disorders of autonomic failure with some caveats. Patients with FD are unable to increase sympathetic activity in association the demands for an adequate blood pressure in response to exercise and warm environments. Similarly, concomitant illness that predisposed an FD patient to volume depletion, anemia, or hypoxia particularly in association with gastrointestinal bleeding, dehydration, mucus plugging, and intercurrent pulmonary infections are at greatest risk for syncope and should be aggressively managed and empirically treated. Breath-holding spells should be considered in the differential diagnosis of syncope, especially in younger children. Nonpharmacological interventions to improve OH should be implemented before drug therapy.

Nonpharmacologic therapy Symptomatic patients with OH should be instructed to avoid potential triggers such as standing immobile for a long time and standing quickly after long periods sitting. Physical counter-maneuvers to raise venous return and increase blood pressure are effective. Typical maneuvers are leg crossing, wiggling the toes, muscle tensing, abdominal compression and even chewing gum can lessen orthostatic hypotension (Hilz et al., 2012). A physical therapy program with emphasis on increasing muscle strength in the extremities can be helpful. Sleeping with the head of the bed elevated raises intravascular volume and blood pressure in the morning on awakening. Adequate hydration and liberal salt intake are essential, although the immediate osmopressor response triggered by water loading appears to be absent in FD (Goulding et al., 2013).

Pharmacologic therapy Midodrine can be used safely in patients with FD as needed rather than on a fixed schedule, and at least 45 min before physical activity, so that the pressor effect coincides with exercise-induced hypotension, but not within 3 h of bedtime or when blood pressure is elevated such as during an autonomic crisis. Fludrocortisone can be highly effective but has some potential adverse effects including the potential worsening of sleep-disordered breathing, decline of glomerular filtration rate, induction of metabolic alkalosis, hypercapnia due to respiratory depression, and a heightened risk of SUDS especially when administered over long periods of time (Norcliffe-Kaufmann et al., 2013b; Palma et al., 2014). Anemia of chronic disease, which can occur in patients with FD responds to erythropoietin treatment by increasing the red blood cell mass and improving the oxygen carrying capacity of blood, and reducing circulating levels of nitric oxide levels, thereby raising blood pressure (Perera et al., 1995). However, such patients should be closely monitored to avoid polycythemia or sustained hypertension.

TREATMENT OF VOMITING ATTACKS AND HYPERTENSIVE CRISES

The normal baroreflex buffering that prevents blood pressure from rising and falling excessively is absent in FD, thus affected patients have both hypertension and hypotension and its management is challenging as aggressive treatment of either one exacerbates the other. Over time, this excessive blood pressure variability is associated with target organ damage, and treatment is geared toward lessening overall blood pressure variability, while the appropriate management of hypertension ultimately depends upon its associated circumstances, that includes transient surges of hypertension and tachycardia with vomiting in everyday activities or attacks of autonomic crises, and even sustained hypertension associated with progressive renal impairment.

Transient surges with everyday activities Such surges in hypertension are common when awakening from sleep, while eating, or with heightened anxious, excitement or frustration from infancy where it can be classically associated with erythematous blotching and diaphoresis (Geltzer et al., 1964) and managed with relaxation and distraction, or at older ages with standing or walking.

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS Vomiting attacks with autonomic crises Hypertensive vomiting attacks are a characteristic of FD when patients are stimulated emotionally, by infection, illness and surgery, constipation or bladder distension and other factors that stimulate increase sympathetic outflow with sustained catecholamine release and excessive circulating plasma dopamine that triggers vomiting (Norcliffe-Kaufmann et al., 2013a). Prevention is always preferable, however early and regular use of pregabalin with its anxiolytic and analgesic action, desired tolerance and bioavailability and rapid absorption makes it an ideal starting point in averting autonomic crises (Axelrod and Berlin, 2009). Once nausea is followed by vomiting antiemetics are seldom useful, and acute treatment with intranasal midazolam can be effective in reducing retching (Lahat et al., 2000), however benzodiazepines can suppress ventilatory drive in patients with FD and caution should be expected in anticipated the need for noninvasive ventilation with their use. Although there have been no controlled trials, the centrally acting a2-adrenergic agonist clonidine can be effective therapy to restrain sympathetic outflow during hypertensive vomiting attacks (Marthol et al., 2003), and worn as a patch to deliver stable blood levels. Carbidopa was found to be a safe and effective antiemetic in patient with FD likely due to its formation of dopamine outside the brain in a randomized, double-blind, placebo-controlled, crossover efficacy study (Norcliffe-Kaufmann et al., 2013c) at the average dose of 480 mg (range 325–600 mg). Unlike other treatment options, carbidopa was not associated with sedation or respiratory depression, making it suitable as a long-term pharmacotherapy to prevent vomiting attacks. The downstream effects of carbidopa on norepinephrine production may also be useful to dampen hypertensive surges and lessen overall blood pressure variability. Finally, as hypertension and tachycardia typically go hand-in-hand, agents that restrain or block sympathetic outflow such as metoprolol can be used safely in patients with cardiac and pulmonary disease to lessen tachycardia and reduce blood pressure through b-1 selective blockade.

DISEASE MODIFYING THERAPY Two therapeutic approaches incorporating small molecules and antisense nucleotides have been investigated.

Small molecules The small molecule kinetin (Gold-von Simson et al., 2009) has a predicted mode of action that experimentally increases the production of ELP1 mRNA in FD cell lines

89

by altering splicing in vivo that underlies the causative intronic splice mutation in the gene that leads to partial skipping of exon 20 and the specific reduction of the encoded protein product (Gold-von Simson et al., 2009). Daily administration of kinetin starting at birth in experimental mice with FD showed phenotypic improvement that correlated with full-length ELP1 mRNA (Morini et al., 2019).

Antisense oligonucleotides Splice-switching antisense oligonucleotides (ASOs) have been effective targeted therapeutics for neurodegenerative diseases, such as nusinersen in spinal muscular atrophy. Using a two-step screen with ASOs targeting the ILP1 exon 20 or the adjoining intronic regions investigators (Sinha et al., 2018) identified a lead ASO that fully restored exon 20 splicing in FD patient fibroblasts. When administered into a transgenic FD mouse model, the lead ASO promoted expression of full-length human ILP1 mRNA and protein product levels in several tissues tested, including the CNS.

AUTOIMMUNE AUTONOMIC FAILURE Background “He is a pink complexioned person, except when he has stood for a long time, when he may get pale and faint. His handshake is warm and dry. … He is thin because his appetite is modest; he never feels hunger pains and his stomach never rumbles. … As old age comes on he will suffer from retention of urine and impotence but frequency, precipitancy, and strangury will not worry him.” Paton’s description of the “hexamethonium man” (Paton, 1954) illustrates the consequences of a pharmacologic blockade of ganglionic transmission. Hexamethonium blocks ganglionic nicotinic acetylcholine receptors (nAChRs), resulting in dysfunction of the efferent sympathetic and parasympathetic pathways. Young et al. (1969, 1975) described acute pandysautonomia characterized by severe sympathetic and parasympathetic failure with relative preservation of motor and sensory function. Their report was remarkable for the relative purity of autonomic involvement and the patient’s complete recovery. However, in subsequent reports the degree of recovery was less complete (Fagius et al., 1983; Low et al., 1983; Stoll et al., 1991) than the original case. Still other reported cases of pandysautonomia (Appenzeller and Kornfeld, 1973; Yahr and Frontera, 1975; Colan et al., 1980; Neville and Sladen, 1984; Taubner and Salanova, 1984; Hart and Kanter, 1990; Feldman et al., 1991; Pavesi et al., 1992) characterized

90

D.S. YOUNGER

a heterogeneity of onset, and autonomic and somatic deficits. The underlying mechanism of acute autonomic neuropathies was unknown, but an immune-mediated pathogenesis was suggested. A case series by Suarez et al. (1994) of so called idiopathic autonomic neuropathy described the characteristic orthostatic, gastrointestinal and sudomotor autonomic features in 27 patients with severe adrenergic and cholinergic failure, manifesting OH, anhidrosis and bowel and bladder disturbances. The majority of patients had a monophasic course with either a progressive phase followed by a plateau and remission, or prolonged stable deficit without remission or recurrences. In a series of publications, Vernino and colleagues identified ganglionic nAChRs in cases of subacute autonomic neuropathy and cancer-related syndromes (Vernino et al., 1998), autoimmune autonomic neuropathies (AANs) (Vernino et al., 2000). This was followed by the report of an animal model of ganglionic nAChR autoimmunity produced by immunizing rabbits with a recombinant a3 neuronal nAChR subunit fusion protein (Lennon et al., 2003), which in response to a single immunization, the animals developed a chronic dysautonomic syndrome so termed, experimental AAN (EAAN) with overt signs of autonomic failure. Using quantitative measures in autonomic function, the same authors (Vernino et al., 2003) demonstrated that EAAN in the rabbit reproduced the phenotype of AAN in humans namely a generalized dysautonomia with prominent cholinergic failure including a prominence of parasympathetic and enteric abnormalities typical of AAN (Klein et al., 2003), with marked loss of postsynaptic nAChR on neurons in autonomic ganglia. A plausible mechanism for the loss of surface nAChR was crosslinking of surface nAChR by ganglionic nAChR-specific antibody leading to accelerated internalization and degradation as had been demonstrated in myasthenia gravis (Drachman et al., 1978) and EAMG (Lennon, 1978). In chronic EAAN, the density of neurons in paravertebral sympathetic ganglia appeared to be normal, but there was a reduction in the number of neurons in enteric plexus suggesting some autonomic neurons sustained immunocytotoxic or apoptotic cell death. Inflammation and loss of ganglionic neurons had previously been documented in biopsied bowel of patients with paraneoplastic AAN (Lennon et al., 1991). In the same year, Vernino and Lennon (2003) described a validated assay to detect ganglionic nAChR antibodies using ganglionic AChR solubilized from membranes of the human neuroblastoma cell line (IMR-32) as antigen, that radiolabeled by complexing with 125 I-epibatidine, detecting one-half of the cases with subacute AAN, often with high titers (>0.2 nmol/L) but also with lower values (0.05–0.2) in up to 10% of cases with limited AAN.

Illustrative case with antemortem and postmortem pathology A patient with AAG is described below from the author’s file illustrating salient antemortem and postmortem histopathological features associated with autonomic failure respectively including unmyelinated peripheral nerve fiber depletion in both sural nerve and epidermal biopsy tissue, clinicopathological features underlying neurocognitive decline, and malignant cancer. In the summer of 2016, a 78-year-old man with numbness, tingling and limb pain had lightheadedness and cognitive decline for 1 year. He was treated for prior Lyme exposures. Neurological examination in June 2016 showed impaired short-term registration, stocking vibratory and cold temperature sensory loss, Romberg sign, tandem imbalance, grade 4+/5 leg distal leg strength, hyporeflexia, and Babinski signs. Autonomic testing (WR-Testworks, Minnesota [MN]) showed a supine SBP of 134 mm Hg and heart rate (HR) of 57 bpm. HUT led to symptomatic orthostasis with a minimum SBP of 70 mm Hg at 3.6 min without a compensatory HR change, followed by a period of prolonged hypotension and near-syncope. Electrodiagnostic studies showed a distal left fibula compound muscle action potential amplitude of 0.3 mV and motor nerve conduction velocity of 30 m/s. Bilateral tibial and right fibular motor, and bilateral superficial fibular and sural sensory responses were absent. Concentric needle electromyography showed chronic neurogenic changes without active or chronic denervation at rest. Epidermal nerve fiber densities (Fig. 4.15A) were consistent with a nonlengthdependent process (mean left calf: 3.2 ENF/mm; mean left thigh 2.3 ENF/mm) consistent with small fiber peripheral neuropathy. Left sural nerve biopsy showed excessive thinly myelinated fibers in epoxy resin sections. Left soleus muscle biopsy (Fig. 4.15B) showed minimal neurogenic changes and perivascular inflammation. Blood studies showed total prostate specific antigen 4.6 ng/mL (normal 4 ng/mL). Whole body 18fluorodeoxyglucose PET showed temporoparietal and frontal lobe hypometabolism, with a focus of intracapsular prostatic cancer, later confirmed on magnetic resonance imaging of the pelvis. However, there were discernible metastatic foci. The CSF protein was 65 mg/dL (normal >50 mg/dL) without inflammation or infection. The Mayo Clinic (Rochester, MN) Autoimmune Dementia panel in serum and CSF showed a serum a3gAChR antibody level of 0.05 nmol/L (normal 0.02 nmol/L) with otherwise normal values. Athena Diagnostics (Marlborough, MA) ADmark® analysis was inconclusive for symptomatic Alzheimer disease. The patient was treated with 6 months of 2 g/kg of intravenous immunoglobulin (IVIg) with stabilization of

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS

91

Fig. 4.15. (A–D) Antemortem and postmortem pathology of autoimmune autonomic ganglionopathy. Top to bottom and left to right: (A) Epidermal nerve fiber analysis. There are reduced densities of epidermal nerve fibers in the calf and thigh (shown) (PGP 9.5 & Eosin, 200
); (B) epoxy resin transverse section of nerve. There is a moderate degree of axonal loss, and occasional regenerative clusters and thinly myelinated fibers in each fascicle suggesting segmental remyelination (Toluidine blue,
600); (C) postmortem hippocampus brain section. Typical plaques and tangles of Alzheimer disease are evident in the CA1 subsector (Bielschowsky silver stain, 40
); (D) postmortem autonomic ganglia. An autonomic ganglion with well-populated neurons (Hematoxylin and eosin, Luxol, 20
).

the autonomic disorder. However, after discontinuation of IVIg there was continued deterioration and death in June 2018 in hospice. Postmortem examination showed foci of intracapsular well-differentiated prostatic adenocarcinoma; neuritic plaques and neurofibrillary tangles (Fig. 4.15C) in sections of hippocampus, amygdala and association cortex without microglial nodules or inflammation. Spinal nerve roots and peripheral nerves showed patchy myelin loss. Autonomic ganglia were unremarkable (Fig. 4.15D).

Neurophysiology Vernino et al. (2009) have reviewed the neurophysiology of the ganglionic nicotinic AChR. Peripheral autonomic components include groups of neurons (ganglia) with extensive synaptic connections outside the central nervous system that project to the periphery and synapse with neurons in autonomic ganglia. Within ganglia, the

peripheral autonomic neurons, especially in the intrinsic ENS, synapse extensively with each other, and project axons (postganglionic unmyelinated C fibers) innervating target organs. Fast synaptic transmission within autonomic ganglia is mediated by nAChRs. In mammalian ganglionic synapses these receptors derive from a family of ligand-gated cation channels comprising two a3 subunits combined with three b4 subunits (Skok et al., 1999). Transgenic mice lacking the a3 subunit have an ionic channel defect leading to profound autonomic failure with prominent bladder distention, gastrointestinal dysmotility and lack of pupillary light reflexes (Xu et al., 1999) suggesting that the a3 subunit is absolutely required for normal autonomic ganglionic neurotransmission. Autonomic ganglia are more than simple relay centers for autonomic information since there is significant signal integration of synaptic inputs. The synaptic strength and the degree of integration varies widely

92

D.S. YOUNGER

among different autonomic ganglia. Autonomic ganglionic transmission depends on the convergence of multiple subthreshold synaptic events. However, antiganglionic AChR antibodies can profoundly affect synaptic transmission by interfering with the interaction of ACh at ganglionic neuronal AChRs that produce fast excitatory postsynaptic potentials and summate to reach threshold in order to propagate action potentials (AP) along postganglionic axon to its target innervation. Although conceptually, autonomic disorders can be categorized as central (contributing preganglionic autonomic fibers) or peripheral (postganglionic) disorders affecting the neurons of the autonomic ganglia and unmyelinated or thinly myelinated peripheral axons, a third category of autonomic disorders in which the pathology lies within the autonomic ganglia itself is suggested by AAG (Vernino et al., 2009).

Serology While ganglionic antibodies in high titers appear to be highly specific and provided a tool for the quantitative and sensitive detection of AAG, with a robust relation between titers and clinical autonomic severity, only 50% of cases of severe diffuse AAG were detected (Vernino et al., 2009), presumably due to antibodies directed at different targets in the nerve including the ganglion (Sandroni and Low, 2009) leading to heterogeneous manifestations of autonomic function. Studies in patients with AAG showed postganglionic sudomotor dysfunction (Kimpinski et al., 2009, 2012), pathological evidence of postganglionic autonomic and somatic nerve fiber loss (Koike et al., 2013) and reduced density of intraepidermal nerve fibers (Manganelli et al., 2011) and multimodal biomarkers (Koay et al., 2021) to quantifying recovery after immunotherapies in tissue specimens, suggesting concomitant postganglionic denervation. Other studies noted AAG in association with reversible cognitive impairment (Gibbons et al., 2012) and late-onset encephalopathy (Baker et al., 2009) with elevated titers of a3-AChR antibodies, in addition to binding at CNS a4 and a7-nAChRs expressed in cerebral cortex and hippocampus (S
egu
ela et al., 1993), and pathophysiologically depleted (a4) in Alzheimer brains (Court et al., 2001; Nordberg, 2001).

Diagnosis and outcome The prototypical AAG case is a previously healthy young or middle-aged subject, more likely to be female, presenting with a severe panautonomic failure that evolves over days to weeks possibly following inadvertent infection, recent immunization, or surgical procedure, with monophasic slow progression and incomplete recovery despite immunotherapy. The presenting and evolving

clinical picture are dominated by OH, Horner syndrome, bradycardia and anhidrosis due to SNS failure. Sicca symptoms with loss of lacrimation and salivation, and Adie pupils are a result of PaNS failure. Gastroparesis, ileus, constipation, and diarrhea, progressing to achalasia or intestinal pseudo-obstruction, are a consequence of ENS failure, with delayed transit time often seen scintigraphically on motility studies, such as esophageal manometry, gastric emptying study, and small bowel transit. Urinary retention, incontinence, erectile and ejaculatory dysfunction due to a combination of PaNS and SNS failure. Impaired late phase II and/or phase IV responses on Valsalva testing signify vascular and cardiac adrenergic impairment while reduced heart rate variation to deep breathing and Valsalva signifies cardiac vagal impairment. Secretomotor findings of global anhidrosis or hypohidrosis on QSART or TST should raise suspicion of AAG. Affected cases with the classic AAG phenotype are likely to manifest high a3gAChR antibody levels that not only correlate with the severity (Vernino et al., 2000; Klein et al., 2003) and spread of autonomic dysfunction, but decline with clinical improvement after immunotherapy. AAG is a rare disease of unknown prevalence that stems from several factors. First, in the lack of an international consensus regarding sufficient clinical and serological criteria for diagnosis. Second, in the serological testing protocol and cohort studied in which ganglionic nAChR antibodies are detected in 50% (Vernino and Lennon, 2003) to 72% (Vernino et al., 2008) of patients. Moreover, while there is absolute consensus that the highest titers of a3-AChR antibodies correspond to cases of panautonomic failure, and lower values unpredictive of AAG; there is divergence in the interpretation of lower values, with some interpreting them as lacking autoimmunity or a frank neurological basis whatsoever (McKeon et al., 2009), and others taking a more cautious view of the difficulty in separating low titer cases from the seronegative, as more likely indicative of an underlying autoimmune response (even one directly related to dysautonomia) yet suggestive of another autoantibody or one yet to be identified (Sandroni and Low, 2009). Koay et al. (2021) described a multimodal approach to the evaluation of a cohort of patients to characterize their phenotype and quantify the underlying immune response. Of the 13 patients seen between 2005 and 2019, 7 were women, age 21–69 years, with autonomic failure and circulating a3gAChR antibodies (>100 pM; 0.1 nmol/L). Patients were longitudinally assessed with cardiovascular, pupillary, urinary, sudomotor, lacrimal and salivary testing, and COMPASS-31 and autonomic symptom questionnaires. At presentation, all patients had widespread sympathetic and parasympathetic cardiovascular autonomic failure with OH, reduced/absent HRRDB,

AUTONOMIC FAILURE: CLINICOPATHOLOGIC, PHYSIOLOGIC, AND GENETIC ASPECTS and impaired heart rate and blood pressure responses to the Valsalva maneuver. One patient had very low levels of supine noradrenaline ( resting tremor predominantly involving the upper limbs, and is considered the most common movement disorder, affecting more than 60 million people worldwide. It is typically associated with abnormal connectivity of the cerebellum with brainstem and basal ganglia structures. Several genetic loci have been associated with ET. There has been one study finding an association between a TREM2

142

Y. MAHJOUB AND D. MARTINO

variant and ET which remains to be replicated (OrtegaCubero et al., 2015). Interestingly, a study of 10 patients with ET undergoing deep brain stimulation (DBS) performed proteomic analysis of compared pre- and post-op CSF and identified downregulation of proteins implicated in complement activation, humoral immune response, and the acute inflammatory response (Zsigmond et al., 2020). There has been growing evidence suggesting a role of the immune system in pathogenesis of Tourette syndrome (TS), a neurodevelopmental disorder characterized by multiple motor and vocal tics, and closely associated with OCD. GWAS have found genetic correlation between TS and allergies (Tylee et al., 2018). Transcriptomic analyses of postmortem TS brains have identified altered expression of immune-related genes within the basal ganglia, including upregulation of IL-1b and IL-2 receptor b transcripts compared to controls (Hong et al., 2004), and an RNA-sequencing study in the striatum of TS subjects identified upregulated genes involved in activation of microglia (Lennington et al., 2016). In live subjects, one small study using PET imaging found increased binding of a ligand binding a transporter protein expressed by activated microglia within the caudates of children with TS compared to controls (Kumar et al., 2015). The presence of antibasal ganglia antibodies in TS has been demonstrated (Kiessling et al., 1993; Rizzo et al., 2006), and more recent studies have identified antidopamine D2 autoantibodies in some children with tic disorders or TS (Dale et al., 2012; Addabbo et al., 2020). While studies attempting to define specific peripheral immunophenotypes of TS have yielded conflicting results, nevertheless these studies do hint at differences between TS and controls, and interpretation may be challenging given the clinically heterogeneous and fluctuating nature of the disorder. Studies of serum proportions of specific immune cell types have had variable results that require replication (Kawikova et al., 2007; M€ oller et al., 2008; Li et al., 2015; Yildirim et al., 2020). Similarly, findings around immune molecule concentrations in TS have been discordant and require replication. For example, while some have reported higher TNF-a levels (Leckman et al., 2005; Li et al., 2015; Yeon et al., 2017) and its elevation during tic exacerbations (Parker-Athill et al., 2015), others have described no difference (BosVeneman et al., 2010) or even lower TNF-a (Matz et al., 2012).

THE IMMUNE SYSTEM IN CEREBELLAR ATAXIAS Moving from the basal ganglia to the cerebellum, ataxias are a heterogeneous group of disorders that are clinically defined by impairments in balance and coordination.

There exist a variety of immune-mediated cerebellar ataxias, with clearly established pathological bases. These include paraneoplastic cerebellar degeneration, gluten ataxia, postinfectious cerebellitis, and neuronal autoantibody-mediated conditions such as anti-DPPX, anti-mGluR1, anti-GABABR, or anti-GAD65 antibodies (Hadjivassiliou et al., 2020). The term primary autoimmune cerebellar ataxia is used to refer to suspected immune-mediated cerebellar ataxias with either no identified autoantibody, or with identification of an antibody not convincingly associated with ataxia (Hadjivassiliou et al., 2020). Nonacquired causes of ataxia, on the other hand, include hereditary and sporadic cases. In a similar vein to previously described conditions, microglia seem to play a role in neurodegeneration within this heterogeneous group of disorders. Activated microglia in the cerebellum have been shown in animal models of spinocerebellar ataxia 6 (SCA6) (Aikawa et al., 2015), SCA21 (Seki et al., 2018). An ataxia-telangiectasia ATM gene knockout mouse model resulted in dysfunctional microglia, with impairment of phagocytosis, neurotrophic factor production, and mitochondrial activity (Levi et al., 2022). There may, therefore, be a particular role for microglia in cerebellar damage in several ataxias (Ferro et al., 2019).

THE IMMUNE SYSTEM IN AMYOTROPHIC LATERAL SCLEROSIS

Neuroinflammation is posited to play a role in the pathogenesis and progression of amyotrophic lateral sclerosis (ALS), which is the quintessential neurodegenerative disorder affecting motor neurons, involving patients between the end of the sixth and the seventh decade of life, is highly fatal and currently lacking a cure. Despite its relatively low prevalence (about 5.4 cases per 100,000), it is acknowledged to have a high public health impact. ALS has a multifactorial pathogenesis, with different lifestyle factors (e.g., lower body mass index, high physical fitness, smoking) and environmental factors (including some neurotoxins) called into play. Several genes linked to ALS encode proteins that influence the immune system. For example, hexanucleotide repeat expansions in the C9orf72 gene are recognized in association with ALS as well as frontotemporal dementia (DeJesus-Hernandez et al., 2011). Although the implication in humans remains uncertain, animal studies have shown deletion of C9orf72 to have several immunealtering effects in myeloid cells (macrophages and macroglia) including increased IL-6 and IL-1b and lysosomal accumulation (O’Rourke et al., 2016). Superoxide dismutase (SOD1) gene variants are also implicated in ALS; SOD1-mutant microglia in vitro enhanced neurotoxicity and led to increased motor neuron injury in cocultures likely through a nitric oxide-mediated

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS mechanism (Xiao et al., 2007). Mutations in the TARDBP gene (encoding the protein TDP-43) have been shown to result in microglial activation with upregulation of NOX-2, TNF-a, and IL-1b via binding to microglial CD-14 receptors and triggering proinflammatory signaling pathways (Zhao et al., 2015). The interplay between microglia and astrocytes, and their neuroprotective and neurotoxic phenotypes, may play a role in ALS. Insights form animal models suggest a shift in the phenotype of microglia from the more neuroprotective M2 early on to the more neurotoxic M1 phenotype in later stages of the disease (Liao et al., 2012). Using PET imaging, presence of activated microglia in the motor cortex, dorsolateral prefrontal cortex, thalamus, and pons in ALS has been suggested (Turner et al., 2004). Moreover, staining of postmortem tissue from ALS subjects has identified greater numbers of activated astrocytes in the motor cortex compared to controls (Liddelow et al., 2017). In a mouse model of ALS, knocking out IL-1a, TNFa, and C1q, which are microglial factors that can activate astrocytes, resulted in extended survival (Guttenplan et al., 2020). Peripheral immune cells are also implicated in ALS. NK cells were shown in two mouse models of ALS to accumulate in the motor cortex and spinal cord (Garofalo et al., 2020). Peripheral blood monocytes in ALS have been shown to exhibit a unique proinflammatory gene profile, with increased mRNA expression of IL-1b, IL-8, FOSb, CXCL1, and CXCL2 (Zhao et al., 2017). While CD4+ Th cells have been found to be protective in animal models of ALS (Beers et al., 2008), CD8+ Tc cells exert cytotoxic effects via Fas ligand and granzyme b pathway (Coque et al., 2019) during the late stages of an animal ALS model. Further, the relative proportion of Treg may also influence disease progression. In animal studies, Treg concentrations were higher during earlier stages (along with neuroprotective M2 microglia) compared to later stages of faster disease progression when Treg numbers dropped; this was corroborated in ALS patients as well who had lower levels of blood FoxP3+ Tregs during rapid progression stages (Beers et al., 2011). Of special therapeutic relevance, a small phase 1 study of autologous Treg infusions in ALS, as well as a small study of combined Treg/IL-2 infusions demonstrated safety and potential benefit; a larger phase 2 study is currently underway to further investigate this potential treatment (Thonhoff et al., 2018, 2022). Dysregulation of Transforming Growth Factor beta (TGF-b) systems has been studied in ALS, and increased plasma levels of this cytokine have been reported in ALS (Houi et al., 2002). Astrocytes in both patients and animal models of ALS have elevated levels of TGF-b1; this overproduction seems to enhance progression in animal models via inhibiting neuroprotective inflammatory processes by microglia and T cells

143

(Endo et al., 2015). While higher levels of TGF-b1 and TGF-b3 have been hypothesized to correlate with increased disease severity, further study is required to establish this observation (Duque et al., 2020; De Marchi et al., 2021) and comparison of ALS patients with fast and slow progression did not reveal any significant differences in TGF-b1 levels (Zubiri et al., 2018).

THE MICROBIOTA-GUT-BRAIN AXIS AND MOTOR SYSTEMS The microbiota-gut-brain axis: Role in physiology and pathology of motor systems The knowledge of the composition of symbiotic microbial flora (predominantly bacteria) and of its complex functional interrelationship with the human host is advancing at very high speed. The thousands of microbial species that form the human gut microbiota exhibit a broad spectrum of metabolic functions—including synthesis and catabolism of amino acids, neurotransmitters, vitamins, bile acids, and lipids—regulation of immune responses, and regulation of the integrity of the intestinal barrier, among other functions. The pathological role of the gut microbiota is related to the phenomenon of gut dysbiosis, a complex concept that encompasses diverse changes to the homeostasis of the gut microbial environment, among which the loss of commensal microbes, decreased microbial diversity, and overgrowth of commensal constituents that can acquire a pathogenic potential. The result of this is a complex series of changes in the functional interactions with the host and the reduction of the ability of the gut microbiota to resist and adapt to microenvironmental changes, which can lead to stabilization and worsening of the dysbiosis. Clinical studies investigating gut dysbiosis in human pathology aim to identify an association between specific enterotypes, i.e., groups of individuals classified based on the bacteriological composition of their gut microbiota, and disease-related phenotypes (either diagnoses or diagnostic subtypes). One major challenge to decipher the physiologic and pathogenic contribution of the gut microbiota in humans is the profound difference in functional interaction with human tissues shown by different species or strains within the same bacterial genus, family, or phylum (Shanahan et al., 2021). The simple taxonomic characterization of the flora quantitatively associated with physiological functions or disease mechanisms is proving to be insufficient to identify key cellular and molecular pathways involved in these processes (Almeida et al., 2021). Moreover, more than 70% of the species identified from genes included in the Unified Human Gastrointestinal Genome catalog have never been cultured or characterized (Almeida et al., 2021).

144

Y. MAHJOUB AND D. MARTINO

The exploration of the microbiota-gut-brain axis is revealing new insight on mechanistic pathways that could be manipulated to prevent, modify, or reverse CNS pathology. The advent of germ-free animal technologies yielded important findings on critical signals that are necessary to guide brain development, including motor systems. Moreover, the manipulation of gut microbiota within toxic and transgenic models of movement disorders like PD is proving highly informative, although neuroscientists and clinicians are still at the foot of this learning curve. Several neurologic conditions affecting the motor systems worsen with aging. Interestingly, the gut microbiota also changes with aging, due to a variety of factors (Kundu et al., 2017). Lifestyle modifications may play a major part in this. Community-dwelling elderly subjects demonstrate greater interindividual variability in the gut microbiota compared to those living in long-term care facilities (Jeffery et al., 2016). Dietary changes, worse nutritional status and changes in physical activity also appear to be relevant (Claesson et al., 2012; DeJong et al., 2020; Shanahan et al., 2021). Additional important factors include increased comorbidity and frailty, increased exposure to medications, and decline of gut physiology (reduced motility, altered intestinal barrier integrity) and of immune responses. The gut microbiota also contributes to the sustained low-grade inflammation and innate immune system activation that occurs during aging (inflammaging) (Franceschi et al., 2018). Healthier elderly individuals, e.g., centenarians, demonstrate greater gut microbial diversity and greater abundance of families or genera thought to exert protective effects, such as Ruminococcaceae and Akkermansia (Kong et al., 2016; Tuikhar et al., 2019). Early insights on the contribution of gut microbiota to sensorimotor systems came from work in germ-free mice. Increased motor activity, associated with altered gene expression in second messenger pathways and synaptic long-term potentiation implicated in motor control, are key phenotypic features of germ-free mice compared to specific pathogen-free mice with typical gut microbiota (Diaz Heijtz et al., 2011). Germ-free mice exposed to maternal immune activation (MIA), known to display features of autism spectrum disorder, exhibited deficits in the sensorimotor gating of the startle reflex, a known biomarker of sensorimotor processing that is associated with neurodevelopmental conditions characterized by motor behavioral features (Hsiao et al., 2013). When the MIA offspring was repopulated with Bacteroides fragilis, researchers observed an improvement in communication, stereotyped behavior, anxiety-like behavior, and sensorimotor gating. Subsequent research showed how specific bacterial metabolites, e.g., the short-chain fatty acid propionate, may mediate stress-modulated

sensory gating function (Wah et al., 2019). These findings are especially relevant to motor control as sensorimotor gating is processed within the corticobasal ganglia circuitry. A modulatory role of microbiota on striatal plasticity derives also from the fact that antibiotic depletion of gut microbiota increases brain-derived neurotrophic factor expression in the ventral striatum of specific pathogen-free mice (Kiraly et al., 2016). Rodent models have started to shed light on the influence of gut microbiota on the development and release of repetitive behaviors, which can range from behavioral patterns that can be advantageous for spatial exploration or social bonding to pathological behaviors that may prevent adequate, contextual responses to the environment or even lead to self-harm. Small intestinal bacterial overgrowth (SIBO), characterized by increased abundance of Gram-negative bacteria (producing lipopolysaccharide) and anaerobes (Bushyhead and Quigley, 2022), may exert a suppressive effect on repetitive behaviors, such as exploratory circling in the open field test. Interestingly, germ-free mice on the other hand demonstrate increased compulsive-like behavior also focused on spatial exploration, e.g., repetitive digging (Nishino et al., 2013). The development and balanced functioning of CNS dopaminergic pathways are critical for the control of voluntary motor behavior and to protect against the development of movement disorders. Gut microbiota may play an important role in maintaining optimal dopamine bioavailability to the brain, through different mechanisms that mediate the microbiota-gut-brain axis, e.g., vagal afferent pathways, immune pathways, hypothalamic–pituitary–adrenal axis-controlled pathways, and the downhill effect of microbial metabolites. Bacteria belonging to the Bacteroides and Prevotella genera (phylum Bacteroidetes), like Bacteroides uniformis and Prevotella copri, have been found to, respectively, increase and decrease striatal dopamine transporter (DAT) binding of dopamine (Hartstra et al., 2020). This binding is a key modulator of the dopaminergic tone in the nigrostriatal pathway, which is essential to motor selection and control, and is profoundly damaged in PD and atypical Parkinsonian syndromes. Like Bacteroides uniformis, also Lactobacillus plantarum PS128 may increase DAT binding and metabolism of dopamine, as well as of norepinephrine (Liao et al., 2019), whereas a different strain (DR7) of the same Lactobacillus species decreases dopamine conversion into norepinephrine by lowering the activity of dopamine-b-hydroxylase, and dopamine biosynthesis by downregulating tyrosine-hydroxylase (Liu et al., 2020). Lactobacillus rhamnosus was found to increase dopamine concentrations by downregulating its degradation by monoamine oxidase B (Srivastav et al., 2019). Different Clostridium species, including

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS Clostridium tetani, have opposite effects, either by inhibiting dopamine synthesis through downregulation of the dopamine-b-hydroxylase enzyme, or by promoting its degradation into homovanillic acid via the 3,4-dihydroxyphenylacetic acid (DOPAC) intermediate (Taj and Jamil, 2018; Hamamah et al., 2022). Finally, Ruminococcaceae and Lachnospiraceae species were found to decrease the expression of D2-type dopamine receptors in the dorsal striatum of a mouse model of compulsive alcohol seeking (Jadhav et al., 2018).

The microbiota-gut-brain axis in Parkinson disease The contribution of the gut microbiota to the pathogenic mechanisms associated with PD has been the objective of extensive research, with remaining challenges that still prevent us to conclude in favor of specific causative mechanisms (Tan et al., 2022). Several preclinical (rodent) models of PD displayed gut microbiota abnormalities, although the pathogenic contribution to these models remains unclear. The chronic administration of the mitochondrial complex I inhibitor rotenone may cause intestinal hyper-permeability and motor dysfunction in conventionally raised, but not germ-free, mice (Johnson et al., 2018; Bhattarai et al., 2021). In another study using the rotenone model, chronic stress contributed to gut dysbiotic changes leading to worsened gut inflammation, hyper-permeability and a-synuclein accumulation. Other studies using rodent models showed that antibiotic treatment attenuate dopaminergic neurotoxicity induced by toxins like MPTP (Pu et al., 2019) and 6-hydroxydopamine (Koutzoumis et al., 2020), but also pathophysiology in adult mice overexpressing a-synuclein. In the latter model, colonization with microbiota from PD patients worsened motor impairments compared to fecal matter transplants from healthy human donors. Overall, at least 42 families, 102 genera and 44 species of gut bacteria appear differentially abundant in PD patients vs control individuals (Tan et al., 2022). Only about 25% of these findings were replicated in one or more studies. A few meta-analyses, however, highlighted large variability of results across studies, which are attributed to differences in study methodology and populations (Nishiwaki et al., 2020; Romano et al., 2021; Toh et al., 2022). These meta-analyses showed a significant increase of a-diversity (i.e., the diversity of species within a single sample) in PD patients, which was detected also between cohorts from different countries and studies. Similarly, differences in countries and studies contribute—almost 100 times more than diagnosis/disease status—to the variance of b-diversity (i.e., the diversity of species between

145

two different communities) of gut microbiota composition in studies involving PD patients. Table 7.3 summarizes the most consistent findings in altered abundance of gut microbial diversity and abundance between PD patients and healthy controls (Tan et al., 2022). Among the most consistent findings of increased abundance, an increase of Bifidobacteriaceae, Christensenellaceae, and Verrumicrobiaceae at the family level, and of Akkermansia, Bifidobacterium, and Lactobacillus at the genus level can be found. In particular, the increased Akkermansia abundance has been reported also in other neurologic diseases (including multiple system atrophy, progressive supranuclear palsy, Alzheimer’s disease, and multiple sclerosis), suggesting that this finding may be related to a more generic difference between neurologically diseased and healthy status (Tan et al., 2021a). Moreover, to date Akkermansia muciniphila is the only bacterial species for which increased abundance in PD patients has been replicated by more than 2 case–control studies using next generation sequencing and including more than 100 patients (Tan et al., 2022). Among the most consistent findings of decreased abundance, a decrease of Lachnospiraceae at the family level, and of Blautia, Faecalibacterium, and Roseburia at the genus level can be found. This is an interesting finding also because Roseburia is an important short-chain fatty acid (SCFA) producer (see below). At the same time, results on correlations of bacterial abundance and severity of motor and nonmotor features and progression of PD remain highly inconsistent. For some of these abundance differences it remains to be determined whether these are primary or secondary to gastrointestinal dysfunction. In addition, several studies have suggested that anti-Parkinsonian medications (L-dopa, catechol-O-methyl-transferase inhibitors, anticholinergics) may affect the gut microbiota composition, although discrepancies exist across reports (Hill-Burns et al., 2017; Weis et al., 2019; Cosma-Grigorov et al., 2020). Microbiome sequencing studies reported several upregulated functional pathways in PD patients vs controls, which are explored to decipher the pathogenic contribution of gut microbiota to PD. Among these, amino acid degradation with increased proteolytic fermentation and production of harmful amino acid catabolites (e.g., p-cresol, phenylacetylglutamine) were associated with constipation and stool consistency (Cirstea et al., 2020; Sankowski et al., 2020; Vascellari et al., 2020). Others included lipid biosynthesis and degradation of inorganic nutrients, whereas PD status was associated with reduced carbohydrate fermentation (Cirstea et al., 2020; Vascellari et al., 2020; Tan et al., 2021a). The direct measurement of bacterial activities through multiple “omics” platforms, e.g., metatranscriptomics,

146

Y. MAHJOUB AND D. MARTINO

Table 7.3 Summary of gut microbiota abnormalities in Parkinson disease. Findings confirmed by all studies

Findings confirmed by at least 5 studies

Findings confirmed by at least 2 studies, without discrepant findings

Increased b-diversitya Increased abundance of: Bifidobacteriaceae Christensenellaceae Verrumicrobiaceae Akkermansia genus Akkermansia muciniphila Bifidobacterium Lactobacillusb

Increased abundance of: Coriobacteriaceae Desulfovibrionaceae Enterobacteriaceae Eubacteriaceae Lactobacillaceae Rikenellaceae Alistipes Bilophila Catabacter Escherichia Porphyromonas

Decreased abundance of: Lachnospiraceaeb Blautia Faecalibacterium Roseburia

Decreased abundance of: Agathobacter Bacteroides Fusicatenibacter Lachnospira Lachnospiraceae UCG-004 Lachnospiraceae unclassified

Findings confirmed by at least 2 studies, with discrepant findings Increased a-diversityc Increased abundance of: Klebsiella Oscillospira Parabacteroides Streptococcus Veillonella

Decreased abundance of: Prevotella

a

This result was confirmed by all 16 studies. At least one study reported the opposite change in abundance (i.e., decreased abundance for Lactobacillus and increased abundance for Lachnospiraceae. c Explored in 13 of the 16 studies: 4 reported increased a-diversity, 1 decreased a-diversity, and 8 did not detect any abnormality in a-diversity. Abbreviations: N/A. Based upon studies that applied next generation sequencing techniques on more than 100 participants (n ¼ 16 studies). This table summarized data reported in Supplementary Material from Tan et al. (2022). b

metaproteomics, or metabolomics, can inform more comprehensively the relationship between bacterial functions and disease status. Similarly, adequate correlational analyses with complex clinical datasets may accelerate the path toward personalized and precision medicine approaches in the design and selection of microbialdirected therapies in a wide variety of diseases. An important, early metabolomic finding in PD was the reduced production of fecal SCFA (butyrate, acetate, propionate) (Shin et al., 2020; Aho et al., 2021). SCFA, especially butyrate, are a major energy source for colon epithelial cells, and their reduction is linked to disrupted integrity of the intestinal barrier, which allows bacterial large molecules like lipopolysaccharide to enter the bloodstream and activate inflammatory mechanisms, among which microglial activation. Butyrate also modulates gene expression in several cell types, through its potent histone deacetylase inhibitory activity (Srivastav et al., 2019).

Both immune and neural cells express receptors for SCFA, and SCFA can also induce in a subpopulation of enteroendocrine cells, L-cells, the secretion of glucagon-like peptide-1 (GLP-1), an incretin hormone that has neuroprotective effects in PD (Manfready et al., 2022). Initial findings showed a correlation between reduced fecal butyrate and severity of constipation, which has also been linked to worse PD progression over time, severity of cognitive deficits, postural and gait abnormalities, and earlier age at symptom onset (Jones et al., 2020; Tan et al., 2021a). Importantly, discrepancies exist among studies exploring the association between SCFA levels and clinical ratings. More research is needed to understand the importance of dosage, type, and ratio of different SCFA in influencing different health outcomes. Another noteworthy finding is the lack of significant differences in abundance of SCFA-producing bacteria between patients with idiopathic REM sleep behavior disorder (iRBD)—a key

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS prodrome of PD—and healthy controls, which led to the hypothesis that decreased SCFA production could be implicated in the conversion from iRBD to PD (HeintzBuschart et al., 2018). Multiomics studies have also explored the association between gut microbiome and host metabolism in PD, suggesting implications of the gut microbiome in amino acid, folate, and homocysteine metabolism, which may be relevant to both neuroprotective and neurodegenerative effects (Cirstea et al., 2020; Rosario et al., 2021). More research is needed to replicate preliminary findings and resolve discrepancies among existing findings. An additional mechanism that has been called into play is the potential effect of proinflammatory bacterial components like lipopolysaccharide (LPS) and PD pathology. A metagenomics analysis reported greater expression of genes involved in LPS biosynthesis and type III bacterial secretion systems in PD patients. LPS can activate microglia and trigger neurodegeneration, effects that can be fostered by intestinal hyper-permeability. In a mouse model, mice exposed to a-synuclein fibrils seeded in the

147

presence of LPS produced a self-perpetuating strain leading to consistent pathology compared to a-synuclein fibrillation in the absence of LPS (Rosario et al., 2021). An expression quantitative trait loci analysis also reported an association between abundance of different opportunistic bacteria commonly observed in the PD gut ecosystem and genetic variation of the gene SNCA, coding for a-synuclein, thus suggesting an inter-relationship between host genetics and gut dysbiosis in this condition (Cui et al., 2021). Fig. 7.2 summarizes the complexity of the host–gut microbiota interactions potentially at play in PD. Several microbiota manipulation strategies (prebiotics, probiotics, fecal matter transplantation, postbiotics and other small molecule drugs) are under scrutiny as potential disease-modifying interventions in PD (reviewed in depth in Lorente-picón and Laguna, 2021; Alfonsetti et al., 2022; Tan et al., 2022; Zhu et al., 2022). However, none of these has led to sufficiently compelling evidence of efficacy and safety. Table 7.4 summarizes the microbiota manipulation strategies that have been tested in a clinical trial to date, and the related findings. Alongside the

Host intrinsic factors Innate immunity

Adaptive immunity

Gender

Age

Genetics

Frailty ns tio

Host–micr obe int er ac

Comorbidites Gut mucosal changes

Slow transit time

Environmental factors Immune function

Local environment

Functional capacity Metabolic capacity

BMI

Geography

Composition

Structural function

Housing and family

Diet Gut microbiome Physical activity Host extrinsic factors

Stochastic effects Medications

Unknown factors

Fig. 7.2. This diagram summarizes the intrinsic and extrinsic factors that can influence the gut microbiome, thus contributing to a profound variability in gut microbial constituents between individuals. Reproduced with permission from Tan AH, Lim SY, Lang AE (2022). The microbiome-gut-brain axis in Parkinson disease—from basic research to the clinic. Nat Rev Neurol 18: 476–495.

148

Y. MAHJOUB AND D. MARTINO

Table 7.4 Summary of clinical trials of microbiota manipulation studies in Parkinson disease patients. Type of microbiota manipulation strategy

Treatment description

Trial design

Findings

Prebiotic

Balanced, ovo-lacto vegetarian diet intervention including short fatty acids for 2 weeks (Hegelmaier et al., 2020)

Open-label, n ¼ 16 PD patients [10 received additionally colon cleansing via enema]

Prebiotic

Diet rich in insoluble fiber (Astarloa et al., 1992)

Open-label, n ¼ 19 PD patients

Prebiotic

Resistant starch (Becker et al., 2022)

Open-label, n ¼ 57 PD patients [32 received resistant starch, 25 received solely dietary instructions], 30 control subjects) RESISTA-PD. ID: NCT02784145

Probiotic and prebiotic

Fermented milk (multistrain probiotics and prebiotic fiber) (Barichella et al., 2016)

Probiotic

Multistrain probiotic (Hexbio®) containing microbial cell preparation MCP®BCMC® (Lactobacillus sp. and Bifidobacterium sp) (Ibrahim et al., 2020) Multistrain probiotics (Tan et al., 2021b)

DB-RCT in PD patients with Rome III-confirmed constipation 120 patients randomized (2:1), treatment once daily for 4 weeks. ID: NCT02459717 DB-RCT in PD patients with Rome III-confirmed constipation 48 patients randomized (1:1), treatment twice daily for 8 weeks DB-RCT in PD patients. 72 patients randomized (1:1), treatment once daily for 4 weeks. ID: NCT03377322

Improved motor symptoms and decrease levodopa-equivalent daily dose. Motor symptom improvement correlated with gut diversity and abundance of Ruminococcaceae. Reduced abundance of Clostridiaceae after colon cleansing Increased plasma L-dopa concentration (at 30 and 60 min after administration), which correlated with constipation improvement, and motor function Increased fecal butyrate and decreased fecal calprotectin concentrations, reduced nonmotor symptom load in PD patients receiving resistant starch (8 weeks). No changes in microbial composition or constipation severity Active treatment resulted in higher increase in the number of complete bowel movements/ week

Probiotic

Probiotic

Multistrain probiotics (Bacillus, Lactobacillus, Bifidobacterium and Enterococcus sp) (Du et al., 2022)

RCT (vs. original drug therapy) in 46 PD patients with Rome III-confirmed constipation

Increased bowel opening frequency and reduced intestinal transit time. No changes in motor and nonmotor symptom load Active treatment resulted in higher increase in the number of spontaneous bowel movements/week, and improvements in stool consistency and quality of life related to constipation Increased number of complete bowel movements/week. Improved stool consistency, constipation symptoms and constipation-related quality of life

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS

149

Table 7.4 Continued Type of microbiota manipulation strategy

Treatment description

Trial design

Findings

FMT

Donor FMT infused via colonoscopy (Segal et al., 2021)

Improvement of motor, nonmotor, and constipation symptoms, sustained at week 24

FMT

Donor FMT infused via colonoscopy (Kuai et al., 2021)

Open-label, n ¼ 6 PD patients with constipation and an indication for screening colonoscopy. ID: NCT03876327 Open-label, n ¼ 11 PD patients with indication for screening colonoscopy

Antimicrobials

Helicobacter pylori standard eradication triple therapy (Tan et al., 2020b)

DB-RCT in PD patients. 67 patients randomized (1:1), treatment once daily for 1 week

Improvement of motor, nonmotor, and constipation symptoms. Decrease of Bacteroidetes and increased abundance of Blautia and Prevotella. Normalization of the lactulose H2 breath test No improvement in any motor, nonmotor, or quality of life outcome at week 12 and week 52

Abbreviations: DB-RCT, Double-blind randomized-controlled trial; FMT, Fecal microbiota transplantation; L-dopa, Levodopa; PD, Parkinson disease; RCT, Randomized-controlled trial; RESISTA-PD, Effect of Resistant Starch on Bowel Habits, Short-Chain Fatty Acids and Gut Microbiota in Parkinson’s Disease trial.

challenges in recapitulating human gut dysbiosis in animal models during preclinical testing of microbiota manipulation strategies, future work should focus on clarifying host–microorganism interactions and how these change with modifications in lifestyle, diet, aging-related factors through the life of an individual.

The microbiota-gut-brain axis in Huntington disease There is considerably less evidence to date on the involvement of gut dysbiosis and alterations of the microbiota-gut-brain axis in other neurodegenerative basal ganglia-related movement disorders. The most investigated of these conditions after PD is HD. Replicating the path to knowledge on the link between the microbiota-gut-brain axis and pathogenesis in PD, the earliest evidence supporting this link also in HD has come from transgenic mouse models in which gut dysbiosis was demonstrated in affected animals, and greater instability of the gut microbiome was observed also during the presymptomatic motor stage of the disease (Kong et al., 2020). Subsequently, gut microbiota characterization in Australian HD gene expansion carriers using 16S rRNA sequencing showed lower a- and b-diversity indices compared to healthy subjects (Wasser et al., 2020), although this finding was not replicated by a more recent study from a Chinese sample (Du et al., 2021). Specific

taxonomic changes in carriers support exaltation of systemic inflammatory processes, specifically the lower abundance of Akkermansiaceae—suggesting altered maintenance of the intestinal barrier—and of Lachnospiraceae and Firmicutes—suggesting reduced production of the “anti-inflammatory” metabolite butyric acid. Lower abundance of Eubacterium hallii was associated with more severe motor signs and, in premanifest HD subjects, with larger estimated proximity to disease onset (Wasser et al., 2020). The evaluation of the contribution of gut dysbiosis to the pathophysiology of HD is in its infancy, and larger studies applying also metagenomic sequencing techniques are necessary before this role can be delineated.

The microbiota-gut-brain axis in amyotrophic lateral sclerosis Similar to other neurodegenerative conditions like PD and Alzheimer’s disease, a potential pathogenic role of the gut microbiota has been hypothesized for ALS and is intensely investigated. The digestive system displays functional abnormalities in both rodent models of ALS and ALS patients (Calvo et al., 2022; Martin et al., 2022). Preliminary, small clinical studies have shown a rise of intestinal inflammatory markers in stool samples from ALS patients (Rowin et al., 2017), whereas at a systemic level sporadic ALS patients displayed a correlation

150

Y. MAHJOUB AND D. MARTINO

between circulating monocyte/macrophage activation and LPS concentration (Zhang et al., 2009), potentially indicating intestinal inflammation and reduced intestinal barrier integrity in advanced ALS. An important murine model of ALS, the hSOD1G93A mice, carries mutations in the Cu, Zn superoxide dismutase (SOD), which normally catalyzes the transformation of superoxide radicals into hydrogen peroxide and O2. Alterations in the intestinal barrier with damage to the tight junctions are observed in this model, with reduced expression of junctional proteins zonulin-1 and E-cadherin (Wu et al., 2015). Likewise, this model shows also increased inflammatory markers, abnormal Paneth cells and other enteric nervous system changes even before the onset of motor changes, as well as subsequent correlation between ENS alterations and increased aggregation of hSOD1G93A in the spinal cord and different intestinal regions. In line with this, the ALS hSOD1G93A mouse model is associated with a reduction in the abundance of butyrateproducing bacteria. Chronic treatment with this SCFA led to improvement of intestinal barrier function, slower weight loss and slower progression to death (Zhang et al., 2017). Importantly, early gut microbial changes are observed in this model, in relation to constituents that can modulate autoimmunity, inflammation and metabolic regulatory control. Whereas antibiotic treatment worsened the progression of these transgenic mice, specific addition of A. muciniphila prolonged their lifespan (Blacher et al., 2019). These findings, associated with those from other transgenic models of ALS (Martin et al., 2022), support the involvement of the gut microbiota in the progression of inflammatory and neurodegenerative changes in this disease. The composition and functional profile of the gut microbiota in ALS patients have been investigated by a relatively limited number of studies that were quite different in design and sample size. Despite this heterogeneity, a few signals from human studies appeared to be more consistent and coherent with findings from transgenic animal models. Gut dysbiosis was reported by several studies, characterized by reduced abundance of Firmicutes, in particular the Clostridium genus, and higher abundance of Enterobacteriaceae (Mazzini et al., 2018; Di Gioia et al., 2020). Early changes in gut microbial constituents were also identified, among which lower abundance of Roseburia intestinalis and Eubacterium rectale, the former belonging to a SCFAproducing genus that was found to be reduced also in PD (Nicholson et al., 2021). Overall, the evidence collected so far indicates an increase in pathogens and a decrease in probiotic organisms in the gut ecosystem of ALS patients. Different metabolic pathways might

be influenced by these dysbiotic modifications. First, a reduced abundance of butyrate-producing species was confirmed as one of the most consistent microbiome profile features in ALS (Calvo et al., 2022). A small, prospective study that administered probiotics to ALS patients led to an increase in propionate and butyrate production (Di Gioia et al., 2020), supporting improvement in energy metabolism and even increased clearance of neurotoxic molecules. This is in line with the aforementioned improvement of intestinal functions, metabolism and progression rate of neurodegeneration obtained by butyrate administration to hSOD1G93A mice (Zhang et al., 2017). More recently, butyrate administration contributed to a shift toward a healthier metabolomic profiling in an ALS mouse model (Zhang et al., 2021). Another metabolic pathway that might be implicated in the link between microbiome and neural pathologic mechanisms of ALS is the tryptophan-nicotinamide pathway, as circulating levels of several key molecules of this pathway differed between ALS patients and healthy subjects (Li et al., 2022a,b). Nicotinamide and other tryptophan metabolites were found to delay, albeit mildly, disease progression in hSOD1G93A mice (Blacher et al., 2019), and supplementing nicotinamide in ALS is currently being tested in a clinical trial (https://clinicaltrials. gov/ct2/show/NCT04562831). Interestingly, tryptophan metabolites play a role in neuroinflammatory mechanisms associated with ALS disease progression, e.g., activation of microglia and astrocytes (Tan and Guillemin, 2019). This initial body of evidence lends promise to microbiota manipulation as a potentially rewarding disease-modifying strategy in ALS, although a substantial amount of work is still needed to identify the most effective approach to microbiota restoration or manipulation in this condition.

The microbiota-gut-brain axis in nondegenerative “network” movement disorders A potential pathogenic role of gut dysbiosis in putatively nonneurodegenerative movement disorders associated with dysfunctional connectivity of sensorimotor networks has been explored only very recently. An observational study conducted in Chinese subjects and comparing ET patients to de novo PD patients and healthy volunteers (sample size 54–67 per group) (Zhang et al., 2022) reported a Bacteroides-dominant enterotype in ET (i.e., more similar to the stratification of human gut microbiota observed in healthy subjects) and a Ruminococcus-dominant enterotype in PD (potentially associated with reduced homeostasis of the gut microenvironment) (Vieira-Silva et al., 2016), as well

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS as the existence of 4 genera whose abundance could help distinguishing between ET and PD individuals. Furthermore, ET patients exhibited reduced abundance of genera known to be SCFA-producers (Romboutsia and Ruminococcus) compared to healthy, and a positive correlation between tremor severity and abundance of Proteus, a genus previous associated with motor deficits and abnormal integrity of the intestinal barrier in a mouse model (Choi et al., 2018). Isolated dystonia is the third most common movement disorder after ET and PD. Like ET, it is considered a network disorder associated with dysfunctional connectivity of the sensorimotor loop of the corticobasal ganglia circuitry and of the cerebellar outflow pathways. Only one study conducted in Chinese individuals has explored metagenomic and metabolomic changes between patients with different forms of isolated dystonia and healthy subjects (Ma et al., 2021). This study reported no difference in a-diversity, but significantly different b-diversity, suggestive of increased heterogeneity in community structure among dystonia patients. Increased abundance of Clostridiales (including Blautia obeum, Dorea longicatena, and E. hallii) and reduced abundance of Bacteroides vulgatus and Bacteroides plebeium were found. The strongest association with the metabolomic profile was observed with the abundance of Clostridiales. Among the associated metabolic pathways were amino acid biosynthesis promoting the synthesis of dopamine, catecholamines, and serotonin. The decreased abundance of Bacteroides species is shared with psychiatric conditions like depressive disorders, with which adult-onset isolated dystonia also shares host gene expression patterns. Like ET and dystonia, there is only very early evidence of gut dysbiosis in TS, which is the most common movement disorder with pediatric onset. Another study from the People’s Republic of China compared 49 children with various types of primary tic disorder to 50 neurotypical individuals (Xi et al., 2021), identifying through shotgun metagenomic sequencing higher abundances of Bacteroides plebeius and Ruminococcus lactaris, and lower abundances of Prevotella stercorea and Streptococcus lutetiensis among children with tic disorders. Moreover, these authors detected an upregulation of GABA catabolic pathways in the functional pathway analysis of gut microbiota of children with tics, and some specific changes (e.g., overgrowth of pathogens like Escherichia coli) associated with use of dopamine receptor-blocking medications. Ongoing studies are addressing the association between gut dysbiosis and the heterogeneous clinical phenotype of tic disorders (Sycuro et al., 2022), in particular their overlap with other neurodevelopmental disorders like OCD, ADHD and autism spectrum disorder.

151

CONCLUSIONS Available data support a relevant contribution of immune-inflammatory mechanisms in neurodegenerative disorders affecting motor systems, particularly PD. There is evidence showing systemic changes suggestive of immune dysregulation in PD. Furthermore, consistent changes in the level of activity of inflammatory cells and the demonstration of neuroinflammation in both experimental models of PD and patients support a crucial pathophysiological link. Despite this, there are major knowledge gaps that need to be bridged by preclinical and clinical research to elucidate the timeline of events leading to the key pathological changes of PD and their translation into clinical symptoms and build-up of disability over time. Where immune-inflammatory mechanisms, both within the CNS and in other organs and systems, intervene to trigger, accelerate, or protect from neurodegeneration in the evolution of PD and its potential subtypes needs to be addressed before adding immune-based interventions to the arsenal of this challenging and socially impacting disorder. ALS is also characterized by neuroinflammatory changes that involve primarily glial cells, both microglia and astrocytes. At a systemic level, ALS patients demonstrate hyperactivation of immune-inflammatory pathways and dysfunction of regulatory cell types in forms that are associated with more rapid progression.

FUTURE DIRECTIONS The exploration of gut dysbiosis in the two prototypical neurodegenerative motor disorders PD and ALS is advancing rapidly. Although enterotyping has not been successful yet in identifying subtypes of these disorders that differ in their basic pathophysiological mechanisms, the increasing knowledge on the taxonomic composition of PD- and ALS-related gut dysbiosis, and the related metabolic and putative functional pathways, is providing enormous insight on which microbiota manipulation strategies should be developed in future research. Preclinical models, especially germ-free, will continue being useful to foster microbial therapeutics in this clinical area. Finally, there is less compelling, but still intriguing evidence suggesting that motor neurodevelopmental disorders, e.g., TS, are associated with abnormal trajectories of maturation that include also immune system development. Microglia seems to play a key role also in these disorders, and new therapeutic avenues aiming at its modulation are exciting prospects. Likewise, preclinical and clinical research on the role of gut dysbiosis in TS and related behavioral disorders is still in its infancy, but early findings support the rationale to delve deeper into its contribution to neural and immune maturation abnormalities associated with the TS spectrum.

152

Y. MAHJOUB AND D. MARTINO

REFERENCES Abdelmoaty MM, Machhi J, Yeapuri P et al. (2022). Monocyte biomarkers define sargramostim treatment outcomes for Parkinson’s disease. Clin Transl Med 12: e958. https:// doi.org/10.1002/ctm2.958. PMID: 35802825; PMCID: PMC9270000. Addabbo F, Baglioni V, Schrag A et al. (2020). Anti-dopamine D2 receptor antibodies in chronic tic disorders. Dev Med Child Neurol 62: 1205–1212. Aho VTE, Houser MC, Pereira PAB et al. (2021). Relationships of gut microbiota, short-chain fatty acids, inflammation, and the gut barrier in Parkinson’s disease. Mol Neurodegener 16: 6. Aikawa T, Mogushi K, Iijima-Tsutsui K et al. (2015). Loss of MyD88 alters neuroinflammatory response and attenuates early Purkinje cell loss in a spinocerebellar ataxia type 6 mouse model. Hum Mol Genet 24: 4780–4791. Alfonsetti M, Castelli V, D’angelo M (2022). Are we what we eat? Impact of diet on the gut-brain axis in Parkinson’s disease. Nutrients 14: 380. Almeida A, Nayfach S, Boland M et al. (2021). A unified catalog of 204,938 reference genomes from the human gut microbiome. Nat Biotechnol 39: 105–114. Astarloa R, Mena MA, Sa´nchez V et al. (1992). Clinical and pharmacokinetic effects of a diet rich in insoluble fiber on Parkinson disease. Clin Neuropharmacol 15: 375–380. https://doi.org/10.1097/00002826-19921000000004. PMID: 1330307. Bae EJ, Lee HJ, Rockenstein E et al. (2012). Antibody-aided clearance of extracellular a-synuclein prevents cell-to-cell aggregate transmission. J Neurosci 32: 13454–13469. Barichella M, Pacchetti C, Bolliri C et al. (2016). Probiotics and prebiotic fiber for constipation associated with Parkinson disease: an RCT. Neurology 87: 1274–1280. https://doi.org/10.1212/WNL.0000000000003127. Epub 2016 Aug 19, PMID: 27543643. Becker A, Schmartz GP, Gr€oger L et al. (2022). Effects of resistant starch on symptoms, fecal markers, and gut microbiota in Parkinson’s disease – the RESISTA-PD trial. Genom Proteom Bioinform 20: 274–287. https://doi.org/ 10.1016/j.gpb.2021.08.009. Epub 2021 Nov 25, PMID: 34839011; PMCID: PMC9684155. Beers DR, Henkel JS, Zhao W et al. (2008). CD4+ T cells support glial neuroprotection, slow disease progression, and modify glial morphology in an animal model of inherited ALS. Proc Natl Acad Sci U S A 105: 15558–15563. Beers DR, Henkel JS, Zhao W et al. (2011). Endogenous regulatory T lymphocytes ameliorate amyotrophic lateral sclerosis in mice and correlate with disease progression in patients with amyotrophic lateral sclerosis. Brain 134: 1293–1314. Benraiss A, Wang S, Herrlinger S et al. (2016). Human glia can both induce and rescue aspects of disease phenotype in Huntington disease. Nat Commun 7: 11758. Bhattarai Y, Si J, Pu M et al. (2021). Role of gut microbiota in regulating gastrointestinal dysfunction and motor symptoms in a mouse model of Parkinson’s disease. Gut Microbes 13: 1866974

Blacher E, Bashiardes S, Shapiro H et al. (2019). Potential roles of gut microbiome and metabolites in modulating ALS in mice. Nature 572: 474–480. Bos-Veneman N, Bijzet J, Limburg PC et al. (2010). Cytokines and soluble adhesion molecules in children and adolescents with a tic disorder. Prog Neuro-Psychopharmacology Biol Psychiatry 34: 1390–1395. Brochard V, Combadie`re B, Prigent A et al. (2009). Infiltration of CD4+ lymphocytes into the brain contributes to neurodegeneration in a mouse model of Parkinson disease. J Clin Invest 119: 182. Bushyhead D, Quigley EMM (2022). Small intestinal bacterial overgrowth-pathophysiology and its implications for definition and management. Gastroenterology 163: 593–607. Calvo AC, Valledor-Martı´n I, Moreno-Martı´nez L et al. (2022). Lessons to learn from the gut microbiota: a focus on amyotrophic lateral sclerosis. Genes (Basel) 13: 865. Cao B, Chen X, Zhang L et al. (2020). Elevated percentage of CD3+ T-cells and CD4+/CD8+ ratios in multiple system atrophy patients. Front Neurol 11: 658. Chang CC, Lin TM, Chang YS et al. (2018). Autoimmune rheumatic diseases and the risk of Parkinson disease: a nationwide population-based cohort study in Taiwan. Ann Med 50: 83–90. Chen SK, Tvrdik P, Peden E et al. (2010). Hematopoietic origin of pathological grooming in Hoxb8 mutant mice. Cell 141: 775–785. Choi JG, Kim N, Ju IG et al. (2018). Oral administration of Proteus mirabilis damages dopaminergic neurons and motor functions in mice. Sci Rep 8: 1275. Cirstea MS, Yu AC, Golz E et al. (2020). Microbiota composition and metabolism are associated with gut function in Parkinson’s disease. Mov Disord 35: 1208–1217. Claesson MJ, Jeffery IB, Conde S et al. (2012). Gut microbiota composition correlates with diet and health in the elderly. Nature 488: 178–184. Codolo G, Plotegher N, Pozzobon T et al. (2013). Triggering of inflammasome by aggregated a-synuclein, an inflammatory response in synucleinopathies. PLoS One 8: e55375. Cook DA, Kannarkat GT, Cintron AF et al. (2017). LRRK2 levels in immune cells are increased in Parkinson’s disease. NPJ Parkinsons Dis 3: 11. https://doi.org/10.1038/s41531017-0010-8PMID: 28649611; PMCID: PMC5459798. Coque E, Salsac C, Espinosa-Carrasco G et al. (2019). Cytotoxic CD8 + T lymphocytes expressing ALS-causing SOD1 mutant selectively trigger death of spinal motoneurons. Proc Natl Acad Sci U S A 116: 2312–2317. Cosma-Grigorov A, Meixner H, Mrochen A et al. (2020). Changes in gastrointestinal microbiome composition in PD: a pivotal role of covariates. Front Neurol 11: 1041. Cui X, Xu C, Zhang L et al. (2021). Identification of Parkinson’s disease-causing genes via omics data. Front Genet 12: 712164. Dale RC, Merheb V, Pillai S et al. (2012). Antibodies to surface dopamine-2 receptor in autoimmune movement and psychiatric disorders. Brain 135: 3453–3468. De Biase LM, Schuebel KE, Fusfeld ZH et al. (2017). Local cues establish and maintain region-specific phenotypes of basal ganglia microglia. Neuron 95: 341–356.

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS De Marchi F, Munitic I, Amedei A et al. (2021). Interplay between immunity and amyotrophic lateral sclerosis: clinical impact. Neurosci Biobehav Rev 127: 958. De S, Van Deren D, Peden E et al. (2018). Two distinct ontogenies confer heterogeneity to mouse brain microglia. Development 145: dev152306. DeJesus-Hernandez M, Mackenzie IR, Boeve BF et al. (2011). Expanded GGGGCC Hexanucleotide repeat in noncoding region of C9ORF72 causes chromosome 9p-linked FTD and ALS. Neuron 72: 245–256. DeJong EN, Surette MG, Bowdish DME (2020). The gut microbiota and unhealthy aging: disentangling cause from consequence. Cell Host Microbe 28: 180–189. Di Filippo M, Tozzi A, Arcangeli S et al. (2016). Interferon-b 1a modulates glutamate neurotransmission in the CNS through CaMKII and GluN2A-containing NMDA receptors. Neuropharmacology 100: 98–105. Di Gioia D, Bozzi Cionci N, Baffoni L et al. (2020). A prospective longitudinal study on the microbiota composition in amyotrophic lateral sclerosis. BMC Med 18: 153. Diaz Heijtz R, Wang S, Anuar F et al. (2011). Normal gut microbiota modulates brain development and behavior. Proc Natl Acad Sci U S A 108: 3047–3052. Du G, Dong W, Yang Q et al. (2021). Altered gut microbiota related to inflammatory responses in patients with Huntington’s disease. Front Immunol 11: 603594. Du Y, Li Y, Xu X et al. (2022). Probiotics for constipation and gut microbiota in Parkinson’s disease. Parkinsonism Relat Disord 103: 92–97. https://doi.org/10.1016/j.parkreldis. 2022.08.022. Epub 2022 Sep 5, PMID: 36087572. Duque T, Gromicho M, Pronto-Laborinho AC et al. (2020). Transforming growth factor-b plasma levels and its role in amyotrophic lateral sclerosis. Med Hypotheses 139: 109632. Endo F, Komine O, Fujimori-Tonou N et al. (2015). Astrocytederived TGF-b1 accelerates disease progression in ALS mice by interfering with the neuroprotective functions of microglia and T cells. Cell Rep 11: 592–604. Fellner L, Irschick R, Schanda K et al. (2013). Toll-like receptor 4 is required for a-synuclein dependent activation of microglia and astroglia. Glia 61: 349–360. Ferguson AR, Christensen RN, Gensel JC et al. (2008). Cell death after spinal cord injury is exacerbated by rapid TNFa-induced trafficking of GluR2-lacking AMPARs to the plasma membrane. J Neurosci 28: 11391. Ferro A, Sheeler C, Rosa JG et al. (2019). Role of microglia in ataxias. J Mol Biol 431: 1792. Fiszer U, Mix E, Fredrikson S et al. (1994). Parkinson’s disease and immunological abnormalities: increase of HLA-DR expression on monocytes in cerebrospinal fluid and of CD45RO+ T cells in peripheral blood. Acta Neurol Scand 90: 160–166. Franceschi C, Garagnani P, Parini P et al. (2018). Inflammaging: a new immune-metabolic viewpoint for age-related diseases. Nat Rev Endocrinol 14: 576–590. Garofalo S, Cocozza G, Porzia A et al. (2020). Natural killer cells modulate motor neuron-immune cell cross talk in models of amyotrophic lateral sclerosis. Nat Commun 11: 1773.

153

Gerhard A, Pavese N, Hotton G et al. (2006). In vivo imaging of microglial activation with [11C](R)-PK11195 PET in idiopathic Parkinson’s disease. Neurobiol Dis 21: 404–412. Ginhoux F, Greter M, Leboeuf M et al. (2010). Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science 330: 841. Grabert K, Michoel T, Karavolos MH et al. (2016). Microglial brain region-dependent diversity and selective regional sensitivities to ageing. Nat Neurosci 19: 504. Grimsley CA, Sanchez JT, Sivaramakrishnan S (2013). Diversity of layer 5 projection neurons in the mouse motor cortex. Front Cell Neurosci 7: 174. Guo H, Callaway JB, Ting JPY (2015). Inflammasomes: mechanism of action, role in disease, and therapeutics. Nat Med 21: 677. Guttenplan KA, Weigel MK, Adler DI et al. (2020). Knockout of reactive astrocyte activating factors slows disease progression in an ALS mouse model. Nat Commun 11: 3753. Hadjivassiliou M, Graus F, Honnorat J et al. (2020). Diagnostic criteria for primary autoimmune cerebellar Ataxia—guidelines from an international task force on immune-mediated cerebellar ataxias. Cerebellum 19: 605. Hamamah S, Aghazarian A, Nazaryan A et al. (2022). Role of microbiota-gut-brain axis in regulating dopaminergic signaling. Biomedicine 10: 436. Harms AS, Cao S, Rowse AL et al. (2013). MHCII is required for a-synuclein-induced activation of microglia, CD4 T cell proliferation, and dopaminergic neurodegeneration. J Neurosci 33: 9592–9600. Harrison JK, Jiang Y, Chen S et al. (1998). Role for neuronally derived fractalkine in mediating interactions between neurons and CX3CR1-expressing microglia. Proc Natl Acad Sci U S A 95: 10896–10901. Hartstra AV, Sch€ uppel V, Imangaliyev S et al. (2020). Infusion of donor feces affects the gut-brain axis in humans with metabolic syndrome. Mol Metab 42: 101076. Hegelmaier T, Lebbing M, Duscha A (2020). Interventional influence of the intestinal microbiome through dietary intervention and bowel cleansing might improve motor symptoms in Parkinson’s disease. Cells 9: 376. https://doi. org/10.3390/cells9020376. PMID: 32041265; PMCID: PMC7072275. Heintz-Buschart A, Pandey U, Wicke T et al. (2018). The nasal and gut microbiome in Parkinson’s disease and idiopathic rapid eye movement sleep behavior disorder. Mov Disord 33: 88–98. Hill-Burns EM, Debelius JW, Morton JT et al. (2017). Parkinson’s disease and Parkinson’s disease medications have distinct signatures of the gut microbiome. Mov Disord 32: 739–749. Ho A, Blum M (1998). Induction of interleukin-1 associated with compensatory dopaminergic sprouting in the denervated striatum of young mice: model of aging and neurodegenerative disease. J Neurosci 18: 5614–5629. Holmans P, Moskvina V, Jones L et al. (2013). A pathway-based analysis provides additional support for an immune-related genetic susceptibility to Parkinson’s disease. Hum Mol Genet 22: 1039–1049.

154

Y. MAHJOUB AND D. MARTINO

Hong JJ, Loiselle CR, Yoon DY et al. (2004). Microarray analysis in Tourette syndrome postmortem putamen. J Neurol Sci 225: 57–64. Houi K, Kobayashi T, Kato S et al. (2002). Increased plasma TGF-beta1 in patients with amyotrophic lateral sclerosis. Acta Neurol Scand 106: 299–301. Hsiao EY, McBride SW, Hsien S et al. (2013). Microbiota modulate behavioral and physiological abnormalities associated with neurodevelopmental disorders. Cell 155: 1451–1463. Hui KY, Fernandez-Hernandez H, Hu J et al. (2018). Functional variants in the LRRK2 gene confer shared effects on risk for Crohn’s disease and Parkinson’s disease. Sci Transl Med 10: eaai7795. Ibrahim A, Ali RAR, Manaf MRA et al. (2020). Multi-strain probiotics (Hexbio) containing MCP BCMC strains improved constipation and gut motility in Parkinson’s disease: a randomised controlled trial. PLoS One 15: e0244680. https://doi.org/10.1371/journal.pone.0244680. PMID: 33382780; PMCID: PMC7774928. Jadhav KS, Peterson VL, Halfon O et al. (2018). Gut microbiome correlates with altered striatal dopamine receptor expression in a model of compulsive alcohol seeking. Neuropharmacology 141: 249–259. Jeffery IB, Lynch DB, O’Toole PW (2016). Composition and temporal stability of the gut microbiota in older persons. ISME J 10: 170–182. Johnson ME, Stringer A, Bobrovskaya L (2018). Rotenone induces gastrointestinal pathology and microbiota alterations in a rat model of Parkinson’s disease. Neurotoxicology 65: 174–185. Jones JD, Rahmani E, Garcia E et al. (2020). Gastrointestinal symptoms are predictive of trajectories of cognitive functioning in de novo Parkinson’s disease. Parkinsonism Relat Disord 72: 7–12. Karpenko MN, Vasilishina AA, Gromova EA et al. (2018). Interleukin-1b, interleukin-1 receptor antagonist, interleukin-6, interleukin-10, and tumor necrosis factor-a levels in CSF and serum in relation to the clinical diversity of Parkinson’s disease. Cell Immunol 327: 77–82. Kaufman E, Hall S, Surova Y et al. (2013). Proinflammatory cytokines are elevated in serum of patients with multiple system atrophy. PLoS One 8. Kawikova I, Leckman JF, Kronig H et al. (2007). Decreased numbers of regulatory T cells suggest impaired immune tolerance in children with Tourette syndrome: a preliminary study. Biol Psychiatry 61: 273–278. Kiessling LS, Marcotte AC, Culpepper L (1993). Antineuronal antibodies in movement disorders. Pediatrics 92: 39–43. Kim R, Kim HJ, Kim A et al. (2019). Does peripheral inflammation contribute to multiple system atrophy? Parkinsonism Relat Disord 64: 340–341. Kiraly DD, Walker DM, Calipari ES et al. (2016). Alterations of the host microbiome affect behavioral responses to cocaine. Sci Rep 6: 35455. Kong F, Hua Y, Zeng B et al. (2016). Gut microbiota signatures of longevity. Curr Biol 26: R832–R833.

Kong G, Cao KAL, Judd LM et al. (2020). Microbiome profiling reveals gut dysbiosis in a transgenic mouse model of Huntington’s disease. Neurobiol Dis 135: 104268. Konishi H, Kiyama H (2018). Microglial TREM2/DAP12 signaling: a double-edged sword in neural diseases. Front Cell Neurosci 12: 206. Koutzoumis DN, Vergara M, Pino J et al. (2020). Alterations of the gut microbiota with antibiotics protects dopamine neuron loss and improve motor deficits in a pharmacological rodent model of Parkinson’s disease. Exp Neurol 325: 113159. Kuai XY, Yao XH, Xu LJ et al. (2021). Evaluation of fecal microbiota transplantation in Parkinson’s disease patients with constipation. Microb Cell Fact 20: 98. https://doi.org/ 10.1186/s12934-021-01589-0. PMID: 33985520; PMCID: PMC8120701. Kumar A, Williams MT, Chugani HT (2015). Evaluation of basal ganglia and thalamic inflammation in children with pediatric autoimmune neuropsychiatric disorders associated with streptococcal infection and Tourette syndrome: a positron emission tomographic (PET) study using 11C-[R]-PK11195. J Child Neurol 30: 749–756. Kundu P, Blacher E, Elinav E et al. (2017). Our gut microbiome: the evolving inner self. Cell 171: 1481–1493. Leckman JF, Katsovich L, Kawikova I et al. (2005). Increased serum levels of interleukin-12 and tumor necrosis factor-alpha in Tourette’s syndrome. Biol Psychiatry 57: 667–673. Lennington JB, Coppola G, Kataoka-Sasaki Y et al. (2016). Transcriptome analysis of the human striatum in Tourette syndrome. Biol Psychiatry 79: 372–382. Levi H, Bar E, Cohen-Adiv S et al. (2022). Dysfunction of cerebellar microglia in Ataxia-telangiectasia. Glia 70: 536–557. Lewitus GM, Pribiag H, Duseja R et al. (2014). An adaptive role of TNFa in the regulation of striatal synapses. J Neurosci 34: 6146. Li X, Sundquist J, Sundquist K (2012). Subsequent risks of Parkinson disease in patients with autoimmune and related disorders: a nationwide epidemiological study from Sweden. Neurodegener Dis 10: 277–284. Li E, Ruan Y, Chen Q et al. (2015). Streptococcal infection and immune response in children with Tourette’s syndrome. Childs Nerv Syst 31: 1157–1163. Li Q, Cheng Z, Zhou L et al. (2019). Developmental heterogeneity of microglia and brain myeloid cells revealed by deep single-cell RNA sequencing. Neuron 101: 207–223. Li H, Wang Y, Zhao C et al. (2022a). Fecal transplantation can alleviate tic severity in a Tourette syndrome mouse model by modulating intestinal flora and promoting serotonin secretion. Chin Med J (Engl) 135: 707–713. Li C, Zhu Y, Chen W et al. (2022b). Circulating NAD+ metabolism-derived genes unveils prognostic and peripheral immune infiltration in amyotrophic lateral sclerosis. Front Cell Dev Biol 10: 831273. Liao B, Zhao W, Beers DR et al. (2012). Transformation from a neuroprotective to a neurotoxic microglial phenotype in a mouse model of ALS. Exp Neurol 237: 147–152.

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS Liao JF, Cheng YF, Li SW et al. (2019). Lactobacillus plantarum PS128 ameliorates 2,5-Dimethoxy-4-iodoamphetamineinduced tic-like behaviors via its influences on the microbiota–gut-brain-axis. Brain Res Bull 153: 59–73. Liddelow SA, Guttenplan KA, Clarke LE et al. (2017). Neurotoxic reactive astrocytes are induced by activated microglia. Nature 541: 481–487. Liu G, Chong HX, Chung FYL et al. (2020). Lactobacillus plantarum DR7 modulated bowel movement and gut microbiota associated with dopamine and serotonin pathways in stressed adults. Int J Mol Sci 21: 1–16. Lorente-pico´n M, Laguna A (2021). New avenues for Parkinson’s disease therapeutics: disease-modifying strategies based on the gut microbiota. Biomolecules 11: 1–39. Ma L, Keng J, Cheng M et al. (2021). Gut microbiome and serum metabolome alterations associated with isolated dystonia. mSphere 6: e0028321. Malpetti M, Passamonti L, Rittman T et al. (2020). Neuroinflammation and tau co-localize in vivo in progressive supranuclear palsy. Ann Neurol 88: 1194. Mancini A, Ghiglieri V, Parnetti L et al. (2021). Neuroimmune cross-talk in the striatum: from basal ganglia physiology to circuit dysfunction. Front Immunol 12: 1044. Manfready RA, Forsyth CB, Voigt RM et al. (2022). Gut-brain communication in Parkinson’s disease: Enteroendocrine regulation by GLP-1. Curr Neurol Neurosci Rep 22: 335–342. Marı´n-Teva JL, Dusart I, Colin C et al. (2004). Microglia promote the death of developing Purkinje cells. Neuron 41: 535–547. Martin S, Battistini C, Sun J (2022). A gut feeling in amyotrophic lateral sclerosis: microbiome of mice and men. Front Cell Infect Microbiol 12: 839526. Martino D, Dale RC, Gilbert DL et al. (2009). Immunopathogenic mechanisms in Tourette syndrome: a critical review. Mov Disord 24: 1267–1279. Matheoud D, Sugiura A, Bellemare-Pelletier A et al. (2016). Parkinson’s disease-related proteins PINK1 and Parkin repress mitochondrial antigen presentation. Cell 166: 314–327. Matz J, Krause DL, Dehning S et al. (2012). Altered monocyte activation markers in Tourette’s syndrome: a case–control study. BMC Psychiatry 12: 29. Mazzini L, Mogna L, De Marchi F et al. (2018). Potential role of gut microbiota in ALS pathogenesis and possible novel therapeutic strategies. J Clin Gastroenterol 52: S68–S70. McGeer PL, Itagaki S, Boyes BE et al. (1988). Reactive microglia are positive for HLA-DR in the substantia nigra of Parkinson’s and Alzheimer’s disease brains. Neurology 38: 1285–1291. M€ oller JC, Tackenberg B, Heinzel-Gutenbrunner M et al. (2008). Immunophenotyping in Tourette syndrome—a pilot study. Eur J Neurol 15: 749–753. Mullin S, Stokholm MG, Hughes D et al. (2021). Brain microglial activation increased in Glucocerebrosidase (GBA) mutation carriers without Parkinson’s disease. Mov Disord 36: 774–779.

155

Nakayama H, Abe M, Morimoto C et al. (2018). Microglia permit climbing fiber elimination by promoting GABAergic inhibition in the developing cerebellum. Nat Commun 9: 2830. Nalls MA, Pankratz N, Lill CM et al. (2014). Large-scale metaanalysis of genome-wide association data identifies six new risk loci for Parkinson’s disease. Nat Genet 46: 989–993. Nicholson K, Bjornevik K, Abu-Ali G et al. (2021). The human gut microbiota in people with amyotrophic lateral sclerosis. Amyotroph Lateral Scler Frontotemporal Degener 22: 186–194. Nishino R, Mikami K, Takahashi H et al. (2013). Commensal microbiota modulate murine behaviors in a strictly contamination-free environment confirmed by culturebased methods. Neurogastroenterol Motil 25: 521–e371. Nishiwaki H, Ito M, Ishida T et al. (2020). Meta-analysis of gut Dysbiosis in Parkinson’s disease. Mov Disord 35: 1626–1635. Nykjær CH, Brudek T, Salvesen L et al. (2017). Changes in the cell population in brain white matter in multiple system atrophy. Mov Disord 32: 1074–1082. Olson KE, Namminga KL, Lu Y et al. (2021). Safety, tolerability, and immune-biomarker profiling for yearlong sargramostim treatment of Parkinson’s disease. EBioMedicine 67: 103380. https://doi.org/10.1016/j.ebiom. 2021.103380. Epub 2021 May 14, PMID: 34000620; PMCID: PMC8138485. O’Rourke JG, Bogdanik L, Ya´n˜ez A et al. (2016). C9orf72 is required for proper macrophage and microglial function in mice. Science 351: 1324–1329. Ortega-Cubero S, Lorenzo-Betancor O, Lorenzo E et al. (2015). TREM2 R47H variant and risk of essential tremor: a cross-sectional international multicenter study. Parkinsonism Relat Disord 21: 306–309. Panicker N, Sarkar S, Harischandra DS et al. (2019). Fyn kinase regulates misfolded a-synuclein uptake and NLRP3 inflammasome activation in microglia. J Exp Med 216: 1411. Paolicelli RC, Bolasco G, Pagani F et al. (2011). Synaptic pruning by microglia is necessary for normal brain development. Science 333: 1456–1458. Parker-Athill EC, Ehrhart J, Tan J et al. (2015). Cytokine correlations in youth with tic disorders. J Child Adolesc Psychopharmacol 25: 86–92. Parkhurst CN, Yang G, Ninan I et al. (2013). Microglia promote learning-dependent synapse formation through BDNF. Cell 155: 1596. Pont-Lezica L, Beumer W, Colasse S et al. (2014). Microglia shape corpus callosum axon tract fasciculation: functional impact of prenatal inflammation. Eur J Neurosci 39: 1551–1557. Prieto GA, Tong L, Smith ED et al. (2019). TNFa and IL-1b but not IL-18 suppresses hippocampal long-term potentiation directly at the synapse. Neurochem Res 44: 49–60. Pu Y, Chang L, Qu Y et al. (2019). Antibiotic-induced microbiome depletion protects against MPTP-induced dopaminergic neurotoxicity in the brain. Aging (Albany NY) 11:

156

Y. MAHJOUB AND D. MARTINO

6915–6929. https://doi.org/10.18632/aging.102221. Epub 2019 Sep 3, PMID: 31479418; PMCID: PMC6756889. Rizzo R, Gulisano M, Pavone P et al. (2006). Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 21: 747–753. Romano S, Savva GM, Bedarf JR et al. (2021). Meta-analysis of the Parkinson’s disease gut microbiome suggests alterations linked to intestinal inflammation. NPJ Parkinsons Dis 7: 27. Rosario D, Bidkhori G, Lee S et al. (2021). Systematic analysis of gut microbiome reveals the role of bacterial folate and homocysteine metabolism in Parkinson’s disease. Cell Rep 34: 108807. Rossi S, Sacchetti L, Napolitano F et al. (2012). Interleukin-1b causes anxiety by interacting with the endocannabinoid system. J Neurosci 32: 13896–13905. Rowin J, Xia Y, Jung B et al. (2017). Gut inflammation and dysbiosis in human motor neuron disease. Physiol Rep 5: e13443. Rugbjerg K, Friis S, Ritz B et al. (2009). Autoimmune disease and risk for Parkinson disease: a population-based casecontrol study. Neurology 73: 1462–1468. Rydbirk R, Elfving B, Folke J et al. (2019). Increased prefrontal cortex interleukin-2 protein levels and shift in the peripheral T cell population in progressive supranuclear palsy patients. Sci Rep 9: 7781. Sa´nchez-Ruiz de Gordoa J, Erro ME, Vicun˜a-Urriza J et al. (2020). Microglia-related gene triggering receptor expressed in myeloid cells 2 (TREM2) is upregulated in the substantia Nigra of progressive Supranuclear palsy. Mov Disord 35: 885–890. Sankowski B, Księzarczyk K, Rackowska E et al. (2020). Higher cerebrospinal fluid to plasma ratio of p-cresol sulfate and indoxyl sulfate in patients with Parkinson’s disease. Clin Chim Acta 501: 165–173. Schafer DP, Lehrman EK, Kautzman AG et al. (2012). Microglia sculpt postnatal neural circuits in an activity and complement-dependent manner. Neuron 74: 691–705. Schr€ oder JB, Pawlowski M, Meyer zu H€orste G et al. (2018). Immune cell activation in the cerebrospinal fluid of patients with Parkinson’s disease. Front Neurol 9: 1081. Scott KM, Kouli A, Yeoh SL et al. (2018). A systematic review and meta-analysis of alpha synuclein auto-antibodies in Parkinson’s disease. Front Neurol 9: 815. Segal A, Zlotnik Y, Moyal-Atias K et al. (2021). Fecal microbiota transplant as a potential treatment for Parkinson’s disease – a case series. Clin Neurol Neurosurg 207: 106791. https://doi.org/10.1016/j.clineuro.2021.106791. Epub 2021 Jun 30, PMID: 34237681. Seki T, Sato M, Kibe Y et al. (2018). Lysosomal dysfunction and early glial activation are involved in the pathogenesis of spinocerebellar ataxia type 21 caused by mutant transmembrane protein 240. Neurobiol Dis 120: 34–50. Shadrin AA, Mucha S, Ellinghaus D et al. (2021). Shared genetics of multiple system atrophy and inflammatory bowel disease. Mov Disord 36: 449–459. 

Shanahan F, Ghosh TS, O’Toole PW (2021). The healthy microbiome-what is the definition of a healthy gut microbiome? Gastroenterology 160: 483–494. Shin C, Lim Y, Lim H et al. (2020). Plasma short-chain fatty acids in patients with Parkinson’s disease. Mov Disord 35: 1021–1027. Simon DK, Simuni T, Elm J et al. (2015). Peripheral biomarkers of Parkinson’s disease progression and pioglitazone effects. J Parkinsons Dis 5: 731–736. https:// doi.org/10.3233/JPD-150666. PMID: 26444095; PMCID: PMC5061495. Siokas V, Aloizou AM, Tsouris Z et al. (2020). Genetic risk factors for essential tremor: a review. Tremor Other Hyperkinet Mov 10: 1–7. Sommer A, Maxreiter F, Krach F et al. (2018). Th17 lymphocytes induce neuronal cell death in a human iPSC-based model of Parkinson’s disease. Cell Stem Cell 23: 123–131.e6. Squarzoni P, Oller G, Hoeffel G et al. (2014). Microglia modulate wiring of the embryonic forebrain. Cell Rep 8: 1271–1279. Srivastav S, Neupane S, Bhurtel S et al. (2019). Probiotics mixture increases butyrate, and subsequently rescues the nigral dopaminergic neurons from MPTP and rotenoneinduced neurotoxicity. J Nutr Biochem 69: 73–86. Stevens B, Allen NJ, Vazquez LE et al. (2007). The classical complement Cascade mediates CNS synapse elimination. Cell 131: 1164–1178. Sulzer D, Alcalay RN, Garretti F et al. (2017). T cells from patients with Parkinson’s disease recognize a-synuclein peptides. Nature 546: 656–661. Sycuro L, Coker O, Ramay H et al. (2022). Metagenomic exploration of the gut microbiome in a sibling-controlled pilot study of children with Tourette syndrome [abstract]. Mov Disord 37: 132. Taj A, Jamil N (2018). Bioconversion of tyrosine and tryptophan derived biogenic amines by Neuropathogenic Bacteria. Biomolecules 8: 10. Takeda K, Akira S (2015). Toll-like receptors. Curr Protoc Immunol 109: 14.12.1-14.12.10. Tan VX, Guillemin GJ (2019). Kynurenine pathway metabolites as biomarkers for amyotrophic lateral sclerosis. Front Neurosci 13: 1013. Tan EK, Chao YX, West A et al. (2020). Parkinson disease and the immune system—associations, mechanisms and therapeutics. Nat Rev Neurol 16: 303–318. Tan AH, Lim SY, Mahadeva S et al. (2020b). Helicobacter pylori eradication in Parkinson’s disease: a randomized placebo-controlled trial. Mov Disord 35: 2250–2260. https://doi.org/10.1002/mds.28248. Epub 2020 Sep 7, PMID: 32894625. Tan AH, Chong CW, Lim SY et al. (2021). Gut microbial ecosystem in Parkinson disease: new clinicobiological insights from multi-omics. Ann Neurol 89: 546–559. Tan AH, Lim SY, Chong KK et al. (2021b). Probiotics for constipation in Parkinson disease: a randomized placebocontrolled study. Neurology 96: e772–e782. https://doi. org/10.1212/WNL.0000000000010998. Epub 2020 Oct 12, PMID: 33046607.

IMMUNOLOGY AND MICROBIOME: IMPLICATIONS FOR MOTOR SYSTEMS Tan AH, Lim SY, Lang AE (2022). The microbiome-gut-brain axis in Parkinson disease—from basic research to the clinic. Nat Rev Neurol 18: 476–495. Thonhoff JR, Beers DR, Zhao W et al. (2018). Expanded autologous regulatory T-lymphocyte infusions in ALS: a phase I, first-in-human study. Neurol Neuroimmunol Neuroinflammation 5: e465. Thonhoff JR, Berry JD, Macklin EA et al. (2022). Combined regulatory T-lymphocyte and IL-2 treatment is safe, tolerable, and biologically active for 1 year in persons with amyotrophic lateral sclerosis. Neurol Neuroimmunol Neuroinflammation 9: e200019. Toh TS, Chong CW, Lim SY et al. (2022). Gut microbiome in Parkinson’s disease: new insights from meta-analysis. Parkinsonism Relat Disord 94: 1–9. Tuikhar N, Keisam S, Labala RK et al. (2019). Comparative analysis of the gut microbiota in centenarians and young adults shows a common signature across genotypically non-related populations. Mech Ageing Dev 179: 23–35. Turner MR, Cagnin A, Turkheimer FE et al. (2004). Evidence of widespread cerebral microglial activation in amyotrophic lateral sclerosis: an [11C](R)-PK11195 positron emission tomography study. Neurobiol Dis 15: 601–609. Tylee DS, Sun J, Hess JL et al. (2018). Genetic correlations among psychiatric and immune-related phenotypes based on genome-wide association data. Am J Med Genet B Neuropsychiatr Genet 177: 641. Ueno M, Fujita Y, Tanaka T et al. (2013). Layer V cortical neurons require microglial support for survival during postnatal development. Nat Neurosci 16: 543–551. Vascellari S, Palmas V, Melis M et al. (2020). Gut microbiota and metabolome alterations associated with Parkinson’s disease. mSystems 5: e00561-20. Vieira-Silva S, Falony G, Darzi Y et al. (2016). Speciesfunction relationships shape ecological properties of the human gut microbiome. Nat Microbiol 1: 16088. Wah DTO, Ossenkopp KP, Bishnoi I et al. (2019). Predator odor exposure in early adolescence influences the effects of the bacterial product, propionic acid, on anxiety, sensorimotor gating, and acoustic startle response in male rats in later adolescence and adulthood. Physiol Behav 199: 35–46. Wasser CI, Mercieca EC, Kong G et al. (2020). Gut dysbiosis in Huntington’s disease: associations among gut microbiota, cognitive performance and clinical outcomes. Brain Commun 2: fcaa110. Weis S, Schwiertz A, Unger MM et al. (2019). Effect of Parkinson’s disease and related medications on the composition of the fecal bacterial microbiota. NPJ Parkinsons Dis 5: 28. Williams GP, Marmion DJ, Schonhoff AM et al. (2020). T cell infiltration in both human multiple system atrophy and a novel mouse model of the disease. Acta Neuropathol 139: 855–874. Witoelar A, Jansen IE, Wang Y et al. (2017). Genome-wide pleiotropy between Parkinson disease and autoimmune diseases. JAMA Neurol 74: 780–792. Wood TE, Barry J, Yang Z et al. (2019). Mutant huntingtin reduction in astrocytes slows disease progression in the

157

BACHD conditional Huntington’s disease mouse model. Hum Mol Genet 28: 487–500. Wu S, Yi J, Zhang YG et al. (2015). Leaky intestine and impaired microbiome in an amyotrophic lateral sclerosis mouse model. Physiol Rep 3: e12356. Xi W, Gao X, Zhao H et al. (2021). Depicting the composition of gut microbiota in children with tic disorders: an exploratory study. J Child Psychol Psychiatry 62: 1246–1254. Xiao Q, Zhao W, Beers DR et al. (2007). Mutant SOD1(G93A) microglia are more neurotoxic relative to wild-type microglia. J Neurochem 102: 2008–2019. Yamada T, Moroo I, Koguchi Y et al. (1994). Increased concentration of C4d complement protein in the cerebrospinal fluids in progressive supranuclear palsy. Acta Neurol Scand 89: 42–46. Yeh FL, Hansen DV, Sheng M (2017). TREM2, microglia, and neurodegenerative diseases. Trends Mol Med 23: 512–533. Yeon S-M, Lee JH, Kang D et al. (2017). A cytokine study of pediatric Tourette’s disorder without obsessive compulsive disorder. Psychiatry Res 247: 90–96. Yildirim Z, Karabekiroglu K, Yildiran A et al. (2020). An examination of the relationship between regulatory T cells and symptom flare-ups in children and adolescents diagnosed with chronic tic disorder and Tourette syndrome. Nord J Psychiatry 75: 18–24. Yun SP, Kam TI, Panicker N et al. (2018). Block of A1 astrocyte conversion by microglia is neuroprotective in models of Parkinson’s disease. Nat Med 24: 931–938. https://doi.org/10.1038/s41591-018-0051-5. Epub 2018 Jun 11, PMID: 29892066; PMCID: PMC6039259. Zhang R, Miller RG, Gascon R et al. (2009). Circulating endotoxin and systemic immune activation in sporadic amyotrophic lateral sclerosis (sALS). J Neuroimmunol 206: 121–124. Zhang Y-G, Wu S, Yi J et al. (2017). Target intestinal microbiota to alleviate disease progression in amyotrophic lateral sclerosis. Clin Ther 39: 322–336. Zhang Y, Ogbu D, Garrett S et al. (2021). Aberrant enteric neuromuscular system and dysbiosis in amyotrophic lateral sclerosis. Gut Microbes 13: 1996848. Zhang P, Huang P, Du J et al. (2022). Specific gut microbiota alterations in essential tremor and its difference from Parkinson’s disease. NPJ Parkinsons Dis 8: 98. Zhao W, Beers DR, Bell S et al. (2015). TDP-43 activates microglia through NF-kB and NLRP3 inflammasome. Exp Neurol 273: 24–35. Zhao W, Beers DR, Hooten KG et al. (2017). Characterization of gene expression phenotype in amyotrophic lateral sclerosis monocytes. JAMA Neurol 74: 677–685. Zhu M, Liu X, Ye Y et al. (2022). Gut microbiota: a novel therapeutic target for Parkinson’s disease. Front Immunol 13: 937555. Zsigmond P, Ljunggren SA, Ghafouri B (2020). Proteomic analysis of the cerebrospinal fluid in patients with essential tremor before and after deep brain stimulation surgery: a pilot study. Neuromodulation 23: 502–508. Zubiri I, Lombardi V, Bremang M et al. (2018). Tissueenhanced plasma proteomic analysis for disease stratification in amyotrophic lateral sclerosis. Mol Neurodegener 13: 60.

This page intentionally left blank

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00014-5 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 8

COVID-19 (novel SARS-CoV-2) neurological illness DAVID S. YOUNGER1,2* 1

Department of Clinical Medicine and Neuroscience, CUNY School of Medicine, New York, NY, United States

2

Department of Medicine, Section of Internal Medicine and Neurology, White Plains Hospital, White Plains, NY, United States

Abstract COVID-19 illness is associated with diverse neurological manifestations. Its exceptionally high prevalence results from unprecedented genetic diversity, genomic recombination, and superspreading. With each new mutation and variant, there are foreseeable risks of rising fatality and novel neurological motor complications in childhood and adult cases. This chapter provides an extensive review of COVID-19 neurological illness, notably the motor manifestations. Innovative treatments have been developed to stem the spread of infectious contagious illness, and attenuate the resultant cytokine storm and other postinfectious immune aspects responsible for postacute COVID-19 syndrome due to the multiplier effect of infection, immunity, and inflammation, termed I3.

BACKGROUND The earliest reports of a pneumonia of unknown origin linked to exposure at a seafood and wet animal market in Wuhan (Hubei Province, China) (Zhu et al., 2020) that identified it as a new beta coronavirus was named severe or novel acute respiratory syndrome-coronavirus-2 (SARS-nCoV-2). These single-stranded ribonucleic acid (RNA)–enveloped viruses have the largest known RNA genome, 26.2–31.7 kilobases, that encodes an important spike (S) glycoprotein that mediates viral entry and determines the range of potential host cell tropism and disease pathogenesis; hence, it has been a major source of vaccine interest (Qiang et al., 2020). Most scientists say that the earliest reports of clusters of patients probably had a natural origin and suggested that it was transmitted from an animal to humans. A lab leak has not been fully ruled out, and many are still calling for a deeper investigation into the hypothesis that the virus emerged from the Wuhan Institute of Virology located in the Chinese city where the first COVID-19 cases originated. The 2019-coronavirus-2 pandemic (COVID-19) affects every continent making it pandemic.

Six coronavirus species cause human disease (Su et al., 2016) types that are widely prevalent in the population and associated with common cold symptoms. Two others, SARS-CoV-1, the causal agent of the SARS outbreaks in 2002 and 2003 of Guangdong Province, China (Zhong et al., 2003) and the Middle East respiratory syndrome or MERS-CoV, responsible for outbreaks in 2012 (Zaki et al., 2012), are zoonotic beta coronaviruses and linked to fatal illness (Cui et al., 2019). SARS-Cov-1 and SARS-nCov-2 use angiotensin-converting enzyme 2 (ACE 2) receptor-binding sites to infect ciliated bronchial epithelial cells and type II pneumocystis, which explains the affinity for pulmonary involvement.

EPIDEMIOLOGY Zoonotic origin With 5 of the 7 known human coronaviruses isolated in this century, coronaviruses assume an important place in the 21st century (Bulut and Kato, 2020). SARS-nCoV-2 originated in bats and reached humans via badgers, Himalayan palm civets, and raccoon dogs, showing a

*Correspondence to: David S. Younger, MD, DrPH, MPH, MS, 333 East 34th Street, Suite 1J, New York, NY 10016, United States. Tel: +1-212-213-3778, Fax: +1-212-213-3779, E-mail: [email protected]

160

D.S. YOUNGER

capacity to infect humans by jumping across species from bat reservoirs. MERS-CoV a decade later originated in bats and utilize camels as intermediate hosts before infecting humans. The zoonotic origin of SARS-nCoV-2 has been confirmed with viral isolation from reservoirs in bats that infected, as intermediate hosts, the Malayan pangolin and other wildlife used for food in China (Lam et al., 2020; Cao et al., 2021). All three outbreaks attest to the high infectivity and lethality of coronaviruses and the serious public health threat it poses.

Animal models Experimental animals provide a model for human infection. Immunodeficient BALB/c and transgenic K18-hACE2 mice experimentally infected by SARSCoV-2 exhibit a clinical syndrome of acute infection similar to humans (McCray et al., 2007) invoking infiltration of macrophages and lymphocytes to the lungs and a local release of proinflammatory cytokines. They convincingly replicate the capacity to enter the central nervous system (CNS) across the blood–brain barrier (BBB) (Subbarao and Roberts, 2006) as do older immunodeficient BALB/c mice, exhibiting a more severe clinical syndrome with increasing age as a risk factor.

Disease definitions An analysis of 425 initial cases of novel coronavirus (2019-nCoV)–infected pneumonia of Wuhan, Hubei Province, China, from December 2019 to January 2020 underscored its early epidemiology (Li et al., 2020). Initially confirmed laboratory cases identified through surveillance of pneumonia of unknown etiology with fever (
38°C), radiographic evidence of pneumonia, low or normal white cell count or low lymphocyte count, and no symptomatic improvement after antimicrobial treatment for 3–5 days after standard therapy, were substantiated by World Health Organization (WHO) laboratory assays by extracting 2019-nCoV RNA by real-time polymerase chain reaction (RT-PCR) using specific primers and probes in upper and lower respiratory tract specimens. Epidemic curves showed an exponential growth rate of 0.10 per day with a doubling time of 7.4 days and a reproductive rate (R0) of 2.2, meaning that on average, each patient spread infection to 2.2 others. The goal of control measures through index case testing, contact tracing, quarantining, and prophylactic social distancing and facial masks in the general population aims to reduce the reproductive number to 70%) is needed to establish herd immunity to protect against epidemic transmission and protect individuals who are either ineligible or have absolute contraindications to receive a COVID vaccine. Vaccination side effects may be immediate, nonspecific, relatively innocuous, and short-lasting such as fever, flushing, and pain at the injection site; or true allergic reactions usually due to an active ingredient namely polyethylene glycol (PEG) in the Moderna and Pfizer vaccines) or polysorbate in the Johnson and Johnson vaccine, and numerous FDA-approved over-the-counter and oral and injectable prescription medications. Each has been associated with true allergic reactions including anaphylaxis requiring acute medical care following vaccination, especially the PEG-containing vaccines. While skin testing for PEG and polysorbate may be helpful in identifying individuals with a higher propensity for allergic reactions in screening for vaccine safety, experts advise avoiding vaccination instead based upon even a history of prior allergic reactions or anaphylaxis. There are potentially devastating neurological side effects of vaccination (Garg and Paliwal, 2022). The most serious neurological postvaccination complication is cerebral venous sinus thrombosis. Cerebral venous sinus is frequently reported in females of childbearing age, generally following adenovector-based vaccination. Another major neurological complication of concern is Bell’s palsy that was reported dominantly following

mRNA vaccine administration. Acute transverse myelitis, acute disseminated encephalomyelitis, and acute demyelinating polyneuropathy are other unexpected neurological adverse events that occur as result of phenomenon of molecular mimicry. Reactivation of herpes zoster in many persons, following administration of mRNA vaccines, has been also recorded. Considering the enormity of recent COVID-19-vaccinated population, the number of serious neurological events is miniscule. Large collaborative prospective studies are needed to prove or disprove causal association between vaccine and neurological adverse events occurring vaccination. To ensure that COVID-19 vaccines lead to widespread acceptance to reach herd immunity, government and public health leaders need to prepare transparent, evidence-based strategies to promote COVID-19 vaccine acceptance and implement equitable and effective vaccine delivery models or fail to contain the COVID-19 pandemic. Because vaccination campaigns require healthy people to seek an intervention, such campaigns require generating demand. Vaccine confidence was already decreasing worldwide before the COVID-19 pandemic for variety of cultural, political, and personal reasons. The WHO’s Increasing Vaccination Model acknowledges that people’s thoughts and feelings about vaccines, including their perceived risk, worry, confidence, trust, and safety concerns, can affect their motivation to get vaccinated. Vaccine hesitancy was cited in 2019 as one of the top 10 threats to global health.

CHRONIC COVID ILLNESS Long-Hauler and Long COVID The scientific community and healthcare professionals are recognizing the propensity of SARS-CoV-2 to manifest symptoms of chronic illness especially in the nervous system long after the acute-phase of infection. Although most individuals prefer to be recognized as COVID Long-Haulers due to Long COVID, the National Institutes of Health (NIH) adopted the term postacute sequelae of SARS-CoV-2 (PASC) for patients who fail to recover from acute COVID-19 or are persistently symptomatic for >30days after onset of infection, with any pattern of tissue injury, including the nervous system. Awaiting the results an NIH SARS-CoV-2 Recovery Cohort and Investigator Consortium (https://covid19. nih.gov/sites/default/files/2021-02/PASC-ROA-OTARecovery-Cohort-Studies.pdf) commissioned to study PASC, hundreds of thousands of individuals have taken to social media and online hosted surveys hoping to provide useful information to guide management, making them potentially useful cohorts for future research. Davis et al. (2021) characterized the symptom profile and time course in patients with Long COVID, along

COVID-19 (NOVEL SARS-CoV-2) NEUROLOGICAL ILLNESS with the impact on daily life, work, and return to baseline health using an international web-based survey design of suspected and confirmed COVID-19 cases with illness lasting over 28 days and onset prior to June 2020 in online COVID-19 support groups and social media from September to November 2020. The study group consisted of 3762 respondents from 56 countries. The most frequent symptoms reported after month 6 were: fatigue (77.7%), postexertional malaise (72.2%), and cognitive dysfunction (55.4%). Respondents with symptoms over 6 months experienced an average of 13.8 symptoms by month 7; 85.9% experienced relapses with exercise, physical or mental activity, and stress as the main triggers. Several others have investigated prolonged COVID symptoms (Sudre et al., 2021; Michelen et al., 2021) with similarly large online cohorts of 4182 and 10,951 respondents composed predominantly of individuals with continued symptoms at 6 months. In these cohorts, the most likely symptoms to persist after month 6 were fatigue, postexertional malaise, cognitive dysfunction (“brain fog”), neurologic sensations (neuralgias, weakness, coldness, electric shock sensations, facial paralysis/pressure/numbness), headaches, memory issues, insomnia, muscle aches, palpitations, shortness of breath, dizziness/balance issues, and speech and language issues. Importantly, the clusters of symptoms that persist the longest and include a combination of the neurological/ cognitive and systemic symptoms suggest the need for a multidisciplinary approach. Dysautonomia, in part manifesting as postural orthostatic tachycardia syndrome (POTS), and so-called myalgic encephalomyelitis/ chronic fatigue syndrome (ME/CFS) appear as likely diagnoses in this population. There are several limitations to online surveys. First, the retrospective nature of the study exposes the possibility of recall bias. Second, as the survey was distributed in online support groups, there may be sampling biases toward Long COVID patients who join support groups and are active participants of the groups at the time the survey is published. Despite translations and inclusive outreach efforts, the demographics are strongly skewed toward English speaking (91.9%), white (85.3%), and higher socioeconomic status. Moreover, these studies require respondents to have stable internet and email addresses, which may exclude participants who lack access and who have low digital literacy. Lastly, the effort to complete the survey can deter some respondents who experience cognitive dysfunction, or are no longer ill and do not have incentives to participate.

Postacute sequela of SARS-CoV-2 (PASC) Encephalopathy, demyelinating and axonal polyneuropathies, painful small fiber sensory polyneuropathy,

173

and dysautonomia established by standard laboratory testing indicate widespread postinfectious inflammatory involvement of the neuroaxis. Several regions of the brain that appear to be important in COVID-19 related encephalopathy include the frontal lobe, mesial temporal lobe and hippocampus. A pattern of hypometabolism in a widespread cerebral network including the prefrontal cortex, anterior cingulate, insula and caudate nucleus was observed in seven adult patients with COVID-19– related encephalopathy that persisted for 6 months after disease onset employing 18Fluorodeoxyglucose (FDG) positron emission tomography (PET) (Kas et al., 2021). By comparison, 18FDG brain PET fused to magnetic resonance imaging (MRI) with 3-dimensional postprocessing displayed prominent hypometabolism of the mesial temporal lobe and hippocampi in adult and pediatric cases of PASC associated with encephalopathy and neuropsychiatric mood disturbances that included obsessive compulsive disorder (OCD), anxiety and depression responsive to psychopharmacologic and immune modulatory therapy with IVIg (Fig. 8.1A and B) (Younger, 2021b, c). The finding of altered metabolism in these structures without morphological changes underscores the usefulness of neuroimaging as a useful biomarker for reversible COVID-19-related brain dysfunction.

Neurodegeneration Neurodegenerative disease is an umbrella concept including a range of conditions that primarily affect the neurons in the human brain and is one of the key factors leading to the decline in quality of life. Whether SARSCoV-2 causes neurodegenerative diseases or accelerates their premature occurrence is still unclear, and it is also difficult to draw conclusions without well designed epidemiological studies. However, the high expression of the ACE2 receptor in a wide range of sites in the brain not only provides an initial target for SARS-CoV-2 to cause acute brain damage, but may also be the basis for later neurodegenerative changes (Lukiw et al., 2022). This possibility is supported by the findings from recent studies that showed the presence of functional inhibition of viral and nicotinic acetylcholine receptor complexes in the pathogenesis of SARS-CoV-2 infection (Stefano et al., 2020).

ALZHEIMER DISEASE Recently published studies have focused on the potential causal relationship between viral infections and AD (Sochocka et al., 2017). Given the identification of damage to the CNS by SARS-CoV-2, there is concern regarding its long-term effects on cognitive function (Calderon-Garciduenas et al., 2020). Long-term studies

174

D.S. YOUNGER

Fig. 8.1. (A) 3D-SSP analysis of 18FDG PET normalized to the whole brain shows mild-to-moderate FDG hypometabolism in the bilateral anterior and medial temporal lobes, superior parietal lobes, lateral occipital lobes, anterior cingulate cortices, and bilateral cerebellar hemispheres. (B) 18FDG PET/MRI-fused image clearly shows the degree of FDG hypometabolism and volume loss in the bilateral hippocampi. Abbreviations: 3D-SSP, three-dimensional stereotactic surface projection; 18FDG, 18Fluorodeoxyglucose; PET, positron emission tomography; MRI, magnetic resonance imaging. Courtesy of Elcin Zan MD, Department of Radiology (Neuroradiology), New York University Grossman School of Medicine, New York.

will be required to identify the relationships among SARS-CoV-2 infection, AD, and other neurodegenerative sequelae. Neuroinflammatory responses, synaptic pruning, and neuronal loss are the structural basis of AD (Heneka et al., 2015), and SARS-CoV-2 infection most likely accelerates these processes. The excitotoxic reaction caused by the imbalance between glutamatergic and GABAergic responses is a potential mechanism that promotes neuronal loss and further cerebral tissue damage (Guo et al., 2020b). The simultaneous expression of ACE2 in glutamatergic and GABAergic neurons indicates that SARS-CoV-2 infection can affect the balance of both signaling pathways in the CNS.

Furthermore, the trans-synaptic transfer and the axonal retrograde or anterograde movement of SARS-CoV-2 make it possible for the virus to slowly and diffusely infiltrate the entire brain, and it also promotes the chronicity and the neurodegenerative changes months and years after the acute infection.

PARKINSON DISEASE Compared with AD, the potential CNS damage that is localized to the substantia nigra striatum that can result in Parkinson disease (PD) seems to be more limited. However, several recent studies have shown that patients

COVID-19 (NOVEL SARS-CoV-2) NEUROLOGICAL ILLNESS with PD not only show motor dysfunction, their cognitive and memory functions are also severely impaired (Das et al., 2019). Also, the pathogenesis of PD is associated with neuroinflammation, synaptic pruning, and neuron loss (Beitz, 2014) sharing commonalities with AD (Bernaus et al., 2020). However, in PD, different CNS sites are damaged, with different types of neurons being more severely affected (Compta et al., 2014). Currently, although there is no direct evidence that SARS-CoV-2 could cause or accelerates PD, it should be noted that the wide expression of ACE2 at different areas in the CNS provides a molecular basis for SARS-CoV-2 to mediate or accelerate the occurrence of PD.

MULTIPLE SCLEROSIS Multiple sclerosis (MS) is associated with focal gray and white matter demyelination and diffuse neurodegeneration of the brain caused by inflammation (Lassmann, 2018). Current knowledge of the neurological changes caused by SARS-CoV-2 shows some similarities with those found in MS. First, the proinflammatory cytokine storm caused by SARS-CoV-2 infection is the initiating factor of CNS neuroinflammatory damage. Second, SARS-CoV-2 may cause demyelination in the brain and spinal cord. A recently published case report showed that SARS-CoV-2 infection was associated with signs and symptoms similar to those of MS (Zanin et al., 2020). Previous studies have shown an association between coronavirus infection and the onset of MS (Arbour et al., 2000).

Neuropsychiatric illness Fears of illness, death, and uncertainty of the future are significant psychological stressors for the population, and social isolation resulting from loss of structured educational and work activities associated with the COVID-19 pandemic threatens to worsen public mental health (Carvalho et al., 2020). For front-line healthcare workers, regular exposure to the illness, protective equipment shortages, and adaptation to rapidly evolving and high-stress work environments are further sources of distress. There have been calls for the development and implementation of mental health screening and intervention programs for both the public and for healthcare workers (Bao et al., 2020). Comparably less attention has focused on SARS-CoV-2 and the host immunologic response to infection on the CNS and related neuropsychiatric outcomes. However, survivors of both SARSCoV-1 were clinically diagnosed with PTSD (54.5%), depression (39%), pain disorder (36.4%), panic disorder (32.5%), and obsessive compulsive disorder (15.6%), a dramatic increase from their preinfection prevalence of

175

any psychiatric diagnoses (of 3%) (Lam et al., 2009) with persistence for up to 4-year follow-up. SARS-CoV-2 likewise appears to contribute to the development of mood and psychotic disorders, with the postulation of the contribution of various biological alterations associated with coronavirus infection including activation of microglia and cytokine signaling (Szczesniak et al., 2021). A surprisingly important source of brain psychopathology may be found in the microbiota-brain axis that includes metabolic byproducts of infection, and connections between the enteric nervous system (ENS) and CNS along the gut–brain neural network via neurotransmitters and neural regulators synthesized by gut microbes and transmitted across the intestinal mucosal barrier and BBB, and along afferent and efferent nerves that link the CNS to the gut (Rao and Gershon, 2016; Mayer et al., 2021). It is of interest that viral shedding in feces of COVID-19 patients is known to occur for at least 5 weeks postinfection (Wu et al., 2020). Although the extent and mechanisms of viral infiltration of gut epithelium by SARS-CoV-2 are currently unknown, ACE2 is expressed by gut epithelial cells, and almost 40% of COVID-19 patients present with GI symptoms, making it a potential source for gut–brain influences.

CONCLUSION Given their high prevalence and wide distribution, prominent genetic diversity, genomic recombination, and human–animal interface activities in certain parts of the world, the COVID-19 pandemic and other novel coronaviruses will likely continue to proliferate. This depends upon multiple factors, not the least of which is super-spreading, which occurs when single patients infect a disproportionate number of contacts across continents enhanced by travel. With them comes the foreseeable risk of rising fatality and expected neurological complications. Adults with SARS-CoV-1 and nCoV-2 have shown inflammatory vasculopathy or vasculitis at postmortem examination, as do many children with MIS in life. This has led to innovative treatments aimed at viral eradication and immunotherapy directed at a heightened postinfectious inflammatory response termed I3 that expresses the multiplier effect of infection, immunity, and inflammation in the context of genetics and other environmental exposures.

REFERENCES Al-Dalahmah O, Thakur KT, Nordvig AS et al. (2020). Neuronophagia and microglial nodules in a SARS-CoV-2 patient with cerebellar hemorrhage. Acta Neuropathol Commun 8: 147.

176

D.S. YOUNGER

Arabi YM, Fowler R, Hayden FG (2020). Critical care management of adults with community-acquired severe respiratory viral infection. Intensive Care Med 46: 315–328. Arbour N, Day R, Newcombe J et al. (2000). Neuroinvasion by human respiratory coronaviruses. J Virol 74: 8913–8921. Arunachalam PS, Wimmers F, Mok CKP et al. (2020). Systems biological assessment of immunity to mild versus severe COVID-19 infection in humans. Science 369: 1210–1220. Baden LR, El Sahly HM, Essink B et al. (2021). Efficacy and safety of the mRNA-1273 SARS-CoV-2 vaccine. N Engl J Med 384: 403–416. Baker AL, Lu M, Minich LL et al. (2009). Associated symptoms in the ten days before diagnosis of Kawasaki disease. J Pediatr 154: e592. Bao Y, Sun Y, Meng S et al. (2020). 2019-nCoV epidemic: address mental health care to empower society. Lancet 395: e37–e38. Barlev-Gross M, Weiss S, Ben-Shmuel A et al. (2021). Spike vs nucleocapsid SARS-CoV-2 antigen detection: application in nasopharyngeal swab specimens. Anal Bioanal Chem 413: 3501–3510. Barton LM, Duval EJ, Stroberg E et al. (2020). COVID-19 autopsies, Oklahoma, USA. Am J Clin Pathol 153: 725–733. Beitz JM (2014). Parkinson’s disease: a review. Front Biosci (Schol Ed) 6: 65–74. Bernaus A, Blanco S, Sevilla A (2020). Glia crosstalk in Neuroinflammatory diseases. Front Cell Neurosci 14: 209. Bestle D, Heindl MR, Limburg H et al. (2020). TMPRSS2 and furin are both essential for proteolytic activation of SARSCoV-2 in human airway cells. Life Sci Alliance 3: 9. Biot C, Daher W, Chavain N et al. (2006). Design and synthesis of hydroxyferroquine derivatives with antimalarial and antiviral activities. J Med Chem 49: 2845–2849. Brown CM, Vostok J, Johnson H et al. (2021). Outbreak of SARS-CoV-2 infections, including COVID-19 vaccine breakthrough infections, associated with large public gatherings—Barnstable County, Massachusetts, July 2021. MMWR Morb Mortal Wkly Rep 70: 1059–1062. Bryce C, Grimes Z, Pujadas E et al. (2021). Pathophysiology of SARS-CoV-2: the Mount Sinai COVID-19 autopsy experience. Mod Pathol 34: 1456–1467. Buja LM, Wolf DA, Zhao B et al. (2020). The emerging spectrum of cardiopulmonary pathology of the coronavirus disease 2019 (COVID-19): report of 3 autopsies from Houston, Texas, and review of autopsy findings from other United States cities. Cardiovasc Pathol 48: 107233. Bulut C, Kato Y (2020). Epidemiology of COVID-19. Turk J Med Sci 50: 563–570. Calderon-Garciduenas L, Torres-Jardon R, Franco-Lira M et al. (2020). Environmental nanoparticles, SARS-CoV-2 brain involvement, and potential acceleration of Alzheimer’s and Parkinson’s diseases in young urbanites exposed to air pollution. J Alzheimers Dis 78: 479–503. Cao W, Liu X, Bai T et al. (2020). High-dose intravenous immunoglobulin as a therapeutic option for deteriorating patients with coronavirus disease 2019. Open Forum Infect Dis 7: ofaa102.

Cao LL, Guan PP, Zhang SQ et al. (2021). Downregulating expression of OPTN elevates neuroinflammation via AIM2 inflammasome- and RIPK1-activating mechanisms in APP/PS1 transgenic mice. J Neuroinflammation 18: 281. Carsana L, Sonzogni A, Nasr A et al. (2020). Pulmonary postmortem findings in a series of COVID-19 cases from northern Italy: a two-Centre descriptive study. Lancet Infect Dis 20: 1135–1140. Carvalho PMM, Moreira MM, de Oliveira MNA et al. (2020). The psychiatric impact of the novel coronavirus outbreak. Psychiatry Res 286: 112902. CDC (2020). Return-to-work criteria for healthcare workers [online], Center for Disease Control. Available. https:// www.cdc.gov/coronavirus/2019-ncov/hcp/return-to-work. htmlAccessed May 3, 2020. Cecere TE, Todd SM, Leroith T (2012). Regulatory T cells in arterivirus and coronavirus infections: do they protect against disease or enhance it? Viruses 4: 833–846. Chen P, Nirula A, Heller B et al. (2021). SARS-CoV-2 neutralizing antibody LY-CoV555 in outpatients with Covid-19. N Engl J Med 384: 229–237. Compta Y, Parkkinen L, Kempster P et al. (2014). The significance of alpha-synuclein, amyloid-beta and tau pathologies in Parkinson’s disease progression and related dementia. Neurodegener Dis 13: 154–156. Copin MC, Parmentier E, Duburcq T et al. (2020). Time to consider histologic pattern of lung injury to treat critically ill patients with COVID-19 infection. Intensive Care Med 46: 1124–1126. Cortegiani A, Ippolito M, Einav S (2021). Rationale and evidence on the use of tocilizumab in COVID-19: a systematic review. Authors’ reply. Pulmonology 27: 87–88. Cui J, Li F, Shi ZL (2019). Origin and evolution of pathogenic coronaviruses. Nat Rev Microbiol 17: 181–192. Das T, Hwang JJ, Poston KL (2019). Episodic recognition memory and the hippocampus in Parkinson’s disease: a review. Cortex 113: 191–209. Davis HE, Assaf GS, McCorkell L et al. (2021). Characterizing long COVID in an international cohort: 7 months of symptoms and their impact. EClinicalMedicine 38: 101019. Deigendesch N, Sironi L, Kutza M et al. (2020). Correlates of critical illness-related encephalopathy predominate postmortem COVID-19 neuropathology. Acta Neuropathol 140: 583–586. Dufort EM, Koumans EH, Chow EJ et al. (2020). Multisystem inflammatory syndrome in children in New York state. N Engl J Med 383: 347–358. Feldstein LR, Rose EB, Horwitz SM et al. (2020). Multisystem inflammatory syndrome in U.S. children and adolescents. N Engl J Med 383: 334–346. Fox SE, Akmatbekov A, Harbert JL et al. (2020). Pulmonary and cardiac pathology in African American patients with COVID-19: an autopsy series from New Orleans. Lancet Respir Med 8: 681–686. Frediani JK, Levy JM, Rao A et al. (2021). Multidisciplinary assessment of the Abbott BinaxNOW SARS-CoV-2 pointof-care antigen test in the context of emerging viral variants and self-administration. Sci Rep 11: 14604.

COVID-19 (NOVEL SARS-CoV-2) NEUROLOGICAL ILLNESS Garg RK, Paliwal VK (2022). Spectrum of neurological complications following COVID-19 vaccination. Neurol Sci 43: 3–40. Geleris J, Sun Y, Platt J et al. (2020). Observational study of hydroxychloroquine in hospitalized patients with Covid19. N Engl J Med 382: 2411–2418. Gottlieb RL, Nirula A, Chen P et al. (2021). Effect of bamlanivimab as monotherapy or in combination with etesevimab on viral load in patients with mild to moderate COVID19: a randomized clinical trial. JAMA 325: 632–644. Griffith JF, Antonio GE, Kumta SM et al. (2005). Osteonecrosis of hip and knee in patients with severe acute respiratory syndrome treated with steroids. Radiology 235: 168–175. Guo XJ, Thomas PG (2017). New fronts emerge in the influenza cytokine storm. Semin Immunopathol 39: 541–550. Guo L, Ren L, Yang S et al. (2020a). Profiling early humoral response to diagnose novel coronavirus disease (COVID19). Clin Infect Dis 71: 778–785. Guo T, Zhang D, Zeng Y et al. (2020b). Molecular and cellular mechanisms underlying the pathogenesis of Alzheimer’s disease. Mol Neurodegener 15: 40. Gupta A, Gonzalez-Rojas Y, Juarez E et al. (2021). Early treatment for Covid-19 with SARS-CoV-2 neutralizing antibody sotrovimab. N Engl J Med 385: 1941–1950. Hammond J, Leister-Tebbe H, Gardner A et al. (2022). Oral nirmatrelvir for high-risk, nonhospitalized adults with Covid-19. N Engl J Med 386: 1397–1408. Hanley B, Naresh KN, Roufosse C et al. (2020). Histopathological findings and viral tropism in UK patients with severe fatal COVID-19: a post-mortem study. Lancet Microbe 1: e245–e253. He S, Chen S, Kong L et al. (2021). Analysis of risk perceptions and related factors concerning COVID-19 epidemic in Chongqing, China. J Community Health 46: 278–285. Helms J, Kremer S, Meziani F (2020). More on neurologic features in severe SARS-CoV-2 infection. Reply. N Engl J Med 382: e110. Heneka MT, Carson MJ, El Khoury J et al. (2015). Neuroinflammation in Alzheimer’s disease. Lancet Neurol 14: 388–405. Jean SS, Lee PI, Hsueh PR (2020). Treatment options for COVID-19: the reality and challenges. J Microbiol Immunol Infect 53: 436–443. Kandeel M, Mohamed MEM, Abd El-Lateef HM et al. (2021). Omicron variant genome evolution and phylogenetics. J Med Virol 94: 4. Kantonen J, Mahzabin S, Mayranpaa MI et al. (2020). Neuropathologic features of four autopsied COVID-19 patients. Brain Pathol 30: 1012–1016. Kas A, Soret M, Pyatigoskaya N et al. (2021). The cerebral network of COVID-19-related encephalopathy: a longitudinal voxel-based 18F-FDG-PET study. Eur J Nucl Med Mol Imaging 48: 2543–2557. Lam MH, Wing YK, Yu MW et al. (2009). Mental morbidities and chronic fatigue in severe acute respiratory syndrome

177

survivors: long-term follow-up. Arch Intern Med 169: 2142–2147. Lam TT, Jia N, Zhang YW et al. (2020). Identifying SARS-CoV-2-related coronaviruses in Malayan pangolins. Nature 583: 282–285. Lassmann H (2018). Multiple sclerosis pathology. Cold Spring Harb Perspect Med 8: 3. Lax SF, Skok K, Zechner P et al. (2020). Pulmonary arterial thrombosis in COVID-19 with fatal outcome: results from a prospective, single-center, clinicopathologic case series. Ann Intern Med 173: 350–361. Lee DW, Gardner R, Porter DL et al. (2014). Current concepts in the diagnosis and management of cytokine release syndrome. Blood 124: 188–195. Levin MJ, Ustianowski A, De Wit S et al. (2021). LB5. PROVENT: phase 3 study of efficacy and safety of AZD7442 (Tixagevimab/Cilgavimab) for pre-exposure prophylaxis of COVID-19 in adults. Open Forum Infect Dis 8: S810. Li Q, Guan X, Wu P et al. (2020). Early transmission dynamics in Wuhan, China, of novel coronavirus-infected pneumonia. N Engl J Med 382: 1199–1207. Lucas C, Wong P, Klein J et al. (2020). Longitudinal analyses reveal immunological misfiring in severe COVID-19. Nature 584: 463–469. Lukiw WJ, Pogue A, Hill JM (2022). SARS-CoV-2 infectivity and neurological targets in the brain. Cell Mol Neurobiol 42: 217–224. Magro C, Mulvey JJ, Berlin D et al. (2020). Complement associated microvascular injury and thrombosis in the pathogenesis of severe COVID-19 infection: a report of five cases. Transl Res 220: 1–13. Malani PN, Golub RM (2021). Neutralizing monoclonal antibody for mild to moderate COVID-19. JAMA 325: 644–645. Mao L, Jin H, Wang M et al. (2020). Neurologic manifestations of hospitalized patients with coronavirus disease 2019 in Wuhan, China. JAMA Neurol 77: 683–690. Matschke J, Lutgehetmann M, Hagel C et al. (2020). Neuropathology of patients with COVID-19 in Germany: a post-mortem case series. Lancet Neurol 19: 919–929. Mayer EA, Nance K, Chen S (2021). The gut-brain axis. Annu Rev Med 73: 439–453. McCray Jr PB, Pewe L, Wohlford-Lenane C et al. (2007). Lethal infection of K18-hACE2 mice infected with severe acute respiratory syndrome coronavirus. J Virol 81: 813–821. Menter T, Haslbauer JD, Nienhold R et al. (2020). Postmortem examination of COVID-19 patients reveals diffuse alveolar damage with severe capillary congestion and variegated findings in lungs and other organs suggesting vascular dysfunction. Histopathology 77: 198–209. Michelen M, Manoharan L, Elkheir N et al. (2021). Characterising long COVID: a living systematic review. BMJ Glob Health 6: 9. Najjar-Debbiny R, Gronich N, Weber G et al. (2023). Effectiveness of Paxlovid in reducing severe coronavirus disease 2019 and mortality in high-risk patients. Clin Infect Dis 76: e342–e349.

178

D.S. YOUNGER

Netland J, Meyerholz DK, Moore S et al. (2008). Severe acute respiratory syndrome coronavirus infection causes neuronal death in the absence of encephalitis in mice transgenic for human ACE2. J Virol 82: 7264–7275. Peiris JS, Chu CM, Cheng VC et al. (2003). Clinical progression and viral load in a community outbreak of coronavirus-associated SARS pneumonia: a prospective study. Lancet 361: 1767–1772. Perico L, Benigni A, Casiraghi F et al. (2021). Immunity, endothelial injury and complement-induced coagulopathy in COVID-19. Nat Rev Nephrol 17: 46–64. Poisson KE, Zygmunt A, Leino D et al. (2022). Lethal pediatric cerebral Vasculitis triggered by severe acute respiratory syndrome coronavirus 2. Pediatr Neurol 127: 1–5. Polack FP, Thomas SJ, Kitchin N et al. (2020). Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine. N Engl J Med 383: 2603–2615. Puelles VG, Lutgehetmann M, Lindenmeyer MT et al. (2020). Multiorgan and renal tropism of SARS-CoV-2. N Engl J Med 383: 590–592. Qiang XL, Xu P, Fang G et al. (2020). Using the spike protein feature to predict infection risk and monitor the evolutionary dynamic of coronavirus. Infect Dis Poverty 9: 33. Qiao J (2020). What are the risks of COVID-19 infection in pregnant women? Lancet 395: 760–762. Radermecker C, Detrembleur N, Guiot J et al. (2020). Neutrophil extracellular traps infiltrate the lung airway, interstitial, and vascular compartments in severe COVID-19. J Exp Med 217. Ramani A, Muller L, Ostermann PN et al. (2020). SARS-CoV2 targets neurons of 3D human brain organoids. EMBO J 39: e106230. Rao M, Gershon MD (2016). The bowel and beyond: the enteric nervous system in neurological disorders. Nat Rev Gastroenterol Hepatol 13: 517–528. Reichard RR, Kashani KB, Boire NA et al. (2020). Neuropathology of COVID-19: a spectrum of vascular and acute disseminated encephalomyelitis (ADEM)-like pathology. Acta Neuropathol 140: 1–6. Russell CD, Millar JE, Baillie JK (2020). Clinical evidence does not support corticosteroid treatment for 2019-nCoV lung injury. Lancet 395: 473–475. Sadoff J, Le Gars M, Shukarev G et al. (2021). Interim results of a phase 1-2a trial of Ad26.COV2.S Covid-19 vaccine. N Engl J Med 384: 1824–1835. Sallusto F, Lenig D, Forster R et al. (1999). Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature 401: 708–712. Schafer A, Muecksch F, Lorenzi JCC et al. (2021). Antibody potency, effector function, and combinations in protection and therapy for SARS-CoV-2 infection in vivo. J Exp Med 218. Schaller T, Hirschbuhl K, Burkhardt K et al. (2020). Postmortem examination of patients with COVID-19. JAMA 323: 2518–2520. Shen C, Wang Z, Zhao F et al. (2020). Treatment of 5 critically ill patients with COVID-19 with convalescent plasma. JAMA 323: 1582–1589.

Sochocka M, Zwolinska K, Leszek J (2017). The infectious etiology of Alzheimer’s disease. Curr Neuropharmacol 15: 996–1009. Solomon IH, Normandin E, Bhattacharyya S et al. (2020). Neuropathological features of Covid-19. N Engl J Med 383: 989–992. Stefano ML, Kream RM, Stefano GB (2020). A novel vaccine employing non-replicating rabies virus expressing chimeric SARS-CoV-2 spike protein domains: functional inhibition of viral/nicotinic acetylcholine receptor complexes. Med Sci Monit 26: e926016. Su S, Wong G, Shi W et al. (2016). Epidemiology, genetic recombination, and pathogenesis of coronaviruses. Trends Microbiol 24: 490–502. Subbarao K, Roberts A (2006). Is there an ideal animal model for SARS? Trends Microbiol 14: 299–303. Sudre CH, Murray B, Varsavsky T et al. (2021). Attributes and predictors of long COVID. Nat Med 27: 626–631. Szczesniak D, Gladka A, Misiak B et al. (2021). The SARSCoV-2 and mental health: from biological mechanisms to social consequences. Prog Neuro-Psychopharmacol Biol Psychiatry 104: 110046. Tahamtan A, Ardebili A (2020). Real-time RT-PCR in COVID-19 detection: issues affecting the results. Expert Rev Mol Diagn 20: 453–454. To KK, Tsang OT, Leung WS et al. (2020). Temporal profiles of viral load in posterior oropharyngeal saliva samples and serum antibody responses during infection by SARS-CoV2: an observational cohort study. Lancet Infect Dis 20: 565–574. Varatharaj A, Thomas N, Ellul MA et al. (2020). Neurological and neuropsychiatric complications of COVID-19 in 153 patients: a UK-wide surveillance study. Lancet Psychiatry 7: 875–882. von Weyhern CH, Kaufmann I, Neff F et al. (2020). Early evidence of pronounced brain involvement in fatal COVID-19 outcomes. Lancet 395: e109. Wang W, Xu Y, Gao R et al. (2020). Detection of SARS-CoV2 in different types of clinical specimens. JAMA 323: 1843–1844. Wang P, Nair MS, Liu L et al. (2021). Antibody resistance of SARS-CoV-2 variants B.1.351 and B.1.1.7. Nature 593: 130–135. Weinreich DM, Sivapalasingam S, Norton T et al. (2021). REGN-COV2, a neutralizing antibody cocktail, in outpatients with Covid-19. N Engl J Med 384: 238–251. Whittaker E, Bamford A, Kenny J et al. (2020). Clinical characteristics of 58 children with a pediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2. JAMA 324: 259–269. Wolfel R, Corman VM, Guggemos W et al. (2020). Virological assessment of hospitalized patients with COVID-2019. Nature 581: 465–469. Wu Y, Guo C, Tang L et al. (2020). Prolonged presence of SARS-CoV-2 viral RNA in faecal samples. Lancet Gastroenterol Hepatol 5: 434–435. Wu L, Zhou L, Mo M et al. (2022). SARS-CoV-2 omicron RBD shows weaker binding affinity than the currently dominant Delta variant to human ACE2. Signal Transduct Target Ther 7: 8.

COVID-19 (NOVEL SARS-CoV-2) NEUROLOGICAL ILLNESS Xu J, Yang X, Yang L et al. (2020a). Clinical course and predictors of 60-day mortality in 239 critically ill patients with COVID-19: a multicenter retrospective study from Wuhan, China. Crit Care 24: 394. Xu K, Chen Y, Yuan J et al. (2020b). Factors associated with prolonged viral RNA shedding in patients with coronavirus disease 2019 (COVID-19). Clin Infect Dis 71: 799–806. Younger DS (2016). I-cubed (infection, immunity, and inflammation) and the human microbiome. Neurol Clin 34: 863–874. Younger DS (2019a). Eleven themes in the history of systemic and nervous system Vasculitides. Neurol Clin 37: 149–170. Younger DS (2019b). Overview of the Vasculitides. Neurol Clin 37: 171–200. Younger DS (2020a). Book review: I—cubed and the autoimmune brain, a five-step plan. World J Neurosci 10: 29–36. Younger DS (2020b). Post-infectious sequela of SARS-COV2 infection in adults and children: an overview of available agents and clinical responsiveness. Arch Neurol 3: e102. Younger DS (2021a). Coronavirus 2019: clinical and neuropathological aspects. Curr Opin Rheumatol 33: 49–57. Younger DS (2021b). Post-acute sequelae of SARS-CoV-2 infection (PASC): peripheral, autonomic, and central nervous system features in a child. Neurol Sci 42: 3959–3963. Younger DS (2021c). Post-acute sequelae of SARS-Cov-2 infection associating peripheral, autonomic and central nervous system disturbances: case report and review of the literature. World J Neurosci 11: 17–21.

179

Younger DS (2021d). Postmortem neuropathology in COVID-19. Brain Pathol 31: 385–386. Zaki AM, van Boheemen S, Bestebroer TM et al. (2012). Isolation of a novel coronavirus from a man with pneumonia in Saudi Arabia. N Engl J Med 367: 1814–1820. Zanin L, Saraceno G, Panciani PP et al. (2020). SARS-CoV-2 can induce brain and spine demyelinating lesions. Acta Neurochir 162: 1491–1494. Zha L, Li S, Pan L et al. (2020). Corticosteroid treatment of patients with coronavirus disease 2019 (COVID-19). Med J Aust 212: 416–420. Zhang Y, Li J, Zhan Y et al. (2004). Analysis of serum cytokines in patients with severe acute respiratory syndrome. Infect Immun 72: 4410–4415. Zhang L, Jackson CB, Mou H et al. (2020). SARS-CoV-2 spike-protein D614G mutation increases virion spike density and infectivity. Nat Commun 11: 6013. Zheng S, Fan J, Yu F et al. (2020). Viral load dynamics and disease severity in patients infected with SARS-CoV-2 in Zhejiang province, China, January-March 2020: retrospective cohort study. BMJ 369: m1443. Zhong NS, Zheng BJ, Li YM et al. (2003). Epidemiology and cause of severe acute respiratory syndrome (SARS) in Guangdong, People’s Republic of China, in February, 2003. Lancet 362: 1353–1358. Zhu N, Zhang D, Wang W et al. (2020). A novel coronavirus from patients with pneumonia in China, 2019. N Engl J Med 382: 727–733. Zinkernagel RM (1996). Immunology taught by viruses. Science 271: 173–178.

This page intentionally left blank

Section 2 Clinical and laboratory diagnosis

This page intentionally left blank

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00003-0 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 9

Neurogenetic motor disorders DAVID S. YOUNGER1,2* 1

Department of Clinical Medicine and Neuroscience, CUNY School of Medicine, New York, NY, United States

2

Department of Medicine, Section of Internal Medicine and Neurology, White Plains Hospital, White Plains, NY, United States

Abstract Advances in the field of neurogenetics have practical applications in rapid diagnosis on blood and body fluids to extract DNA, obviating the need for invasive investigations. The ability to obtain a presymptomatic diagnosis through genetic screening and biomarkers can be a guide to life-saving disease-modifying therapy or enzyme replacement therapy to compensate for the deficient disease-causing enzyme. The benefits of a comprehensive neurogenetic evaluation extend to family members in whom identification of the causal gene defect ensures carrier detection and at-risk counseling for future generations. This chapter explores the many facets of the neurogenetic evaluation in adult and pediatric motor disorders as a primer for later chapters in this volume and a roadmap for the future applications of genetics in neurology.

INTRODUCTION Genetics is the study of inheritance with many overlapping concepts. Classic genetics is the study of genes that form the basis for inherited physical traits such as eye color. Cytogenetics deals with chromosomes in a karyotype obtained before or after birth in amniotic fluid or blood. Molecular genetics examines the fine structure of alleles encoded in nucleic acid base pairs of deoxyribonucleic acid (DNA). The latter is transcribed into ribonucleic acid (RNA) forming a mirror image of the genetic blueprint, which, like a manufacturing code for the production of all body proteins, gives precise directions for protein molecule production and assembly. Abolition of gene product function by a causative mutation is associated with the most severe form of an illness often characterized by early symptom onset and rapid progression. In some instances, the gene is altered but the product is not completely lost and there may even be residual function leading to an attenuated form of the disease with later age of onset and a more insidious course. Once thought that characterization of the genotype would assist in accurately predicting the phenotype, there is often a lack of perfect concordance

suggesting the influence of other factors in disease expression. These modifiers may be other genes, gene products and other epigenetic influences that interact with the primary defect.

MITOCHONDRIAL GENETICS An understanding of mitochondrial (mt) genetics beginning in the 1960s focused attention on ultrastructurally abnormal organelles (Shy et al., 1966) and myofibers that stained ragged red (RRF) with a modified G€om€ori trichrome stain (Engel and Cunningham, 1963) with evidence of mitochondrial proliferation with succinate dehydrogenase (SDH) and cytochrome-c-oxidase (COX) enzymes. However, the histologic abnormalities are neither specific nor sensitive enough to define all mitochondrial diseases. As a general rule, mutations in structural genes are not associated with RRFs. Mitochondrial diseases are highly heterogeneous with varying clinical features caused by impaired function of the mitochondrial respiratory chain. The underlying cause can be gene mutations in nuclear or mitochondrial mtDNA, both of which contain genes encoding components of the oxidative phosphorylation machinery that generate

*Correspondence to: David S. Younger, MD, DrPH, MPH, MS, 333 East 34th Street, Suite 1J, New York, NY 10016, United States. Tel: +1-212-213-3778, Fax: +1-212-213-3779, E-mail: [email protected]

184

D.S. YOUNGER

adenosine triphosphate (ATP), the currency of cellular energy (Gorman et al., 2016). Since the initial discoveries of mtDNA point mutation and large-scale deletions (Holt et al., 1988; Lestienne and Ponsot, 1988; Wallace et al., 1988; Zeviani et al., 1988), more than 250 distinct mitochondrial diseases have been linked to a variety of pathogenic variants occurring in the nuclear or mtDNA genomes sometimes manifesting as severe and often lethal multisystemic disorders (Falkenberg and Hirano, 2020). However, modifiers and variation in mtDNA influence phenotypic expression as in Leber hereditary optic atrophy (LHON) characterized by bilateral, painless, subacute central vision loss in young adults resulting from primary degeneration of retinal ganglion cells (RGCs) and optic atrophy (Yu et al., 2020). Although three common mutations in mitochondrial complex I, subunit ND4 (MTND4) at 11778G>A, 14484T>C, or 3460G>A are responsible for over 90% of cases and affect genes encoding complex I subunits of the respiratory chain, their influence on bioenergetic properties of the cell do not fully explain the pathology of the disease. As LHON is characterized by low penetrance and sex bias, the participation of the other factorsgenetic and environmental-beside mtDNA mutations have been studied showing the impact of mitochondrial haplotypes and cigarette smoking in the complex etiology of the disorder. Moreover, concomitant hemizygous or heterozygous mutation in the prickle planar cell polarity protein 3 (PRICKLE3) on chromosome Xp11 synonymous with the Leber optic atrophy modifier (LOAM), combines resulting in higher penetrance and younger age of onset of optic neuropathy than MTND4 alone. Cells in the brain and muscle with an increased demand for aerobic metabolism can be compromised by mtDNA mutation impairing ATP generation. Those with exclusively mutant mtDNA termed homoplasmy, may demonstrate a more severe phenotype, in contrast to those with a mixed population of mutant and normal mtDNA termed heteroplasmy. Whereas a high load of the mtDNA T8993G mutation leads to fatal childhood maternally inherited Leigh syndrome (MILS), a lower mutant threshold causes Neuropathy, Ataxia and Retinitis pigmentosa (NARP) (D’Aurelio et al., 2010). Investigations have shown that the mtDNA background play an important role in modulating the biochemical defects and clinical outcome in NARP/MILS (D’Aurelio et al., 2010). There are many large-scale deletions and point mutations of mtDNA (Lott et al., 2013). Moreover, the threshold for biochemical deficiency varies for each mutation and cell type and may even be different for the same mutation in different patients. A pathogenic mutation that is normally required to reach a high heteroplasmy (>70%) before a deleterious biochemical phenotype is expressed at a single-cell level may be associated with a clinical phenotype that will also vary in patients with the same mtDNA

mutation even at apparently similar levels of heteroplasmy (Pickett et al., 2018). Inherited and spontaneous genetic mutation leading to errors in transcription, translation, RNA processing, posttranslational modifications and environmental or epigenetic influences may be factors in the cause of neurogenetic illness. Society’s desire to learn about our human nature that led to the human genome project by the National Human Genome Research Institute (Green et al., 2015) paved the way for more than a thousand consortium-based Genomics Projects, culminating in numerous crucial genomic technologies, and substantial innovations in molecular biology and the understanding of diverse neurogenetic disorders. Notwithstanding, the ability of individuals and their physicians to see the entire gene pool in a printout of alleles has brought about new insights and limitations to concepts of health and illness.

CLINICAL CLUES OF A NEUROGENETIC MOTOR DISORDER The history and examination are important first steps in the diagnosis of a genetic motor disorder of the peripheral (PNS), central (CNS) and autonomic nervous systems (ANS). The goal is to establish the tempo, distribution, severity, and diversity of motor symptoms and signs, along with the associated findings including nonmotor symptoms in order formulate an accurate presumptive diagnosis and localize the disease process. A detailed neurologic examination is the next step in the elucidation of a likely neurogenetic disorder that begins with assessment of mental status for memory loss or dementia. The examination of cranial function includes assessment of ocular motility, strength of facial and neck muscles, and tests for auditory, vestibular, speech and lingual function. Individual limb muscles should be graded on a scale of 0–5 according to the criteria of the Medical Research Council (MRC) (1982). Balance and coordination should be ascertained with eyes open and closed with feet standing together and with tandem gait for axial and limb coordination and sensory ataxia, further amplified by rapid successive movements and finger-to-nose pointing. Thermal, pinprick, light touch and proprioceptive sensation and cortical sensory perception should be tested and rated by the patient. Tendon reflexes are best tested in the seated position with hands folded in the lap and legs dangling. The inclusion of nonmotor symptoms and signs to the assessment adds precision to the presumptive diagnosis since sensory symptoms, when they occur, suggests an underlying cause other than myopathy, for example, in a lesion along the peripheral nerves, dorsal root ganglia (DRG) and dorsal columns of the spinal cord. The rule that proximal or girdle weakness equals myopathy and

NEUROGENETIC MOTOR DISORDERS distal weakness implies neuropathy is useful only when the latter is accompanied by sensory deficits and early loss of tendon reflexes. Facial weakness, when it occurs, may be so slowly progressive and symmetric that it escapes notice in facioscapulohumeral muscular dystrophy 1 (FSHD1), type 1 myotonic dystrophy-1 (DM1), and centronuclear myopathy-1 (CNM1). The latter disorder caused by heterozygous mutation in the gene encoding dynamin-2 (DNM2), is often associated with ptosis and limitation of eye movement, and slowly progressive limb weakness and wasting that remain unobvious until well into the third decade of life.

Peripheral nervous system The genetic neuromuscular disorders comprise the inherited myopathies, congenital myasthenia gravis and familial polyneuropathies. These disorders are easily separable by the history and neurological examination and family pedigree to elucidate the pattern of inheritance, supplemented by findings on nerve conduction studies (NCS) and concentric needle electromyography (EMG) and other relevant laboratory studies including sampling of blood, epidermis, muscle and nerve tissue, and cerebrospinal fluid (CSF) for routine and detailed serologic and molecular neurogenetic studies. Muscle and nerve biopsies should only be performed in institutions that are particularly equipped to process the tissue and have the expertise to interpret the results. Three sets of procedures are required for the microscopic analysis of skeletal muscle chosen from a clinically involved limb such as the biceps brachialis, vastus lateralis or medial gastrocnemius depending upon the distribution of weakness. Fresh tissue is placed into 10% buffered formalin and 2.5% buffered glutaraldehyde, and a third piece is snap frozen in isopentane cooled by liquid nitrogen. Paraffin and cryostat sections of snap-frozen muscle cut in cross-section and the longitudinal plane are stained with standard hematoxylin and eosin (H&E) and with the modified Gomori trichrome stain as the initial survey sections. This is followed by a battery of enzyme histochemical reactions including ATP reactions at acid pH 4.3 and 4.6, and alkaline pH 9.4, to evaluate the proportion of type 1 and type 2 fibers and subtypes; as well as, their shape, size and distribution. Specific histochemical reactions are performed to assess lipid content using Oil Red O and Sudan Black stains; while glycogen content is assessed by reactions with the Periodic acidSchiff (PAS) stains with and without diastase; in muscle fibers, as well as mitochondrial and oxidative enzyme activities employing nicotinamide adenine dinucleotidetetrazolium (NADH-TR), SDH, and COX reactions. Frozen sections are used for the immunohistochemical localization of membrane-associated proteins using a standard panel that includes dystrophin, as well as

185

dystroglycan, sarcoglycan and merosin reactivity, and any other pertinent genetic analyses. While the presence of dystrophic muscle pathology prompts the performance of biochemical and molecular genetic analysis on an involved site of muscle biopsy tissue for definitive evidence of sarcoglycanopathy, caveolinopathy, calpainopathy, dysferlinopathy, or a-dystroglycanopathy (Bushby, 2009), the evaluation of suspected familial polyneuropathy that defies a simple diagnosis may necessitate open biopsy of the sural or superficial fibular sensory cutaneous nerves and underlying muscle tissue. The resection of a segment of nerve 3–5 cm in length is divided into three portions and placed into separate mediums: 10% buffered formalin, 2.5% buffered glutaraldehyde, fresh tissue that is snap frozen in isopentane cooled by liquid nitrogen, respectively for paraffin and frozen cryosections stained by hematoxylin and eosin (H&E) and basic histochemical analysis for myelin, axons, connective tissues and immunofluorescence (IF); 1 mm semithin sections (STS); electron microscopy (EM), teased nerve fiber analysis, and biochemical or genetic studies. Many investigators of peripheral neuropathy have found skin biopsy for the analysis of epidermal nerve fibers (ENFs) to be a useful and predictive tool, particularly in patients who are not candidates for open nerve biopsy. In this analysis, a 3-mm punch biopsy of skin taken from the lateral thigh and calf of one leg and placed in paraformaldehyde-lysine-periodic acid fixative for a density measure of epidermal nerve fibers with comparison to age-matched controls (McArthur et al., 1998; Lauria et al., 2010), to which can be added sweat gland density, qualitative aspects of ENF histology and Congo red IF to detect amyloid deposits. The genetic analysis of the congenital myasthenic syndromes (CMS) and related ion channel abnormalities has been achieved with sophistical molecular genetics to detect mutations in the acetylcholine receptor (AChR) subunit genes, accompanied by sophisticated morphologic and electrophysiological studies of the neuromuscular junction (NMJ) (Engel, 2018b). Clinical clues of a possible CMS include the history of weakness and fatigue ability in a neonate, infant or young child with progression into adolescence or adulthood; lack of a significant response to acetylcholinesterase (AChE) drugs, absent AChR antibodies and inconsistent findings on repetitive nerve stimulation (RNS) at 2 Hz. Symptomatic CMS results from one or more specific etiopathogenic mechanisms residing at the presynaptic nerve terminal, synaptic basal lamina, or the postsynaptic region. The consequence is a loss of the safety factor for NMJ transmission defined as the difference between the postsynaptic depolarization caused by the end plate potential (EPP) and the depolarization required to active the postsynaptic voltage-gated Nav1.4 channel to trigger

186

D.S. YOUNGER

a muscle fiber action potential (AP). Targeted mutational analysis by Sanger or exome sequencing methods can be used to test for known disease-causing genes of which approximately 10% are presynaptic, 15% synaptic, and 75% postsynaptic, the majority of which are caused by AChR deficiency. A meaningful investigation of the CMS requires sophisticated in vitro electrophysiologic studies usually available at only a few centers to assess the quantal release of ACh and morphology of the NMJ such as motor point biopsy with cytochemical localization of AChE and immune deposits at end plates, followed by EM and immunocytochemical studies to determine the size and density of synaptic vesicles and the morphology of nerve terminals and postsynaptic region. Quantitative assessment of AChR binding is performed using peroxidase-labeled a-bungarotoxin whereas in vitro microelectrode studies including noise analysis and patch-clamp recordings provide additional information about the kinetic properties of abnormal AChR channels.

Central nervous system Amyotrophic lateral sclerosis (ALS), the commonest adult-onset motor neuron disease (MND), results from motor neuron degeneration in the motor cortex, brainstem and spinal cord, and culminates in death typically from respiratory failure within 3–5 years. It is defined as familial when there are more than one first- or second-degree relatives in a given pedigree. From a clinical perspective, there are few significant differences between typical sporadic (SALS) and familial (FALS) forms other than a slightly earlier age of onset in FALS as compared to SALS. Rarely, and devastatingly, some patients with FALS show onset in early teen years. Clinical, pathological, and genetic studies of FALS families suggest three characteristic phenotypes, all apparently of autosomal dominant (AD) transmission (Horton et al., 1976). The first is a rapid decline of 10–20 years. Meaningful clues in a given patient can be gleaned from careful inspection of the involved limbs, keeping in mind the basic clinical construct divergent ends of the spectrum of a single heterogeneous disease manifesting a combination of lower (LMN) and upper motor neuron (UMN) weakness, wasting and fasciculation; and hyperreflexia, Hoffman, Babinski signs and clonus (Younger et al., 1991). Some cases of ALS with frontotemporal dementia (FTD) (FTDALS) occur with

other multisystem manifestations, including pathology in muscle and bone. Rarely, “ALS-plus” syndromes are encountered in which ALS patients also develop ANS involvement.

HEREDITARY CEREBELLAR ATAXIAS Two groups of hereditary cerebellar ataxia (HCA), autosomal recessive (AR) and AD (ADCA) are differentiated by age at onset, mode of inheritance, and associated neurologic and nonneurologic features. ADCA is generally referred to as “spinocerebellar ataxia,” (SCA) even though “spinocerebellar” is a hybrid term, referring to both clinical signs and neuroanatomical regions (Margolis, 2003). Neuropathologists define SCA as cerebellar ataxia with variable involvement of the brainstem and spinal cord, and the clinical features of the disorders are caused by degeneration of the cerebellum and its afferent and efferent connections, which involve the brainstem and spinal cord (Schols et al., 2004; Taroni and DiDonato, 2004) and collectively lead to impaired motor coordination. One common diseasecausing mechanism of ADCA is an expanded (CAG)n trinucleotide repeat in the respective disease genes. However, the repeat is not only in genomic DNA but also in the messenger (m)RNA transcript as in the case of ataxin-1 similar to other repeat expansion diseases (REDs) including fragile X syndrome (FXS), DM1, Kennedy spinobulbar muscular atrophy (SBMA), and Huntington disease (HD). The encoded mutant ataxin proteins have abnormally long polyglutamine stretches which lead to SCA pathology by a toxic gain of function (GOF). Protein aggregates are also hallmarks of neurodegeneration in the brains of patients with SCA, but a direct link between these aggregates and neuronal death is not well understood. There is evidence that perturbed gene transcription contributes to neurodegeneration. Recessive ataxias are often multisystemic with inactivating mutations resulting in a loss of protein function, and a mutated protein that compromises the regulation of energy output, oxidative stress, DNA maintenance and the cell cycle. This is exemplified by two relatively common AR HCAs, Friedreich ataxia (FRDA), due to point mutation in the frataxin (FXN) gene or a GAA trinucleotide repeat expansion that perturb mitochondrial iron metabolism and the cellular response to oxidative stress; and ataxia telangiectasia (AT) due to a mutation in the ATM serine/threonine kinasegene (ATM) gene encoding a protein that coordinates the cellular response to DNA damage. Clues to the origin of an HCA will often be found in a three-generation family pedigree to establish the inheritance pattern, while empiric testing for trinucleotide repeat expansion in ADCA is informative in up to 50% of cases. When the pedigree reveals affected siblings only, an AR pattern is suggested, and FRDA,

NEUROGENETIC MOTOR DISORDERS AT, and vitamin E deficiency, should all be considered and confirmed by biochemical and molecular genetic analysis. Prenatal testing in HCA for known diseasecausing alleles is available using fetal DNA cells extracted by chorionic villus sampling at 10–12 weeks gestation, and amniocentesis performed at 15–18. By comparison the hereditary spinal paraplegias (HSPs) are genetically determined neurodegenerative disorders suggested by a positive family history of progressive weakness and spasticity of the legs. However, HSP remains a diagnosis of exclusion in sporadic cases. They are among the most clinically and genetically heterogeneous human diseases. More than 100 loci/88 spastic paraplegia genes (SPG) and new patterns of inheritance combine with wide ranging clinical phenotypes and all modes of monogenic AD, AR, recessive and dominant X-linked (XL), and mtDNA transmission. Autosomal recessive HSP is usually associated with diverse additional features referred to as complicated (c)HSP compared to mostly pure (p)HSP. The postmortem findings of pHSP and cHSP include axonal degeneration of crossed and uncrossed CST, with variable similar involvement of other long tracts such as the dorsal columns and spinocerebellar tracts. The identification of additional mutations and affected families has enlarged the clinical spectra of HSP, and complicated forms have been described for primary pHSP subtypes adding further complexity to genotype–phenotype correlations. In addition, the introduction of next generation sequencing in has revealed a genetic and phenotypic overlap with other neurodegenerative disorders such as ALS, neuropathies and SCA and neurodevelopmental disorders, including some with intellectual disability. With the advent of next-generation sequencing (NGS)–based techniques, the genetic basis of HSP appears more complicated than previously thought. HSP forms with mixed inheritance (AD/AR) modes have been observed, in addition to subtle mixed inheritance patterns with a dominating mode of inheritance and allele-dose-dependent variability in the expressed clinical phenotype. This is the case in the kinesin encoding gene (KIF1C) underlying HSP type 58 (SPG58) that presents a mild or subclinical dominant phenotype in heterozygous carriers, however the homozygous mutation results in a more severe recessive phenotypes in their homozygous states (Caballero Oteyza et al., 2014).

GENETIC NEUROLOGICAL DISORDERS Muscular dystrophy In the premolecular era, there were very few clues that different clinical and genetic forms of muscular dystrophy shared a common pathogenesis of membrane instability. Since the cloning of the gene for Duchenne muscular dystrophy (DMD) in late 1987 (Koenig et al.,

187

1987), concepts have dramatically changed along with identification of novel skeletal muscle genes including those encoding the extracellular matrix, sarcolemmal, cytoskeletal, cytosolic, and nuclear membrane proteins (Cohn and Campbell, 2000). These advances have translated into improved neurogenetic diagnosis and treatment approaches. A large number of genes encode components of the dystrophin-glycoprotein complex (DGC) that link the intracellular cytoskeleton to the extracellular matrix, mutations of which lead to a loss of sarcolemma integrity and render myofiber susceptible to injury. Prototypical examples of muscular dystrophies include those associated with nuclear (Emery–Driefuss muscular dystrophy [EDMD], laminopathy), cytosolic (calpainopathy, dystroglycanopathy, Fukyama congenital muscular dystrophy (CMD) [FCMD]), cytoskeleton (dystrophinopathy, telethoninopathy), sarcolemma (calveolinoapthy, dysferlinopathy), extracellular matrix (dystroglycanopthy a-2 laminin and merosin CMD [LAMA2]), and the sarcoglycan–sarcospan complex (sarcoglycanopathies). The associated underlying genetic defect results in weakness or altered muscle function at rest or with voluntary contraction, that are clinically separable by the distribution, inheritance pattern in a family pedigree, genetic studies, and distinctive findings on serum creatine kinase (CK), EMG and NCS, and muscle biopsy. Frank enlargement of the calves in toe walking boys and young men with a positive Gower’s sign and greater than a 10-fold elevation in serum CK is likely due to deletion or duplication in the DMD gene encoding dystrophin on chromosome Xp21 (Kunkel et al., 1986), but can be phenotypically indistinguishable in still ambulatory children with limb-girdle muscular dystrophy-1 (LGMDR1) (Angelini, 2020) due to homozygous or compound heterozygous mutation in the calpain-3 (CAPN3) gene (Chae et al., 2001). The genetic diagnosis of DMD can be ascertained by NGS in >90% of cases (Okubo et al., 2016). Both homozygous AR Becker disease and heterozygous AD cases of Thomsen disease (Lossin and George, 2008) manifesting myotonia derive from mutations in the chloride channel 1 (CLCN1) gene (Colding-Jorgensen, 2005) regulating the electric excitability of the skeletal muscle membrane. Affected individuals with myotonia congenita show stiff skeletal musculature that fail to relax after voluntary contraction contrasting with the distinctive presentation of DM1 (Johnson, 2019) that causes progressive weakness and myotonia associated with cataracts, hypogonadism, frontal balding, and electrocardiography (ECG). There is a corresponding amplified CTG trinucleotide repeat in the 3-prime untranslated region of the dystrophia myotonica protein kinase (DMPK) gene (Fu et al., 1993). Both AD myotonic dystrophy-2 (DM2) and nondystrophy proximal myotonic myopathy (PROMM) manifest nonprogressive

188

D.S. YOUNGER

weakness, myotonia, myalgia, stiffness, male hypogonadism, cardiac arrhythmias, diabetes, and early cataracts due to heterozygous expansion of a CCTG repeat in intron 1 of the zinc finger protein-9 (ZNF9) gene (Liquori et al., 2001).

Congenital myasthenic syndrome Engel (2018a) classified the CMS according to the anatomic site of the diseased protein as presynaptic, synaptic space, and postsynaptic, and due to defects in endplate development and maintenance, respectively accounting for 5.6%, 12.85%, 51.4%, and 25.7% of cases with known genetic defects (95.5%), and the remainder due to defects in glycosylation or other mechanisms. Given the heterogeneity of causes and disease presentations, it is obviously important to recognize appropriate cases and to screen the genome most appropriately for the likeliest disease-causing mutation. The CMS phenotypes are enumerated CMS 1–24 (Online Mendelian Inheritance in Man, OMIM®. McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University (Baltimore, MD) (2021), World Wide Web URL: https://omim.org/).

PRESYNAPTIC DEFECTS A presynaptic defect leading to choline acetyltransferase (CHAT) deficiency due to homozygous or compound heterozygous mutation in the CHAT gene is the commonest cause of presynaptic CMS. Affected patients with CHAT mutations (CMS6) experience abrupt episodic crises with increased weakness, bulbar paralysis, and apnea precipitated by undue exertion, fever, or excitement from birth or early infancy with negative tests for anti-AChR antibodies and an affected sib (Ohno et al., 2001). NMJ studies show a stimulation-dependent decrease of the amplitude of the miniature EPPs (MEPP) without a deficiency of the AChR. RNS can be negative while there can be a decrement of successive compound muscle action potentials (CMAPs) with exercise. Direct sequencing of CHAT demonstrated 10 AR mutations in 5 affected patients, including a frameshifting null mutation, 3 missense, and others that markedly reduced CHAT expression, or affected catalytic activity, or impaired catalytic efficiency. Treatment with AChE inhibitors may be beneficial, however, the most severely ill patients may remain respirator dependent and unable to move since birth (Engel et al., 2015).

SYNAPTIC SPACE DEFECTS In skeletal muscle, AChE exists in homomeric globular forms of type T catalytic subunits (ACHET) encoded by ACHET and heteromeric asymmetric forms composed of 1, 2, or 3 tetrameric ACHET attached to a collagenic

tail (ColQ) encoded by COLQ (Ohno et al., 1998). Asymmetric AChE is concentrated at the EP, where its collagenic tail anchors it into the basal lamina. Synaptic CMS5 is caused by mutations in ColQ of the endplate species of AChE that prevent the tail subunit from associating with catalytic subunits or from becoming inserted into the synaptic basal lamina (Engel et al., 2003). The synaptic space defect leading to endplate AChE deficiency is caused by homozygous or compound heterozygous mutation in the COLQ gene on chromosome 3p25. Clinically apparent cases show onset of generalized weakness soon after birth that increases with exertion and fatigability. RNS shows a decremental response to stimulation and MEPPs show prolonged duration and decreased frequency. Muscle EM shows decreased nerve terminal size, with reduced postsynaptic membrane density, and focal degeneration of the postsynaptic folds. Histochemical analysis does not detect EP AChE. Absence of AChE from the synaptic space prolongs the EPP leading to a second (or repetitive) muscle AP. Cholinergic overactivity injures the postsynaptic region causing degeneration of the junctional folds with loss of AChR. The nerve terminals are abnormally small which reduce the synaptic vesicles available for release which may protect the postsynaptic region from overexposure to ACh. Neuromuscular transmission is compromised by decreased quantal release, degeneration of junctional folds, and desensitization of AChR by overexposure to ACh. This disease can be improved by ephedrine or albuterol.

POSTSYNAPTIC DEFECTS Primary kinetic defects of the AChR referred to a slow (CMS1A)- and fast-channel (CMS1B) of the postsynaptic NMJ are characterized by early-onset of progressive muscle weakness are respectively due to mutations in the cholinergic receptor, nicotinic, alpha polypeptide 1 (CHRNA1) gene on chromosome 2q31. This gene encodes the alpha subunit of the muscle AChR, which is the main target of pathogenic autoantibodies in autoimmune myasthenia gravis (MG), and also controls electrical signaling between nerve and muscle cells by opening and closing a gate, membrane-spanning pore (Giraud et al., 2007). The slow-channel disorder results from kinetic abnormalities of the AChR channel, specifically prolonged opening and activity of the channel, which causes prolonged synaptic currents resulting in a depolarization block. This is associated with calcium overload, which may contribute to subsequent degeneration of the EP and postsynaptic membrane. Treatment with quinine, quinidine, or fluoxetine may be helpful in those with slow-channel defects. Slow-channel CMS (CMS2A) is caused by heterozygous mutation in the cholinergic receptor, nicotinic, beta polypeptide 1 (CHRNB1) gene

NEUROGENETIC MOTOR DISORDERS on chromosome 17p13 (Engel et al., 1996). Compound heterozygous mutations in the CHRNA gene (Wang et al., 1999) lead to fast-channel disorder which leads to abnormally brief opening and activity of the channel, with a rapid decay in EP current and a failure to reach the threshold for depolarization can be treated with pyridostigmine but not quinine, quinidine, or fluoxetine. AChR deficiency (CMS2A) leading to severe postsynaptic CMS with marked EP AChR deficiency is caused by two heteroallelic mutations in CHRNB1 (Quiram et al., 1999). One mutation causes skipping of exon 8, truncating the beta subunit before its M1 transmembrane domain, and abolishing surface expression of pentameric AChR. The other mutation, a 3-codon deletion (beta426delEQE) in the long cytoplasmic loop between the M3 and M4 domains, curtails but does not abolish expression. Affected patients are clinically characterized by early-onset muscle weakness typically from birth with variable severity often requiring frequent ventilation and enteric alimentation through a gastrostomy. There is a decremental response to RNS, however AChR antibodies are negative; and electrophysiologic studies show a decrease in MEPPs with normal ACh quantal release (Engel et al., 2015). Analysis of affected muscle fibers show an increased number of small endplate regions distributed over a 3-fold increased span of the muscle fiber surface. Nerve terminal size and the postsynaptic area of folds and clefts are both decreased compared to normal. Treatment with cholinesterase inhibitors may be helpful.

ENDPLATE DEVELOPMENT AND MAINTENANCE DEFECTS CMS11 associated with AChR deficiency is caused by homozygous or compound heterozygous mutation in the receptor-associated protein of the synapse, 43-KD (RAPSN) gene that encodes rapsyn, a postsynaptic protein that connects and stabilizes AChRs at the EP. Dystroglycan colocalizes with AChR-rapsyn clusters on the cell surface in vitro however rapsyn can cluster dystroglycan and AChR independently suggesting that rapsyn is a molecular link connecting the AChR to the cytoskeleton-anchored DGC at the NMJ thereby stabilizing AChR clustering. Pathogenic missense mutations in the RAPSN gene disrupt RAPSN function through different intracellular mechanisms (Cossins et al., 2006). An N88K mutation interferes with AChR clustering, whereas the other mutations either truncates the protein or alters membrane attachment. In compound heterozygotes, the combination of disruptive mechanisms leads to a more severe phenotype (Dunne and Maselli, 2003). Transgenic mice with targeted disruption of the Rapsn gene and homozygous mutants are both born in

189

expected numbers and similar in appearance to normal littermates, but die within hours (Gautam et al., 1995). The mutant mice have difficulty breathing and are unable to support themselves on all fours or to lift their heads. Analysis of the neuromuscular junction shows no detectable AChR clusters along the length of muscle fibers, indicating that rapsn is essential for AChR aggregation at the developing NMJ. Isolated gene knockout studies in transgenic mice indicate involvement of agrin, muscle-specific kinase (MuSK), and eapsn in clustering AChR at the neuromuscular junction (Lin et al., 2001). One other disorder, (CMS10) that results from a postsynaptic defect affecting endplate maintenance of the NMJ is AR inherited due to homozygous or compound heterozygous mutation in the tyrosine kinase 7 (DOK7) gene. Affected patients present with a characteristic limb girdle pattern of muscle weakness, in which the muscles have small, simplified neuromuscular junctions but normal AChR and AChE function (Beeson et al., 2006). Such patients have weak facial muscles or ptosis sparing eye movements. Anticholinesterase medication either has no effect or makes the weakness worse, although a short-lived initial response is occasionally seen. A frameshift mutation in the DOK7 gene in patients with limb-girdle type cases alone or in combination with a nonsense or splice site mutation, or missense change of a conserved residue is associated with AR inheritance. Other investigators (Selcen et al., 2008) noted highly variable in clinical features with severe progressive disease, and complex mutations identifiable only in cloned complementary DNA. Dok-7 is essential for maintaining not only the size but also the structural integrity of the EP, and the profound structural alterations at the EPs likely contribute importantly to the reduced safety margin of neuromuscular transmission. In a mouse model of DOK7 myasthenia (Arimura et al., 2014), therapeutic administration of an adeno-associated virus (AAV) vector encoding human DOK7 results in enlargement of NMJs and substantial increases in muscle strength and life span. Three other CMS loci occur in 100 mg/dL (Filosto et al., 2003). Other clinical features include ptosis, cardiomyopathy, short stature, weakness of muscles of the face, pharynx, trunk, or extremities and progressive hearing loss (Tsang et al., 2018). Full-field ERG shows evidence of generalized retinal dysfunction, involving both rods and cones. The mtDNA deletions are rarely identified in blood so muscle biopsy is necessary to identify the molecular genetic defect that leads to RRF on modified Gomori trichrome stained tissue sections, as well as COX-deficient myofibers. Biochemical studies of respiratory chain enzymes in muscle extracts usually show decreased activities of mitochondrial respiratory chain (MRC) complexes (I, IV) containing mtDNA encoded subunits. KSS and PEO should be differentiated from other disorders associated with ophthalmoplegia including myasthenia gravis, oculopharyngeal muscular dystrophy, myotonic dystrophy, and AD and AR PEO with multiple deletions of mtDNA. Neuropathological changes include basal ganglia calcifications and spongy changes of the brain white matter. Typically, KSS patients are sporadic as mtDNA rearrangements seem to originate in oogenesis or early zygote formation. Long

NEUROGENETIC MOTOR DISORDERS PCR analysis that detects large-scale mt-DNA4977bp common deletions (nt8482–nt13460) have a high sensitivity and low specificity (Lee et al., 2018), but may be of limited use and contribute to misdiagnosis in older individuals who accumulate mtDNA deletions with age. As a result, Southern blot is the preferred method of molecular diagnosis. Myoclonus epilepsy and ragged red fibers MERRF is also a multisystemic syndrome characterized by myoclonus which is often the first symptom, followed by generalized epilepsy, ataxia, weakness, and dementia. Onset is usually in childhood after normal early development. Common findings include hearing loss, short stature, optic atrophy, peripheral neuropathy, exercise intolerance, lipomas, lactic acidosis, cardiomyopathy, and Wolff-Parkinson-White arrhythmia. Most MERRF patients have a history of affected maternally related family members, although not all have the full syndrome. The diagnosis is based upon the presence of the following four elements: myoclonus, generalized epilepsy, ataxia, and RRF in the muscle biopsy. Diagnostic evaluation includes measurement of pyruvate and lactate in the blood and CSF, which are commonly elevated at rest, and increase excessively after moderate activity. The CSF protein is typically elevated but rarely surpasses 100 mg/dL. Electroencephalography (EEG) shows generalized spike and wave discharges with background slowing, but focal epileptiform discharges can be seen. The EMG and NCS are consistent with myopathy but often show coexisting axonal neuropathy. Brain magnetic resonance imaging (MRI) shows atrophy and basal ganglia calcification. Muscle biopsy typically shows RRF with the modified Gomori trichrome stain and fibers that fail to stain with COX. Biochemical studies of respiratory chain enzymes in muscle extracts usually show decreased activities of respiratory chain complexes containing mtDNA encoded subunits, especially COX. An heteroplasmic point mutation m.8344A>G in the transfer RNA, mitochondrial, lysine (MTTK) gene is identified in 80% of MERRF cases (Yoneda et al., 1990). At least five other mtDNA point mutations cause MERRF including 3 additional MTTK point mutations (m.8356T>C, m.8363G>A, m.8361G>A) and 2 mutations in other tRNA genes (m.611G>A in MT-TF and m.15967G>A in MT-TP) (Silvestri et al., 1992; Ozawa et al., 1997; Rossmanith et al., 2003; Mancuso et al., 2004). There is no clear correlation between the genotype and clinical phenotype for individual patients. Prenatal testing is possible by extraction of DNA from fetal cells obtained at amniocentesis and chorionic villus biopsy sampling, however the specific mtDNA mutation in the mother must be identified before prenatal

195

diagnosis is performed. In addition, as a result of mitotic segregation, the mtDNA mutational load in amniocytes and chorionic villi may not correspond with that of other fetal or adult tissues, nor is prediction of phenotype, age of onset, severity, or rate of progression possible. Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes MELAS has the defining clinical features of stroke-like episodes typically before age 40, encephalopathy manifested as seizures and dementia, and mitochondrial dysfunction with lactic acidosis and RRF in muscle tissue. It is uncommon for more than one family member to have the full syndrome. In most pedigrees, there is only 1 MELAS patient with an oligosymptomatic or asymptomatic relative in the maternal lineage. In typical cases, there is normal early psychomotor development with short stature is common. Onset of symptoms occurs frequently between the ages of 2 and 10 years with seizures, recurrent headaches, anorexia, and vomiting. Exercise intolerance and proximal limb weakness can be an initial manifestation followed by generalized epilepsy. Seizures can be associated with stroke-like episodes of transient hemiparesis, cortical blindness, altered mental consciousness, migraine headache, and residual cumulative motor impairment in adolescence and young adulthood. Other symptoms can include myoclonus, ataxia, episodic coma, optic atrophy, cardiomyopathy, pigmentary retinopathy, diabetes mellitus, hirsutism, gastrointestinal dysmotility, nephropathy, fever, drop attached, impaired gait, Wolf Parkinson White (WPW) pre-excitation syndrome, cardiac conduction block, PEO, and sensorineural hearing loss. The typical range of death is from 10 to 35 years and is due to intercurrent infection and gastrointestinal obstruction, however some individuals live into their sixth decade. Laboratory testing should include determination of lactate and pyruvate in blood and CSF along with protein content, which rarely surpasses 100 mg/dL. During stroke-like episodes, brain MRI typically shows lesions with increased fluid-attenuated inversion recovery (FLAIR)/T2 signals that do not conform to vascular territories along with basal ganglia calcification. Pathophysiologically, they are most likely due to a regional disruption of the blood–brain barrier triggered by the underlying metabolic defect, epileptic activity, drugs, or other factors, that manifest clinically not only as an ischemic stroke, but with ataxia, visual impairment, vomiting, and psychiatric disturbances. Muscle biopsy demonstrates RRF with modified Gomori trichrome that also stain positively for COX, in contrast to other mtDNA related disorders such as KSS and MERRF in which RRF do not react with COX histochemical stain. There may be an overabundance of

196

D.S. YOUNGER

mitochondria n smooth muscle and endothelial cells of intramuscular blood vessels best revealed with SDH stain. Biochemical analysis of respiratory chain enzymes in muscle extracts shows multiple partial defects especially involving complex I and IV. There are no clear correlations between genotype and clinical phenotype for individual patients. There are at 30 causative mtDNA mutations, the commonest of which is the A-to-G transition at nucleotide 3243 in MTTL1 encoding tRNALeu (UUR) (King and Attardi, 1989), so noted in 80% of MELAS cases. Although the common MELAS mutation, m3243A>G was identified more than two decades ago, the molecular consequences of the mutation are still incompletely understood. Muscle cells with the m.3243A>G mutation grown in tissue culture demonstrate respiratory deficiency (Kobayashi et al., 1990).

Hereditary cerebellar ataxia AUTOSOMAL RECESSIVE CEREBELLAR ATAXIA There are eight etiopathogenetic mechanisms for the AR HCAs (Table 9.1) including those associated with (1) nuclear DNA repair, replication and genome stability, exemplified by aprataxin (APTX), ATM serine/threonine kinase (ATM), and senataxin (SETX) genetic defects, respectively in ataxia-oculomotor apraxia 1 (AOA1),

ataxia-telangiectasia (AT), and AOA2; (2) mitochondrial metabolism due to the FXN, SACSIN (SACS), and twinkle mtDNA helicase (TWNK) genes, respectively associated with FRDA, AR spastic ataxia of CharlevoixSaguenay (ARSACS), and infantile-onset spinocerebellar ataxia (IOSCA); (3) peroxisomal metabolism exemplified by peroxisome biogenesis factor 7 (PEX7) genetic involvement in peroxisome biogenesis disorder 9b (Refsum disorder, PBD9B); (4) lysosomal metabolism as exemplified by the sorting nexin 14 (SNX14) and tripeptidyl peptidase 1 (TPP1) genetic defects, respectively associated with type 20 AR spinocerebellar ataxia (SCAR20); (5) protein translation due to involvement of the ring finger protein 216 (RNF216), SIL1 nucleotide exchange factor (SIL1), STIP1 homologous and U boxcontaining protein 1 (STUB1), and ubiquitin-like modifier activating enzyme 5 (UBA5) genes, respectively associated with cerebellar ataxia and hypogonadotropic hypogonadism (CAHH), Marinesco–Sjogren syndrome (MSS), SCAR16 and SCAR24; (6) endosome and membrane vesicle trafficking in the run domain- and cysteinerich domain-containing beclin-1–interacting protein (RUBCN), synaptotagmin (SYT14), and Abelson helper integration site 1 (AH11) genetic defects resulting in SCAR16, SCAR11, and Joubert syndrome 3 (JBTS3); (7) lipid and lipoprotein cell membrane and intracellular signaling exemplified by glucosidase, beta, acid 2

Table 9.1 Recessive hereditary cerebellar ataxias Subtyped by organelle function and disrupted cellular or metabolic function Nuclear DNA repair, replication, genome stability: APTX (AOA1)a, ATM (AT), SETX (AOA2) Mitochondrial metabolism: FXN (FRDA), SACS (SACS), TWNK/C10orf2 (IOSCA) Peroxisomal Metabolism: PEX7 (PBD9B) Lysosomal metabolism: SNX14 (SCAR20), TPP1 (SCR7) Protein translation: RNF216 (CAHH), SIL1 (MSS), STUB1 (SCAR16) Endosome and membrane vesicle trafficking: RUBCN (SCAR15), SYT14 (SCAR11) Lipid and lipoprotein cell membrane and intracellular signaling: GBA2 (SPG42), PNPLA6 (SPG39) Signal transduction: ANO10 (SCAR10), ATCAY (ATCAY), GRID2 (SCAR18), GRM1 (SCAR13), PIK3R5 (AOA3), SPTBN2 (SCAR14), WWOX (SCAR12) a

Gene (SCA phenotype). Abbreviations: DNA, deoxyribonucleic acid; APTX, aprataxin; AOA, ataxia-oculomotor apraxia 1; ATM, ATM serine/threonine kinase; AT, Ataxia-telangiectasia; SETX, senataxin; FXN, frataxin; FRDA, Friedrich ataxia; SACS, SACSIN (gene); SACS, spastic ataxia, CharlevoixSaguenay type; TWNK/C10orf2, twinkle mtDNA helicase/chromosome 10 open reading frame 2; IOSCA, spinocerebellar ataxia, infantile-onset; PEX7; peroxisome biogenesis factor 7; PBD9B (Adult Refsum disease), peroxisome biogenesis disorder 9b; SNX14, sorting nexin 14; SCAR20, spinocerebellar ataxia, autosomal recessive 20; TPP1, tripeptidyl peptidase 1; SCAR7, spinocerebellar ataxia, autosomal recessive 7; RNF216, ring finger protein 216; CAHH, cerebellar ataxia and hypogonadotropic hypogonadism; SIL1, SIL1 nucleotide exchange factor; MSS, Marinesco– Sjogren syndrome; STUB1, STIP1 homologous and U box-containing protein 1; RUBCN, run domain- and cysteine-rich domain-containing beclin-1-interacting protein; SYT14, synaptotagmin 14; AH11, Abelson helper integration site 1; JBTS3, Joubert syndrome 3; GBA2, glucosidase, beta, acid 2; PNPLA6, patatin-like phospholipase domain-containing protein 6; ANO10, anoctamin 10; ATCAY, caytaxin; cerebellar ataxia, Cayman type; ATCAY, cerebellar ataxia, Cayman type; GRID2, glutamate receptor, ionotropic, delta 2; GRM1, glutamate receptor, metabotropic, 1; PIK3R5, phosphatidylinositol 3-kinase, regulatory subunit 5; AOA3, ataxia-oculomotor apraxia 3; SPTBN2, spectrin, beta, nonerythrocytic; WWOX, WW domain–containing oxidoreductase.

NEUROGENETIC MOTOR DISORDERS (GBA2), patatin-like phospholipase domain-containing protein 6 (PNPLA6), and tocopherol transfer protein, alpha (TTPA), respective resulting in SPG42, SPG39, and ataxia with vitamin 3 deficiency (AVED, vitamin E deficiency); and (8) cellular signal transduction as noted in defects in the anoctamin 10 (ANO10), cerebellar ataxia, Cayman type (ATCAY), glutamate receptor, ionotropic, delta 2 (GRID2), glutamate receptor, metabotropic, 1 (GRM1), phosphatidylinositol 3-kinase, regulatory subunit 5 (PIK3RS), spectrin beta, nonerythrocytic (SPTBN2), and WW domain–containing oxidoreductase (WWOX) genes, respectively in SCAR10, Cayman cerebellar ataxia, SCAR18, SCAR13, AOA3, SCAR14, and SCAR12. Nuclear DNA repair, replication, and genome stability Oculomotor apraxia is a disorder characterized by limitation of ocular movements on command dissociated from movements of pursuit. It occurs in a variety of syndromes, including Joubert syndrome, AOA1 and 2, and AT. Mutations in the APTX gene encoding aprataxin, cause AOA1. Aprataxin is a nuclear protein with a role in DNA repair; and APTX-defective cell lines are sensitive to agents that cause single-strand breaks and exhibit an increased incidence of inducted chromosomal aberrations. Neurological disorders associated with APTX mutation may be caused by gradual accumulation of unrepaired DNA strand breaks resulting from abortive DNA ligation events (Ahel et al., 2006). Ataxiaoculomotor apraxia-2, which is instead due to mutation in the SETX gene (Moreira et al., 2004) encoding senataxin-interacting proteins play a role in coordinating transcriptional events and DNA repair (Suraweera et al., 2009). The clinical features of AT include ataxia, telangiectasia, dystonia, tremor, myoclonus, cellular and humoral immune deficiencies, growth retardation, progeria, high serum a-fetoprotein, chromosomal instability, predisposition to lymphoreticular malignancy, and sensitivity to ionizing radiation. The AT gene encoding the ATM protein responds to DNA damage by phosphorylating key substrates involved in DNA repair and/or cell cycle control. The stable association of the ATM repair protein with chromatin is a likely critical step in triggering, amplifying and maintaining the DNA damage repair signal even in the absence of DNA lesions (Soutoglou and Misteli, 2008).

197

clinical diagnosis. The most common molecular abnormality in FRDA is a GAA trinucleotide repeat expansion the FXN gene encoding frataxin, which associates with mitochondrial membranes and crests. A yeast homolog of frataxin is involved in iron homeostasis and respiratory function, raising the intriguing possibility that abnormal iron mitochondrial metabolism in FRDA produces toxic free radicals (Campanella et al., 2009). The spastic ataxia unique to Charlevoix-Saguenay was identified more than two decades ago by Bouchard and coworkers (Bouchard et al., 1978); all ARSACS patients exhibit signs of spasticity in the legs with a tendency to fall especially at the onset of gait initiation from 12 to 18 months age when there is little sign of cerebellar dysfunction. The disorder is caused by pathogenic mutations in the SACS gene encoding the sacsin protein, which is most highly expressed in large neurons, including cerebellar Purkinje cells (PC) with cytoplasmic localization in the amitochondrial component in neuroblastoma cells, integrating ubiquitin-proteasome Hsp70 functions and protecting against polyglutamine-expanded ataxin-1. Previously normal infants age 1–2 years with onset of clumsiness, gait ataxia, athetosis, muscle hypotonia, and areflexia followed by ophthalmoplegia, hearing loss and later sensory neuropathy and acute status epilepticus characterizes the mtDNA depletion syndrome-7, IOSCA due to mutation of the TWNK/C10ORF2 gene encoding twinkle and twinky proteins that colocalizes with mtDNA in mitochondrial nucleoids. It interacts with both single-stranded (ss) DNA (ssDNA) and double-stranded (ds) DNA functioning as a helicase with both unwinding and annealing activities (Sen et al., 2012). Peroxisomal metabolism The disorder of inborn error of lipid metabolism described by Refsum (1949) classically presents with retinitis pigmentosa, peripheral neuropathy, cerebellar ataxia, and acellular CSF with increased protein levels. Adult Refsum disease is ascribed to pathogenic mutations in the phytanoyl-coA hydroxylase (PHYH) gene encoding phytanoyl-CoA hydroxylase, a peroxisomal protein that catalyzes the first step in the b-oxidation of phytanic acid while other mutations in the PEX7 gene, the receptor for type 2 peroxisomal targeting signal (PTS2) encoding peroxin-7, one of a set of peroxisomal assembly proteins, is required for import of matrix proteins into peroxisomes.

Mitochondrial metabolism

Lysosomal metabolism

McLeod (1971) regarded the triad of hypoactive knee and ankle deep tendon reflexes, progressive cerebellar dysfunction, and preadolescent onset as sufficient for

AR spinocerebellar ataxia-20 (SCAR20) is clinically characterized by severely delayed psychomotor development with poor or absent speech, wide-based or absent

198

D.S. YOUNGER

gait, coarse facies, and cerebellar atrophy due to homozygous mutation in the SNX14 gene. Loss of SNX14 results in increased accumulation of autophagic organelles and disruption of intracellular cholesterol homeostasis (Bryant et al., 2018). AR spinocerebellar ataxia-7 (SCAR7) characterized by onset of progressive gait difficulties, eye movement abnormalities, and dysarthria in the first or second decade of life (Dy et al., 2015) is caused by compound heterozygous mutation in the TPP1 gene; biallelic mutation in TPP1 can cause neuronal ceroid lipofuscinosis-2 (CLN2). Experimental mice with targeted homozygous disruption of the Tpp1 gene (Sleat et al., 2004) are viable and healthy at birth, but develop progressive neurologic deterioration around 7 weeks of age with features of tremor and ataxia. Postmortem examination shows extensive neuronal pathology with accumulation of autofluorescent cytoplasmic storage material within the lysosomal-endosomal compartment, as well as loss of cerebellar PCs, and widespread axonal degeneration. Protein translation Patients with CAHH are clinically characterized by progressive cognitive decline, dementia, ataxia, and chorea due to homozygous or compound heterozygous mutation in the RNF216 gene (Santens et al., 2015). Encoded RING finger proteins called TRIADS mediate complex protein and DNA interactions (van der Reijden et al., 1999). First described by Marinesco and colleagues in four Romanian twins (Marinesco et al., 1931) and later by Sjogren (Sjogren, 1950), MSS is an early onset cerebellar ataxia with congenital cataracts, mental retardation, delayed development and short stature. There are four disease-associated predicted loss-of-function (LOF) mutations in SIL1, which encodes a nucleotide exchange factor that interacts with the ATPase domain of heatshock protein 70 (HSP70) chaperone HSPA5 to enhance nucleotide exchange, disturbance of which leads to abnormal protein folding as the primary pathology in MSS. A murine model of MSS depicts affected cells with intracellular protein accumulations reminiscent of protein inclusions in both the endoplasmic reticulum (ER) and the nucleus; in addition, upregulation of the unfolded protein response, suggestive of ER stress, occurs in mutant PCs (Zhao et al., 2005). SCAR16 is caused by homozygous or compound heterozygous mutation in the STUB1 or C terminus of the HSC70-interacting protein (CHIP) gene, an E3 ubiquitin ligase/cochaperone that participates in protein quality control by targeting a broad range of misfolded chaperone protein substrates for proteasomal degradation (Min et al., 2008).

Endosome and membrane vesicle trafficking SCAR15, clinically characterized clinically by earlyonset ataxia, cognitive impairment, dysarthria, and developmental delay with variable seizures, nystagmus, and abnormal reflexes (Seidahmed et al., 2020) is due to homozygous mutation in the RUBCN gene encoding rubicon (Assoum et al., 2013), a negative regulator protein of autophagy and endocytic trafficking. SCAR11, manifested by ataxia and psychomotor disturbances, is caused by homozygous mutation in the SYT14 that specifically localizes to PCs of the cerebellum, and encodes synaptotagmin 14, associated with exocytosis of secretory and synaptic vesicles alteration of which disturbs membrane trafficking (Doi et al., 2011). Lipid and lipoprotein cell membrane and intracellular signaling SPG46, clinically manifesting slowly progressive spastic paraplegia with cerebellar signs, cognitive impairment, cataracts, and cerebral, cerebellar, and corpus callosum atrophy on brain imaging (Boukhris et al., 2010) is caused by homozygous or compound heterozygous mutation in the GBA2 gene that encodes a microsomal b-glucosidase enzyme that catalyzes the conversion of glucosylceramide to free glucose and ceramide, as well as the reverse reaction consisting in the transfer of glucose to different lipid substrates. As an enzyme of sphingolipid metabolism, it plays a role in a variety of cell signaling responses and in structural components of the plasma membrane (Martin et al., 2013). SPG39, clinically characterized by progressive spastic gait ataxia and paraplegia associated with distal upper and lower extremity wasting, is caused by homozygous or compound heterozygous mutation in the PNPLA6 gene mutation of which inhibit the catalytic activity PNPLA6 that provides precursors of the neurotransmitten Ach and catalyzes the deesterification of membrane phosphatidylcholine into fatty acids and glycerophosphocholine (Synofzik et al., 2014). Signal transduction SCAR10, clinically characterized by onset of gait and limb ataxia, dysarthria, and nystagmus in teenage years, is caused by homozygous or compound heterozygous mutation in the calcium-activated chloride channel anoctamin 10 (ANO10) gene (Vermeer et al., 2010) that encodes the protein ANO10 with highest expression in frontal, occipital lobe and cerebellum where it may be engaged in calcium activation of chloride channels for signal transduction (Hartzell et al., 2009). Cyaman type of cerebellar ataxia (ATCAY), which is clinically characterized by hypotonia from birth with

NEUROGENETIC MOTOR DISORDERS variable psychomotor retardation, cerebellar dysfunction, nystagmus, intention tremor, dysarthria, ataxic gait, and truncal ataxia, is caused by homozygous mutation in the caytaxin (ATCAY) gene that encodes the neuronrestricted protein caytaxin responsible for the recruitment of cholinergic machinery at neurite terminals to promote ACh signaling and neurite outgrowth (Sun et al., 2015). SCAR18, clinically characterized by delayed psychomotor development, severely impaired gait due to cerebellar ataxia, ocular movement abnormalities, and intellectual disability due to homozygous mutation in the GRID2 gene (Hills et al., 2013) encoding the glutamate receptor channel delta-2 subunit, selectively expressed in PCs (Takayama et al., 1995) where it plays important roles in motor coordination, formation of parallel fiber-Purkinje cell synapses and climbing fiber-PC synapses, and longterm depression of parallel fiber-PC synaptic transmission (Kashiwabuchi et al., 1995). SCAR13 is characterized by delayed psychomotor development beginning in infancy with affected individuals showing mildly to profoundly impaired intellectual development, poor or absent speech as well as gait and stance ataxia and hyperreflexia (Guergueltcheva et al., 2012) due to homozygous mutation in the GRM1 gene encoding a group 1 metabotropic glutamate receptor (Stephan et al., 1996) that experimentally protects neurons from apoptotic death (Maiese et al., 2000) and evokes slow excitatory postsynaptic conductance (Kim et al., 2003) mediated by a transient receptor potential (TRP) superfamily of cation channels (TRPC1) (Zhang et al., 2009) expressed in perisynaptic regions of the cerebellar parallel fiber-PC synapse. AOA3 is caused by homozygous mutation in the PIK3R5 gene that encodes the class 1 receptor-regulated phosphoinositide 3-kinase (PI3K) subunit PIK3R5 (Brock et al., 2003) which under the tight control of cell surface receptors, including the G protein-coupled receptor Gbg and the noncatalytic 101 subunit function as a downstream regulator of G protein-coupled receptors (GPCR)–dependent signal transduction pathways in the brain (Roldan-Sastre et al., 2021) potentially in the cerebellar cortex where it could contributes to the regulation of neurotransmission and neuronal excitability (Oldham and Hamm, 2008). SCAR14, clinically characterized by delayed psychomotor development, severe early-onset gait ataxia, eye movement abnormalities (Lise et al., 2012) is caused by homozygous mutation in the SPTBN2 gene that encodes the spectrin protein SPTBN2 expressed predominantly in cerebellar PC soma and dendrites where it presumably form flexible tetramers to stabilize cell contacts, channels, and adhesion molecules along the cytoplasmic face of membrane bilayers (Jackson et al., 2001).

199

SCAR12, clinically characterized by onset of generalized seizures in infancy, delayed psychomotor development with mental retardation, and cerebellar ataxia is caused by homozygous mutation in the WWOX gene where its expressin is comparatively higher in the human cerebellum notably in GABAergic basket cells and granule cells pointing to a direct link to cerebellar ataxia due to LOF (Aldaz and Hussain, 2020).

AUTOSOMAL DOMINANT CEREBELLAR ATAXIA Historically, Harding (1982) proposed a clinical classification for ADCAs categorizing ADCA I by cerebellar ataxia in combination with various associated neurologic features, such as ophthalmoplegia, pyramidal and extrapyramidal signs, peripheral neuropathy, and dementia, among others (SCA1–3), ADCA II characterized by the cerebellar ataxia, associated neurologic features, and the additional findings of macular and retinal degeneration (SCA7); and ADCA III with pure forms of lateonset cerebellar ataxia without additional features (SCA5, SCA6, SCA11, and SCA31). The known ADCAs have since grown with many additional subtypes enumerated SCA1–46, dentatorubralpallidoluysian atrophy (DRPLA) and episodic ataxias (EA) types 1–6. The etiopathogenetic mechanisms of ADCAs can be categorized into four groups as follows: (1) polyglutamine-encoding CAG REDs (SCA1–3, 6, 7, 17, and DRPLA (considered later in this chapter); (2) noncoding repeat expansion and RNA toxicity (SCA8, 10, 12, 31, and 36); (3) ion channel dysfunction (SCA5, 13, 15, 19/22, 27); and signal transduction (SCA11, 14, 23, and 40). Noncoding repeat expansion/RNA toxicity SCA8: Koob et al. (1999) reported eight pedigrees with slowly progressive, severely affected members with nystagmus, spastic and ataxic dysarthria, incoordination, gait ataxia, spasticity, and vibratory sensory loss with onset from age 18 to 65 year, with a mean age at onset of 40 years, with most nonambulatory by the fifth decade. Polyglutamine expansions in IC2-immunoreactive intranuclear inclusions from SCA8 is found in human brain tissue encoded by an expanded CAG repeat in ataxin 8 (ATXN8) gene complimentary to the expanded CTG repeat of the ataxin 8 opposite strand (ATNX8OS) on the opposite strand indicating bidirectional transcription with expression of both a polyglutamine protein and a CUG expansion transcript (Moseley et al., 2006). This molecular defect is referred to as a “CTG*CAG” repeat expansion, referring to the complementary basepairs of the ATXN8OS and ATXN8 genes, reading 5-prime to 3-prime (Ikeda et al., 2008). Humans and mice with SCA8, ATXN8OS mRNA containing the expanded repeat

200

D.S. YOUNGER

accumulate as ribonuclear inclusions, or RNA foci, that colocalize with the RNA-binding protein in cerebellar cortical neurons (Daughters et al., 2009). SCA10: Grewal et al. (1998) described fourthgeneration pedigree that segregated a distinct form of SCA characterized by cerebellar ataxia and seizures; anticipation was observed in available parent child pairs suggesting that a dinucleotide repeat expansion could be the mutagenic mechanism. SCA10 varies clinically from pure progressive cerebellar ataxia to ataxia associated with seizures, polyneuropathy, pyramidal signs, and cognitive and neuropsychiatric impairment caused by a heterozygous expanded 5-bp repeat (ATTCT) in the ATXN10 gene. Normal alleles have 10–29 repeats while pathologic alleles have 400–4500 repeats; however, rare cases can have as few as 280 repeats (Alonso et al., 2006). SCA12: This disorder presents from age 8 to 55 years with upper extremity tremor, progressing over several decades to include head tremor, gait ataxia, dysmetria, dysdiadokinesis, hyperreflexia, paucity of movement, abnormal eye movements, and, in the oldest subjects, dementia (Holmes et al., 1999). The disorder is due to expansion of a CAG repeat in the protein phosphatase 2, regulatory subunit B, beta (PPP2R2B) gene encoding a brain-specific regulatory subunit B of protein phosphatase 2 (PP2A) which has been implicated in a number of cellular function including modulation of cell cycle progression, tau phosphorylation, and apoptosis. SCA31: This disorder is considered one of a growing number of neuromuscular disease with RNA-mediated gain-of-function (GOF) mechanism caused by a 2.5–3.8-kb insertion containing pentanucleotide repeats including (TGGAA)n with an intron at the 16q21 locus of the BEAN gene. Ishikawa and colleagues (Ishikawa et al., 2011) showed an association of SCA31 with diverse pentanucleotide repeats, that included the prototypical (TAAAA)n; (TGGAA)n and (TGAAA)n in Japanese; whereas (TACAA)n, (GAAA)n, (TAACA)n, and (TGAAA)n exclusively in Caucasians, each potentially arising from a single-nucleotide mutation in TAAAA before their expansion, suggesting founder effect. Neuropathological study of an involved kindred (Owada et al., 2005) demonstrated shrinkage of Purkinje cell bodies surrounded by synaptophysin-immunoreactive amorphous material containing calbindin- and ubiquitinpositive granules, indicating that the amorphous material was formed in association both with the degeneration of PC processes and the increase of presynaptic terminals innervated either from basket cells, inferior olivary neurons, or other neurons. Alteration of calbindinimmunoreactivity in PCs may indicate that the intracellular calcium buffering system, which is one of the important roles of calbindin, may be altered in this disorder (Owada et al., 2005).

SCA36: Ohta and colleagues (Ohta et al., 2007) reported two unrelated patients who showed ataxia as the first symptom later resembling ALS. Kobayashi and colleagues (Kobayashi et al., 2011) undertook a genome-wide linkage analysis and subsequent mapping of five unrelated Japanese families with onset of cerebellar truncal and gait ataxia, ataxic dysarthria, and limb incoordination at a mean age of 53 years, with signs of MND and longer disease duration exhibiting lingual atrophy, fasciculation, skeletal muscle atrophy and fasciculation, and hyperactive reflexes. Electrophysiology showed neurogenic features indicative of LMN involvement typically limited to the tongue, which differentiated it from typical ALS. Expansions of the hexanucleotide repeat GGCCTG (rs68063608) were found in intron 1 of the NOP56 ribonuclear protein (NOP56) gene in all five index cases through the use of a repeat-primer PCR method, and complete segregation of the expanded hexanucleotide was confirmed in all pedigrees. A founder haplotype was found in these cases, and RNA foci formation was detected in lymphoblastoid cells from affected subjects by fluorescence in situ hybridization. Double staining and gel-shift assay showed that (GGCCUG)n bound protein SRSF2, and transcription of MIR1292, a neighboring miRNA, was significantly decreased in lymphoblastoid cells of SCA patients. The findings suggested that SCA36 is caused by hexanucleotide repeat expansions through RNA gain of function. SCA36 is caused by heterozygous expansion of an intronic GGCCTG hexanucleotide repeat at the 20p13 locus of the NOP56 gene. Unaffected individuals carry 3–8 repeats, whereas affected patients carry 1500–2000 repeats. The NOP56 gene functions in an early to middle step in prerRNA processing. Ion channel dysfunction SCA5: Ranum et al. (1994) described a kindred with slowly progressive cerebellar dysfunction, CST signs, and bulbar dysfunction. All four juvenile-onset patients of age 10–18 years resulted from maternal transmission suggesting maternal anticipation bias. Linkage analysis mapped the disease locus, designated SCA5, to the centromeric region of chromosome 11. The most consistent clinical feature is downbeat nystagmus, imbalance of stance and gait, dysarthria, intention and resting tremor, impaired smooth muscle pursuit, and gaze-evoked nystagmus. There is slow progression of symptoms yet all patients are ambulatory despite disease duration of up to 31 years (Burk et al., 2004). Ikeda et al. (2006) found a 39-bp deletion in exon 12 of the spectrin beta nonerythrocytic 2 (SPTBN2) gene that caused an in-frame 13 amino acid deletion within the third of 17 spectrin repeats in all 90 affected individuals and 35 presymptomatic carriers from a large 11 generation American

NEUROGENETIC MOTOR DISORDERS kindred, as well as, a short in-frame deletion in the same spectrin repeat of the SPTBN2 gene of a French family; and a T-to-C transition in exon 7 of the SPTBN2 gene of a German family, that caused a leucine-to-proline change in the calponin homology domain containing the actin/ ARP1-binding site. SCA5 is caused by pathogenic mutations at the 11q13.2 locus of SPTBN2 (Clarkson et al., 2010). SCA13: Durr, Herman-Bret, and colleagues (Herman-Bert et al., 2000) examined a large French family with ADCA, members of which displayed mental retardation, cerebellar dysarthria, moderate mental retardation in the IQ range of 62–76, mild developmental motor delays, as well as horizontal and vertical nystagmus, square wave jerks, and pyramidal signs, generalized bradykinesia, upward gaze palsy, swallowing difficulty, urinary urgency, short stature, slight facial dysmorphia, and petit mal epilepsy in occasional individuals. SCA13 is caused by heterozygous mutation in the potassium channel, voltage-gated, shaw-related subfamily, member 3 (KCNC3) gene. SCA15: Storey et al. (2001) described an Australian family in which 8 members had slowly progressive cerebellar ataxia with disabling action and postural tremor, and some CST and dorsal column involvement and gaze palsy. Van de Leemput and colleagues (2007) identified heterozygous deletions involving the inositol 1,4,5triphosphate receptor, type 1 (ITPR1) gene in affected members of three unrelated families including SCA15 families of Australian origin used to map the locus (Storey et al., 2001), as well as, a large deletion removing the first 10 exons of the ITPR1 gene in the family reported by Knight and colleagues (2003). The cause of SCA15 is due to heterozygous mutation and deletions at the 3p26.1 locus of the ITPR1 gene. Inositol 1,4,5triphosphate is an intracellular second messenger produced by phospholipase C through a G proteindependent mechanism that releases ER calcium by binding to specific receptors coupled to calcium channels that is abundant in neuronal tissues including the cerebellum, particularly the perikarya of PCs. SCA19/22: Schelhaas and colleagues (2001) studied 12 patients in a four-generation Dutch family with earlyonset of cerebellar ataxia, intentional and postural irregular low frequency tremors, myoclonus, and cognitive impairment, followed by dysarthria, limb ataxia, fine saccadic eye movements, and horizontal nystagmus. Anticipation as evidenced by earlier age of onset in successive generation was encountered, and the disease onset ranged from the first to fourth decade. SCA19 is caused by heterozygous mutation in the KCND3 gene on chromosome 1p13 encoding Kv4.3, an alpha subunit of the Shal family of A-type voltage-gated potassium channels important in membrane repolarization in excitable

201

cells (Lee et al., 2012). The disease locus mapped to chromosome 1p21-q23 and was designated SCA22 by other investigators (Chung et al., 2003). As the locus of SCA22 overlapped with that of SCA19 on 1p21-q21 (Schelhaas et al., 2001), the two disorders are allelic with a worldwide distribution. KCND3 encodes the voltage-gated potassium channel KV4.3 that is highly expressed in the cerebellum, where it regulates dendritic excitability and calcium influx. Loss-offunction KV4.3 mutations have been associated with SCA19/22 (Zanni et al., 2021). SCA27: A large third-generation Dutch family in which 14 members had trembling of the hands exacerbated by emotional stress and exercise beginning in childhood followed by slowly progressive cerebellar gait and upper limb ataxia beginning between the ages of 28 and 40 years (Wang et al., 2002). Neurological examination from age 24 to 79 years, showed dysmetric saccades, disrupted ocular pursuit movements, gaze-evoked nystagmus, cerebellar dysarthria, and small-amplitude high-frequency hand tremor, head tremor, orofacial dyskinesia, hyperreflexia and vibratory sensory loss, and variably impaired cognition, memory and cortical language disturbances. Linkage analysis and recombination analysis in affected individuals demonstrated a critical region on chromosome 13q34 that included the fibroblast growth factor-14 (FGF14) gene. Recognizing a murine model of ataxia and paroxysmal dyskinesia led to candidate-gene mutation analysis of FGF14 and the eventual elucidation of a pathogenic mutation in FGF14 leading to a T-to-C transition at position 434 of the FGF14a open reading frame (ORF) resulting in an amino acid substitution of a serine for a phenylalanine at position 145 (F145S). Dalski et al. (2005) reported ataxia in an 18-year-old male with mental retardation, inborn strabismus, redgreen vision defect, and normal motor development until age 12 when he developed slowly progressive gait disturbance, memory loss, and depressed mood. Examination showed truncal and gait ataxia, small-amplitude tremor in the hands, gaze-evoked nystagmus, and pes cavus. The patient’s father reportedly had gait disturbances, memory loss, and pes cavus. Genetic analysis revealed six different DNA variations, two of which resulted in amino acid level changes, including a single base pain deletion in exon 4 (c.487delA) creating a frameshift mutation, and DNA polymorphisms in exon 1a, 4, and 5, an amino-acid exchanged at position 124, as well as, single-nucleotide polymorphism in the 30 -untranslated region of exon 5 of the FGF14 gene. The AD FGF14 mutations were in contrast to the recessive Fgf14 knockout murine model of ataxia and paroxysmal dyskinesia. Misceo et al. (2009), who reported a daughter and mother with karyotype XXt (5;13) (q31.2;q33.1), and a

202

D.S. YOUNGER

translocation breakpoint identical in both patients disrupting the gene encoding the isoform FGF14-1b, showed clinical signs of SCA including early onset in the daughter, with gait ataxia, dysarthria, writing disability, dyskinesia, and titubation; and pes cavus and UMN involvement in both mother and daughter. The authors suggested that truncation of one allele could lead to FGF14 haploinsufficiency in turn, causing SCA27. SCA27 is caused by a mutation at the 13q33.1 locus in the FGF14 gene encoding FGF14 which appears to function in neuronal signaling, axonal trafficking, and synaptosomal function. Signal transduction SCA11: Worth et al. (1999) and Houlden et al. (2007) described multigenerational British families with relatively benign late-onset slowly progressive cerebellar ataxia, and characterized the causative gene for SCA11, tau tubulin kinase 2 (TTBK2), encoding the TTBK2 protein, a member of the casein kinase (CK1) group of protein kinases with an ability to phosphorylate tau and tubulin in vitro. However beyond tau phosphorylation, TTBK2 is involved in multiple other important cellular processes including regulating the growth of axonemal microtubules in ciliogenesis (Liao et al., 2015) and its assembly by controlling the final step of cilia initiation (Bernatik et al., 2020). SCA14: Initial evidence for the assignment of the protein kinase C, gamma (PKCG) gene derived from informative RFLP studies (Johnson et al., 1988) and fluorescence in situ hybridization (Trask et al., 1993). Yamashita et al. (2000) described a third-generation Japanese family with onset of cerebellar ataxia at age 27 years or less of cerebellar ataxia and intermittent axial myoclonus. Systematic linkage traced established a novel mutation and locus to a 10.2-cM interval on chromosome 19q13.4-qter, in agreement with the observations of Brkanac and colleagues (Brkanac et al., 2002) in a description of a four-generation family of English and Dutch descent of 14 members, 10 of whom available for analysis showed gait ataxia, dysarthria, dysmetria, abnormal eye movement, and onset in the third decade. Chen et al. (2003) identified 3 mutations in the protein kinase C, gamma (PKCG) gene, 2 of which cosegregated with the disorder. Yabe et al. (2003) identified a mutation in the PRKCG gene in all 11 affected members of the family reported by Yamashita et al. (2000) with SCA14, as did Klebe et al. (2007) who described six different mutations including 5 novel ones, in the PRKCG gene among 15 affected members of six French families. Stevanin et al. (2004) studied a family of French origin with 20 affected members with SCA14 in four generations. Age of onset ranged from childhood to 60 years,

with mild to moderate cerebellar signs, as well as dysphagia, facial myokymia, decreased vibratory sensation in the feet, chorea of the hands, head tremor, memory and attention deficits. The PRKCG gene encodes aprataxin (APTX), a DNA repair protein. Decreased nuclear APTX increases oxidative stress-induced DNA damage and cell death. SCA23: Verbeek and colleagues (2004) reported five members of a third-generation Dutch family with onset at age 43 to 56 years of gait or simultaneous cycling difficulties and speech disturbances in association with dysarthria, slow saccades, ocular dysmetria, decreased vibratory sensation, and hyperreflexia. Neuropathological examination of the brain in one affected subject showed generalized moderate to severe generalized atrophy, most pronounced in the frontotemporal region, cerebellar vermis, basis pontis, and spinal cord. Pronounced neuronal loss was evident in the Purkinje cell layer especially the vermis, dentate nuclei, and inferior olives accompanied by gliosis and myelin loss in surrounding white matter. There were ubiquitin-positive, IC2 and ataxin-3–negative intranuclear inclusions in substantial nigra neurons interpreted as Marinesco bodies. Bakalkin et al. (2010) identified missense mutations in the prodynorphin (PDYN) gene in the original reported SCA23 family (Verbeek et al., 2004) and in three families from a Dutch ataxia cohort. PDYN is the precursor protein for the opioid neuropeptides, and dynorphins A (Dyn A) and B (Dyn B). Alterations in Dyn A activity and impairment of secretory pathways by mutant PDYN leads to glutamate neurotoxicity which may underlie PC degeneration and ataxia. More recent insights suggest converging mechanisms of the loss of opioid-mediated neuroprotection and an N-methyl-D-aspartate (NMDA) nonopioid mechanism of Dyn A peptide-mediated excitotoxicity underlying the pathology of SCA23.

Hereditary spastic paraplegia Blackstone (2012) attributed the etiopathogenesis of HSP to several cellular processes: axon pathfinding, myelination, endoplasmic reticulum network morphology, lipid synthesis and metabolism, endosomal dynamics, motor-based transport, mitochondrial function, and related disorders (Table 9.2).

AXON PATHFINDING The first HSP mutations were described in the L1 cell adhesion molecule (L1CAM) gene, encoding, LICAM, a cell surface glycoprotein of the immunoglobulin (Ig) superfamily. LOF mutations in L1CAM are implicated in XL, early-onset, cHSP (SPG1), as well as in the XL disorder, MASA (for mental retardation, aphasia, shuffling gait, and adducted thumbs), with hydrocephalus,

NEUROGENETIC MOTOR DISORDERS

203

Table 9.2 Hereditary spastic paraplegias Subtyped by mechanism of disease and inheritance (encoded protein, gene symbol) Axon Pathfinding: XLR: SPG 1 (L1CAM, LICAM) Demyelination: XLR: SGP 2 (PLP1, PLP1); AR: SPG 35 (FA2H, FA2H), SGP 44 (Connexin 47, GJC2) ER network morphology: AD: SPG 3A (ATL1, ATL1), SPG 4 (Spastin, SPAST), SPG 12 (reticulon 2, RTN2), SPG31 (REEP1, REEP1); AR: SPG61 (ARL6IP1, ARL6IP1) Lipid synthesis and metabolism: AD: SPG42 (AT1, SLC33A1) Endosomal dynamics: AR: SPG 11 (spatacsin), SPG 15 (spastizin, ZFYVE26), SPG 48 (AP5, AP5Z1) BMP signaling: AD: SGP 6 (NIPA1, NIPA1) Motor transport: AD: SPG 10 (KIF5A, KIF5A) Mitochondrial function: AR: SPG 7 (paraplegin, SPG7) Abbreviations: XLR, X-linked recessive; SPG: SPG, spastic paraplegia type; L1CAM, L1 cell adhesion molecule; PLP1, proteolipid protein; GJC2, gap junction protein, gamma-2; ATL1, atlastin-1; ATL1, atlastin GTPase 1; SPAST, spastin; RTN2, reticulon; REEP1/REEP1, receptor expressionenhancing protein 1; ARL6IP1/, ARL6IP1, ADP-ribosylation factor-like GTPase6-interacting protein 1; AT1, acetyl-CoA transporter; SLC33A1, solute carrier family 33 (acetyl-CoA transporter), member 1; ZFYVE26, zinc finger FYVE domain-containing protein 26; AP5, adaptor-related protein complex 5; AP5Z1, adaptor-related protein complex 5, zeta-1 subunit; NIPA1, NIPA magnesium transporter 1; KIF5A, kinesin family member 5A.

and agenesis of the corpus callosum (Weller and Gartner, 2001). Both display clinical and pathological evidence of CST impairment and are considered in the disease spectrum known as L1 disease or CRASH syndrome (for corpus callosum hypoplasia, retardation, adducted thumbs, spastic paraplegia or shuffling gait, and hydrocephalus) (Soderblom and Blackstone, 2006). A compelling role of L1CAM in axon pathfinding during development is consistent with the early onset of SPG1.

DEMYELINATION Mutations in the PLP1 gene encoding the tetraspan integral membrane proteolipid protein (PLP) and its smaller DM20 isoform give rise to two major diseases along a clinical spectrum: a pure or complicated HSP (SPG2) and Pelizaeus–Merzbacher disease (PMD) (Inoue, 2005). Plp1 null mice have been widely studied as a model for SPG2. In these mice the myelin sheath maintains its normal thickness, although with subtle anomalies of the intraperiod lines. In the underlying axons, anterograde transport is impaired, and cargoes undergoing retrograde transport become stuck at distal juxtaparanodal regions (Edgar et al., 2004). Thus, it seems reasonable to postulate that oligodendrocytes modulate the activity of motor proteins involved in intracellular cargo transport via signaling cascades in the underlying axon and that this modulation is sensitive to PLP/DM20 (Gruenenfelder et al., 2011). Mutations in a more recently identified HSP gene similarly define a disease spectrum comprising HSP and PMD-like disease where cell–cell communication is altered. Slowly progressive, complicated SPG44 is caused by homozygous mutations in the gap junction protein, gamma-2

(GJC2) gene encoding connexin 47 (CX47). As X47 forms connections between astrocytes and oligodendrocytes in concert with CX43. Because CX47/CX43 heterotypic channels appear essential for the maintenance of CNS myelin, alterations in CX47 that result in CX47/CX43 channel dysfunction likely underlie SPG44 (Orthmann-Murphy et al., 2009). A third HSP with a link to dysmyelination is AR SPG35, which spans neurodegeneration and brain iron accumulation, leukodystrophy, and HSP and results from LOF mutations in the fatty acid-2 hydroxylase (FA2H) gene (Dick et al., 2010).

ENDOPLASMIC RETICULUM NETWORK MORPHOLOGY Several HSP genes and encoded proteins are involved in intracellular trafficking, distribution, biogenesis, and shaping of membrane compartments of microtubules and tubular ER of CST motor neurons including: atlastin-1 encoded by ATL1 in SPG3A; spastin encoded by SPAST in SPG4; and reticulon 1 (RTN1) and RTN2 encoded respectively by Reep1 and Reep2 in SPG31 and SPG12; and ADP-ribosylation factor-like GTPase 6-interacting protein 1 (ARL6IP1) encoded by ARL6IP1 in SPG61. SPG4 is the most common form of AD SPG, comprising up to 45% of cases. A role for spastin in microtubule dynamics of the cytoskeleton is suggested by constitutive binding in transinfected cells mediated through ATPase activity of the AAA domain with the formation of thick perinuclear bundles and microtubule disassembly. Furthermore, cells lacking spastin have increased tubulation of endosomal tubular recycling compartments, with resulting defects in receptor sorting. These functions

204

D.S. YOUNGER

may be particularly relevant, since loss of spastin decreases axon branching in cultured neurons, and this process also involves coordinated microtubule regulation and membrane modeling (Riano et al., 2009). Although partial LOF in microtubule severing from loss of spastin function is widely postulated, neurotoxicity of mutant spastin proteins, particularly the M1 spastin isoform that binds other common HSP proteins within the tubular ER, such as atlastin-1 and REEP1 is another possible mechanism of injury to the CST (Solowska and Baas, 2015). Dynamin-related GTPase atlastin-1 plays a role in formation of the tubular ER) network and in axon elongation in neurons (Orso et al., 2009). Knockdown of atlastin-1 using shRNA in cultured cortical cells inhibited axonal growth. Overall, the findings suggested that atlastin-1 has diverse functions in neurons, likely acting both in intracellular membrane trafficking as well as in expansion at the axonal growth cone. These functional studies suggested that the early-onset axonopathy observed in SPG3A may result from abnormal development of axons (Zhu et al., 2006). SPG31 is the third most common HSP; and its encoded protein REEP1, along with REEP2 (Esteves et al., 2014) and ARL6IP1 (SPG61) (Pettersson et al., 2000) participate in the ER tubular network forming paired hairpin domains that insert into the tubular ER membranes, generating high curvature through hydrophobic wedging and scaffolding (Voeltz et al., 2006). The interactions among REEPs, atlastin, and spastin via hydrophobic hairpins provide an attractive mechanism for coupling ER membrane remodeling to cytoskeletal dynamics.

LIPID SYNTHESIS AND METABOLISM A key function of the tubular ER is the synthesis, metabolism, and distribution of lipids and sterols, and their vesicular and nonvesicular storage in lipid droplet (LD) organelles. Overexpression of atlastin-1 and possibly REEP1 and spastin may be related to a LOF mutations in determining the number and size of LDs contributing to the pathogenesis of SPG3A; while missense mutations of SLC33A1 encoding the acetyl-CoA transporter interrupt the outgrowth and maintenance of long motor axons in human heterozygotes (Lin et al., 2008) in AD SPG42.

ENDOSOMAL DYNAMICS An emerging HSP-related complex relates to endocytic trafficking in SPG15, SPG11, and the SPG48. Mutations in the SPG11 gene encoding spatacsin (Perez-Branguli et al., 2014) and SPG15 encoding zinc finger FYVE domain-containing protein 26 (ZFYVE26) in SPG15

(Hanein et al., 2008), both essential components for initiation of lysosomal tabulation, lead to truncated deficient proteins with resultant depletion of free lysosomes that can fuse with autophagosomes and an accumulation of autolysosomes reflecting failure in autophagic lysosome reformation (Chang et al., 2014). Moreover, spastizin and spatacsin coprecipitate with the SPG48 gene, adaptor-related protein complex-5, ZETA-1 subunit (AP5Z1) encoded protein, adaptor-related protein complex-5 (AP5), that facilitates vesicle-mediated intracellular sorting and trafficking of selected transmembrane cargo proteins, the loss of which results in accumulation of aberrant endolysosomes and association with the cHSP phenotype of spastic paraplegia accompanied by neuropathy, parkinsonism and/or cognitive impairment (Hirst et al., 2015). Mutations in the wash complex, subunit 5 (WASHC5/KIAA0196) encoding strumpellin, ubiquitously expressed in cytosolic and endoplasmic reticulum cell fractions, caused the pure motor phenotype of SPG8 in three large families of North American, British, and Brazilian descent (Valdmanis et al., 2007) due to LOF or haploinsufficiency of the strumpellin protein. The expression level of wild-type strumpellin seems to be of critical importance in the pathogenesis of strumpellin associated HSP. The presence of strumpellin in the Wiskott–Aldrich Syndrome protein and scar homolog complex at endosomes (Derivery et al., 2009) in conjunction with its observed presence in endoplasmic reticulum fractions suggests that the pathophysiology of strumpellin-related HSP is primarily associated with defective actin dynamics in membrane organization and trafficking (Clemen et al., 2010).

BMP SIGNALING Several proteins regulate signaling pathways known to be important for axon function in particular transforming growth factor-beta (TGF-b) family of cytokines, including TGF-b, bone morphogenic proteins (BMPs), and activin/inhibin, that plays crucial roles in embryonic development, adult tissue homeostasis and the pathogenesis of a variety of diseases (Guo and Wang, 2009). The Drosophila NMJ recently provided new insights into the roles of various proteins in neurodegenerative diseases and the importance of BMP/TGF-b signaling. Six proteins, atlastin-1, NIPA1 (SPG6), acetyl-CoA transporter (SPG42), spastin, and spartin, function as inhibitors of BMP signaling. The earliest studies of spastin function in vivo were performed at the Drosophila NMJ. Spastin was found to localize to the NMJ and to be present both in neuronal and muscle cytoplasm. Loss of spastin causes an increase in the number of boutons that are smaller than normal and clustered together, in a pattern reminiscent of the “satellite boutons” seen in some endocytic mutants.

NEUROGENETIC MOTOR DISORDERS Spastin mutants also display a thickened microtubule (MT) network, while tissue-specific overexpression results in a loss of the MT network. This work, along with other data and in vitro experiments, provided further evidence that Spastin is a microtubule-severing protein (Roll-Mecak and Vale, 2008). Of the HSP proteins known to inhibit BMP signaling, among the best characterized mechanistically is NIPA1, a protein with multiple transmembrane domains that localizes to endosomes and the plasma membrane and functions in Mg2 + transport (Goytain et al., 2007). NIPA1 normally encodes Mg2+ transporter and the LOF of NIPA1 (SPG6) due to abnormal trafficking of the mutated protein provides the basis of the HSP phenotype. NIPA1 missense changes in SPG6 patients interfere with this process, thus upregulating signaling. The mechanism by which NIPA1 inhibits BMP signaling involves downregulation of BMP receptors by promoting their endocytosis and lysosomal degradation. Disease-associated mutant versions of NIPA1 alter the trafficking of the type II BMP receptor (BMPRII) and are less efficient at promoting BMPRII degradation than wild-type NIPA1. In conjunction with the observation that spastin and spartin inhibit BMP signaling. As BMP signaling is important for distal axonal function, dysregulation of BMP signaling is a likely unifying pathological component in this endosomal group of HSPs (Tsang et al., 2009).

MOTOR TRANSPORT The kinesin-1 family of cytoskeletal motor proteins use a remarkably versatile cargo recognition mechanism to enable the binding and transport of protein and ribonuclear protein complexes, viruses, microtubules, and many different membrane-bound organelles (Hirokawa et al., 2009). The kinesin thereby powers intracellular movement of membranous organelles and other macromolecular cargo from the cell body to the distal tip of the axon especially in the longest axons of the CST. Neuron-specific kinesin family member 5A (KIF5A) is expressed exclusively in neurons (Niclas et al., 1994). Missense mutations in KIF5A gene leads to the SPG10 phenotype (Reid et al., 2002) of AD HSP with variable manifestations. Some patients have onset of a pHSP with lower limb spasticity, hyperreflexia, extensor plantar responses, and variable involvement of the upper limbs beginning in childhood or young adulthood including distal sensory impairment. However, some patients also show additional neurologic features, including Parkinsonism or cognitive decline, consistent with a cHSP phenotype. The KIF1C gene by contrast represents a member of the Unc104 subfamily of kinesin-like proteins that are involved in the transport of mitochondria or synaptic vesicles in axons and using

205

immunofluorescence, it localizes primarily at the Golgi apparatus where it regulates Golgi to ER membrane flow. Homozygous mutation in the KIF1C gene lead to AR spastic ataxia-2 (SPAX2) characterized by onset in the first two decades manifesting cerebellar ataxia, dysarthria, and variable spasticity of the lower limbs sparing cognition (Dor et al., 2014).

MITOCHONDRIAL FUNCTION The m-AAA protease, an ATP-dependent proteolytic complex in the mitochondrial inner membrane, controls protein quality and regulates ribosome assembly, thus exerting essential housekeeping functions within mitochondria. Mutations in the SPG7 matrix AAA peptidase subunit, paraplegin (SPG7) gene causes axonal degeneration in AR HSP. Such patients can have optic atrophy, cortical atrophy, cerebellar atrophy, cognitive defects in executive function and attention, and supranuclear palsy with spinal cord, frontal lobe, and midbrain white matter changes. Consistent with a role for paraplegin in mitochondrial function, muscle biopsies obtained from some patients with SPG7 mutations showed typical signs of mitochondrial disease (Casari et al., 1998). These included RRFs, intense SDH stained areas, and COX negative fibers. Furthermore, the degree of mitochondrial abnormality correlated with the severity of the disease. Taken together, these findings suggested that a mitochondrial-based mechanism underlies spastic paraplegia. An animal model (Ferreirinha et al., 2004) of AR HSP developed a mouse model for AR HSP due to mutation in the SPG7 gene are affected by a distal axonopathy of spinal and peripheral axons, characterized by axonal swelling and degeneration with mitochondrial morphologic abnormalities in synaptic terminals and distal regions of axons long before the first signs of swelling and degeneration that correlate with onset of motor impairment. Axonal swellings also occur through massive accumulation of organelles and NFs, suggesting impairment of anterograde axonal transport. The investigators (Ferreirinha et al., 2004) speculated that local failure of mitochondrial function may affect axonal transport and cause axonal degeneration.

Familial ALS Approximately 60%–70% of FALS and 10% of SALS cases are caused by mutations in six genes: superoxide dismutase 1 (SOD1), TAR DNA-binding protein (TARDBP), fused in sarcoma (FUS), valosin-containing protein (VCP), chromosome 9 open reading frame 72 (C9orf72), and optineurin (OPTN) implicating disruption of RNA metabolism toxic to motor neurons (C9orf72, TARDBP, FUS) with aberrant cytosolic protein

206

D.S. YOUNGER

misfolding, aggregation, and ubiquitinated deposition (SOD1, OPTN, VCP), disturbed cytoskeletal architectural and dynamic function of distal axons in dynactin 1 (DCTN1). These primary disturbances converge on multiple secondary, downstream pathological processes that result in neuroinflammation, activation of ER stress and autophagy, proteasomal dysfunction, altered mitochondrial function, disturbed axonal transport, dendritic morphology, and excitotoxicity (Ghasemi and Brown, 2018).

C9ORF72 Heterozygous hexanucleotide repeat expansion (HRE) of GGGGCC in a noncoding region of the C9orf72 gene on chromosome 9p21 causes FTD and/or ALS (FTDALS). Unaffected individuals have 2–19 repeats, whereas affected individuals have 250 to over 2000 repeats. However, some individuals can show symptoms with as few as 20–22 repeats (Reddy et al., 2013). The HRE forms DNA and RNA G-quadruplexes with distinct structures and promotes RNA/DNA hybrids (R-loops). The structural polymorphism causes a repeat lengthdependent accumulation of transcripts aborted in the HRE region that bind to ribonucleoproteins (RNPs) in a conformation-dependent manner. Nucleolin, an abundantly expressed acidic phosphoprotein of exponentially growing cells located mainly in dense fibrillar regions of the nucleolus where it is involved in the control of transcription of ribosomal RNA (rRNA) genes, binds the HRE G-quadruplex causing nucleolar stress and incipient neurodegeneration. A mouse model expressing (G4C2)66 throughout the murine CNS using somatic brain transgenesis mediated by adeno-associated mimics the neuropathological and clinical FTDALS phenotype caused by C9orf72 mutations including hyperactivity, anxiety, antisocial behavior, and motor neuron deficits (Chew et al., 2015). Nonetheless, the molecular mechanisms underlying neurodegeneration in C9orf72-related diseases are still debated.

TAR DNA-BINDING PROTEIN 43 (TARDBP) In 2006, the 43 kDa transactive response (TAR) DNAbinding protein 43-KD (TDP-43), a ubiquitous nuclear protein of 414 amino acids involved in transcription regulation and splicing process among other functions, was identified as the major component of the cytosolic ubiquitinated inclusions that are observed in cortical neurons in the vast majority of ALS patients and in patients with tau-negative FTD (Neumann et al., 2006). Research has focused on the formation and consequences of cytosolic protein aggregates as drivers of ALS pathology through both GOF and LOF mechanisms. Not only does

aggregation sequester the normal function of TDP-43, but these aggregates also actively block normal cellular processes inevitably leading to cellular demise in a short time span. Additionally, mutations in TARDBP confer a baseline increase in cytoplasmic TDP-43 thus suggesting that small changes in the subcellular localization of TDP43 could in fact drive early pathology. These pathological findings, and the coexistence of AD ALS and FTD in some patients, underscore that these two diseases are part of the same clinic-pathological spectrum. In 2008, TARDBP mutations were identified in both FALS (ALS10) and SALS patients (Sreedharan et al., 2008). All such mutations identified to date map to exon 6, except Asp169Gly in exon 4 encoding a highly conserved region of the C-terminal domain of TDP-43 protein. This domain is involved in the inhibition of the splicing of the cystic fibrosis transmembrane conductance regulator mRNA (Buratti et al., 2005). This C-terminal domain has been observed in a phosphorylated form in the neuronal cytosolic ubiquitinated inclusions of ALS and FTD patients, together with a TDP-43 nuclear loss, suggesting that cleavage, phosphorylation and cytoplasmic accumulation of this C-terminal fragment underlie the neurodegenerative process involved in ALS and FTD. The frequency of TARDBP mutations in ALS dramatically varies among different studies; the highest frequencies are estimated to be about 5%–6% of FALS and SALS cohorts. On the other hand, little is known about phenotype–genotype correlations in patients with TARDBP mutations. Only one study (Corcia et al., 2012) investigated the phenotypes of 28 patients with TARDBP mutations (9 were SALS) and reviewed the phenotypes of 117 patients with TARDBP mutations reported in the literature (21 were SALS). In that report (Corcia et al., 2012), individuals with TARDBP mutations had an earlier onset and longer survival than SALS patients without TARDBP mutations. Moreover, the site of onset in this group was predominantly upper limb. Additionally, the G298S mutation was associated with shorter survival. Given that ALS and FTD coexist in some individuals, and that there are cytosolic ubiquitinated TDP43 inclusions in both diseases, it is reasonable to anticipate that TARDBP mutations could also account for FTD cases without MND. In fact, one TARDBP mutation (N267S) has been reported in a single patient with FTD without MND (Borroni et al., 2009). A more common observation is that TARDBP mutations may cause both ALS and FTD in the same individual (Benajiba et al., 2009). Increasing evidence suggests that nuclear-tocytoplasmic mislocalization of TDP-43 induces toxicity through both LOF and GOF mechanisms. Classic roles for TDP-43 pertain to mRNA maturation in the nucleus, specifically acting as a repressor of alternate splicing,

NEUROGENETIC MOTOR DISORDERS cryptic exon splicing, and alternate polyadenylation (Polymenidou et al., 2011). TDP-43 is involved in mRNA transport, a mechanism that is dysregulated within ALS, as well as local translational regulation. Disruption of either of these mechanisms may effectively trap TDP-43 in the cytoplasm, inhibiting its normal functions. This hypothesis is substantiated by transcriptomic evidence showing that diseased neurons and mouse models of ALS demonstrate increases in alternative splicing events, cryptic exon inclusion and alternate polyadenylated sequences (Suk and Rousseaux, 2020). Measuring TDP-43 in accessible biofluids such as blood or CSF might reduce diagnostic delay and offer a readout for use as a biomarker and for future drug trials (Feneberg et al., 2018). However, attempts at measuring disease-specific forms of TDP-43 in peripheral biofluids of ALS and FTLD patients have not yielded consistent results, and only some of the pathological biochemical features of TDP-43 found in human brain tissue have been detected in clinical biofluids to date. Moreover, although there may be some benefit to therapeutically targeting TDP-43 aggregation, this step may be too late in disease development to have substantial therapeutic benefit.

FUSED IN SARCOMA (FUS) Mutations in FUS at the 16p11.2 chromosome locus cause FALS and SALS, and rarely FTD (Rademakers et al., 2010). Products of the FUS, EWS RNA-binding protein 1 (EWS) and the TAF15 RNA polymerase II, TATA Box-binding protein-associated factor, 68-kd (TAF15) genes, are structurally similar multifunctional proteins first discovered upon characterization of fusion oncogenes in human sarcomas and leukemias. The proteins belong to the FET (previously TET) family of RNA-binding proteins and are implicated in central cellular processes such as regulation of gene expression, maintenance of genomic integrity, RNA metabolism, mRNA/microRNA processing including mRNA splicing (Law et al., 2006). However, the impact of ALS-causative mutations on splicing has not been fully characterized, as most disease models have been based on overexpressing mutant FUS, which will alter RNA processing due to FUS autoregulation. FUS-ALS mutations induce widespread LOF on expression and splicing (Humphrey et al., 2020). Specifically, mutations in FUS directly alter intron retention levels in RNA-binding proteins and identify an intron retention event in FUS itself that is associated with its autoregulation. The highest levels of FUS are found in the nucleus, driven by a highly conserved carboxyl (C) terminal PY nuclear localization signal (PY-NLS) (Zakaryan and Gehring, 2006). FUS is

207

found in the cytoplasm at lower levels and can shuttle rapidly between the nucleus and the cytoplasm (Zinszner et al., 1997). However, the majority of disease-linked FUS mutations cluster in the C-terminus and disrupt nuclear import, but the precise pathogenic mechanism of FUS mutations is currently unknown.

CU/ZN-SUPEROXIDE DISMUTASE (SOD1) Mutations in the SOD1 gene cause ALS1 through toxic GOF however, the nature of this toxic function remains largely uncertain. Ubiquitylated aggregates of mutant SOD1 proteins in affected brain lesions are pathological hallmarks of the disease and are suggested to be involved in several proposed mechanisms of motor neuron death. SOD1 is a major cytoplasmic antioxidant enzyme that metabolizes superoxide radicals to molecular oxygen and hydrogen peroxide, thus providing a defense against oxygen toxicity (Niwa et al., 2007). Most cases of ALS1 follow AD inheritance, however there are rare AR cases. Superoxide dismutase catalyzes the oxidation/reduction conversion of superoxide radicals to molecular oxygen and hydrogen peroxide (McCord and Fridovich, 1969). The name “superoxide dismutase” comes from the fact that the reaction is a “dismutation” of superoxide anions. The protein had been known for over 30 years as a copper-containing, low molecular weight cytoplasmic protein. Using immunohistochemistry, SOD1 localized to motor neurons, interneurons, and sensory neurons of mouse and human spinal cord, distributed in a punctate pattern throughout neuronal perikarya, in proximal dendrites, and in terminal axons (Pardo et al., 1995). In the brain, SOD1 is present in motor and sensory cranial nerve nuclei, as well as diffusely through the brain in the neurons of the cortex, certain regions of the hippocampus, and amygdala. The intracellular localization was primarily cytoplasmic, but also included nuclei and membranous organelles, presumably peroxisomes. Due to the diffuse and abundant SOD1 expression, it was concluded that pathogenic SOD1 mutations resulted in a toxic GOF rather than haploinsufficiency. SOD1 is a powerful antioxidant enzyme that protects cells from the damaging effects of superoxide radicals. The enzyme binds both copper and zinc ions that are directly involved in the deactivation of toxic superoxide radicals. More than 170 mutations have now been detected in the SOD1 gene in FALS (Abel et al., 2012) together accounting for 20% of FALS cases. The most commonly identified mutations in SOD1 that affect protein activity are D90A, G93A, and A4V. The latter was the commonest mutation in a large registry and correlated with short survival, and severe abnormalities of LMNs with absent or mild UMN involvement clinically and

208

D.S. YOUNGER

pathologically, and with abnormalities in systems other than the MNs more frequent in the FALS A4V subjects. The short survival in A4V is contrasts with AR D90A (Asp-Ala) mutation, with a survival greater than 14 years, and AD G41D (Gly-Asp), G37R (Gly-Arg), G93C (GlyCys), and G93D (Gly-Asp) mutations, wherein survival was >10 years (Cudkowicz et al., 1997). The shorter survival in SOD1 subjects is associated with the stability of the SOD1 protein wherein the propensity to aggregate SOD1 mutations decreases the protein structural stability, acting as a trigger for protein misfolding as well as the appearance of toxic species. Almost all of these mutations are missense changes, scattered across the coding sequence without focal mutation hotspots, however none are predicted to eliminate production of the protein. This strongly suggests that the mutant protein must be present to initiate motor neuron death. Deleterious mutations modify SOD1 activity, which leads to the accumulation of highly toxic hydroxyl radicals. Accumulation of these free radicals causes degradation of both nuclear and mitochondrial DNA and protein misfolding, features which can be used as pathological indicators associated with ALS. Since pathogenicity of mutant SOD1 is proportional to the dose of the toxic protein, a rational approach to treating SOD1-related ALS is to reduce levels of the toxic SOD1 species. This strategy obviates intervening in multiple, downstream pathological cascades. In recent years, several strategies to silence gene expression have been developed. Although mutations in SOD1 account for a minority of ALS cases, the discovery of this gene influenced ALS research leading to the first mouse model of the disease (Gurney et al., 1994) and numerous pathophysiologic and therapeutic investigations. The most clinically promising are predicated on approaches that enhance degradation of RNA, such as antisense oligonucleotides (ASO) and RNA interference (RNAi); the latter include small inhibitory RNA (siRNA), short hairpin RNA (shRNA), and microRNA (miR), which are capable of permeating the CNS and efficiently silencing genes in the brain and spinal cord.

OPTINEURIN (OPTN) Mutations in the OPTN gene have been implicated in both FALS (ALS12) with or without frontotemporal dementia due to homozygous or heterozygous mutation on chromosome 10p13. Both AD and AR inheritance patterns have been reported as well as SALS occurrences (Feng et al., 2019). Optineurin was first identified as a binding partner of an adenoviral E3 14.7 kDa protein and named “FIP-2” (for E3–14.7K-interacting protein) but after renamed to “optineurin” (for optic neuropathy inducing) since mutations in the OPTN gene had been

identified in patients with primary open-angle glaucoma. Later, OPTN mutations were also identified in other human pathologies including Paget disease of bone, ALS and FTD (Maruyama et al., 2010). Optineurin has been characterized as a multifunctional protein regulating multiple cellular processes such as vesicular trafficking, cell division, inflammatory and antiviral signaling, antibacterial responses, and autophagy. OPTN can bind multiple partners; hence, disease-causing mutations may alter these interactions disturbing normal signaling. The clinical phenotypes in OPTN mutated ALS have not been homogeneous, with some individuals showing a relatively slow progression and a long duration, and others an aggressive progression and a short survival. OPTN-immunoreactive skein-like inclusions were found in AHC neurons and neurites in spinal cord sections from a cohort of 32 patients with SALS and in all eight patients with FALS who did not have mutations in the SOD1 gene (Deng et al., 2011). OPTN immunoreactivity was absent in all six patients with familial ALS due to SOD1 mutations and in tissue from two mouse models of ALS due to Sod1 mutations. The findings suggested that OPTN may play a role in the pathogenesis of non-SOD1 ALS, and that SOD1-linked ALS has a distinct disease pathogenesis. ALS-associated OPTN mutations include deletions, missense, frameshift, and nonsense mutations. Functional studies identified three major neuroprotective mechanisms of optineurin: regulation of autophagy, mitigation of inflammatory signaling, and blockade of necroptosis. Deletions, nonsense and frameshift mutations in OPTN are detected in either homozygous or heterozygous states, suggesting that complete LOF or haploinsufficiency of optineurin may be sufficient to cause or contribute to ALS. Most ALS-linked missense optineurin mutations are disproportionally enriched in the C-terminus coiled coil domain, ubiquitin-binding domain, and zinc finger domain. The strongest evidences for pathogenicity exists for the heterozygous missense mutation p.E478G in the ubiquitin-binding domain of OPTN suggesting that it causes disease by GOF or haploinsufficiency (Markovinovic et al., 2017). Optineurin is an adaptor protein that interacts with numerous proteins and is involved in regulating many cellular functions, including vesicular trafficking from the Golgi to plasma membrane, endocytic trafficking, and signaling leading to NF-kappa-B activation (Vaibhava et al., 2012). OPTN actively suppresses receptor-interacting kinase-1 (RIPK1; 603,453)-dependent signaling by regulating its turnover (Ito et al., 2016). Loss of OPTN leads to progressive demyelination and axonal degeneration through engagement of necroptotic machinery in the CNS, including RIPK1, RIPK3, and mixed lineage kinase domain-like protein (MLKL). Furthermore,

NEUROGENETIC MOTOR DISORDERS RIPK1- and RIPK3-mediated axonal pathology was commonly observed in SOD1(G93A) transgenic mice and pathological samples from human ALS patients. In Alzheimer disease, which is partly caused by inadequate mitophagy (Cao et al., 2021), downregulation of OPTN results in activation of AIM2 inflammasomes due to a deficiency in mitophagy in APP/PS1 Tg mice. By ectopic expression, OPTN blocked the effects of Ab oligomer (Abo) on activating AIM2 inflammasomes by inhibiting mRNA expression of AIM2 and apoptosisassociated speck-like protein containing a C-terminal caspase recruitment domain (ASC), leading to a reduction in the active form of caspase-1 and interleukin (IL)-1b in microglial cells.

VALSOLIN-CONTAINING PROTEIN (VCP) VCP encodes valsolin-containing protein, a ubiquitously expressed multifunctional protein that is a member of the AAA+ (ATPase associated with various activities) protein family and implicated in multiple cellular functions ranging from organelle biogenesis to ubiquitin-dependent protein degradation (Weihl et al., 2009). Mutations in VCP cause inclusion body myopathy (IBM) associated with Paget’s disease of the bone (PDB) and FTD (IBMPFD1). In some families with a VCP mutation, family members may have ALS, FTD, or IBMPFD. Patients with frontotemporal dementia and/or amyotrophic lateral sclerosis-6 (FTDALS6) present with AD inheritance of and highly variable manifestations including the behavioral variant of FTD, characterized by reduced empathy, impulsive behavior, personality changes, and reduced verbal output; and others with features of UMN and LMN features of ALS. In both ALS and FTD, there are ubiquitinpositive inclusions within surviving neurons as well as deposition of pathologic TDP43 and pathologic tau aggregates. The primary finding in a cohort of 231 patients were the general lack of genotype–phenotype correlations because of the enormous phenotypic heterogeneity within and between families such that among 36 families of European, Brazilian, Hispanic/Apache, and an African-American ethnicity carrying 15 different heterozygous VCP mutations (Al-Obeidi et al., 2018), 187 were clinically symptomatic and 44 were presymptomatic carriers. Most (90%) of symptomatic patients presented with myopathy (90%), with others Paget disease (42%), dementia (30%), and ALS (8%) associated with UMN and LMN degeneration. Others were diagnosed with Parkinson disease (PD) (4%) and AD (2%). Autopsy data available on one individual with ALS revealed loss of brainstem and spinal cord motor neurons with Bunina

209

bodies in surviving anterior horn cells and TDP-43 immunostaining, consistent with the diagnosis of ALS. There are 17 exons in the VCP gene and mutations have been reported in 11 of them—the vast majority occurring in exon 5. The mutations associated with IBMPFD and/or familial ALS are all exonic missense mutations. In particular, the R155 locus is a mutation hotspot. Transgenic mice with the VCP D395G mutation (601,023.0014) do not spontaneously develop a neurodegenerative phenotype and their brains do not show abnormal tau accumulation (Darwich et al., 2020). However, when stimulated with pathologic tau derived from patients with AD, transgenic mice developed pathologic tau aggregation in several brain regions. The findings suggested that neurons with this VCP mutation have increased susceptibility to pathologic tau aggregation under certain circumstances, resulting in downstream neurodegeneration.

DYNACTIN (DCTN1) Regulation of microtubule dynamics in neurons is critical, as defects in the microtubule-based transport of axonal organelles lead to neurodegenerative disease. The microtubule motor cytoplasmic dynein and its partner complex dynactin drive retrograde transport from the distal axon. The DCTN1 gene encodes p150(Glued), the largest polypeptide of the dynactin complex that binds directly to microtubules and to cytoplasmic dynein, a microtubule-based biologic motor protein (Holzbaur and Tokito, 1996). The p150(Glued) subunit of dynactin promotes the initiation of dynein-driven cargo motility from the microtubule plus-end. As plus end-localized microtubule-associated proteins like p150(Glued) also modulate the dynamics of microtubules, it was hypothesized that p150(Glued) might promote cargo initiation by stabilizing the microtubule track (Lazarus et al., 2013). In vitro assembly assays and total internal reflection fluorescence (TIRF) microscopy in primary neurons using live-cell imaging found that p150(Glued) was a potent anticatastrophe factor for microtubules by binding both to microtubules and to tubulin dimers. Depletion of p150(Glued) in neurons led to a dramatic increase in microtubule catastrophe due to absence of anticatastrophe activity. Disruption of the two functions of dynactin in neurons, in activating dynein-mediated retrograde axonal transport or enhancing microtubule stability through a novel anticatastrophe mechanism regulated by tissue-specific p150(Glued) or both, can contribute to susceptibility of FALS. Neurons possess a striking asymmetric morphology in which elongated axonal processes extend from the cell

210

D.S. YOUNGER

body. The growth and maintenance of the axon, as well as the movement of materials between the cell body and the distal tip of the axon, rely on the mechanism of fast axonal transport. Impaired axonal transport in motor neurons is a mechanism for neuronal degeneration in MND. Cytoplasmic dynein is necessary but not sufficient for retrograde transport directed from the synapse to the cell body. Dynactin is a heteromultimeric protein complex, enriched in neurons, that binds to both microtubules and cytoplasmic dynein. The dynactin complex is required for dynein-mediated retrograde fast axonal transport of vesicles and organelles along microtubules. The complex provides a link between specific cargos, the microtubule and cytoplasmic dynein during vesicle transport. Overexpression of the dynamitin (p50) subunit of dynactin causes a dissociation of dynactin at the junction of p150Glued and the Arp1 filament that effectively renders dynactin nonfunctional (Echeverri et al., 1996) and disrupts the complex and produces a late-onset progressive motor neuron disease in transgenic mice (LaMonte et al., 2002). Dynamitin overexpression is a powerful tool in dissecting the roles of dynein and dynactin in the cell. A genome-wide screen showed linkage of the DCTN1 gene to chromosome 2q13 between the flanking markers D2S291 and D2S2114 in a family with adult-onset progressive AD LMN MND characterized by onset of breathing difficulty due to vocal fold paralysis followed by facial and limb weakness sparing sensation (Puls et al., 2003). Mutation analysis shows a single base-pair change resulting in an amino-acid substitution of serine for glycine at position 59 (G59S) in all affected family members. Modeling of the G59S substitution which occurred in a highly conserved CAP-Gly motif of the p150Glued subunit of dynactin was postulated to reduce the affinity of DCTN1 for microtubules and cause steric hindrance introduced by the larger side chain of serine that distorts its folding compared to the wild-type, that distorts the folding of the microtubulebinding domain. A mutation that severely inhibits the ability of dynactin to bind to microtubules would probably result in a more pronounced degeneration of motor neurons, as is predicted by the transgenic mouse model in which dynamitin is overexpressed (LaMonte et al., 2002).

have enabled the discovery of the genetic basis of a number of Mendelian disorders, notably one form of congenital mental retardation disorder known as Kabuki syndrome due to heterozygous mutation in the MLL2 gene characterized clinically by postnatal dwarfism and characteristic facial dysmorphism, cleft or higharched palate, scoliosis, short fifth finger, persistence of fingerpads, radiographic abnormalities of the vertebrae, hands, and hip joints; and recurrent infantile otitis media (Niikawa et al., 1981). Whole-genome sequencing (WGS) followed by HIT identified seven families with a rare lethal AR disorder clinically characterized by hydranencephaly and progressive CNS glomerular ischemic vasculopathy affecting the retina, brain stem, basal ganglia, and spinal cord, known as Fowlers syndrome, due to homozygous or compound heterozygous mutation in the feline leukemia virus subgroup c receptor 2 (FLVCR2) gene (Thomas et al., 2010). Such strategies eventually enhance the ability to make a rapid diagnosis, provide therapy if available, and enable appropriate counseling regarding prognosis and reproductive implications. It is anticipated that technological advances will reduce the cost of the testing of panels of relevant genes in those with a suspected inherited neurologic disorders. A range of defects can arise in particular genes that lead to disease manifestations. These include nucleotide substitutions that result in a missense or nonsense mutation that encodes part of a gene sequence that can generate a transcript of mRNA that is prematurely degraded and lost via nonsense mediated decay, or a nonfunctional protein product that is significantly truncated and missing key components (Nicholson and Muhlemann, 2010). In instances where the mutation results in the generation of a misfolded protein, a quality control system within the ER may mark the protein for ultimate degradation via the ubiquitin-proteasome pathway (Mehnert et al., 2010). A gene sequence can also be subject to a complete or partial deletion, or its sequence altered by a recombination event or the use of a cryptic splice site which leads to the generation of a dysfunctional protein.

FACTORS INFLUENCING PHENOTYPICAL GENE EXPRESSION

GENETIC SEQUENCING STRATEGIES

Dominant negative effects and genetic heterogeneity

High-throughput (HIT) sequence capture methods and NGS technologies are proving to be productive means for identifying causal gene defects, particularly in sporadic and rare disorders not amenable to linkage type study analyses (Moorhouse and Sharma, 2011). Whole-exome sequencing (WES) and target region sequencing (TRS)

A dominant negative effect occurs when a mutation leads to a defective gene product that effectively abrogates the function of the normal gene product or its interacting partner. Genetic heterogeneity refers to the manifestation of a single phenotype despite different underlying genes mutations.

NEUROGENETIC MOTOR DISORDERS

211

LIMB GIRDLE MUSCULAR DYSTROPHY

TAY-SACHS AND JUVENILE GM2 GANGLIOSIDOSIS

Patients with LGMD2A and CAPN3 gene defects can harbor mutations at D705G and R448H associated with retained proteolytic activity that can eventuate in a muscular dystrophy attributed to reduction in CAPN3 ability to bind in vitro to titin resulting in myofiber structural instability (Ermolova et al., 2011). The equivalent phenotypes of LGMD1 and LGMD2A that derive respectively from mutations in the myotillin (MYOT) gene, leads to a defective structural protein important in stabilizing and anchoring thin filaments to the Z-disc during myofibrillogenesis, and in others with mutation in the CAPN3 gene that encode a different muscle-specific calcium-activated neutral protease that plays a role in disassembling sarcomeric proteins (Broglio et al., 2010). When testing for a limited number of genes, a negative result may not exclude the clinical diagnosis under consideration. As new gene defects are described on a regular basis, it is generally worthwhile to re-examine a patient without an etiologic diagnosis, if several years have passed since testing was last performed.

Tay-Sachs disease (TSD) is an AR progressive neurodegenerative disorder due homozygous or compound heterozygous mutation in the alpha subunit of the hexosaminidase A (HEXA) gene. The most frequent DNA lesion in TSD in Ashkenazi Jews is a 4-bp insertion in exon 11, which leads to the classic infantile form characterized clinically by developmental retardation, followed by paralysis, dementia and blindness, and death by age 2 years. In contrast, non-Jewish patients with the variant of juvenile-onset GM2-gangliosidosis have an onset of ataxia by age 6 years and thereafter show decerebrate rigidity and blindness with death by age 15 years. The defect in classical TSD is a deficiency in HEXA while patients with the GM2-gangliosidosis variant produce HEXA that appears catalytically normal when tested with substrates but is defective against substrates that are hydrolyzed by the active site on the alpha subunit of normal HEXA that is inactivated in patients’ enzyme (Kytzia and Sandhoff, 1985). An analysis of 33 missense mutations in the HEXA gene reveal a close correlation both between large structural alterations in functionally important regions in infantile cases along with smaller ones located in less functionally important regions in later-onset cases both with reduced red blood cell (RBC) sphingomyelin. The latter is useful in carrier detection, and in predicting who may be unaffected because of sufficient residual Hex A enzyme activity from the corresponding normal allele.

Haploinsufficiency Haploinsufficiency presides when a deletion or nonsense mutation in one copy of a gene leads to no protein being expressed from that allele, or a missense mutation in one copy of a gene results in the allelic expression of a nonfunctional protein even though the total amount in the cell may be normal.

ACUTE INTERMITTENT PORPHYRIA Acute intermittent porphyria (AIP) is caused by the haploinsufficiency of the porphobilinogen deaminase (PBGD) gene that results in a marked loss of activity of PGBD, the third enzyme of the heme biosynthesis pathway (Chen et al., 2016). Acute neurovisceral attacks of abdominal pain, nausea, constipation, vomiting result from external or internal factors that increase the demand for heme or strongly induce ALAS1 in the liver of individuals carrying the inherited deficiency of PBGD. The consequence is a marked overproduction and accumulation of ALA and PBG, the intermediate products between both enzymes. Aminolevulinic acid induces the formation of free radicals that causes oxidative damage compounded by inadequate heme synthesis that impairs mitochondrial electron transport, energy dependent Na+/K+ ATPase, and P450 drug and mitochondrialbased oxidative damage repair. A reduction of the relevant enzyme activity to just one-half of normal is not adequate to prevent development of symptoms of peripheral neuropathy (Lin et al., 2011; Younger and Tanji, 2015).

Penetrance Penetrance can be a factor in nonmanifesting carriers of the AD disease phenotype in primary torsion dystonia (DYT1) due to heterozygous mutation in the torsin 1A (DYT1) gene encoding the ATP-binding protein torsin1A. Although a GAG deletion in the DYT1 gene is the major cause of early-onset dystonia, expression as clinical disease occurs in only 30% of mutation carriers due reduced penetrance. Investigators (Risch et al., 2007) examined the genetic factors influencing penetrance in DYT1 torsion dystonia noting the presence of a single-nucleotide polymorphism, D216H codingsequence variation the moderated the effect of the DYT1 GAG deletion in cellular models. The frequency of the 216H allele was increased in GAG-deletion carriers without dystonia and decreased in carriers with dystonia compared to controls. There was a highly protective effect of the H allele in trans with the GAG deletion and suggestive evidence that the D216 allele in cis was required for the disease to be penetrant establishing for the first time a clinically relevant genet modifier of DYT1. All relatives suspected of carrying

212

D.S. YOUNGER

the same gene defect should undergo appropriate genetic counseling particularly if they are symptomatic, and if not, they can still pass the genetic trait on to future generations.

Lyonization Lyonization (Lyon, 1961; Emery, 1965) that results in random inactivation of the X chromosome early in development can cause aberrant expression of the DMD phenotype in female carriers.

DUCHENNE MUSCULAR DYSTROPHY CARRIERS One study (Bonilla et al., 1988) used antibodies directed against dystrophin to study the protein in individual fibers of carriers of the gene. In transverse sections of normal human muscle, the immunocytochemical reaction for dystrophin was evident in the sarcolemma of all muscle fibers. In longitudinal sections, the protein was distributed homogeneously in the sarcolemma, all along the length of the fiber. In patients with typical DMD, the protein was either totally absent or there were only faint nonhomogeneous spots of immunoreactive material in the sarcolemma. Moreover, muscle from carriers of the gene contained two populations of fibers, some containing dystrophin and others with partial deficiency of dystrophin, the proportion of which was higher (18%–32%) in manifesting carriers than in asymptomatic carriers, where the proportion of dystrophin-deficient fibers was 0%–4.3%. These results were compatible with the Lyon hypothesis stating that there was random inactivation of the X chromosome early in development (Lyon, 1961; Emery, 1965) resulting in the random fusion of two populations of myoblasts, one carrying the mutated X-chromosome, the other containing the normal X. The proportion of nuclei containing the mutated DMD gene within any muscle fiber of a carrier determined whether the fiber will have a normal amount, partial deficiency or total lack of dystrophin at the sarcolemma. The authors (Bonilla et al., 1988) suggest something long believed by clinicians, that manifesting carriers of the DMD gene probably have a higher proportion of affected muscle fibers than asymptomatic carriers, and that symptoms result even when fewer than half of the fibers are affected.

Loss-of-function Discovery of gene defects can lead to the understanding of novel mechanisms of disease. For example, mutations in the ab hydrolase domain-containing protein 12, lysophospholipase (ABHD12) gene encoding an hydrolase domain containing 12, causes AR polyneuropathy, hearing loss, ataxia, retinitis pigmentosa, and cataract

(PHARC) (Fiskerstrand et al., 2010), due to enzymatic defect of 2-arachidonoyl glycerol (2-AG) hydrolysis, an endogenous transmitter acting on cannabinoid receptors. While dysregulation of the endocannabinoid system was not previously implicated in ataxia, to date it not have associated with any other neurodegenerative disorders. This discovery may ultimately enable development of rationale therapies based on understanding of this disease producing mechanism.

Toxic gain-of-function A gene defect can result in a GOF or a novel property for the resulting gene product, which then causes a disruption in a particular cellular\biochemical pathway. Infrequently, the defect may not be an alteration of the gene sequence but a consequence of changes in its expression pattern as occurs in FSHD. The distinctive clinical pattern of skeletal muscle weakness is associated with a wide spectrum of disease severity. The pathophysiologic consequences of the genetic lesion, the loss of a critical number of macrosatellite repeats in D4Z4 in the subtelomeric region of chromosome 4q35, remained unexplained for almost two decades until recent studies demonstrated that contraction in the number of D4Z4 repeats results in chromatin relaxation and transcriptional derepression of DUX4, a gene normally expressed only in the germline. It is now recognized that normal individuals have >10 D4Z4 repeats and normally methylated chromatin, with DUX4 expression turned off, whereas those with FSHD type 1 (FSHD1), contraction of D4Z4 to A and c.1965-2A>C, suggests that they operate via a haploinsufficiency mechanism. Specifically, the c.209C>A

218

D.S. YOUNGER

nonsense mutation results in degradation of c.209C>A mRNA by the nonsense mediated decay (NMD) pathway reducing the levels of hGle1 both at the nuclear pore complex (NPC) (Isoform B) and in the cytoplasm (Isoform A). It is intriguing that hGle1, like TDP-43 and FUS, plays roles at multiple stages of the mRNA life-cycle and that all three are linked to motor neuron pathologies. While there are no apparent functional links between hGle1 and either TDP-43 or FUS, hGle1 could be required for the nuclear export of the same mRNAs that need TDP-43 and FUS function.

Immunoglobulin m-binding protein 2 In contrast to SMN, mutations in the immunoglobulin m-binding protein 2 (IGHMBP2) gene on chromosome 11q13-q21 are associated with the AR disorder type 1 SMA with respiratory distress (SMARD) also called diaphragmatic SMA, distal hereditary motor neuropathy type VI, or severe infantile axonal neuropathy with respiratory failure. This clinically and genetically distinct form of SMA1 concomitantly involves AHCs and DRG pathologically with evidence of severe peripheral motor and sensory axonopathy at postmortem examination. Among 29 infants affected by SMARD1 with 26 novel IGHMBP2 mutations described by Grohman and colleagues (Grohmann et al., 2003), intrauterine growth retardation, weak cry, and foot deformities were the earliest symptoms in this cohort who presented from age 1 to 6 months with respiratory distress due to diaphragmatic paralysis. All had progressive muscle weakness with predominantly distal lower limb muscle, and sensory and autonomic involvement. Because of the poor prognosis, there is a demand for prenatal diagnosis, and clear diagnostic criteria for infantile SMARD1. Patients with non-5q SMA or unknown neuropathy and the consanguineous parents of a child with sudden infant death syndrome should be examined for IGHMBP2 mutations.

Glycyl-tRNA synthetase 1 Aminoacyl-tRNA synthetases perform an essential function in protein synthesis by catalyzing the esterification of an amino acid to its cognate tRNA. These enzymes are necessarily present in each cell and must properly recognize the tRNA and the amino acid in order to maintain fidelity of translation. The nuclear glycyl-tRNA synthetase gene (GARS1) and the encoded enzyme glycyl-tRNA synthetase covalently attach glycine to its cognate tRNA, which is essential for protein translation (Boczonadi et al., 2018). Unlike most other tRNA synthetase genes, GARS1 encodes both the cytoplasmic and mitochondrial isoforms of the enzyme. James type

of infantile spinal muscular atrophy (SMAJI) is caused by heterozygous mutations in the GARS1 gene on chromosome 7p15 (James et al., 2006; Markovitz et al., 2020). James et al. (2006) reported a 7-year-old girl, born of unrelated parents who presented at 6 months of age with lower extremity weakness and floppy feet and normal early motor development until that time, but thereafter showed motor difficulties and never achieved independent ambulation. The disorder was progressive with distal muscle weakness, wasting and areflexia of the legs and hands with weak eye closure, cough, decreased voice volume, and mild respiratory insufficiency. Skeletal features included hip flexion contractures, foot deformities, marked lumbar lordosis, and scoliosis. Nerve conduction studies were normal but EMG and muscle biopsy were consistent with AHC disease. Functional studies of the variant were not performed, but the mutation affected a residue in the anticodon binding domain; the authors postulated a dominant-negative effect. A de novo heterozygous missense mutation in GARS1 was found and while functional studies of the variant were not performed, the mutation affected a residue in the anticodon binding domain and the authors postulated a dominantnegative effect. In a series of three unrelated cases, Markovitz et al. (2020) identified de novo heterozygous missense mutations in GARS1 by trio-based exome sequencing. In vitro functional complementation studies showed that the I334N variant was unable to rescue the growth defect phenotype in yeast, consistent with a LOF effect. A case series of pathogenic variants in the GlycyltRNA synthetase gene causing the allelic disorders CMT2D and distal HMN type V. Other similar cases have been described, the youngest being 3 months demonstrated the clinical utility of NGS (Forrester et al., 2020). Certain aminoacyl-tRNA synthetases are autoantigens in patients with the idiopathic inflammatory myopathies, polymyositis, and dermatomyositis. Autoantibodies reactive with synthetases are found almost exclusively in these conditions, with individuals usually having autoantibodies to only a single synthetase. Most commonly they are directed at histidyl-tRNA synthetase labeled anti–Jo-1 autoantibodies (Ge et al., 1994).

Nova1 Patients with breast and lung cancer may develop paraneoplastic opsoclonus-myoclonus-ataxia (POMA), a disorder of motor control and harbor an antibody to Ri that recognizes the tumor and nuclear neuronal protein of 55 kd. The nova alternative splicing regulator 1 (Nova1) gene encodes a highly conserved protein

NEUROGENETIC MOTOR DISORDERS homologous to the RNA-binding protein (hnRNP K) that is inhibited by the Ri antibodies. Northern blot analysis detects Nova1 transcripts only in brain, and in E18 experimental mice in the developing ventral brain stem and spinal cord (Buckanovich et al., 1993). Using antisera from patients with POMA, neuron-specific RNA-binding protein Nova-1 binds to mRNA with high affinity in the subcortical nervous system (Buckanovich et al., 1996). Younger and colleagues (Younger et al., 2013) described a 49-year-old woman with elevated serum and CSF Ri antibodies associated with malignant breast cancer who after curative surgery, radiation therapy and chemotherapy developed MND with LMN and UMN signs. Nova was present in the patient’s tumor cells and reactive with mouse cerebellar neurons, staining the developing mouse spinal cord, as did a 1:100 dilution of CSF. Moreover, patient antisera reacted to 55 kD antigen present in tumor nuclei and Nova1 and Nova2 antigens. In the same way that the production of Ri antibodies that inhibit Nova-1-RNA interactions appear to be the cause of POMA, these same antibodies

219

present in serum and CSF were the likely cause of paraneoplastic MND. The patient improved with 3 months of immune modulatory therapy of intravenous immune globulin and plasma exchange.

REPEAT EXPANSIONS DISEASES Investigation of putative disease mechanisms may reveal disease pathways that explain exceptional genetic heterogeneity as in the REDs. These simple sequence repeat disorders account for more than 50 human diseases, the majority of which are affect the nervous system, and are severe, degenerative, and not currently treatable or preventable. There are also at least 13 different types of tandem repeats whose expansions cause various human diseases (Table 9.3). Beyond these monogenic diseases, some specific repeat expansions might contribute to the etiology of various complex polygenic psychiatric and brain disorders, such as autism spectrum disorder, bipolar spectrum disorders, schizophrenia, and others

Table 9.3 Neurological repeat expansions diseases Grouped by the repeat expansion CAG: HD (HTT), SBMA (AR), DRPLA (ATN1), and SCA 1 (ATXN1), 2 (ATXN2), 3 (ATXN3), 6 (CACNA1A), 7 (ATXN7), 12 (PPP2R2B), 17 (TBP) CGG: FXS, (FMR1), FXTAS (FMR1) CTG: DM1 (DMPK), HDL2 (JPH3), SCA 8 (ATXN8) GAA-FRDA (FXN) GCG: OPMD (PABPN1) ATTCT: SCA 10 (ATXN10), SCA 37 (DAB1) TGGAA, TAAAA, TACAA, GAAA, TAAC, TGAAA: SCA 31 (BEAN1) TTTTA: TTTCA: FAME3 (MARCHF6) GGCCTG: SCA36 (NOP56) CCCTCT: XDP (TAF-1) GGCCTG: SCA 36 (NOP56) GGGGCC: FTD/ALS1 (C9orf72) CCCCGCCCCGCG: EPM1 (CSTB) Grouped by etiopathogenetic mechanism of the repeat expansion LOF of the gene containing the repeat: FRDA, FXS, EPM1A Toxic GOF due to production of a protein containing a polyglutamine tract expansion: HD, SBMA, SCA1, 2, 3, 6, 7, 17, DRPLA Toxic GOF due to production of RNA containing an expanded CUG tract: SCA8, C9orf72 ALSFTD, DM1/2 Toxic GOF due to production of a protein containing a polyalanine tract expansion: OPMD Abbreviations: HD, Huntingtin disease, HTT, huntingtin; SBMA, spinal and bulbar muscular atrophy, X-linked; AR, androgen receptor; DRPLA, Dentatorubral-Pallidoluysian atrophy; ATN1, atrophin 1; ATXN1, ataxin-1; ATXN2, ataxin-2; ATXN3, ataxin-3; CACNA1A, calcium channel voltage-dependent, P/Q type, alpha 1A subunit; ATXN7, ataxin-7; PPP2R2B, protein phosphatase 2, regulatory subunit B, beta; TBP, TATA box-binding protein-like protein 1; FXS, fragile X syndrome; FMR1, FMRP translational regulator 1; FXTAS, fragile X tremor/ataxia syndrome; DM1, myotonic dystrophy; DMPK, dystrophia myotonica protein kinase; HDL2, Huntington disease-like 2; JPH3, junctophilin-3; ATXN8, ataxin 8; FRDA, Friedrich ataxia; FXN, frataxin; OPMD, oculopharyngeal muscular dystrophy; PABPN1, MARCHF6, Membrane-associated ring-ch finger protein 6 Polyadenylate-binding protein, nuclear, 1; ATXN10, ataxin 10; DAB1, DAB adaptor protein 1; BEAN1, brain-expressed, associated with NEDD4, 1; MARCF6, membrane-associated ring-CH finger protein 6; NOP56, NOP56 ribonuclear protein; C9orf72, chromosome 9 open reading frame 72; CSTB, cystatin B; LOF, loss-of-function; GOF, gain-of-function.

220

D.S. YOUNGER

(Hannan, 2010). The number of known REDs is likely to grow as more than 100 human genes contain DNA repeats that are known to expand in some REDs. The majority of REDs share two common features. First, the number of inherited repeats positively correlates with disease severity and negatively correlates with age of onset. Such disorders with strong correlations between the number of repeats and age of onset of CAG repeats in SCA7 (Gouw et al., 1998) and SCA3 (Kawaguchi et al., 1994), SCA2 (Giuffrida et al., 1999a), ATTTC repeat expansion in SCA37 (Seixas et al., 2017), CAG repeat expansion in HD (Vital et al., 2016), CTG repeat exapansion in DM1 (Morales et al., 2012) and DRPLA (Morales et al., 2012), (CCCTCT)n repeat expansion in X-linked dystonia parkinsonism (XDP) (Westenberger et al., 2019), TTTTA/TTTCA repeat expansion in familial adult myoclonic epilepsy 3 (FAME3) (Florian et al., 2019), and GAA repeat expansion in FRDA (Al-Mahdawi et al., 2018). For some disorders, the correlation between the number of repeats and symptom manifestation is less clear, although there typically is a marginally significant trend as in GGGGCC-repeat expansion in C9orf72 gene in the major cause of FTD and ALS (Nordin et al., 2015), CAG repeat expansion in SCA6 (Wiethoff et al., 2018), and a 12-nucleotide homozygous dodecamer repeat expansion mutation in the cystatin B (CSTB) gene in Unverricht–Lundborg disease (progressive myoclonus epilepsy type 1 (EPM1) (Hypponen et al., 2015). Second, expansion of one disease-causing repeat does not promote expansions of other repeats in the patient’s genome. In other words, each RED patient typically has only a single repeat type expanded (Ida et al., 2018) so noted among cases of HD in whom C9orf72 repeat expansion did not coexist with HTT repeat expansion. Since their discovery three decades ago, a variety of different repeat expansion mutations have been identified, reflecting four possible etiopathogenetic mechanisms: LOF of the gene containing the repeat (FRDA, FXS, EPM1A); toxic GOF due to production of a protein containing a polyglutamine tract expansion (HD, SBMA, SCA1, 2, 3, 6, 7, 17, DRPLA); toxic GOF due to production of RNA containing an expanded CUG tract (SCA8, C9orf72 FTDALS, DM1, DM2) or the toxic GOF due to production of a protein containing a polyalanine tract expansion (OPMD). Classification of the repeat expansion diseases into one of these four mechanistic categories typically reflects both the sequence composition of the repeat and the location of the repeat within a gene.

Loss-of-function in the repeat expansion Friedreich ataxia: The majority of FRDA patients are homozygous for GAA repeat expansion mutations

within intron 1 of the FXN gene on chromosome 9q13; normal individuals have 5–30 GAA repeat expansions, whereas affected expansions associated with FRDA vary from 44 to 1700 repeats, with most abnormal alleles ranging from 600 to 900 GAAs. The length of the shorter allele is negatively correlated with the age of onset. Longer expansions usually result in a more severe phenotype with an earlier onset, faster progression and a higher rate of nonneurological features (Durr et al., 1996). There are resultant decreased levels of frataxin, subsequent oxidative stress, iron deposition, and ultimately neurodegeneration, primarily in the large sensory neurons of the dorsal root ganglia, with hypertrophic cardiomyopathy (Al-Mahdawi et al., 2006). The triad of hypoactive knee and ankle jerks, signs of progressive cerebellar dysfunction, and preadolescent onset is commonly regarded as sufficient for diagnosis. A comprehensive characterization of the spatial profile and progressive evolution of structural brain abnormalities in people with FRDA (Harding et al., 2021) revealed regional brain volume deficits in the brainstem and cerebellar subcortex as core features of FRDA, and volumetric measures of the superior cerebellar peduncles and dentate regions in particular representing, excellent candidates for longitudinal follow-up and validation as T1-weighted MRI biomarkers in this disease (Harding et al., 2020). The clinicopathological features of FRDA disease are recapitulated in model mice that express frataxin only from GAA repeat expansioncontaining FXN transgenes (Al-Mahdawi et al., 2006). It is believed that the GAA triplet repeat expansion may result in an unusual yet stable DNA structure that interferes with transcription, ultimately leading to a cellular deficiency of frataxin. Pathogenic GAA repeat expansions in the FXN gene cause decreased mRNA expression of FXN by inhibiting transcription. Frataxin localizes to the mitochondrial matrix where it is associated with the inner mitochondrial membrane. In FRDA patients, frataxin is reduced to 5%–35% of the levels of healthy individuals while levels in asymptomatic heterozygotes are reduced by 50% (Campuzano et al., 1997). Expression levels are correlated to the repeat length and disease severity. Frataxin normally activates mitochondrial iron–sulfur-cluster (ISC) protein assembly as part of a multiprotein complex. Frataxin is thought to be involved in the synthesis of heme-containing proteins involved in a variety of cellular processes such as oxygen metabolism and electron transfer. In essence, the demonstration that the human pathology of FRDA is also characterized by mitochondrial iron accumulation, deficit of respiratory chain complex activities and in vivo deficit of tissue energy metabolism establishes FRDA as a “new” nuclear encoded mitochondrial disease (Lodi et al., 2001).

NEUROGENETIC MOTOR DISORDERS Fragile X syndrome: FXS is also a neurological disorder with profound genetic variability and a leading cause of inherited intellectual disability and autism. Variable expressivity, which refers to the different patterns of clinical manifestations that occur with the same mutation suggests in FXS, the influence of other factors and modifiers that interacting with the gene defect to a differing disease course. The majority of people with this condition have an allele with an expansion of more than 200 repeats in a tract of CGGs within the 50 untranslated region; this expansion is associated with a hypermethylated state of the gene promoter. FXS has incomplete penetrance and variable expressivity. Intellectual disability is present in 100% of males and 60% of females. Autism spectrum disorder symptoms appear in 50% to 60% of males and 20% of females. Other characteristics such as behavioral and physical alterations have significant variations in presentation frequency. The molecular causes of the variable phenotype in FXS patients are becoming clear: these causes are related to the FMR1 gene itself and to secondary, modifying gene effects. In FXS patients, size and methylation mosaicisms are common. Secondary to mosaicism, there is variation in the quantity of FMR1 mRNA and the encoded Fragile Mental Retardation Protein (FMRP). Characterizing patients according to CGG expansion, methylation status, concentration of mRNA and FMRP, and genotypification for possible modifier genes in a clinical setting offers an opportunity to identify predictors for treatment response evaluation. When intervention strategies become available to modulate the course of the disease they could be crucial for selecting patients and identifying the best therapeutic intervention. Epilepsy, progressive myoclonic 1A: Myoclonic epilepsy of Unverricht and Lundborg (ULD), also known as progressive myoclonic epilepsy-1A (EPM1A) is an AR neurodegenerative disorder. Affected children present with progressive myoclonic epilepsy (PME) between 6 and 13 years of age and later develop ataxia with minimal or no cognitive decline into adults often with well-controlled epilepsy (Ramachandran et al., 2009). Clinicopathological data was provided in several large families (Carr et al., 2007). A survey of 15 United States (US) families (Eldridge et al., 1983) showed 27 affected members with onset at age 10 of photosensitive, occasionally violent, myoclonus, usually worse upon waking; generalized tonic–clonic seizures, sometimes associated with absence attacks; and light-sensitive, generally synchronous, spike-andwave discharges on EEG that preceded clinical manifestations. Necropsy revealed marked loss of Purkinje cells (PCs) of the cerebellum, but no inclusion bodies. The major gene involved in ULD is CSTB that encodes stefin B, also called cystatin B, a member of

221

the superfamily of cysteine inhibitors that resides intracellularly. EPM1 is caused by LOF mutations in CSTB. There are at least 10 identified mutations with an unstable expanded polymorphic dodecamer repeat in the promoter region of CSTB accounting for >90% of mutated alleles (Joensuu et al., 2008), and practically every ULD case shows the dodecamer repeat expansion in one of the alleles. Mutations lead to dramatic reduction in CSTB mRNA that leave only about 10% indicating that ULD is the phenotypic outcome of vastly reduced CSTB. The latter is a ubiquitously expressed intracellular inhibitor of cysteine proteases, including cathepsins B, H, S, K, and L (Turk et al., 2008). In addition, it is reported to protect cells against apoptosis (Caballero Oteyza et al., 2014) and oxidative stress (Lehtinen et al., 2009), and play a role in cell cycle regulation (Ceru et al., 2010), suggesting a neuroprotective role for cystatin B, quenching excess lysosomal hydrolases that leach into the cytoplasm during times of cellular stress. Experimental cystatin B-deficient mice develop myoclonic seizures and ataxia, similar to symptoms seen in the human disease. The resulting cytopathology is a loss of cerebellar granule cells which displays condensed nuclei, fragmented DNA and other cellular changes characteristic of apoptosis suggesting that cystatin B in fact has a role in preventing cerebellar apoptosis,. However, knockout Cstb+/+ mice crossed with Cstb/ mice show no improvement in the ataxia or PME, but have reduced cerebellar granule cell apoptosis. CSTB deficiency contributes to presymptomatic GABAergic signaling and early neuroinflammation. A recent proteomic approach in presymptomatic Cstb-deficient mouse cerebellar synaptosomes noted a decrease in sodium- and chloride dependent GABA transporter 1 (GAT-1) suggesting early mitochondrial dysfunction may contribute to altered synaptic function (Gorski et al., 2020).

Gain-of-function due a protein containing the polyglutamine tract expansion Huntington disease: HD is caused by a heterozygous expanded trinucleotide repeat (CAG)n encoding glutamine in the huntingtin (HTT) gene at the 4p16.3 chromosome locus. In normal individuals, the range of repeat numbers is 9–36 whereas in HD, the repeat number exceeds >37. Affected patients present in adulthood with a distinctive phenotype of chorea, dystonia, incoordination, cognitive decline, and behavioral difficulties due to progressive and selective neural cell loss and atrophy in the caudate and putamen. Juvenile-onset HD, typically defined as onset 50%) can be associated with a normal CMAP and normal clinical strength (see Fig. 10.7; Ferrante and Tsao, 2020). In addition to the number of innervated muscle fibers, the CMAP amplitude also reflects the synchrony of the arrival times of the contributing MUAPs. With pathologic temporal dispersion, CMAP amplitude declines. Other important causes of CMAP amplitude reduction include muscle endplate dispersion and excessive adipose or edema. Muscle endplate dispersion occurs with skeletal changes (e.g., arthritis) disperse the muscle endplates over a larger surface area. Excess adipose or edema between increases the distance between the muscle fibers and the E1 electrode, thereby increasing signal loss. Negative area under the curve The area located between the baseline (the x-axis) and the trace, which is referred to as the negative area under the curve (AUC), is calculated by the EMG machine and reported in mV ms. Like amplitude, negative AUC normally reflects the total number of innervated muscle fibers and the number of functioning motor axons. Unlike the amplitude, negative AUC is much less affected by pathologic temporal dispersion and, therefore, is preferred over amplitude when the waveform is dispersed (DMCS) to avoid overestimating lesion severity.

DISTAL LATENCY The onset latency is the time measurement value, in ms, between the stimulus and the onset of the CMAP. The

NEUROMUSCULAR ELECTRODIAGNOSIS

257

onset latency value, thereby approximating the nerve conduction time between the two stimulation sites.

CONDUCTION VELOCITY Like other velocities, the NCV, in m/s, is calculated by dividing the change in distance (the cutaneous distance between the two stimulation sites) by the change in time (the onset latency difference between the proximal and distal motor responses).

NEGATIVE PHASE DURATION The duration of the negative phase is the time interval, in ms, from the onset of the negative phase to it termination. It reflects the range of conduction velocities among the conducting motor axons and increases with the distance between the stimulation site and the E1 electrode.

THE UTILITY OF MOTOR NCS

Fig. 10.7. The illustration depicts a 4-axon nerve with an innervation ratio of 5. Top: The normal nerve will generate a motor response that reflects 20 muscle fiber APs. Middle: Following the loss of 2 AHCs, the motor response will reflect 10 muscle fiber APs, which would correlate with the degree of severity. Bottom: Following reinnervation, the motor response will again reflect 20 muscle fiber APs, despite 50% of the motor units being nonfunctional (pseudonormalization). Although clinical strength would also normalize, easier fatigability would likely be present. Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 30.

onset latency of the distal response is termed the distal latency. Because the latency value only reflects the CV of the fastest conducting axon, it provides no information about the other axons. Hence, it is insensitive to axon loss, the pathophysiology underlying most PNS lesions. The proximal latency is used to compute the NCV between the stimulation sites. In addition to reflecting nerve conduction time, the onset latency reflects a number of events, including nerve activation time, nerve conduction time, terminal branch conduction time, NMJ transmission time, muscle fiber activation time, muscle fiber conduction time, and muscle tissue transit time. Thus, the onset latency severely underestimates the NCV. To eliminate these contaminants, the nerve is stimulated at two sites and the distal onset latency value is subtracted from the proximal

Motor NCS are able to assess long nerve segments for focal demyelination and they are invaluable for estimating lesion severity. The presence of focal dispersion identifies nonuniform DMCS, whereas focal amplitude and negative AUC decrements identify focal DMCB. Early axon disruption (prior to Wallerian degeneration) mimics DMCB and is localized in the same way (i.e., by stimulating the nerve more proximally while looking for a change in response morphology). The NCS manifestations of these pathophysiologies are discussed below.

Sensory nerve conduction studies TECHNIQUE For the sensory NCS, the E1 an E2 electrodes are placed over the nerve, approximately 3 cm apart, and the nerve is stimulated proximally (antidromic technique) or distally (orthodromic technique). Because SNAPs are composed of nerve fiber APs (rather than large numbers of muscle fiber APs), they are much smaller (measured in microvolts) than motor responses. Because the E1 and E2 recording electrodes are both positioned over the nerve under study, both electrodes record a triphasic response. Because differential amplification is used to remove unwanted signal, the distance between the E1 and E2 electrodes has a major effect on SNAP morphology and, hence, the electrodes must be precisely placed— desired signal is lost when they are too close and undesired signal is amplified when they are too distant.

MEASUREMENTS The important measurements for sensory responses are amplitude and latency. There are a large number of

258

M.A. FERRANTE

concepts pertinent to these two measurements that must be understood by the EDX provider to avoid making significant errors. Amplitude The SNAP amplitude reflects the functioning sensory axons and, to a lesser degree, their synchrony. The amplitude is measured from the baseline to first negative peak with biphasic responses (the baseline-to-peak amplitude) and from the first positive peak to the first negative peak with triphasic responses (the peak-to-peak amplitude) (Fig. 10.8; Ferrante and Tsao, 2020). The first positive peak represents SNAP onset as this point represents when moment when positive ions change from moving toward the electrode to moving away from it. It is a serious mistake to measure the amplitude from the first negative peak to the second positive peak because this portion of the response represents repolarization rather than depolarization and is the portion of the curve where the E2 electrode has its greatest influence. Latency Latency measurements show the greatest variation among EMG laboratories, including whether onset latencies or peak latencies are used, whether the distance between the stimulation site and the E1 electrode is fixed or landmark-based, and whether the latency value is converted into a conduction velocity value. In the past, due to technical limitations, peak latencies were used because onset latencies were not easily visualized. With modern EMG machines, this is not an issue. The onset latency value is taken where the trace first departs from the baseline (biphasic response) or at the

peak of the first positive phase (triphasic response), whereas the peak latency is taken at the peak of the first negative phase. The onset latency value reflects the AP propagation speed of the fastest conducting fiber and provides no information about the other functioning sensory axons. The peak latency value does not reflect the fastest fiber but, instead, more closely approximates the average conduction speed among the functioning sensory axons. Like motor latencies, these values are contaminated by phenomena unrelated to nerve conduction time, including depolarization and tissue transit times. Nonetheless, no study has shown that onset latency is more sensitive than peak latency. In addition, peak latency may be more sensitive than onset latency because it represents the all of the functioning axons (Nandedkar, 2010). When fixed distances are used, the distance between the stimulating cathode and the E1 electrode is predefined, whereas with landmark-based distances, the stimulating cathode and the E1 electrode are placed at predefined landmarks. Because limb lengths vary among individuals, landmark-based distances also vary and, thus, require normal control values for every possible distance. To overcome this, the latency value is converted into a conduction velocity value so that only a single control value is required. However, this approach introduces problems. Because peak latency values do not represent the fastest conducting fiber, the calculated CV underestimates the fastest CV. Given that the range of CVs among the sensory axons contributing to the SNAP is about 25 m/s (Kimura et al., 1986) the average CV is about 12.5 m/s slower, which is a significant difference. In our EMG laboratories, we use fixed distances and peak latency values and compare the recorded value to age-dependent normal control values. Also, because

20 mV 20 mV

1 ms

1 ms

Fig. 10.8. Amplitude measurements. With biphasic responses (left), the amplitude is measured from the baseline to the first negative peak, whereas with triphasic responses (right), the amplitude is measured from the first positive peak to the first negative peak. Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 33.

NEUROMUSCULAR ELECTRODIAGNOSIS the distal segments of sensory axons are thinner, their myelinated segments are shorter, and distal extremity temperature is cooler, physiological slowing (Gilliatt and Thomas, 1960), especially among taller individuals (Campbell et al., 1981), must be considered before concluding that a mildly reduced CV is pathologic (especially when it is an isolated finding).

ORTHODROMIC VS ANTIDROMIC TECHNIQUES The terms orthodromic and antidromic refer to the direction of AP propagation following nerve stimulation in relation to the direction they propagate physiologically. With orthodromic conduction, the APs propagate as they would physiologically, whereas with antidromic techniques, they propagate in the direction opposite to their physiological direction. These terms only apply to sensory NCS because motor NCS can only be performed orthodromically. Except for the digital sensory NCS, most sensory NCS are performed antidromically. Digital sensory responses can be collected orthodromically (stimulate digit and record wrist) or antidromically (stimulate wrist and record digit). With mixed NCS (palmar NCS and plantar NCS), the sensory axons are assessed orthodromically and the motor axons are assessed antidromically. When the interelectrode distance is constant, the peak latency value is identical regardless of whether an antidromic or orthodromic technique is utilized (Cohn et al., 1990). However, the amplitude varies with the distance between the axons and the E1 electrode. Thus, regarding digital sensory responses, the amplitude is much larger using an antidromic technique because there is much less tissue between the digital nerves and the E1 electrode as opposed to wrist recording. One minor disadvantage of the antidromic technique is that wrist stimulation evokes both sensory and motor nerve fiber APs and the resultant CMAP (motor artifact) can interfere with SNAP recording. However, this is easily remedied by shifting the recording electrodes distally 0.5–1.0 cm and adjusting the normal latency value accordingly (0.2 ms per cm). Although the orthodromic technique does not generate motor artifacts, the lower SNAP amplitude is a major disadvantage.

SENSORY NCS UTILITY Because the sensory NCS assess the cell bodies and postganglionic axons but not the preganglionic axons, they are normal with radiculopathies and myelopathies. Thus, SNAPs have localizing value—when abnormal, they indicate a ganglionic (sensory neuronopathy) or postganglionic (plexopathy; neuropathy) localization. Their small size renders them more sensitive to axon loss than CMAPs

259

and their poor recovery makes them useful for identifying remote lesions following CMAP normalization. The sensory NCS have a significant number of limitations. Due to their smaller size, triphasic morphology, and wider range of conduction velocities (Kimura et al., 1986), they are more susceptible to physiologic temporal dispersion and, thus, cannot as easily assess long nerve segments. Also due to their small size, they are much more susceptible to technical errors and they overestimate lesion severity. Acutely, when 50% of the axons of a nerve are damaged, although the CMAP amplitude is about 50% smaller, the SNAP amplitude is usually about 90% smaller or even absent (Ferrante, 2018). Finally, their small size renders them susceptible to body habitus issues (adipose; thick digits; edema). This explains why antidromic digital sensory responses recorded from females (thinner digits) are higher in amplitude than those recorded from males (thicker digits) (Bolton and Carter, 1980). Consequently, mildly reduced antidromic SNAPs in a male with thick digits should be considered normal, assuming the other digits display a similar reduction. Due to their superficial location, sensory response abnormalities can arise from long forgotten, trivial trauma. After the age of 60 years, the lower extremity SNAPs may be absent. After the age of 70 years, this is quite common (Tavee et al., 2014). Also, because sensory NCS do not assess the nerve segment distal to the surface electrodes, when the sensor symptoms of a polyneuropathy are limited to the distal foot, clinically, the sural and superficial fibular SNAPs can be normal.

Mixed nerve conduction studies With mixed NCS (palmar and plantar NCS), motor and sensory axons are simultaneously stimulated, thereby generating compound electrical potentials composed of both motor and sensory nerve fiber APs. Because the stimulating electrodes are positioned distal to the recording electrodes, the sensory fiber contribution is orthodromic and the motor fiber contribution is antidromic. In our EMG laboratories, we record peak latency values and either baseline-to-peak or peak-to-peak amplitudes, depending on response morphology.

The EDX manifestations of demyelination and axon loss The EDX examination assesses the larger, more heavily myelinated axons. It does not assess the lightly myelinated or unmyelinated axons. In order to identify focal nerve lesions, current must past through the lesion. With focal demyelination, the lesion remains focal and, therefore the stimulating and recording electrodes must straddle the lesion for it to manifest. This allows the focal

260

M.A. FERRANTE

demyelination to be localized by stimulating the nerve at multiple sites. With focal axon disruption, however, the lesion does not remain focal. Rather, it extends distally due to Wallerian degeneration. As a result, all of the stimulation sites generate a similar appearing response. These two pathologies (demyelination and axon loss) are associated with three pathophysiologies, all of which have unique EDX manifestations. In order of severity, they are: (1) demyelinating conduction slowing (DMCS), both uniform and nonuniform, (2) demyelinating conduction block (DMCB), and (3) axonal conduction failure. In the presence of myelin, the TMC is significantly reduced (because only the nodal membrane is charged) and, hence, AP propagation is significantly accelerated (termed saltatory conduction). The myelin also limits Na+ efflux through nongated channels, thereby allowing the AP to advance further down the axon between rejuvenations (nerve safety factor). Because focal myelin loss increases the amount of exposed membrane, it increases the TMC and, hence, decreases AP propagation speed (DMCS). When the axons of a nerve are demyelinated to the same degree, they are equally slowed (uniform DMCS) and, therefore, the individual APs composing the response maintain their temporal relationship with each other. Consequently, the waveform morphology is essentially unaffected. Uniform DMCS is commonly seen in early carpal tunnel syndrome. Conversely, when DMCS only involves some of the axons or involves them to different degrees, nonuniform slowing (nonuniform DMCS) results waveform morphology dispersion. Temporal dispersion affects the amplitude to a greater extent than the negative AUC. Therefore, with focal nonuniform DMCS, negative AUC is a more accurate severity assessor than amplitude. The dispersion also increases the overlap of the negative and positive phases of individual APs, thereby causing some signal loss. Nonuniform demyelination is commonly observed with ulnar neuropathies at the elbow. With more profound myelin loss, Na+ current leakage increases to the point that the advancing AP is lost (DMCB). As a result, the amplitude and negative AUC response values recorded from the affected nerve are reduced in proportion to the number of blocked APs. With DMCB, temporal dispersion is not observed (signal is lost, not dispersed). By stimulating the nerve at multiple sites, focal DMCS and DMCB can be localized (the responses appear normal with stimulation below the lesion and abnormal with stimulation above the lesion) (see Fig. 10.9; Ferrante and Tsao, 2020). When the DMCB lies proximal to the most proximal stimulation site, it goes unrecognized because current does not traverse it. When the DMCB lies distal to the most distal stimulation site, all the responses demonstrate

5 mV

5 ms

Amp mV 4.0 3.4 3.5 .750

Fig. 10.9. Focal DMCB of the radial nerve at the spiral groove. Note that stimulation below the spiral groove (forearm-trace 1; elbow-trace 2; below spiral groove-trace 3) generates identical-appearing responses (except for onset latency differences), whereas stimulation above the spiral groove (trace 4, the lowest trace) generates a response with reduced signal (amplitude, negative AUC, and negative phase duration). Thus, the lesion is located between the third and fourth stimulation sites. Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 37.

5 mV

5 ms

Lat ms

Amp mV

2.7

4.5

6.9

.750

13.4

.667

Fig. 10.10. Carpal tunnel syndrome (CTS) with elements of axon loss (the distal motor response is reduced; 4.5 mV), DMCS (the two proximal responses show dispersion), and DMCB (indicated by the waveform) morphology differences between the below carpal tunnel response and the above carpal tunnel response. Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 38.

equivalent amounts of signal loss, mimicking axon loss. When it is possible to stimulate below the lesion, these two possibilities can be differentiated. For example, although carpal tunnel syndrome typically begins as DMCS and progresses to axon loss, infrequently there is a component of DMCB. When this occurs, stimulation below the carpal tunnel identifies the DMCB component, thereby localizing the lesion to the carpal tunnel (see Fig. 10.10; Ferrante and Tsao, 2020).

NEUROMUSCULAR ELECTRODIAGNOSIS

R

S

S

R

S

261

S

Fig. 10.11. Wallerian degeneration causes distal expansion of the lesion. Prior to Wallerian degeneration (left), the lesion is focal (shown in red) and the distal stumps conduct normally (solid lines), whereas after Wallerian degeneration (right), the lesion expands distally (shown in red) and the distal stumps are no longer excitable (broken lines). Consequently, prior to Wallerian degeneration, axon loss is localizable in the same way that DMCB is localizable (the proximal and distal responses appear differently). Once Wallerian degeneration is complete, both responses are equally diminished. Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 39.

With axon disruption, the APs cannot traverse the lesion (similar to DMCB). Initially, because the distal axon stumps do not undergo Wallerian degeneration instantaneously, the lesion is transiently focal and mimics DMCB because stimulation below the lesion produces a normal response, whereas stimulation above the lesion produced a smaller response (see Fig. 10.11). Because the NMJs degenerate before the axons, CMAPs show response decrement before the SNAPs (Gilliatt and Hjorth, 1972). In general, the size of the CMAP begins to decrease around day 3 and reaches its trough by day 7, whereas the size of the SNAP begins to decrease around day 6 and reaches its trough by day 11 (Wilbourn, 1983). To avoid mislocalization, NCS are typically not performed before day 11. For example, when a patient with numbness and weakness related to a brachial plexopathy involving the lower trunk is studied on day 6, the pattern of low amplitude ulnar (recording abductor digiti minimi) and radial (recording extensor indicis) CMAPs and normal ulnar SNAPs would mislocalize the lesion to the intraspinal canal (e.g., C8 radiculopathy).

SPECIAL STUDIES H reflex studies During an H reflex study, two CMAPs are collected, an M wave and an H wave. Both are elicited by stimulating the tibial nerve in the popliteal fossa, with the cathode positioned proximal to the anode, while recording from the gastrocnemius-soleus complex. The use of longer duration, lower intensity stimuli favor the activation of 1a sensory axons over motor axons. Like other stimulator-induced APs, propagation is bidirectional. Because the recording electrodes are positioned over the gastrocnemius-soleus muscle complex, the distally propagating sensory APs do not generate a SNAP on

the monitor. The proximally propagating sensory APs reach the S-1 segment of the spinal cord and activate LMNs within that segment that, in turn, generate APs that propagate distally along their motor axons to produce a gastrocnemius-soleus CMAP. This is the H-wave and it normally has an amplitude value exceeding 1 mV and an onset latency below 35 ms. Like all CMAPs, the amplitude of the H wave is more sensitive for identifying axon loss than is its latency. However, once the stimulus intensity exceeds the activation threshold for the tibial motor axons, APs also propagate bidirectionally along the tibial motor axons. The distally propagating motor APs generate a gastrocnemius-soleus CMAP, whereas the proximally propagating motor APs collide with the distally propagating motor APs (generated by the LMNs activated by the sensory Ia fibers). The collisions result in diminution of the H-wave and, as the stimulus intensity is increased further, eventually results in complete loss. Thus, as the stimulus intensity increases, there is initially no response, followed by the appearance of an H-wave, followed by a maximal H-wave response, followed by the presence of both an H-wave diminishing size and an M-wave, followed by solely an M-wave (Fig. 10.12). The M wave is the gastrocnemius-soleus CMAP and has the same utility as other CMAPs and, hence, should also be maximized. It is useful in estimating the severity of tibial neuropathies, sciatic neuropathies, and sacral plexopathies involving S1-derived plexus fibers. Clinically, H wave studies are especially helpful in detecting early polyneuropathies and S1 radiculopathies. Unfortunately, they may be bilaterally absent among normal individuals over the age of 60 years and, for this reason, their bilateral absence is of unclear significance. They may also be absent in the setting of obesity or large body habitus.

262

M.A. FERRANTE R

S1 root (tibial nerve)

2 mV

Fig. 10.12. H-reflex study technique with resultant H and M waves. The use of longer duration, lower intensity stimuli favor the activation of 1a sensory axons, which initially generates the H wave. As stimulation intensity increases, both sensory and motor axons are activated. The distally propagating motor axon APs generate an M wave, whereas the proximally propagating motor axon APs collide with the distally propagating motor axon APs generated by the proximally propagating sensory axon APs, thereby decreasing the size of the H wave. With further stimulation increases, the H wave disappears and the M wave is maximized. See text for further details. Reproduced with permission from Ferrante MA (2012). What we measure and what it means, American Association of Neuromuscular & Electrodiagnostic Medicine [AANEM], p. 74.

F wave studies F waves are elicited by the supramaximal stimulation of a motor nerve, distally, with the cathode oriented proximal to the anode. This induces bidirectionally propagating APs along the motor axons. Those traveling distally produce an M wave, whereas those traveling proximally reach the LMN cell bodies located within the spinal cord. Depending on their state of excitation at the time of arrival of the motor axon APs, backfiring of the LMN may occur. In general, 5% to 10% of the LMN pool is activated, resulting in a delayed CMAP (the F wave) that is about 5% to 10% of the size of the M wave (Fig. 10.13). Unlike H waves, F waves can be elicited from any motor nerve from which an M wave is elicitable. Theoretically,

F waves should be useful for detecting proximal nerve lesions. However, because only the latency is recorded, only the fastest conducting fiber is reflected. Thus, like other latency measurements, they are insensitive to focal axon loss. They are also insensitive to focal demyelination because sparing of just one of the faster conducting axons would generate an F wave with a normal latency. Consequently, they typically do not identify processes that have not already been detected during basic NCS. Their primary value is in the setting of early Guillain– Barre syndrome although, even then, the presence of focal dispersion (DMCS), low amplitude sensory and motor responses (DMCB and axon loss), and neurogenic recruitment (DMCB and axon loss) still render them superfluous.

NEUROMUSCULAR ELECTRODIAGNOSIS 5 mV

500 mV

RECORD

263

5 ms

Fig. 10.13. F waves. The 8 traces show supramaximal M waves all with the same morphologies and F waves, all of which have different morphologies. Reproduced with permission from Ferrante MA (2012). What we measure and what it means, American Association of Neuromuscular & Electrodiagnostic Medicine [AANEM], p. 77.

5 mV

RECORD Stimulator impedance high

Fig. 10.14. 2-Hz repetitive nerve stimulation study in a patient with anti-MuSK AB-positive myasthenia gravis showing significant decrement on the baseline train.

Repetitive nerve stimulation studies Repetitive nerve stimulation (RNS) studies are performed in patients with suspected NMJ disorders. Low-frequency RNS (e.g., 2–3 Hz) identifies postsynaptic disorders (e.g., myasthenia gravis), whereas high-frequency RNS (e.g., 40–50Hz) identifies presynaptic disorders (e.g., Lambert–Eaton myasthenic syndrome). The stimulation and recording techniques are identical to those of individual motor NCS except that trains of motor responses are collected and each train is assessed for decrement (postsynaptic disorders of NMJ transmission) and increment (presynaptic disorders of NMJ transmission) (Figs. 10.14 and 10.15). In our EMG laboratories, when screening for postsynaptic disorders, we use a stimulation rate of 2 Hz to record a baseline CMAP train, after which the patient is exercised and subsequent trains are collected over the next 5 min, seeking response decrement. We use an exercise period of 60s if the baseline train appears normal and

15 s if the baseline train shows decrement (to avoid missing postexercise facilitation). In our EMG laboratories, we exercise the muscle generating the first low motor response for 10 s and then immediately restimulate the nerve looking for an amplitude increment (Lambert test). When an increment is identified, we perform high-frequency stimulation (40Hz) for several seconds.

THE NEEDLE EMG EXAMINATION The needle EMG assesses the larger, more heavily myelinated axons and the electrical potentials it records are muscle fiber APs, including those from single muscle fibers (e.g., endplate spikes; fibrillation potentials) and from groups of muscle fibers (e.g., complex repetitive discharges), MUAPs, and groups of motor units (e.g., grouped repetitive discharges; myokymia). The AHCs innervating the commonly studied limb muscles are arranged in vertical columns that span at

264

M.A. FERRANTE Table 10.1 Nerve root innervation of commonly studied upper extremity muscles Anterior primary rami Proximal nerve innervation Rhomboids (dorsal scapular) Spinati (suprascapular) Deltoid (axillary) Biceps (musculocutaneous) Brachialis (musculocutaneous)

Fig. 10.15. 40-Hz repetitive nerve stimulation study in a patient with Lambert–Eaton myasthenic syndrome, a presynaptic neuromuscular transmission disorder, showing significant increment. Reproduced with permission from Ferrante MA (2012). What we measure and what it means, American Association of Neuromuscular & Electrodiagnostic Medicine [AANEM], p. 99.

least two spinal cord segments. Consequently, each muscle is innervated by at least two nerve roots and, therefore, these muscles belong to more than one myotome (myotomal overlap). The nerve root innervation of commonly studied upper and lower extremity muscles is provided in myotomal charts (Tables 10.1 and 10.2; Ferrante and Tsao, 2020). During the NEE, three types of muscle activity are assessed: (1) insertional activity, which follows needle electrode advancement into muscle tissue in the relaxed state, (2) spontaneous activity, which is sought between advancements in relaxed muscle, and (3) voluntary activity, which is assessed during muscle activation. By far, the most important and most challenging part of the needle EMG is MUAP analysis (morphology, recruitment, and stability). For discussion purposes, these three types of activity are best discussed separately although, in practice, they are not performed sequentially. For example, with a screen sensitivity of 50 mV/division, the needle electrode is repeatedly advanced (in 1.5–2 mm increments) and then held motionless for approximately 2 s. With each advancement, insertional activity is assessed, and with each pause, spontaneous activity is sought. The screen sensitivity is then changed to 200 mV/division and the patient gently activates the muscle so that voluntary activity can be assessed. Alternatively, especially with deeper muscles, the muscle can be gently activated and the electrode inserted until voluntary activity is identified, which is then assessed first. Whenever spontaneous activity is perceived during the activation phase (e.g., the sound of fibrillation potentials) or whenever an abnormal

C5 C6 C7 C8 T1

Radial nerve innervation Brachioradialis Triceps Anconeus Extensor carpi radialis Extensor pollicis brevis Extensor indicis Median nerve innervation Pronator teres Flexor carpi radialis Flexor pollicis longus Pronator quadratus Abductor pollicis brevis Ulnar nerve innervation Flexor carpi ulnaris Flexor digitorum profundus (D4,D5) Abductor digiti minimi Adductor pollicis First dorsal interosseous Posterior primary rami Cervical paraspinal muscles High thoracic paraspinal muscles Predominant contribution Sometimes significant contribution Minor contribution

appearing MUAP appears during the resting phase, the screen sensitivity is changed and the intruding activity assessed.

Resting activity INSERTIONAL ACTIVITY Insertional activity is assessed during the insertional phase of the needle EMG study. The burst of activity

NEUROMUSCULAR ELECTRODIAGNOSIS

265

Table 10.2 Nerve root innervation of commonly studied upper extremity muscles Anterior primary rami Proximal nerve innervation Iliacus (femoral nerve) Adductor longus (obturator nerve) Vastus lateralis (femoral) nerve) Rectus femoris (femoral nerve) Tensor fascia lata (superior gluteal nerve) Gluteus medius (superior gluteal nerve) Gluteus maximus (inferior gluteal nerve)

L2

L3

L4

L5

S1

S2

Sciatic nerve innervation Semitendinosus/semimembranosus (tibial division) Biceps femoris, long head (tibial division) Biceps femoris, short head (peroneal division) Common peroneal nerve innervation Tibialis anterior (deep peroneal nerve) Extensor hallucis (deep peroneal nerve) Peroneus longus (superficial peroneal nerve) Extensor digitorum brevis (deep peroneal nerve) Tibial nerve innervation Tibialis posterior Flexor digitorum longus Gastrocnemius, lateral head Gastrocnemius, medial head Soleus Abductor hallucis Flexor hallucis brevis Abductor digiti quinti pedis Posterior primary rami Lumbar paraspinal muscles High sacral paraspinal muscles Predominant contribution Sometimes significant contribution

precipitated by each needle electrode advancement within the muscle is termed insertional activity and reflects mechanical depolarization of muscle fibers by the electrode. Its presence indicates muscle tissue viability. Its quantity which is proportional to the number of muscles depolarized, normally ceases within 300–400 ms following electrode advancement. Decreased insertional activity can be seen with disorders producing muscle fiber loss (fibrofatty replacement), silent contractures (muscle cramps without associated electrical activity), or loss of muscle excitability (e.g., periodic paralysis). The term increased insertional activity is misleading because the activity referred to actually follows the burst of insertional activity (i.e., it occurs during the pause phase). Because this activity is provoked by electrode advancement, it may better be termed provoked activity. Physiologic

examples include snap-crackle-pop and Wiechers– Johnson syndrome; insertional positive sharp waves are a pathologic example.

Snap-crackle-pop Snap-crackle-pop is a form of normal insertional activity that follows the insertional activity described above (Wilbourn, 1982). It typical lasts just a few seconds and consists of a series of individual electrical potentials, each of which has a different morphology and sound. Because it appears during the initial portion of the pause period, it is provoked activity. It is most commonly observed in the calf muscles of younger, muscular males. The different waveform morphologies and its transient

266

M.A. FERRANTE

nature allow this benign activity to be distinguished from pathologic forms of spontaneous activity. Wiechers–Johnson syndrome This type of electrical activity, which is characterized by runs of insertional positive waves provoked by electrode advancement, tends to have a wide distribution (Wiechers and Johnson, 1982) and may represent a forme fruste of myotonia congenita (Mitchell and Bertorini, 2007).

SPONTANEOUS ACTIVITY Spontaneous activity is assessed during the resting phase with the needle held motionless. Unlike the transient nature of insertional activity, spontaneous activity is sustained. Some forms of spontaneous activity are physiologic (e.g., endplate activity), whereas other forms are pathologic (e.g., fibrillation potentials). The waveform morphology of spontaneous activity reflects its site of generation. When the activity is generated above the arborization point (i.e., where the intramuscular motor axon arborizes), the activity resembles MUAPs (e.g., fasciculation potentials), whereas when it is generated below this point, it resembles single muscle fiber APs (e.g., fibrillation potentials). Due to the scope of this chapter, only endplate activity and fibrillation potentials are reviewed. For discussions of other forms of spontaneous activity, the interested reader is referred to any of a large number of comprehensive EMG textbooks (Ferrante, 2018).

ENDPLATE ACTIVITY Endplate activity (endplate noise), a form of physiologic spontaneous activity, occurs in the endplate region and is composed of miniature endplate potentials (MEPPs) and endplate spikes. The MEPPs reflect the spontaneous release of ACh vesicles from the presynaptic membrane, whereas endplate spikes reflect mechanical depolarizations by the needle electrode of intramuscular terminal nerves (Blight and Precht, 1980; Brown, 1984). Unlike fibrillation potentials, endplate spikes are never regular and never observed in isolation. MEPPs generate a hissing sound akin to the sound of an empty sea shell or to a radio tuned between stations, whereas endplate spikes generate a sound similar to the sputtering sound of fat frying in a pan. Clinically, when the needle electrode is in the endplate region, patients typically report pain.

FIBRILLATION POTENTIALS Following denervation, extrajunctional acetylcholine receptors (AChRs) appear, the RMP decreases and

begins to oscillate, and the oscillations slowly increase in size. Around day 21, the RMP is around
60 mV (about 5 mV below the depolarization threshold) and the oscillations are about 5 mV in size. Consequently, each oscillation triggers a muscle fiber AP, termed a fibrillation potential (Thesleff, 1974). Fibrillation potentials have two waveform morphologies—the spike form and the positive sharp wave form—both of which indicate muscle fiber denervation (Fig. 10.16; Ferrante, 2012). The spike form typically has a triphasic morphology, but can be biphasic (negative–positive or positive– negative). The positive sharp wave form is biphasic (positive–negative) and has a higher amplitude and a longer duration (10–30 ms or longer). Both forms indicate muscle fiber denervation that occurred at least 21 days earlier. A third form of denervation potential precedes these two forms by approximately 1 week. Around day 14, before the RMP reaches
60 mV and the oscillations reach 5 mV, the mechanical depolarization associated with needle electrode advancement may produce transient depolarization resulting in nonsustained denervation potentials with a positive sharp wave form termed insertional positive sharp waves. Because one motor axon innervates a large number of muscle fibers, disruption of a single axon results in a large number of fibrillation potentials. Consequently, needle EMG is much more sensitive than motor NCS for identifying motor axon loss. Once reinnervation occurs, however, this sensitivity is lost. Finally, the quantity of fibrillation potentials does not reflect lesion severity but, rather, the timing of the study. The amplitude of the muscle fiber AP is proportional to the diameter of the muscle fiber. Thus, in the acute setting, prior to muscle fiber atrophy, fibrillation potential amplitude may be identical to MUAP amplitude. Over time, as the denervated muscle fiber atrophies, fibrillation amplitude decreases. By 1 year, they are typically below 100 mV (Kraft, 1990).

VOLUNTARY ACTIVITY Voluntary activity is assessed during the activation phase, when the patient lightly activates the muscle under study so that 2–4 MUAPs appear on the monitor. The morphology (amplitude; duration; phases and turns) of approximately 20 MUAPs are assessed; MUAP recruitment and stability are also evaluated. Because MUAP analysis requires considerable knowledge and experience, novice providers tend to spend more time seeking spontaneous activity and less time in the activation phase, whereas more seasoned providers spend most of their time in the activation phase.

NEUROMUSCULAR ELECTRODIAGNOSIS

267

Fig. 10.16. Upper 2 traces show the spike form of fibrillation potential, whereas the lower 2 traces show the positive sharp wave form of fibrillation potential. Modified and reproduced with permission from Ferrante MA (2012). What we measure and what it means, American Association of Neuromuscular & Electrodiagnostic Medicine [AANEM], pp. 106–107.

Morphology Both extrinsic (amplitude; duration) and intrinsic (turns; phases) properties are assessed. Most MUAPs are 5–15 ms in duration (typically 8–10 ms), 1–3 mV in amplitude, and have 4 or fewer phases. Because MUAP size is proportional to the number of muscle fibers composing its motor units and because motor unit size varies among muscle, MUAP size also varies with the muscle under study. Because normal aging is associated with AHC loss, followed by reinnervation of the denervated muscle fibers through collateral sprouting, MUAP size increases with age. Thus, when assessing MUAP size, patient age and the specific muscle under study must be considered. MUAP amplitude The MUAP amplitude (measured peak-to-peak) is the most variable and, therefore, the least reliable measurement. Its value reflects a number of factors, including technical factors and nontechnical factors, such as the number, diameter, and density of the muscle fibers composing the motor unit; patient age; and muscle fiber AP synchrony. Of these, muscle fiber density is the most important. Smaller MUAPs associated with lower levels of contractile force may have amplitudes below 1 mV,

whereas those associated with greater levels of contractile force are larger. In general, the latter do not exceed 3 mV. Because electrical signals exponentially decrease in amplitude as they travel toward the recording electrode, only those muscle fibers adjacent to the needle electrode contribute to the MUAP. This is why the density of the muscle fibers adjacent to the needle electrode primarily dictates MUAP amplitude (Swash and Schwartz, 1981). Because the muscle fibers of an individual motor unit are intermingled with those of up to 50 other motor units (Burke et al., 1973), MUAP amplitude typically reflects only 1 to 6 muscle fibers (Nandedkar et al., 1985; Stalberg and Sanders, 2009). Following reinnervation, even significant increases in the innervation ratio do not change MUAP amplitude because the territory in which the muscle fibers of the motor unit are located is intermingled with so many other motor units that muscle fiber density is often unaffected (Burke et al., 1973). MUAP duration Because the muscle fiber APs composing an MUAP arrive at the electrode at slightly different times, the duration of an MUAP (5–15 ms) is much longer than a muscle fiber AP (2–3 ms). Unlike MUAP amplitude, MUAP

268

M.A. FERRANTE

duration does not exponentially decay over distance and, therefore, even distant muscle fibers of the motor unit, including those on the other side of the beveled surface of the electrode, contribute to MUAP duration (King et al., 1997). Unlike MUAP amplitude, which is nonGaussian, MUAP duration has a Gaussian distribution, which makes it much easier to identify MUAPs with abnormally short or long durations. Because the innervation ratio of a given muscle is relatively constant, its MUAP duration is also relatively constant. In general, regarding upper extremity muscles, the longest duration MUAPs are found in the triceps, the shortest duration MUAPs are found in the brachioradialis and flexor digitorum profundus, and the MUAP durations of the other muscles are intermediate to these extremes (the biceps muscle is often at the shorter end of the intermediate muscle group). Whenever the observed MUAP duration deviates from this lineup, the same muscle on the contralateral side should be assessed to determine if the deviation is abnormal. This relationship is easily appreciated (Fig. 10.17; Ferrante and Tsao, 2020). Among lower extremity muscles, the vastus lateralis typically shows the longest duration MUAPs, the iliacus and the short head of the biceps femoris show the shortest duration MUAPs, and the other commonly studied muscles show intermediate values (the gluteus medius is at the shorter end of the intermediate group).

200 mV 10 ms

Fig. 10.17. This illustration shows the spectrum of MUAP duration values among commonly studied upper extremity muscles (taken from the author). The brachioradialis (left) has the shortest value (about 6 ms), the triceps (right) has the longest value (about 20 ms), and the remainder have an intermediate value (first dorsal interosseous, center; about 12 ms). Reproduced with permission from Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization. Electromyography, Demos Medical, p. 51.

MUAP phases and turns A phase is defined as the portion of the MUAP between two baseline crossings and is either negative (above the baseline) or positive (below the baseline), whereas a turn is a change in direction that does not cross the baseline and is at least 100 mV in size. These two features reflect the number of muscle fiber APs contributing to the MUAP and their synchrony. Up to 15% of normal MUAPs may be polyphasic (more than 4 phases); this value is higher (up to 25%) for the deltoid and tibialis anterior muscles (Buchthal, 1977; Campbell, 1999).

Recruitment The innervation ratio of a muscle fiber is proportional to the contractile force that it generates. The firing frequency of the motor unit also contributes to the amount of contractile force it generates. Spatial recruitment refers an increase in the number of motor units activated and temporal recruitment refers to an increase in their firing frequency. The Henneman size principal states that motor units are recruited based on their size, beginning with the smallest one; this ensures a smooth increment in force production. For the same reason, the firing frequency also starts low and increases slowly. The firing frequency of the first recruited motor unit is termed the onset frequency (5–8 Hz) (Conwit et al., 1998). As effort is increased, the motor unit fires faster (temporal recruitment). For most limb muscles, once the firing frequency reaches 10–12 Hz, a second motor unit appears (spatial recruitment). The firing frequency of the first MUAP when the second MUAP appears is termed the recruitment frequency (Petajan and Phillip, 1969). Spatial and temporal recruitment increase in unison until spatial recruitment maximizes. This occurs when there are no more motor units left to recruit. At this point, temporal recruitment increases in isolation (up to about 40 Hz). The contractile force when temporal recruitment maximizes is three or more times greater than the value when spatial recruitment maximizes (Ferrante and Wilbourn, 2015). As more MUAPs are recruited, MUAP overlap causes individual MUAPs to become less discernible (interference pattern). When MUAP overlap is complete (individual MUAPs are no longer discernible), the interference pattern is complete. At 30% maximum contraction, MUAP firing frequencies above 20Hz are not discernible because increases in spatial recruitment result in full interference, rendering individual MUAPs undiscernible (Dorfman et al., 1988; Petajan, 1991). When individual MUAPs are fully discernible, the interference pattern is discrete. With mixtures of the two, the interference pattern is incomplete (reduced). Importantly, all three of these

NEUROMUSCULAR ELECTRODIAGNOSIS interference patterns are seen among normal individuals and in all three, spatial recruitment and temporal recruitment are concordant. With pathology, however, this relationship becomes discordant. The relationship between temporal and spatial MUAP recruitment can be assessed by calculating the recruitment ratio (the frequency of the fastest firing MUAP divided by the total number of MUAPs). This value is normally 5 to 7. Another way to assess MUAP recruitment is based on the rule of fives (Daube, 1991). In general, the first MUAP appears on the screen at approximately 5 Hz and when its firing frequency reaches about 10 Hz, a second MUAP appears. At this point, the recruitment ratio is 5 (10 Hz/2 MUAPs ¼ 5). With further effort, the frequencies of the two recruited MUAPs increase further and, around 15 Hz, a third MUAP appears (the recruitment ratio ¼ 15 Hz/3 MUAPs ¼ 5). Thus, the recruitment ratio is maintained around 5. Using this method, a recruitment ratio value exceeding 10 implies that spatial recruitment is not keeping pace with temporal recruitment (MUAP dropout). This is termed neurogenic recruitment (discussed below). Although some authors refer to this as decreased or reduced recruitment, poor effort (either voluntary or involuntary) also generates a limited number of MUAPs. With poor effort, however, spatial recruitment and temporal recruitment are both reduced (concordance), whereas with neurogenic recruitment, spatial recruitment is reduced and temporal recruitment is normal (discordance). Because neurogenic recruitment signifies motor unit loss, it can be observed with lesions producing AHC loss, axon loss, and DMCB, all of which reflect disease proximal the arborization point of the motor axon. Conversely, with disorders distal to the arborization point (those involving the terminal motor branches, the NMJs, or the muscle fibers), the innervation ratio of the motor unit is reduced. This is termed disintegration of the motor unit. As a result, these motor units generate smaller contractile forces and, hence, a greater number of them are required to generate a given force. Thus, disorders associated with disintegration of the motor unit show more rapid MUAP recruitment, termed early recruitment. With upper motor neuron (UMN) disorders, there is a reduction of both spatial and temporal recruitment (concordance), similar to poor effort. However, with UMN disorders, the examiner usually appreciates that the degree of force being generated by the patient is better than expected for the degree of recruitment. Stability NMJ transmission time normally varies among the NMJs of a motor unit, even for sequential discharges of the same NMJ. As a result, the MUAP morphology of a given motor unit normally changes from discharge to

269

discharge. Although these changes are apparent during single fiber EMG, they are not apparent during routine needle EMG, hence, MUAP morphology does not appreciably change with sequential firing of a normal motor unit. However, with impaired NMJ transmission, routine needle EMG may disclose MUAP morphology changes during sequential firing, termed moment-to-moment variation (MMV) (Lindsley, 1935; Lambert, 1966). Thus, when NMJ transmission disorders are suspected, the sequential firing of individual motor units should be assessed for variability.

REFERENCES Blight AR, Precht W (1980). “Spontaneous” quantal release of transmitter absent in vivo. Abst Soc Neurol Soc 6: 601. Bolton CF, Carter K (1980). Human sensory nerve compound action potential amplitude. Variation with sex and finger circumference. J Neurol Neurosurg Psychiatry 43: 925–928. Brown WF (1984). The physiological and technical basis of electromyography, Butterworth, London, pp. 339–368. Buchthal F (1977). Diagnostic significance of the myopathic EMG. In: P Rowland (Ed.), Pathogenesis of human muscular dystrophies. Proceedings of the fifth international scientific conference of the Muscular Dystrophy Association, Durango, Colorado. Excerpta Medica, Oxford, Amsterdam, pp. 205–218. Burke RE, Levine DN, Tsairis P et al. (1973). Physiological types and histochemical profiles in motor units of the cat gastrocnemius. J Physiol 234: 723–748. Campbell WW (1999). Essentials of electrodiagnostic medicine, Williams and Wilkins, Baltimore, 107. Campbell WW, Ward LC, Swift TR (1981). Nerve conduction velocity varies inversely with height. Muscle Nerve 4: 520–523. Cohn TG, Wertsch JJ, Pasupuleti DV et al. (1990). Nerve conduction studies: orthodromic versus antidromic latencies. Arch Phys Med Rehabil 71: 579–582. Conwit RA, Tracy B, Cowl A et al. (1998). Firing analysis using decomposition-enhanced spike triggered averaging in the quadriceps femoris. Muscle Nerve 21: 1338–1340. Daube JR (1991). AAEM Minimonograph #11: needle examination in clinical electromyography. Muscle Nerve 14: 685–700. Dorfman LJ, Howard JE, McGill KC (1988). Influence of contractile force on properties of motor unit action potentials: ADEMG analysis. J Neurol Sci 86: 125–136. Feinstein B, Lindegaard B, Nyman E et al. (1955). Morphologic studies of motor units in normal human muscles. Acta Anat 23: 127–142. Ferrante MA (2004). Brachial plexopathies: classification, causes, and consequences. Muscle Nerve 30: 547–568. Ferrante MA (2012). EMG: what we measure and what it means, American Association of Neuromuscular and Electrodiagnostic Medicine Publishing, Rochester, Minnesota.

270

M.A. FERRANTE

Ferrante MA (2018). Comprehensive electrodiagnostic medicine: principles and concepts with clinical correlations and case studies, Cambridge University Press, Cambridge, United Kingdom. Ferrante MA, Tsao BE (2020). EMG lesion localization and characterization: a case studies approach, Demos Medical (an imprint of Springer Publishing), New York. Ferrante MA, Wilbourn AJ (2015). The electrodiagnostic examination of peripheral nerve injuries. In: SE Mackinnon (Ed.), Nerve surgery. Thieme Medical Publishers, New York City, pp. 59–74. Gilliatt RW, Hjorth R (1972). Nerve conduction during Wallerian degeneration in the baboon. J Neurol Neurosurg Psychiatry 35: 335–341. Gilliatt RW, Thomas PK (1960). Changes in nerve conduction with ulnar lesions at the elbow. J Neurol Neurosurg Psychiatry 23: 312–320. Kimura J, Machida M, Ishida T et al. (1986). Relationship between size of compound sensory or muscle action potentials and length of nerve segment. Neurology 36: 647–652. King JC, Dumitru D, Nandedkar S (1997). Concentric and single fiber electrode spatial recording characteristics. Muscle Nerve 20: 1525–1533. Kraft GH (1990). Fibrillation potential amplitude and muscle atrophy following peripheral nerve injury. Muscle Nerve 13: 814–821. Lambert EH (1966). Defects of neuromuscular transmission in syndromes other than myasthenia gravis. Ann N Y Acad Sci 135: 367–384. Lindsley DB (1935). Electrical activity of human motor units during voluntary contraction. Am J Phys 114: 90–99. Mitchell CW, Bertorini TE (2007). Diffusely increased insertional activity: “EMG disease” or asymptomatic myotonia

congenita? A report of 2 cases. Arch Phys Med Rehabil 88: 1212–1213. Nandedkar S (2010). Motor and sensory nerve conduction: technique, measurements, and anatomic correlation. In: Neurophysiology and instrumentation, 57th annual meeting of the American Association of neuromuscular and electrodiagnostic medicine, Quebec City, Quebec, Canada. 1–8. Nandedkar SD, Sander DB, Stalberg EV (1985). Selectivity of electromyographic recording techniques: a simulation study. IEEE Trans Biomed Eng 23: 536–538. Petajan JH (1991). AAEM Minimonograph #3: motor unit recruitment. Muscle Nerve 14: 489–502. Petajan JH, Phillip BA (1969). Frequency control of motor unit action potentials. Electroencephlogr Clin Neurophysiol 27: 66–72. Stalberg EV, Sanders DB (2009). Jitter recordings with concentric needle electrodes. Muscle Nerve 40: 331–339. Swash M, Schwartz MS (1981). Neuromuscular diseases, Springer-Verlag, Berlin. Tavee JO, Polston D, Zhou L et al. (2014). Sural sensory nerve action potential, epidermal nerve fiber density, and quantitative sudomotor axon reflex in the healthy elderly. Muscle Nerve 49: 564–569. Thesleff S (1974). Physiological effects of denervation of muscles. Ann NY Acad Sci 228: 89–103. Wiechers DO, Johnson EW (1982). Syndrome of diffuse abnormal insertional activity. Arch Phys Med Rehabil 63: 538–539. Wilbourn AJ (1982). An unreported, distinctive type of increased insertional activity. Muscle Nerve 5: S101–S105. Wilbourn AJ (1983). How can electromyography help you? Postgrad Med 73: 187–195.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00016-9 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 11

Quantitative electrodiagnosis of the motor unit MARK B. BROMBERG* Department of Neurology, University of Utah, Salt Lake City, UT, United States

Abstract Electromyography (EMG) focuses on assessment of the motor unit (MU), and a given muscle has several hundred MUs, each innervating hundreds of muscle fibers. Assessment is limited by the recording radius of electrodes, 1–2 fibers with single-fiber electrodes and 7–15 fibers with concentric or monopolar electrodes. Routine qualitative EMG studies rely on observing MUs in free-run mode and qualitatively estimating common metrics. In contrast, quantitative EMG (QEMG) applied to routine studies includes assessment of individual MUs by software available in modern EMG machines with extraction of discrete values for common metrics, and also derived metrics. This results in greater precision and statistical interpretation. Other QEMG techniques assess muscle fiber density within the MU and time variability at the neuromuscular junction. The interference pattern can also be assessed. The number of MUs innervating a muscle can be estimated. Advanced signal processing, called near-fiber EMG, allows for extraction of underlying muscle fiber contributions to MU waveforms. It is also possible to use QEMG to make statistical probabilities of the state of a muscle as to whether normal, myopathic, or neuropathic. Time to acquire QEMG data is minimal. QEMG is most useful in situations where pathology is uncertain.

INTRODUCTION The role of electromyography (EMG) is assessment of the motor unit (MU), the final common pathway for muscle activation. Defined by Sherrington, the MU is the alpha motor neuron cell body, its axon, intramuscular terminal branches, and innervated muscle fibers (Sherrington, 1925) (Fig. 11.1). While simple in concept, it is challenging to study the MU directly. The anatomic MU (A-MU) as defined above cannot be readily viewed, except in experimental animal preparations, which has been done for a limited number of leg muscles (Edstrom and Kugelberg, 1968). Extrapolation of basic findings to human MUs is reasonable, but there are no data for MU features in large size muscles. The number of MUs innervating a muscle cannot be readily measured, but estimates in humans for distal muscles indicates 100–300 MUs, and likely more in large proximal muscles (Bromberg, 2020). Similarly, the number of

muscle fibers innervated by an MU cannot be easily measured, but are in the hundreds to thousands. EMG assessment focuses on whether a muscle is normal or has pathologic changes due to nerve fiber loss (neuropathic disorder) or damage to muscle fibers (myopathic disorder). Needle EMG studies are performed using a variety of electrode types to record an electric MU (E-MU), which is a limited view of the A-MU (Fig. 11.2). The most common EMG study (routine qualitative EMG) uses concentric or monopolar electrodes, and separation of one E-MU from another and analysis of waveform features (metrics) is by visualization on the EMG machine screen in real time mode, and metrics are reported in qualitative terms (increases or decreases, scaled as 1+ to 4+ in severity). While this approach is expedient and practical, there are limitations in diagnosing subtle clinical conditions. Most waveform metrics are within normal limits in both neuropathologic and myopathic

*Correspondence to: Mark B. Bromberg, MD, PhD, Department of Neurology, University of Utah, Salt Lake City, UT 84132, United States. Tel: +1-801-585-7575, Fax: +1-810-581-4192, E-mail: [email protected]

272

M.B. BROMBERG

Fig. 11.1. Composite figure showing: (A) Motor unit (MU) composed of alpha motor neuron, axon, terminal arborization, and innervated muscle fibers. (B) Muscle fibers from an MU based on glycogen depletion studies in cat; perimeter of a typical MU and distribution of glycogen negative fibers. (C) Area occupied by MU in muscle. Redrawn and modified with permission from Bromberg MB (2020). The motor unit and quantitative electromyography. Muscle Nerve 61: 131–142.

Fig. 11.2. Commonly used intramuscular EMG electrodes with uptake shapes and recording radiuses: (A) Concentric needle electrode; various sizes (diameters and lengths) available with corresponding small differences in recording radiuses. (B) Monopolar needle electrode; various sizes (diameters and lengths) with corresponding small differences in radiuses. (C) Single-fiber needle electrode. Modified with permission from Bromberg MB (2020). The motor unit and quantitative electromyography. Muscle Nerve 61: 131–142.

conditions, and thus aids to making a diagnosis include discrete metric values, additional derived metrics, and the ability to identify outlying values (Stewart et al., 1989; Stalberg et al., 1994). Quantitative EMG (QEMG)

represents several techniques where metrics are expressed as discrete values (millivolts, milliseconds, numbers). The most common QEMG technique is viewing E-MUs in a manner similar to routine EMG, but individual E-MU waveforms are captured by computer software, and metrics are listed in tables as discrete values. QEMG is possible because contemporary EMG machines acquire analogue voltage data and convert it to digital data for display, and have the capability of processing MU waveform signals in a variety of ways. The most basic operation is isolation of E-MUs and making measurements (Tankisi et al., 2020). With discrete data, statistical approaches can be used in the analysis to provide a probability as to whether E-MUs, and hence the whole muscle, is normal or pathologic (and whether neuropathic or myopathic). QEMG capabilities are available on all modern EMG machines. Another form of QEMG is focused study of elements of the E-MU using a restricted electrode, the single-fiber electrode. This allows for assessment of the density of innervated muscle fibers in an MU as fiber density (FD), and neuromuscular junction (NMJ) transmission variability in the form of jitter. While both of these studies require a special electrode and a focused study, recent signal processing capabilities allow extraction of muscle fiber contribution to E-MU waveforms, called near fiber EMG, to provide similar information during routine QEMG studies. The interference pattern at maximal contraction effort is frequently assessed qualitatively as full or reduced. A third form of QEMG assesses the composite contribution of MU recruitment (number of MUs) and E-MU waveforms in the form of analysis of turns and amplitude as the interference pattern increases with greater patient effort. Analysis of turns and amplitude allows for discrete data points to support neuropathic or myopathic changes.

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT There is no routine nerve conduction or EMG study that provides estimates of the number of MU innervating a muscle or group of muscles, as the compound muscle action potential (CMAP) amplitude does not fall linearly with motor axon loss as collateral reinnervation slows the decline, and amplitude may be at the lower limit of normal when
80% of axons are lost in slowly progressive neuropathic disorders. During EMG studies, when only a few (1–2) MU appear on the screen, there may be others in the muscle too distant to record. The technique of motor unit number estimation (MUNE) was developed to provide an assessment of the number of innervating motor axons to a muscle or group of muscles, but the value is an estimate and not a count. This chapter first reviews the physiology and pathology of MUs and how QEMG techniques enhance diagnostic information beyond what is obtained with routine qualitative EMG studies. Second, is a review of single-fiber techniques. Recent efforts using signal processing allows for collection of single nerve fiber contributions to the E-MU and the third section show how this can be incorporated into routine QEMG studies. Analysis of turns and amplitude are reviewed in the fourth section. The various MUNE techniques are reviewed, and an alternative technique, MUNIX, is included. The final section suggests clinical uses of these QEMG techniques.

ANATOMIC MOTOR UNIT The A-MU can only be assessed in experimental animals by glycogen depletion studies, where a single ventral root is dissected and stimulated repeatedly to deplete glycogen stores in muscle fibers of the innervated A-MU (Fig. 11.1). Sacrifice of the animal and processing of the muscle for histology with a glycogen stain indicates the perimeter, number of fibers and their distribution within a single A-MU. In rat and cat hindlimb muscles, the cross-sectional area of an A-MU occupies only a portion of the area of the whole muscle. Within an A-MU, few muscle fibers lie adjacent to each other, and the density of fibers varies somewhat within the MU’s territory (Edstrom and Kugelberg, 1968; Bodine-Fowler et al., 1990). At any given site in a human muscle (i.e., position of an EMG electrode’s recording surface), the territories of
20 A-MUs overlap. Studies in some human muscle using a special EMG electrode with multiple recording sites along the shaft supports a similar regional distribution of motor units within muscles (
10 mm in long and cross axes) as from the animal studies (Buchthal et al., 1959). Recent studies using special MRI techniques in humans confirm in the few muscles studied that A-MUs have a variety of shapes that occupy limited areas within the muscle (Birkbeck et al., 2020).

273

ELECTROPHYSIOLOGIC MOTOR UNIT The E-MU represents a portion of the A-MU dependent upon the recording (uptake) radius of the electrode used (Tankisi et al., 2020). The uptake radius reflects the distance over which contributing voltages from muscle fibers can be distinguished, and the recording radius is partially based on the geometry of the active electrode (E1) surface (Fig. 11.2). Since the distribution of muscle fibers in the A-MU varies over its area, random positioning of the E1 recording surface affects the E-MU waveform. Electrode types used for QEMG studies are singlefiber electrodes and more commonly concentric and monopolar electrodes. Single-fiber electrodes have a very small recording surface area and a recording radius of
300 mm, which includes action potentials from 1 to 3 muscle fibers of an MU. Concentric and monopolar electrodes differ in the geometry and recording surface areas, but have a similar recording radius of
1500 mm, which includes action potentials from 7 to 15 muscle fibers of an MU, but amplitude is influenced by the nearest 1 to 3 fibers within 0.5 mm of the electrode (Thiele and Bohle, 1978; Nandedkar et al., 1988a) (Fig. 11.1). Other electrodes are the macro-electrode and surface electrodes, used for quantitative MU research, but uncommonly in the clinic, and not considered.

MOTOR UNIT VIEWING AND ASSESSMENT During routine EMG studies, E-MUs are viewed at relatively weak patient contraction efforts, 10% to 15% of maximum contractions. E-MU assessments can be made visually in real time or quantitatively after MU isolation and analysis of digital data.

Routine E-MU viewing E-MUs are viewed from continuous sweeps across the computer screen, commonly at 10 ms/div along a 10 or 20 division screen (Fig. 11.3). This results in three to five defined higher amplitude E-MUs with relatively steep rise times, intermixed with less well defined MUs with shallow rise times. Although EMG machines convert analogue voltage data to digital data, waveforms appear on the screen as continuous data; thus, routine E-MU observation are under “analogue” condition (Tankisi et al., 2020). It is a challenge to reliably visualize and estimate E-MU metrics of amplitude, duration and complexity (number of turns and phases), and descriptions of increased or decreased values expressed as 1 to 4+ are highly subjective, and subject to a degree of prestudy bias (while a suspected diagnosis is necessary before starting an EMG study, it can bias assignment of 1 to 4 values).

274

M.B. BROMBERG

Fig. 11.3. Free-run EMG traces, 10 ms/div and 200 mV/div: (A) 10 division screen and (B) 20 division screen. Showing difficulty in discerning duration and complexity metrics; may be harder in 20 division screen as eyes are only able to visually focus on a portion of screen. Author’s figure.

E-MUs are commonly “focused” by moving the electrode to optimize rise time, resulting in a bias to viewing and hearing higher amplitude/louder sounding MUs, and lower amplitudes MUs which may have pathologic significance (low amplitude  complex MUs) can be missed (Barkhaus and Nandedkar, 1996).

E-MU isolation E-MU waveforms can be isolated for more detailed viewing by three methods; photographing individual analogue waveforms, using a voltage trigger and delay line to view individual analogue waveforms, or by matching waveform templates to isolate digital waveforms.

ISOLATION BY PHOTOGRAPHY With early analogue oscilloscopes, clear and separate E-MUs could only be made from very weak interference patterns. Visually separate E-MUs were photographed and measurements made manually, and resultant metric values are commonly used as reference limiting values during routine qualitative EMG studies (Buchthal et al., 1954; Buchthal, 1975). However, some MU metrics increase in value at higher and more common contraction efforts, and thus reference values from photographed E-MUs are biased to low amplitude MUs. The disadvantage of this method in a quantitative manner is that individual waveforms must be measured from photographs.

ISOLATION BY DELAY LINE An adjustable negative or positive voltage level can be set or adjusted such that only the negative or positive waveform component of one E-MU crosses the voltage level, and then triggers the rest of the waveform to be

displayed on another screen (Tankisi et al., 2020). However, the early portion of the waveform before the trigger would not be observed without a delay line that allows the whole triggered signal to be observed (Fig. 11.4). Since current digital EMG machines convert analogue voltages to digital format stored in memory, the delay represents looking into memory for earlier features of the waveform (Czekajewski et al., 1969). The advantage of this method is that individual waveforms can be observed from the screen during a study. The disadvantage of using this quantitative technique is that E-MUs are assessed individually; however, this methods can be used in a semiquantitative manner (discussed below).

ISOLATION BY TEMPLATE MATCHING All E-MUs recorded during a contraction are stored in digital format in the EMG machine. A template of each MU can be generated and compared to subsequent templates, and matching templates collected to isolate and identify individual E-MUs from an interference pattern at higher levels of contraction than can be assessed visually (as for photographing or use of delay line) (Stalberg et al., 1996) (Fig. 11.5). For each E-MU isolated, various statistical assessments can be carried out to ensure that the waveform is not contaminated by other waveforms. An averaged waveform from 50+ similar templates is obtained for each isolated E-MU, and quantitative metric values determined (amplitude, duration, phases/turns), and there is the option for the electromyographer to review the ensemble of waveforms for each E-MU. Advantages of this method are that three to six individual E-MU can be isolated from each contraction site within the muscle. In addition to routine metrics, derived and extrapolated metrics can be calculated. Quantitative data from 20+ MUs can be averaged for a muscle, and directly compared to normative data (discussed below). There are a number of techniques

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT

A

275

B1

Trigger Voltage

Progressive Time Delay 0 msec

B2 2 msec

B3 4 msec

B4 6 msec

Fig. 11.4. Trigger and delay line: (A) Free-run trace at 10 ms/div with trigger voltage set so that peak voltage of motor unit (MU) crosses trigger line. (B1) Segment of MU displayed after trigger, trace at 2 ms/div. (B2–B4) Display of longer MU segments from computer memory to show full MU. Author’s figure.

Templates

10sec ~100 sweeps 3-6 E-MUs/ sweep

New MU waveforms must match

Averaged MU

A

B

Fig. 11.5. Schematic of template matching method to decompose individual motor units (MU) from free-run traces. (A) Free-run sweeps with computer identifies MU templates 1–15. (B) Showing for MUs 5 and 1 how
50 templates are extracted and averaged. Author’s figure.

used to isolate and extract E-MUs which vary among EMG manufactures, but achieve similar isolation and metric measurement values (McGill, 2009).

ELECTROPHYSIOLOGIC MOTOR UNIT METRICS E-MU waveforms can be characterized by a number of metrics, some basic or traditional and obvious, and others derived from signal processing (Stalberg et al., 2019).

Metrics from routine qualitative studies are limited in number, and values are scaled subjectively (increased or decreased and 1 to 4+ in severity), while metrics from QEMG studies are derived from stored digital data and are expressed as discrete values (millivolts or milliseconds, numbers), which can be used for statistical analysis. Since most MU metrics in pathologic muscle are with normal limits, discrete values are helpful to separate ambiguities or biases, and additional derived metrics can help with diagnoses. Extrapolated metrics represents E-MU waveform features that are made visible by special

276

M.B. BROMBERG

filtering and detection algorithms, and are available from routine QEMG studies with special software (discussed below).

Routine metrics During routine studies, basic E-MU metrics are assessed visually from moving waveforms and also aurally from the loud speaker (Fig. 11.6 and Table 11.1). Assessment is difficult as waveforms may overlap, and especially problematic for estimates of duration and number of phases.

RECRUITMENT MU recruitment is commonly assessed at weak patient efforts varies (10%–15% of maximal voluntary force), and attempts at rating include recruitment frequency and recruitment ratio are subjective (1 to 4+, increased or decreased), and there are no quantitative EMG techniques (Stalberg et al., 2019). Increased effort activates larger MUs (size principle of Henneman), which can introduce a bias to higher amplitude values (Henneman, 1957; Boe et al., 2005). Amplitude: Peak-peak amplitude values reflect the distance between the electrode recording surface and the nearest 1–3 muscle fibers, but amplitude varies based on rise time of the negative E-MU waveform slope as the electrode is moved to the closest fiber. MU amplitude values also vary with type of electrode used, and amplitudes are
25% to 30% higher with monopolar compared to concentric electrodes (Howard et al., 1988).

With qualitative EMG, the practice of moving the electrode to “focus” on an MU biases to high values and misses low amplitude MUs, while with QEMG a wider range of MU amplitudes are captured (Barkhaus and Nandedkar, 1996). At greater levels of patient effort, amplitudes are higher (Boe et al., 2005).

DURATION Duration reflects the number of muscle fibers up to 2.5 mm from the recording surface (Nandedkar et al., 1988a). Duration measurements differ based on display gain and between concentric and monopolar electrodes (Dumitru et al., 1997). Qualitative MU duration measurements from moving waveforms are highly subjective, especially for determining the termination point, and more likely visually represent only the duration of the main spike. With QEMG, duration markers are set by an algorithm in a consistent manner (Stalberg et al., 1986).

PHASES AND TURNS Very small electrode movements cause phases to bifurcate into turns or turns combine to become phases, and the term MU “complexity” incorporates both phases and turns (Stewart et al., 1989). From early quantitative efforts in normal muscle, up to
5% of MU are polyphasic (>4 phases), and with qualitative EMG there can be a bias to over emphasizing the significance of a few complex MUs.

+

RECEFTIIED AREA

Fig. 11.6. Common motor unit (MU) metrics. Amplitude: peak-peak (+ to ) voltage. Duration: onset to termination, but termination may be difficult to discern. Phases: portions above or below isoelectric baseline. Turns: changes in voltage, but must extend 50–100 mV between changes. Satellite potential: not part of MU duration or phases/turns. Rectified area: MU portions above and below baseline; used to calculate area: amplitude ratio. Author’s figure.

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT

277

Table 11.1 List of electrophysiologic motor unit metrics (routine subjective and quantitative metrics), their attributes, and caveats and features Metric

Attributes

Caveats/features

Routine metrics: Subjective scaling Recruitment Subjective assessment Amplitude Peak-peak voltage Duration Clinical duration Phases/turns Phases >4; turns>5 QEMG metrics: Numeric values Amplitude Peak-peak voltage Duration Clinical duration Phases/turns Phases >4; turns>5 Area:amplitude ratio Lower values favor myopathies Fiber density Measure of reinnervation Jitter Measure of NMJ transmission Apparent fiber density Proxy for fiber density Apparent jitter Proxy for jitter Apparent jiggle Proxy for jiggle

Varies with contraction force Bias to higher amplitude MUs Subjective assessment Rarely counted, over called Includes lower amplitude MUs Consistent assessment by algorithm Phases/turns counted Derived metric SFEMG electrode, dedicated test SFEMG electrode, dedicated test Derived metric from filtered data Derived metric from filtered data Derived metric from filtered data

QEMG: quantitative EMG; NMJ: neuromuscular junction; SFEMG: single fiber EMG.

DERIVED METRICS Derived metrics can only be determined from digitized waveform data as they require calculations based on discrete data (Table 11.1). Area Area reflects the number of muscle fibers up to 2.5 mm from the recording surface (Nandedkar et al., 1988b).

SINGLE-FIBER EMG The restricted recording uptake area of single-fiber EMG electrodes allows for recording muscle fiber action potentials from an uptake area of
300 mm, which includes 1–3 fibers of a normal A-MU. EMG machine low-pass filters are reset from 10 to 1000 Hz to reduce low frequency waveform contamination, thus achieving sharp waveform spikes.

Fiber density Area:amplitude ratio MU area is calculated from the rectified E-MUP waveform. The calculation of the area:amplitude ratio represents the “thickness” of the E-MUP, and is a feature of myopathic MUs (Nandedkar et al., 1988a).

SIZE INDEX This value is calculated as 2  log(amp) + area/amp, and has been found to discriminate neurogenic from normal MUs (Sonoo and Stalberg, 1993). Jiggle Discharge-to-discharge variability in the E-MU waveform is attributed to neuromuscular junction (NMJ) transmission variability (jitter) and variations in conduction velocity of muscle fiber action potentials from NMJs to the electrode. Jiggle is a statistical method to measure this degree of variability among all fibers of the E-MUP (Stalberg and Sonoo, 1994). Jiggle increases in pathologic states, both neuropathic and myopathic conditions.

Fiber density measurements require use of a single-fiber electrode and a trigger and delay line with successive waveform viewed in a superimposed screen, and is performed as a dedicated fiber density study (Fig. 11.7). Under weak voluntary activity by the subject, the electrode is moved to record a clear, consistent single-fiber waveform, verified by uncontaminated signal in the superimposed screen and whose amplitude is >200 mV (Stalberg and Thiele, 1975). Fiber density measurements are indexed to “1,”—the isolated waveform; and the number, if any, of linked discharges from the same MU are counted. Twenty observations of different isolated single-fiber waveforms at different sites in the muscle are made to provide an average fiber density number for a muscle. The normal range of density values has been determined empirically, and values differ somewhat among muscles and with age, and values are available (Bromberg and Scott, 1994). Higher fiber density values reflect pathology, but values are increased in both neuropathic and myopathic muscle. With neuropathic disorders, loss of MUs leads to collateral reinnervation with

278

M.B. BROMBERG

Fig. 11.7. Measurement of fiber density. (A) Muscle fibers of a motor unit. (B) Single-fiber electrode adjusted so recording radius includes at least one muscle fiber; (B1) recording from two muscle fibers, with C1 showing fiber density ¼ 2; (B2) recording from three muscle fibers, with C2 showing fiber density ¼ 3 (note, 2 potentials not included because too small, as waveforms must have steep rise times and amplitude >200 mV s). Author’s figure.

more muscle fibers innervated in the reinnervated MUs territory; with myopathic disorders, there will be atrophy and loss of muscle fibers resulting in less space between fibers in the MU.

Jitter Jitter measurements are traditionally made with a singlefiber electrode (which is sterilizable and reusable due to high electrode cost), but with concerns for possible transmissible infections, a disposable pediatric/facial concentric electrode with a smaller recording radius can be used. EMG machine filter settings and display format are similar to fiber density settings. Jitter measurements require a dedicated study. NMJ jitter is commonly measured during weak voluntary activation of the muscle by the subject, and a pair of muscle fiber action potentials from the same MU are recorded—voluntary jitter study (Stalberg et al., 1971; Sanders et al., 2019) (Fig. 11.8). One muscle fiber action potential is the triggered potential, and is fixed in time, and the time variability of the other potential reflects the relative variability of two NMJs, measured over 80–100 discharges. Twenty observations of fiber pairs are made to provide an average jitter number for a muscle. Occasionally, the electrode records from three or four muscle fibers of the MU. The normal range of jitter values has been determined empirically, and differs somewhat with among muscles and with age (Bromberg and Scott, 1994; Stalberg et al., 2016). When studies are made with a pediatric/facial concentric electrode, the larger uptake radius means that a potential that “looks like” a single fiber but may be made up of two fibers, one superimposed on top of the other, and care must be taken to inspect waveform for inflections indicating a slight time displacement of one potential to the other. Thus, when using a concentric electrode

to measure jitter, even with care and attention to waveform detail, the more accurate term is “apparent single-fiber EMG” versus “single-fiber EMG” when using a single-fiber electrode. The normal range of jitter values has been determined empirically, and importantly, values differ between use of single-fiber and concentric electrodes, and values are available (Trontelj et al., 1986; Stalberg et al., 2016). Jitter measurements can also be made by electrically stimulating a branch of a motor nerve, and obviates the need for patient activation—stimulation jitter study (Trontelj et al., 1986; Trontelj and Stalberg, 1992). A monopolar needle stimulating electrode is moved so as to activate a small portion of the muscle under study when using threshold currents (Fig. 11.9). A single-fiber or pediatric/facial concentric needle is inserted in the segment of activated muscle. With muscle stimulation frequencies of 5–10 Hz, the recording electrode is moved to record a clear muscle fiber waveform; the stimulating current is gradually increased to above threshold for the nerve fiber activating the MU to ensure that time variability observed is not due to threshold changes at the stimulation site and is true NMJ jitter. Measurements are made from 20 fibers and averaged. The jitter values from single NMJs differ from values derived from voluntary jitter technique, and values are available (Trontelj and Stalberg, 1992).

Extrapolated metrics Extrapolated metrics represent measurements from stored data from isolated E-MUs. It is possible to identify individual muscle fiber contributions to E-MU waveforms recorded with concentric electrodes by signal processing using special filters and assessment of peak waveform signal acceleration, and is called

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT

279

Fig. 11.8. Measurement of jitter, voluntary method. (A) Muscle fibers of a motor unit. (B) Single-fiber electrode adjusted so recording radius includes at least two muscle fibers. (C) Triggered waveforms from one fiber and variability (jitter) of second fiber; note, jitter abnormal with some potentials blocked. Author’s figure.

- + Stimulator Stimulus Artifact

A

B

C

Fig. 11.9. Measurement of jitter, stimulation method. (A) Muscle fibers of a motor unit with stimulating electrode activating motor nerve branch(s). (B) Single-fiber electrode adjusted so recording radius includes at least one muscle fiber. (C) Stimulus artifact and muscle fiber waveforms and variability (jitter); note, jitter normal. Author’s figure.

“near-fiber EMG” (Stashuk, 1999; AbdelMaseeh and Stashuk, 2017; Piasecki et al., 2021). Within a normal E-MU there is a small discharge–discharge change in waveform shape (jiggle) due mostly to variability in NMJ transmission (represented by jitter in single-fiber studies), and to a lesser degree, time variability of conduction of nerve fiber action potentials along terminal branches to the NMJs and conduction of muscle fiber action potentials from the NMJs to the electrode. Variability in these elements will change with disease state, and near-fiber EMG is a method to measure these changes. Near-fiber EMG measurements include “apparent fiber density,” “apparent jitter,” and “apparent jiggle” (Fig. 11.10). While not equivalent to fiber density as measured with single-fiber electrode or jitter as measured with single-fiber or concentric electrode during dedicated studies, near-fiber metrics can be obtained during a routine QEMG study with concentric electrodes.

NEAR-FIBER COUNT This represents the number of fibers contributions to the E-MU, as “apparent fiber density” (Stashuk, 1999).

NEAR-FIBER JIGGLE With identification of near fibers, time covariability between pairs of muscle fibers represents “apparent jitter” (Piasecki et al., 2021).

NEAR-FIBER MOTOR UNIT JIGGLE Time variability of the whole MU (AbdelMaseeh and Stashuk, 2017). The stability of the whole E-MUP can be calculated as “MUP jiggle” (Stalberg et al., 1994).

NORMATIVE DATA Normative E-MU data for basic and derived metrics should be generated for each laboratory based on muscle, patient age, type of electrode used and level of patient effort requested, rather than relying on historic values likely gathered under different circumstances (photographed waveforms). Traditional methods for gathering normal data are to recruit subjects with no neuromuscular disease, but this is impractical, and the likely reason for relying on values from early studies. However, E-MUs were photographed at lower levels of contraction than

280

M.B. BROMBERG

Near-fiber EMG examples • Normal

• Neurogenic

• Myopathic

Fig. 11.10. Near-fiber EMG signals and metrics. Modified with permission from Piasecki M, Garnes-Camarena O, Stashuk DW (2021). Near-fiber electromyography. Clin Neurophysiol 132: 1089–1104. 4500 4000

MU Amplitude (μV)

currently used and values may not apply at higher levels of activation. Early values were obtained with concentric electrodes and amplitudes will be higher with monopolar electrodes. Newer techniques allow for determining limiting metric values from a full spectrum of patients, independent of whether the muscle is normal or with disease. The technique, called “E-norm or E-reference” methodology is based on compiling stored QEMG data and plotting for each metric all quantitative values in ascending order; the resultant curve will have inflection points where pathologic values deviate from the range of normal values (Jabre et al., 2015; Nandedkar et al., 2018) (Fig. 11.11). The stored QEMG MU data comes from files in the EMG machine for each patient, each muscle, each MU, and for each metric. Data files can be extracted and grouped by muscle and metric, independent of the diagnosis. It is practical to obtain normative data for “informative” muscles (deltoid, biceps brachii, vastus for myopathic disorders; first dorsal interosseous, tibial anterior for neuropathic disorders) and ages below and above 65 years.

3500 3000 2500 2000 1500 1000 500 0

Inflecon at LLN 700μV

Inflecon at ULN 2,200μV

Fig. 11.11. Method to determine normal range of motor unit (MU) metrics (MU amplitude displayed) from plotting values in ascending order data from patients independent of diagnosis; inflection points suggest lower limit of normal (LLN) and upper limit of normal (ULN).

based on specific clinical applications. MUNE is the only method to provide this estimate, but because of the time and training needed, it remains a research technique.

MOTOR UNIT NUMBER ESTIMATES Interest in MUNE led to the development of a number of methods, based on dividing the maximal CMAP amplitude by the average single MU amplitude (de Carvalho et al., 2018). The techniques are applicable to muscles where nerve stimulation can be performed, limited to biceps brachii, thenar and hypothenar, anterior tibialis, and extensor digitorum muscles. There are a number of issues with each technique, leading to selection

Incremental stimulation technique The incremental stimulation technique is based on initial low stimulation current to activate the lowest threshold axon, then increasing stimulation current to activate successive axons resulting in step increases in the CMAP; after
10 steps are discerned, the highest amplitude is divided by the number of steps for the average MU amplitude (McComas et al., 1971).

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT

The multipoint stimulation technique The multipoint stimulation technique is a derivate of the incremental stimulation technique in that a single axon’s response is obtained at a stimulation site, and the stimulating electrode is moved to different sites along the nerve to obtain more single axon’s responses, and the responses are averaged for the average MU amplitude (Doherty and Brown, 1993).

The F-wave technique The F-wave technique is based on an F-wave representing a single axon’s response; to ensure single axon responses, the same F-wave (amplitude, latency, shape) is sought three times from a series of F-waves, and confirmed single responses are averaged for the average MU amplitude (Stashuk et al., 1994).

The Poisson distribution technique The Poisson distribution technique is based on a low and constant stimulation level evoking a response whose amplitude varies from stimulus to stimulus based on overlap of axon threshold levels (activating varying numbers of axons); the variability is treated statistically similar to how quantile release of acetylcholine from end plate potential variability was determined, and the unit change in amplitude reflects the average MU amplitude (Daube, 1995).

The spike triggered averaging technique The spike triggered averaging technique differs in that the patient actives motor axons at weak contraction levels; a concentric or monopolar electrode is adjusted to record from a single E-MU, and the surface EMG signal from the same muscle is time-linked to the isolated E-MU and the surface response is averaged by spiketriggering to average out contaminating signals due to surface activity from other MUs to yield the amplitude of a single MU; a number of such responses are averaged for the average MU amplitude (Brown et al., 1988).

MUNIX MUNIX provides derived numeric values proportional to the number of remaining MU in a muscle or group of muscles. It is based on recording the interference pattern at different levels of force, and is based on an underlying model of the power of the response. MUNIX can be performed rapidly (Nandedkar et al., 2004).

MU METRIC CHANGES WITH PATHOLOGY The utility of EMG studies is to assess MUs for pathologic metrics. Since most MU metrics are within normal

281

ranges, the diagnostic challenge is assessing for outlying values associated with pathology. Complicating interpretation is that MUs from neuropathic and myopathic muscle can have similar outlying values, and thus looking for patterns of outlying values is more discriminative than relying on single metrics.

Normal aging MU loss occurs with normal aging, and collateral reinnervation of remaining MU results in changes in E-MU metrics. A comparison using MUNE and QEMG techniques in normal subjects in the 2nd decade versus 7th decades indicate increases in E-MU amplitude by
30%–40%, area:amplitude ratio by 6%–14%, apparent fiber density by 5%–25%, and jiggle 0%–6% (Power et al., 2012; Hourigan et al., 2015). Duration values increase by 4%–7%, which is in contrast to
22% increase in older studies using manual isolation techniques (Buchthal et al., 1954). NMJ jitter and fiber density also increases linearly with age (Bromberg and Scott, 1994).

Pathology A spectrum of E-MU changes occurs with both neuropathic and myopathic pathology, while some metrics change in the same direction. One approach is to narrow tolerance limits of individual metrics by averaging metric values from 20 E-MUs in a muscle (Engstrom and Olney, 1992). Another approach is to rely on statistical limits based on standard deviations (SD) of metrics to define threshold values from normative data (gathered as above). Another approach is based on defining a muscle as abnormal when >10% of E-MUs exceed normative metric values (Stalberg et al., 1994). Efforts have been made to identify clusters of metrics that distinguish between normal and pathologic muscles and between neurogenic from myopathic muscles (Bromberg, 2020). Distinguishing different muscle pathologies is accomplished by considering clustered metrics; “size” based on amplitude, duration, area, “shape” based on area:amplitude ratio, and complexity based on number of phases/turns (Abdelmaseeh et al., 2014).

STATISTICAL MOTOR UNIT AND MUSCLE CHARACTERIZATION The overlap of E-MU metric values has led to the concept of determining the diagnostic probability, “definite,” “probable,” and “possible,” for the state of an E-MU, and collectively the state of a muscle. The concept has been extended to the use of Bayesian statistics, utilizing prior knowledge (characterization of E-MU metrics in

282

M.B. BROMBERG Motor Units (Probabilities each class) Neuropathic

Normal

Muscle Characterization Confidence: 0.602

Myopathic

+ + +

Equations

25% Normal

25% Neuropathic

Computations Bayesian Aggregation Equations

Motor Unit Characteristics Amplitude Duration

Apparent FD

20+ Motor Units

Apparent Jitter

50% Myopathic

Phases Turns

Fig. 11.12. Steps in categorizing the pathologic state of a muscle: (Left) individual motor units with probability of being normal or pathologic. (Middle) Use of Bayesian aggregation statistics to estimate the probability of the state of the who muscle. (Right) Final statistical characterization (probabilities) of the state of the whole muscle. With permission from Bromberg MB (2020). The motor unit and quantitative electromyography. Muscle Nerve 61: 131–142.

know muscle conditions—normal, neuropathic, myopathic—a training set) and applying these metric features to the metrics obtained from the muscle understudy (Bromberg, 2020). Thus, for each E-MU, basic and derived metric values can be used to determine the probability that it is normal, neuropathic or myopathic; and a similar process can be applied to all E-MUs in the muscle to determine the probability that the whole muscle is normal, neuropathic or myopathic (Fig. 11.12).

ANALYSIS OF TURNS AND AMPLITUDE Some electromyographers assess the “fullness” of the interference pattern when the patient maximally activates the muscle in an attempt to estimate recruitment and overlapping E-MU waveforms. This is listed in a qualitative manner as normal or full pattern, reduced, pattern, discrete pattern, and maybe increased pattern. Interference assessment is an observation usually made at the end of the routine EMG study (Nandedkar and Sanders, 1990). As an interference pattern increases from weak to full, the changes observed are more MUs discharging, and larger MUs recruited, and the signal shows more turns (changes in voltage direction—positive/negative) and higher amplitudes. This can be quantified for a given level of activation as the number of turns/s and average amplitude/turn, and graphically as a point on an x–y plot of turns/s versus amplitude/turn (Stalberg et al., 1983)

(Fig. 11.13). If points are obtained at varying levels of activation from minimal to maximal, the points increase along both axes. Empiric normal data have been obtained for different muscles and ages, for males and females, and for concentric and monopolar electrodes, and the statistical limiting values as “turns and amplitude cloud plots.” About 20 observations are made over a range of contraction efforts, from weak to maximal, with a brief rest between each effort during which the needle is moved several millimeters to a new area in the muscle. A muscle is considered abnormal when >10% of values are outside of the cloud; for neuropathic disorders, points tend to be above the cloud (greater amplitude/turn and fewer turns/s), and for myopathic disorders points tend to be below the cloud (more turns/s and lower amplitude/turn). Turns and amplitude analysis requires special software, available on EMG machines.

PRACTICAL CONSIDERATIONS Routine qualitative EMG studies in real time mode from free-run sweeps represent pattern recognition, and rely on a training apprenticeship. There is bias to larger MUs and smaller pathologic MUs may be missed, but studies are generally diagnostically reliable. There is a practical time competition between performing routine qualitative EMG studies and quantitative EMG studies. However, time for quantitative MU data processing has become negligible, and it is argued that detailed analysis of a few key muscles by QEMG is more informative

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT

0 – 2K

Neuropathic Pattern

283

N

M M M M MM M M M M MM M

Normal Pattern

B

0 – 2K

Amplitude (mV)

0 – 2K

NN N NN N N N NN N

Myopathic Pattern 0

A

100

Time (mSec)

Fig. 11.13. Analysis of turns and amplitude. (A) Single free-run sweeps at moderate effort showing normal interference pattern complexity (middle trace); reduced pattern with neurogenic disorder (upper trace); increased pattern with myopathic disorder (lower trace). (B) Curved enclosed area represents limit of turns/s vs. amplitude/turn plots from normal muscle at different levels of contraction, from minimum to maximum. “N” represents plots from neuropathic muscle with fewer turns/s and higher amplitude/turn. “M” represents plots from myopathic muscle with more turns/s and lower amplitude/turn. Author’s figure.

than qualitative study of a large number of muscles. It is somewhat paradoxical that qualitative EMG stands apart from the quantitative aspects of nerve conduction studies where discrete values are relied upon. Current EMG machines have the capability of isolating MUs by use of a voltage trigger and delay line, and this technique, which can be referred to as “semiquantitative” EMG can be easily used to more clearly estimate basic metrics (Bromberg, 1993). The triggered and isolated MUs can be viewed in expanded time scale (2 ms/div), allowing for clear inspection of basic metrics, which can be scaled semiquantitatively against the voltage and time axes. If the triggered voltages are also viewed in a superimposed screen, the stability of the MU can be viewed.

Routine QEMG Current EMG machines have QEMG software packages available that include methods to isolate E-MUs and extract routine and derived metrics. There may be differences among manufactures in the techniques of MU isolation, but metric values are similar. The time to acquire QEMG data in a muscle occurs without delay (seconds), and several such observations in a muscle takes no more time than with qualitative studies. There is an option to view and edit acquired data for each E-MU isolated, and this may take a few minutes for
20 MUs per muscle. With respect to the overall time for an EMG study, some electromyographers study up to 40 muscles in a patient! It will be argued that detailed study with QEMG of a limited number of index or informative muscles to diagnose a condition is more informative than briefly studying a large number of muscles, and takes no more

time (probably less time) and causes less patient discomfort. A reasonable approach is a combination of QEMG for informative muscles and routine qualitative EMG as a screen for diffuseness of involvement. Neuropathic disorders tend to affect distal or focal muscles while myopathic disorders tend to affect proximal muscles,

AMYOTROPHIC LATERAL SCLEROSIS Index muscles for the diagnosis of ALS include first dorsal interosseous, biceps brachii, deltoid, anterior tibialis, gastrocnemius, vastus (Babu et al., 2017). QEMG frequently documents greater changes than routine EMG in these muscles, supporting subclinical denervation and the diagnosis. Other muscles can be studied by routine EMG for the presence of active denervation (positive waves and fibrillation potentials). Inclusion body myositis: There is overlap of involved muscles in inclusion body myositis (IBM) and ALS, leading to occasional misdiagnosis. IBM has a mixture of neuropathic-like E-MUs that may mask observation of myopathic E-MUs, and QEMG can clarify the diagnosis (Barkhaus et al., 1999).

RADICULOPATHY Assessment for chronic and subtle denervation in a root distribution can be challenging, especially in an elderly patients with age-related MU changes. Comparison of QEMG metrics between the contralateral (normal) muscles innervated by the root under consideration to the affected side can aid in showing a quantitative difference between the two sides.

284

M.B. BROMBERG

MYOPATHY VS. FATIGUE It can be difficult to determine if there is true muscle weakness versus apparent weakness due to fatigue and pain. Subtle MU changes can be missed or over diagnosed, and determining whether quantitative E-MU values are outside of the normal range can be helpful.

Analysis of turns and amplitude This study is helpful in subtle cases of either neuropathic or myopathic disorders. Since it is usually performed after a routine EMG study, it can be used to support or confirm findings from the EMG studies.

Motor unit number estimation Single-fiber EMG studies FIBER DENSITY STUDIES Fiber density studies require a single-fiber electrode, which have high costs and need for periodic maintenance to ensure clear signals. The information must be interpreted in the clinical context as increases in fiber density occur both in neuropathic disorders due to collateral reinnervation, and myopathic disorders due to reduced space between muscle fibers. Fiber density is perhaps more sensitive to changes in the MU than routine EMG metrics, e.g., in the diagnosis ALS. However, the increase in fiber density has not been compared to increases in near-fiber count obtained from near-fiber EMG signal processing. Overall, measurement of fiber density is rarely used in clinical practice.

JITTER STUDIES Jitter studies are the most sensitive test for NMJ pathology, but jitter values are increased in both presynaptic and post synaptic disorders, and the clinical context is essential. They have great utility in confirming the diagnosis of myasthenia gravis (MG). Repetitive nerve stimulation can document NMJ transmission failure, but jitter values can be increased when there is an insufficient number of fibers that are not activated (blocked). Further, jitter studies in patients with only ocular or bulbar symptoms can be performed in facial muscles with less patient discomfort and artifacts than with repetitive stimulation of the facial nerve. It is important to query patients for previous botulinum toxin use as it is commonly injected in facial muscles for therapeutic and cosmetic reasons, and in other muscles for focal dystonias. With neuropathic denervation and reinnervation, newly formed neuromuscular junctions include the fetal gamma acetylcholine receptor subunit (before replacement by the adult epsilon subunit), and the fetal subunit leads to less secure opening of the receptor’s ligandgated sodium channels, causing increased jitter. With myopathic disorder, jitter can be increased due to atrophic muscle fibers which have slower and more variable conduction velocities from the neuromuscular junction to the electrode.

MUNE is primarily a research tool, although in the setting of a diagnosis of ALS, the multiple point stimulation technique can reveal very large amplitude MUs and support extensive denervation and reinnervation.

CONCLUDING COMMENTS Routine EMG studies based on qualitative metrics have served electromyographers with ease of use, but have not changed since its development in the 1950s. EMG machines have evolved into sophisticated instruments with the capability of acquiring precise metric values and new metrics, but routine qualitative studies have not taken up the advantages of digital analysis. Use of discrete metric values has clear utility. The ability to use near-fiber EMG to detect underlying features contributing to the E-MU that are not visible in the waveform, and statistical methods to give the probability of the state of a muscle, represent advances in signal processing that are not being taken advantage of. This is in contrast to the use of similar data interpretations in other fields of medicine, such as machine reads of ECG traces based on machine learning and evolving MRI sequences that are used clinically. Proficiency in EMG interpretation is taught by observation, and adoption of QEMG techniques should start in teaching EMG laboratories.

REFERENCES AbdelMaseeh M, Stashuk DW (2017). Motor unit potential jitter: a new measure of neuromuscular transmission instability. IEEE Trans Neural Syst Rehabil Eng 25: 1018–1025. Abdelmaseeh M, Smith B, Stashuk D (2014). Feature selection for motor unit potential train characterization. Muscle Nerve 49: 680–690. Babu S, Pioro EP, Li J et al. (2017). Optimizing muscle selection for electromyography in amyotrophic lateral sclerosis. Muscle Nerve 56: 36–44. Barkhaus PE, Nandedkar SD (1996). On the selection of concentric needle electromyogram motor unit action potentials: is the rise time criterion too restrictive? Muscle Nerve 19: 1554–1560. Barkhaus PE, Periquet MI, Nandedkar SD (1999). Quantitative electrophysiologic studies in sporadic inclusion body myositis. Muscle Nerve 22: 480–487. Birkbeck MG, Heskamp L, Schofield IS et al. (2020). Non-invasive imaging of single human motor units. Clin Neurophysiol 131: 1399–1406.

QUANTITATIVE ELECTRODIAGNOSIS OF THE MOTOR UNIT Bodine-Fowler S, Garfinkel A, Roy RR et al. (1990). Spatial distribution of muscle fibers within the territory of a motor unit. Muscle Nerve 13: 1133–1145. Boe SG, Stashuk DW, Brown WF et al. (2005). Decomposition-based quantitative electromyography: effect of force on motor unit potentials and motor unit number estimates. Muscle Nerve 31: 365–373. Bromberg M (1993). Electromyographic (EMG) findings in denervation. Crit Rev Phy Rehabil Med 5: 83–127. Bromberg MB (2020). The motor unit and quantitative electromyography. Muscle Nerve 61: 131–142. Bromberg MB, Scott DM (1994). Single fiber EMG reference values: reformatted in tabular form. AD HOC committee of the AAEM single fiber special interest group. Muscle Nerve 17: 820–821. Brown WF, Strong MJ, Snow R (1988). Methods for estimating numbers of motor units in biceps-brachialis muscles and losses of motor units with aging. Muscle Nerve 11: 423–432. Buchthal F (1975). Electromyography—sensory and motor conduction; findings in normal subjects. In: C Rigshospitalet (Ed.), Laboratory of clinical neurophysiology. Rigshospitalet, Copenhagen. Buchthal F, Guld C, Rosenfalck P (1954). Action potential parameters in normal human muscle and their dependence on physical variables. Acta Physiol Scand 32: 200–218. Buchthal F, Erminio F, Rosenfalck P (1959). Motor unit territory in different human muscles. Acta Physiol Scand 45: 72–87. Czekajewski J, Ekstedt J, Stalberg E (1969). Oscilloscopic recording of muscle fiber action potentials. The window trigger and the delay unit. Electroencephalogr Clin Neurophysiol 27: 536–539. Daube JR (1995). Estimating the number of motor units in a muscle. J Clin Neurophysiol 12: 585–594. de Carvalho M, Barkhaus PE, Nandedkar SD et al. (2018). Motor unit number estimation (MUNE): where are we now? Clin Neurophysiol 129: 1507–1516. Doherty TJ, Brown WF (1993). The estimated numbers and relative sizes of thenar motor units as selected by multiple point stimulation in young and older adults. Muscle Nerve 16: 355–366. Dumitru D, King JC, Nandedkar SD (1997). Motor unit action potential duration recorded by monopolar and concentric needle electrodes. Physiologic implications. Am J Phys Med Rehabil 76: 488–493. Edstrom L, Kugelberg E (1968). Properties of motor units in the rat anterior tibial muscle. Acta Physiol Scand 73: 543–544. Engstrom JW, Olney RK (1992). Quantitative motor unit analysis: the effect of sample size. Muscle Nerve 15: 277–281. Henneman E (1957). Relation between size of neurons and their susceptibility to discharge. Science 126: 1345–1347. Hourigan ML, McKinnon NB, Johnson M et al. (2015). Increased motor unit potential shape variability across consecutive motor unit discharges in the tibialis anterior and vastus medialis muscles of healthy older subjects. Clin Neurophysiol 126: 2381–2389.

285

Howard JE, McGill KC, Dorfman LJ (1988). Properties of motor unit action potentials recorded with concentric and monopolar needle electrodes: ADEMG analysis. Muscle Nerve 11: 1051–1055. Jabre JF, Pitt MC, Deeb J et al. (2015). E-norms: a method to extrapolate reference values from a laboratory population. J Clin Neurophysiol 32: 265–270. McComas AJ, Fawcett PR, Campbell MJ et al. (1971). Electrophysiological estimation of the number of motor units within a human muscle. J Neurol Neurosurg Psychiatry 34: 121–131. McGill KC (2009). A comparison of three quantitative motor unit analysis algorithms. Suppl Clin Neurophysiol 60: 273–278. Nandedkar SD, Sanders DB (1990). Measurement of the amplitude of the EMG envelope. Muscle Nerve 13: 933–938. Nandedkar SD, Barkhaus PE, Sanders DB et al. (1988a). Analysis of amplitude and area of concentric needle EMG motor unit action potentials. Electroencephalogr Clin Neurophysiol 69: 561–567. Nandedkar SD, Sanders DB, Stalberg EV et al. (1988b). Simulation of concentric needle EMG motor unit action potentials. Muscle Nerve 11: 151–159. Nandedkar SD, Nandedkar DS, Barkhaus PE et al. (2004). Motor unit number index (MUNIX). IEEE Trans Biomed Eng 51: 2209–2211. Nandedkar SD, Sanders DB, Hobson-Webb LD et al. (2018). The extrapolated reference values procedure: theory, algorithm, and results in patients and control subjects. Muscle Nerve 57: 90–95. Piasecki M, Garnes-Camarena O, Stashuk DW (2021). Near-fiber electromyography. Clin Neurophysiol 132: 1089–1104. Power GA, Dalton BH, Behm DG et al. (2012). Motor unit survival in lifelong runners is muscle dependent. Med Sci Sports Exerc 44: 1235–1242. Sanders DB, Arimura K, Cui L et al. (2019). Guidelines for single fiber EMG. Clin Neurophysiol 130: 1417–1439. Sherrington C (1925). Remarks on some aspects of reflex inhibition. Proc Roy Soc Lond A 97: 519–545. Sonoo M, Stalberg E (1993). The ability of MUP parameters to discriminate between normal and neurogenic MUPs in concentric EMG: analysis of the MUP “thickness” and the proposal of “size index”. Electroencephalogr Clin Neurophysiol 89: 291–303. Stalberg EV, Sonoo M (1994). Assessment of variability in the shape of the motor unit action potential, the “jiggle,” at consecutive discharges. Muscle Nerve 17: 1135–1144. Stalberg E, Thiele B (1975). Motor unit fibre density in the extensor digitorum communis muscle. Single fibre electromyographic study in normal subjects at different ages. J Neurol Neurosurg Psychiatry 38: 874–880. Stalberg E, Ekstedt J, Broman A (1971). The electromyographic jitter in normal human muscles. Electroencephalogr Clin Neurophysiol 31: 429–438.

286

M.B. BROMBERG

Stalberg E, Chu J, Bril V et al. (1983). Automatic analysis of the EMG interference pattern. Electroencephalogr Clin Neurophysiol 56: 672–681. Stalberg E, Andreassen S, Falck B et al. (1986). Quantitative analysis of individual motor unit potentials: a proposition for standardized terminology and criteria for measurement. J Clin Neurophysiol 3: 313–348. Stalberg E, Bischoff C, Falck B (1994). Outliers, a way to detect abnormality in quantitative EMG. Muscle Nerve 17: 392–399. Stalberg E, Nandedkar SD, Sanders DB et al. (1996). Quantitative motor unit potential analysis. J Clin Neurophysiol 13: 401–422. Stalberg E, Sanders DB, Ali S et al. (2016). Reference values for jitter recorded by concentric needle electrodes in healthy controls: a multicenter study. Muscle Nerve 53: 351–362. Stalberg E, van Dijk H, Falck B et al. (2019). Standards for quantification of EMG and neurography. Clin Neurophysiol 130: 1688–1729.

Stashuk DW (1999). Detecting single fiber contributions to motor unit action potentials. Muscle Nerve 22: 218–229. Stashuk DW, Doherty TJ, Kassam A et al. (1994). Motor unit number estimates based on the automated analysis of F-responses. Muscle Nerve 17: 881–890. Stewart CR, Nandedkar SD, Massey JM et al. (1989). Evaluation of an automatic method of measuring features of motor unit action potentials. Muscle Nerve 12: 141–148. Tankisi H, Burke D, Cui L et al. (2020). Standards of instrumentation of EMG. Clin Neurophysiol 131: 243–258. Thiele B, Bohle A (1978). Number of spike-components contributing to the motor unit potential (author’s transl). EEG EMG Z Elektroenzephalogr Elektromyogr Verwandte Geb 9: 125–130. Trontelj JV, Stalberg E (1992). Jitter measurement by axonal micro-stimulation. Guidelines and technical notes. Electroencephalogr Clin Neurophysiol 85: 30–37. Trontelj JV, Mihelin M, Fernandez JM et al. (1986). Axonal stimulation for end-plate jitter studies. J Neurol Neurosurg Psychiatry 49: 677–685.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00005-4 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 12

Neuromuscular pathology ANTHONY A. AMATO1 AND UMBERTO DE GIROLAMI2* 1

Department of Neurology, Division of Neuromuscular Diseases, Neuropathology Laboratory, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA, United States

2

Department of Pathology, Neuropathology Laboratory, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA, United States

Abstract In this chapter, we discuss the indications for muscle, nerve, and skin biopsies, the techniques and normal processing of biopsy specimens, normal histological appearance, and the commonest histopathological abnormalities of different myopathies and neuropathies.

INTRODUCTION Muscle and nerve biopsies can be extremely useful in the evaluation of patients with myopathies and neuropathies, but the indications and limitations of these procedures must be carefully considered, especially as regards the biopsy site selection, proper handling of the specimens, and the techniques selected to study the tissue. The role of skin biopsy to assess intraepidermal nerve fibers in the evaluation of patients with peripheral neuropathy will also be discussed briefly. Most importantly in neuromuscular disease, the assessment histopathologic findings on muscle and nerve biopsy must be considered in concert with the whole of the clinical context, the neuromuscular examination, and the electrodiagnostic findings. To this aim, our Neurology and Neuropathology Services should conduct, as we do, regular joint meetings to review a patient who undergoes muscle and/or nerve biopsy or other relevant neuropathological interventions. This chapter reviews the clinical indications for muscle and nerve biopsy and basic concepts on the normal and abnormal light and electron microscopic (EM) findings of skeletal muscle and of peripheral nerve. The scope is not to be all-inclusive, rather the aim is to stress fundamental principles and illustrate them with key

figures. Other chapters will deal with specific diseases in depth and include morphologic findings.

MUSCLE BIOPSIES The past few decades have witnessed enormous advances in the understanding of muscle disease based on morphologic, biochemical, and genetic studies of normal and abnormal skeletal muscle tissue obtained by open biopsy as well as tissue obtained at postmortem examination. Our aim here will be to confine our considerations to biopsy of muscle studied through a combination of standard light microscopy, enzyme histochemistry, electron microscopy (EM), and molecular biology (Carpenter and Karpati, 2001; de Girolami et al., 2003; Engel, 2004; Banker and Engel, 2004; Engel and Banker, 2004; Amato and Russell, 2016; Gherardi et al., 2019; Nix and Moore, 2020; Chkheidze and Pytel, 2020; Nathani et al., 2021).

Indications for muscle biopsy A muscle biopsy may be requested by the attending physician when evaluating a patient objective muscle weakness, abnormal muscle enzymes (e.g., elevated

*Correspondence to: Umberto De Girolami, MD, Brigham and Women’s Hospital, Department of Pathology, 75 Francis Street, Boston, MA 02115, United States. Tel: +1-617-732-7510, Fax: +1-617-975-0944, E-mail: [email protected]

288

A.A. AMATO AND U. DE GIROLAMI

serum creatine kinase levels), abnormal skeletal muscle imaging, or myopathic electromyographic findings (EMG). The findings from these studies may point to a myopathy but not to the exact etiology; a muscle biopsy may then help identify the nature of the disease. If the diagnosis is suspected based on less invasive means, one need not proceed to muscle biopsy. For example, in the case of a patient strongly suspected to have a muscular dystrophy or congenital myopathy based on clinical and laboratory findings, the usual practice is to first proceed to genetic testing. Also, the experienced clinician will be in a position to evaluate and treat selected patients with myalgias, subjective weakness, or mild laboratory abnormalities (e.g., elevations of the serum creatine kinase (CK) without the needed to resort muscle biopsy to aid in establishing the diagnosis (Filosto et al., 2007).

Techniques The surgical procedure should be performed by a neurologist or surgeon experienced in the performance of biopsy surgery, at centers with a neuromuscular pathologist knowledgeable in the processing and interpretation of neuromuscular disorders. The quadriceps, deltoid, biceps brachii, or triceps (at some institutions) muscles are preferred sites of biopsy for disorders that have a clinical presentation characterized by proximal weakness; the extensor digitorum longus, tibialis anterior, or gastrocnemius muscles are sampled in patients where the clinical presentation indicates a distal myopathy. Communication between the physician who requested the biopsy and will be following the patient, the individual who performs the biopsy, and neuromuscular pathologist is important to ensure that the correct muscle is biopsied and appropriately processed in light of the clinical issues to help resolve. Muscle tissue can be obtained through an open (minor surgical procedure) or needle biopsy. A larger sample of tissue can be biopsied by the open surgery technique, and we prefer this method in most cases (e.g., patients with inflammatory myopathies) or in patients suspected of having myopathies that require metabolic analysis (e.g., mitochondrial disorders or glycogen storage diseases), molecular studies (e.g., Western blotting and direct genetic analysis), or electron microscopy (EM). We avoid needle biopsies, particularly in patients with abnormal amounts of subcutaneous tissue or those whose muscles are atrophic and/or fibrotic. However, opinions vary with institutions experienced in handling the small amount of tissue obtained by this technique (Edwards et al., 1983; Heckmatt et al., 1984; Cote et al., 1992; Magistris et al., 1998). Inasmuch as it is often difficult to discern the underlying process in end-stage muscle disease, the choice of

biopsy site should select a mildly weak muscle in the Medical Research Council grade 4/5 range to increase the yield. In adults, biopsy is performed under local anesthesia: young children ordinarily require sedation or general anesthesia. Sampling should be taken from the belly of the muscle, avoiding the tendinous insertion because the structure varies considerably at the ends of the fiber. Each specimen should be about 1–2 cm in length and 0.5 cm in width. After the procedure it should be wrapped in slightly moist gauze (not immersed in saline) and placed in separate, labeled. Sterile containers and brought immediately to the laboratory for processing.

Tissue processing In the laboratory, the tissue is divided into three or four fragments to allow for paraffin embedding, cryostat frozen sections including enzyme histochemistry, plastic embedding for electron microscopy, and snap freezing biochemical/molecular studies. Paraffin-embedded sections oriented for cross- and longitudinal sections are useful to assess the extent of disease in the connective tissue compartment, including inflammatory reactions involving the muscle and supporting connective tissues; these are also useful to determine vessel abnormalities and infiltration by tumor or abnormal deposits. Cryostat sections of snap-frozen muscle in the cross-sectional plane, and in some centers, the longitudinal plane as well, are also used routinely. The frozen sections are stained with hematoxylin and eosin (H&E) and with the modified Gomori trichrome stain as the initial survey sections. Also, a battery of enzyme histochemical reactions is run. The adenosine triphosphatase (ATP) reactions at acid pH 4.3 and 4.6, and alkaline pH 9.4, are used to evaluate the proportion of type 1 and type 2 fibers and subtypes: as well as their shape, size, and distribution. Specific histochemical reactions are performed to assess lipid content using Oil Red O and/or Sudan Black stains, while glycogen content is assessed by reactions with the Periodic acid-Schiff (PAS) stains with and without diastase. Mitochondrial and oxidative enzyme activities are assessed employing nicotinamide adenine dinucleotide-tetrazolium (NADH-TR), succinate dehydrogenase (SDH), and cytochrome-c-oxidase (COX) reactions. Frozen sections can be used for the immunohistochemical localization of membrane-associated proteins using a standard panel that includes dystrophin, as well as dystroglycan, sarcoglycan, and merosin reactivity, among others. Immunoperoxidase studies using the large number of commercially available antibodies can be performed on paraffin sections, and sometimes frozen sections as well. For example, the panoply of monoclonal antibodies used

NEUROMUSCULAR PATHOLOGY in hematopathology can be used to identify lymphocyte subsets or membranolytic complement activity. The third procedure is plastic embedding for lightmicroscopic examination of 1-mm semithin sections, and ultrastructural study of ultrathin sections mounted on copper grids. These are particularly useful to identify abnormalities in the integrity of the myofibrillar contractile apparatus and organelles, and to assess the presence of abnormal accumulations of proteins, lipids, or glycogen. Morphometric analysis is performed only in specialized centers possessing the required equipment and expertise to evaluate the tissue. The techniques are intended to demonstrate statistically important differences in the size, shape and distribution of fiber types using the battery of histochemical stains described above. They can be especially useful in evaluating muscle biopsies of children, where even the well-trained observer may have difficulty with fine discrimination of relative proportions of affected fibers. Age-matched control comparisons can be extremely valuable. Morphometric analysis of cross-sections of muscle may be carried out either after photography or directly by image analysis. Finally, the definitive identification of specific mitochondrial electron-transport abnormalities, genetically determined disorders of membrane-associated proteins, or metabolic derangements requires that frozen tissue be sent out to specialized centers that can perform the required biochemical or molecular/genetic analysis.

STRUCTURE OF NORMAL SKELETAL MUSCLE In cross-section, the muscle extrafusal fibers in adults are polygonally shaped, with little intervening space between them and rather uniform in size. Neuromuscular spindles are recognized as rounded structures (about 50–100 mm in diameter) containing intrafusal fibers and bounded by a connective tissue capsule of multiple layers. The nuclei of muscle extrafusal fibers are ordinarily located next to the sarcolemma and as many as 3%–5% of fibers may have “internalized” nuclei. Muscle fibers are embedded in connective tissue compartments. Individual muscle fibers are surrounded by the endomysium, which normally is a thin, delicate network of strands between the fibers, also containing the microcirculation network. Individual muscle fascicles are enclosed in the perimysium comprised of concentric strands of connective tissue enclosing the fascicle. Muscle fascicles are separated from each other by the epimysium, which contains adipose tissue, the arterial and venous supply to the fascicle and larger peripheral nerve branches. The fascia and tendons are contiguous to the epimysium. The term motor unit refers to a physiologic concept where a single motor neuron in the spinal cord and its

289

Fig. 12.1. Muscle biopsy reveals a mosaic of fiber types (type 1 dark staining, type 2A pale staining, and type 2B intermediate staining. ATPase pH 4.5.

motor axon is responsible for the innervation of a population of myocytes. The motor neuron, furthermore, determines the histochemical type of the muscle innervated, be they Type 1 or 2 fibers. Normally, in human beings, the muscle fibers belonging to a given motor unit are spatially dispersed in a muscle fascicle over several millimeters. Using histochemical reactions that type the fibers, the muscle appears as a mosaic checkerboard of the two fiber type prototypes (Fig. 12.1). In the adult, the number of type 1, 2A, and 2B fibers is roughly comparable in the muscles that are usually studied: the proportion is type 1: 30%–40%, type 2A: 20%–30%, type 2B 40%–50%, and type 2C 1%–2%. However, the percentages of each fiber type will vary not only with the specific muscle studied, but also according to sex, age, and physical state, therefore making it necessary to evaluate the results in comparison to normative data of matched controls. The percentages of type 1, 2A, and 2B fibers differ in various muscle groups, and it is important to be aware of the normal percentage (Johnson et al., 1973). The most commonly biopsied muscles (i.e., biceps brachii, triceps, and quadriceps) have approximately equal amounts of the three major fiber types, although the deltoid muscle has more type 1 fibers than type 2A and 2B. Because muscle fibers from a single motor unit are randomly distributed among muscle fibers of different motor units and fiber types, a checkerboard or mosaic pattern is appreciated on ATPase stains. The number of fibers within each motor unit is proportional to the refinement of movement, so that the ratio is low in the lumbrical and extraocular muscles and high in the gluteus and limb-girdle muscles. This description of normal muscle applies to adults; in infants and very young children, muscle fibers are rounded and only take on a polygonal cross-sectional shape later in development, from age 3 to 6 years.

290

A.A. AMATO AND U. DE GIROLAMI

Furthermore, as the mean fiber diameter is a function of age, analysis of muscle biopsies in infants and children requires comparison to normal values for age. Nevertheless, variability of fiber diameter is not a function of age, i.e., excessive variation in fiber diameter is abnormal, even in infants. Normally, there may be increased variability in fiber size and shape near tendinous insertions, sites that should be avoided in biopsy; this is also the case with certain muscles, such as the extraocular muscles, diaphragm, and paraspinal muscles. Satellite cells, best visualized by EM, are present next to the sarcolemma and are enveloped by basement membrane that surrounds the muscle fibers. Most of the sarcoplasm of the muscle fiber contains myofilaments, which form the contractile apparatus and supporting structures. Individual muscle fibers contain repeating units (sarcomeres) of interlaced, longitudinally directed thin filaments and thick filaments and perpendicularly oriented Z bands to which the thin filaments are connected. The sarcomere is connected to the sarcolemma via filamentous actin. The sarcolemma is composed of various protein complexes and is connected to the extracellular matrix. The T tubules are composed of invaginations of the sarcolemmal membrane into the interior of the muscle fibers. Their course is parallel to the Z bands, and they are surrounded on each site by the sarcoplasmic reticulum. The T tubules allow for rapid depolarization of muscle membrane deep within muscle fiber cells and the accelerated release of calcium from the sarcoplasmic reticulum during excitation. Quantitative analysis is performed by measuring the mean and range of the diameters for each different fiber type (Brooke and Engel, 1969a,b,c,d). Importantly, the diameters of muscle fibers increase to a point during childhood until the early teens. At 1 year of age the mean muscle fiber diameter is approximately 16 mm. The size increases by about 2 mm/yr until the age of 5 years and subsequently by 3 mm/yr until 9 years of age. By 10 years of age, mean muscle diameters range from 38 to 42 mm. Normal adult size is reached between the ages of 12 and 15 years. There is usually less than 12% difference in the largest mean fiber diameters between the major fiber types. Both types 1 and 2 adult muscle fibers are larger in men than in women. Type 2 fibers are usually larger than type 1 fibers in men; type 1 fibers are larger than type 2 fibers in women. The diameter of muscle fibers is also dependent on the specific muscle biopsied. For example, in the biceps brachii, the diameters of muscle fibers are as follows: type 1 fibers 64.3  3.7 mm and type 2 fibers 72.7  5.3 mm in males and type 1 fibers 56.8  4.8 mm and type 2 fibers 54.6  7.0 mm in females. In the vastus lateralis, the diameters of muscle fibers are slightly different: type 1 fibers 59.5  6.4 mm and type 2 fibers

64.8  8.1 mm in males and type 1 fibers 58.8  6.1 mm and type 2 fibers 49.9  6.2 mm in females (Brooke and Engel, 1969a).

REACTIONS TO INJURY Before considering the basic reactions to injury in in muscle fibers, it is important to recognize the histologic artifacts that frequently occur. First are those due to improper tissue freezing; these are manifest as small, rounded, empty, evenly distributed spaces within the muscle fiber the result of intracellular microcrystal ice formation. Tissue fixed in formalin destined for paraffin-embedding and not appropriately stretched with a muscle biopsy clamp or other device can become distorted during fixation and is rarely properly oriented. Paraffin sections, under these circumstances, commonly show contraction band artifacts that preclude fine morphological analysis of muscle fibers. Artifacts are also encountered in the preparation of sections for electron microscopy; these are related to delayed fixation and improper processing of tissue. Muscle abnormalities may be classified on histopathologic and etiologic grounds into three major categories: (1) neurogenic atrophy: a pattern of muscle pathology consequent to denervation and reinnervation; (2) myopathies: inherited and acquired diseases characterized by abnormalities in the muscle fiber itself; these include dystrophies, congenital, inflammatory, metabolic, and toxic myopathies; and (3) disorders of the neuromuscular junction. Patients with neuromuscular junction defects usually have only slight and nonspecific alterations apparent on routine light microscopy and are rarely biopsied except at specialized centers; when studied these have shown EM abnormalities at the neuromuscular junction (Carpenter and Karpati, 2001; de Girolami et al., 2003; Engel, 2004; Banker and Engel, 2004; Engel and Banker, 2004; Amato and Russell, 2016; Gherardi et al., 2019). The initial analysis of the muscle biopsy slides evaluated with the battery of preparations indicated above assesses the cross-sectional size and range of size variability of the fibers, the shape of the fibers (polygonal, rounded or “angulated”), the topographic distribution of fiber major fiber types and subtypes, the size and location of the myonuclei, the presence of necrosis, other alterations in the cytoarchitecture and organelles (e.g., the presence of target fibers, cores, vacuoles, tubular aggregates, and ragged red fibers), and any abnormal accumulation of glycogen or lipid. Also important is to study the supporting connective tissue and vasculature in the endomysial and epimysial compartment. The evaluation includes, for example, a search for evidence of

NEUROMUSCULAR PATHOLOGY vasculitis, inflammation in the connective tissue, abnormal deposits (such as amyloid). The inflammatory cell infiltrates can be further characterized using the armamentarium of immunohistochemical markers used by hematopathologists (e.g., typing of macrophages, lymphocytes, plasma cells, eosinophils) and the location (endomysial, perimysial, and perivascular). As regards inflammatory infiltrates the location and distribution of these, and their relation to diseases muscle fibers is also assessed. The detailed analysis of these findings reveals the histopathologic differential diagnosis if inflammatory myopathies are covered in other chapters of this text. In the setting of motor neuron/axonal degeneration, the muscle fibers within that motor unit lose their neural input and undergo denervation atrophy. This leads to decreased synthesis of myofilaments, degeneration of myofibrils, and a reduction in the size of the muscle fiber (Brooke and Engel, 1969b). The atrophic fibers lose their polygonal appearance and are smaller than normal and have sharp edges (“angulated”) (Fig. 12.2). Neurogenic disorders affect motor nerves that innervate both type 1 or 2 fibers. Therefore, in early denervation, muscle biopsies reveal randomly distributed, atrophic angulated muscle fibers of both fiber types. As adjoining surviving neurons and their axons reinnervate the denervated muscle, the checkerboard pattern is lost, and there are aggregates of contiguous muscle fibers of near normal size, or larger than normal, that have the same histochemical profile (usually separate patches of both fiber type prototypes). This phenomenon is referred to as type grouping and has important neurophysiologic correlates since there has been an increase in the number of fibers within a given motor unit (Fig. 12.3). In time, if the denervating process is progressive, the reinnervated

Fig. 12.2. Muscle biopsy demonstrates ongoing neurogenic denervation with large group of atrophic, angulated fibers. H&E, paraffin section.

291

Fig. 12.3. Muscle biopsy revealed fiber type grouping indicated of a chronic neurogenic process with reinnervation. ATPase pH 4.2.

fibers undergo atrophy and cluster together (group atrophy). Another feature of denervation is the presence of the so-called target fibers. Reorganization of the cytoarchitecture within muscle cells results in a rounded central zone of disorganized filaments that contain fewer mitochondria and less glycogen. Target fibers have three zones that are circumferentially oriented, which are best seen on NADH-TR staining. The innermost zone is devoid of mitochondrial, glycogen, phosphorylase, and ATPase enzymatic activity; the second zone has increased enzymatic activity, while the third zone exhibits intermediate enzymatic activity. Targetoid fibers refer to a similar phenomenon without a distinct intermediate zone of enzyme activity. As with central cores, target and targetoid fibers preferentially affect type 1 fibers. In contrast to central core myopathy in which the cores are present in most type 1 fibers, target and targetoid fibers are less abundant. These occur in neurogenic disorders during reinnervation. Target and targetoid fibers can also be appreciated on other stains such as the ATPase and modified Gomori-trichrome stains. The second large category disease assessed by muscle biopsy includes all those conditions where primary abnormality is not in the innervation of the muscle but in the muscle fiber itself. These are generally grouped together under the generic term “myopathy.” The abnormalities found in the muscle biopsy include those associated with destruction of the fiber, as seen in either toxic or inflammatory processes directed toward the cell. There is subsequent myophagocytosis of the injured portions of the cell and concomitant regeneration originating the adjacent noninjured portion of the cell. These myopathic processes may also occur secondary to intrinsic problems in the fiber, be they metabolic or genetically determined. In addition, there are abnormalities that affect

292

A.A. AMATO AND U. DE GIROLAMI

Fig. 12.4. Muscle biopsy in a patient with a mitochondrial myopathy reveals ragged red fibers on modified Gomori trichrome stain (A) and scattered cytochrome oxidases negative fibers (B).

the vascular supply or connective tissue framework of muscle and result in seconder injury to the muscle fibers themselves. Muscle is a syncytium formed from the fusion of thousands of myoblasts. Because of its syncytial nature, histopathological abnormalities may be focal rather than occurring along the entire length of a muscle fiber (e.g., segmental necrosis). Genetic disorders can manifest discrete abnormalities, with other regions of the single fiber appearing relatively normal. An example of this can be seen in mitochondrial myopathies in which the histopathological alterations are dependent on the degree of abnormal mitochondria, which in turn reflects the percentage of mutated mitochondrial DNA in the region. Thus, when cut longitudinally, one may appreciate segments of the muscle fiber with a ragged red appearance (Fig. 12.4A), which may not stain with cytochrome oxidase (Fig. 12.4B), while other nearby segments of the same fiber may be normal. In dystrophies, randomly distributed necrotic fibers are seen on the cross section. If the tissue is cut longitudinally, one sees that necrosis is often segmental in along the length of the fiber. Likewise, the cellular infiltrate seen in inflammatory myopathies is multifocal, surrounding, and invading segments of muscle fibers along their length. Myopathies are usually associated with a random loss of muscle fibers belonging to different motor units. Damaged fibers may appear small than or manifest a wide range of morphologic changes, including necrosis, muscle fiber splitting, and regeneration. Preferential atrophy or hypotrophy of type 1 fibers is seen in certain myopathic disorders (e.g., myotonic dystrophy and various congenital myopathies). On the other hand, preferential type 2 fiber atrophy can be seen in certain endocrine disorders (e.g., steroid myopathy), as well as a complication of

Fig. 12.5. Muscle biopsy reveals segmental necrosis with myophagocytosis. Paraffin, H&E.

disuse. Muscle fiber hypertrophy can develop in response to increased load, either in the setting of exercise or in pathologic conditions where other muscle fibers are injured. Large fibers may divide along a segment (muscle fiber splitting) so that, in cross section, a single large fiber contains a cell membrane traversing its diameter. Necrosis is a feature more common in myopathies, but destruction of muscle but may be seen in the late stages of denervating diseases. A single muscle fiber can undergo either total necrosis or segmental necrosis, but destruction of the entire fiber length is rare. Segmental necrosis is best appreciated on paraffin or semithin sections (STS) of muscle fibers cut longitudinally (Fig. 12.5). With segmental necrosis, the affected portion of the single muscle fiber becomes more rounded, and the sarcoplasm begins to have a featureless ground-glass appearance. EM sections reveal degeneration of the Z disk and myofibrillar

NEUROMUSCULAR PATHOLOGY

293

Fig. 12.6. Muscle biopsy in patient with inclusion body myositis reveals endomysial inflammation, muscle fibers with rimmed vacuoles and eosinophilic inclusions (A), H&E (B). Immunoperoxidase stain reveals positive p62 reactive inclusions.

network as well as abnormal mitochondria. Macrophages are recruited into the area and infiltrate the necrotic segments to digest the disintegrating muscle tissue and damaged tissue. In certain diseases (inclusion body myositis), macrophages and lymphocytes may invade nonnecrotic tissue such that a muscle fiber can be “severed” into distinct segments. In inclusion body myositis, aside from endomysial inflammation, muscle fibers can show rimmed vacuoles and p62-positive inclusions (Fig. 12.6). Repair of necrotic segments can occur and begins with the proliferation of adjacent satellite cells in the region of the destroyed portion of the fiber (Brooke and Engel, 1969c). The satellite cells align next to each other to form myotubes. Several myotubes form per segment and adhere to the surrounding basal lamina. The expansion of myotubes occurs laterally and longitudinally, eventually reaching and fusing with the healthy muscle tissue stumps. The regenerating muscle fibers can be appreciated by their large, internalized nuclei with prominent nucleoli, and their basophilic cytoplasm that is laden with ribonucleic acid (RNA).

NERVE BIOPSIES Peripheral neuropathy is one of the most common disorders in the practice of Neurology. Considering the many advances in clinical electrophysiology and molecular diagnostics, the clinical indications to perform a nerve biopsy in patients with peripheral neuropathy have decreased in comparison to those for a muscle biopsy. Furthermore, the structural abnormalities seen in many neuropathies are not sufficiently specific for the results of the biopsy to be of major clinical utility. It is also important to consider is that peripheral nerve most often

chosen for biopsy is a sensory nerve, the invasive procedure results in a focal permanent sensory deficit and, not uncommonly, neuralgia. Despite these considerations, many cases of peripheral neuropathy remain unexplained and nerve biopsy has been used to attempt to better understand the disease process. As is the case for muscle biopsies, the interpretation of a nerve biopsy requires correlation of histological changes with clinical information, including the results of electrophysiological investigations. Important indications for a nerve biopsy in adults include the evaluation of a patient where there is a strong clinical suspicion of vasculitis or amyloid polyneuropathy based on the neurological examination and laboratory studies. The possibility of amyloidosis should be considered in patients with a monoclonal gammopathy, autonomic neuropathy, systemic signs of amyloidosis (e.g., renal insufficiency or cardiomyopathy), or those individuals with a family history of amyloidosis. Vasculitic neuropathy enters the differential diagnosis in patients presenting with a history of multiple mononeuropathies, particularly when of acute onset and painful, an underlying connective tissue disease (e.g., systemic lupus erythematosus and rheumatoid arthritis), eosinophilia or late-onset asthma (Churg–Strauss syndrome), renal failure or chronic sinusitis, hepatitis B or C, an elevated erythrocyte sedimentation rate, or antinuclear cytoplasmic antibody. Additional indications that raise the option of a nerve biopsy include considerations of other autoimmune inflammatory conditions (e.g., sarcoidosis), possible infectious processes (e.g., leprosy), and tumor infiltration (e.g., lymphoma and leukemia). Also, a nerve biopsy may be required for diagnosis of a tumor of the peripheral nerve (e.g., perineurioma). Less commonly, nerve biopsy may be warranted to diagnose uncommon forms of hereditary neuropathy when

294

A.A. AMATO AND U. DE GIROLAMI

DNA testing is not available or is negative (e.g., giant axonal neuropathy and polyglucosan body neuropathy). Directed nerve biopsy is sometimes performed in cases of asymmetric poly/mononeuropathy with focal signal changes on neuroimaging, searching for inflammation as in focal CIDP, sarcoidosis, perineuritis, and tumor infiltrates in neurolymphomatosis and leukemia, or peripheral nerve tumors. In parts of the world where leprosy is endemic, nerve biopsy, in conjunction with skin biopsy is used for diagnosis. Molecular studies performed in commercial laboratories are now used for the diagnosis of certain hereditary neuropathies, for example, Charcot–Marie–Tooth disease. Nerve biopsy should preferably be performed in laboratories where there is technical and professional expertise in the performance and interpretation of paraffin embedded sections of nerve and appropriate staining techniques, frozen section histology, plastic-embedding, and preparation of semithin sections and EM (Midroni and Bilbao, 1995; Richardson and de Girolami, 1995; de Girolami et al., 2003; Dyck et al., 2005; Amato and Russell, 2016; Chkheidze and Pytel, 2020; Nathani et al., 2021). In children, analysis of nerve biopsies is greatly aided by the use of morphometric methods. As indicated above for muscle biopsy, careful coordination of efforts between the clinical team and the laboratory is required for the proper evaluation of nerve biopsies. The peripheral nerve chosen for biopsy is commonly a purely sensory nerve that is affected clinically indicated by nerve conduction studies. The sural nerve is ordinarily chosen for biopsy. We recommend biopsying the sural nerve in the mid-shin approximately one-third to onefourth of the distance from ankle to knee, approximately where the nerve is stimulated on nerve conduction studies. Patients should be informed that complications of nerve biopsy a permanent loss of sensation on the lateral aspect of the ankle and foot as well as pain that may last for several months after the procedure. Biopsy of the superficial peroneal nerve biopsy is recommended when vasculitic neuropathy is suspected and there is involvement of the peroneal nerve, because the underlying peroneus brevis muscle can also be biopsied through the same incision site, thereby increasing the diagnostic yield.

TECHNIQUES The Neuropathology Laboratory should be alerted prior to the biopsy to allow preparation for the special handling that is required. Care should be taken by the neurologist or surgeon performing the biopsy to handle the nerve during dissection as little as possible and to avoid infiltration by local anesthetic. A resection of a segment

of nerve 3–5 cm in length is recommended. To prevent distortion by coiling, immediately after removal the specimen should be gently stretched over a tongue depressor or other suitable device. The nerve biopsy specimen is then allocated to be able to perform five types of studies. For paraffin embedding, the distal portions of the nerve are most appropriate as the technique is not as sensitive to artifactual changes from manipulation. These are fixed in 10% buffered formalin and processed for cross and longitudinal sections. Paraffin sections are especially useful to evaluate the connective tissue components of nerve including epineurium, perineurium, and endoneurium and also the analysis of blood vessels and search for any abnormal deposits, such as amyloid. The conventional stains used on paraffin sections include H&E; elastic tissue stains, Masson trichrome, silver impregnation or neurofilament stains for axons, and stains for myelin. The sections can also be processed with stains that will demonstrate micro-organisms such as methenamine silver stains for fungi, Gram stains for bacteria, and those for mycobacteria. Various markers can be employed to identify different types of inflammatory cells including T- and B-lymphocyte, and macrophages. Immunoperoxidase studies with the armamentarium of commercially available antibodies can be applied to the sections for demonstration of neurofilaments, myelin, and other proteins. The middle third of the nerve tissue specimen is fixed in 2.5% buffered glutaraldehyde and embed in plastic after fixation and staining with osmium tetroxide. Plastic embedded 1 mm semithin sections (STS) cut in the transverse and longitudinal plain and stained with toluidine blue are the best method to examine the fine structural details of both myelinated and unmyelinated fibers and their distribution. The use of STS also allows for quantitative assessment of axonal density, myelin thickness, and subcellular structures that cannot easily be visualized with routine histology (Fig. 12.7). Plastic-embedding of the tissue is also the only technique that allows for ultrastructural (EM) evaluation (Fig. 12.8). In specialized laboratories around the world, immuno-EM is performed in order to identify specific proteins or other substances at the ultrastructural level. Quantitative and semiquantitative analysis is typically done on STS or EM sections. Morphometric analysis is time-consuming and requires a substantial investment in equipment. From a clinical point of view, it is often not necessary, and is usually reserved for research purposes. A trained observer can reliably assess the density of myelinated fibers, an estimate of the degree of loss of large or small fibers, and the presence and the severity of demyelinating and/or axonal lesions, without the expense of

NEUROMUSCULAR PATHOLOGY

295

Fig. 12.7. Semithin section of normal nerve reveals a normal amount of large and small myelinated axons and small unmyelinated nerve fibers (A), a mild reduction of large myelinated axons in chronic axonal neuropathy (B), active axonal degeneration with myeloid debris (yellow arrow) and a cluster of small diameter, thinly myelinated fibers represented regenerating axonal sprouts (red arrow) in active axonal neuropathy (C), and scattered thinly myelinated fibers in a primary demyelinating polyneuropathy (D).

Fig. 12.8. Electron microscopy reveals an axon undergoing Wallerian degeneration (A) in an active axonopathy. Thinly myelinated fiber with early Schwann cell proliferation (onion bulbs). Yellow arrow may be seen in Charcot–Marie–Tooth disease type 1 and chronic inflammatory demyelinating polyneuropathy (B).

quantitative assessment. Morphometric analysis of cross sections of the nerve may be carried out either after photography or directly by image analysis. To perform teased fiber analysis, after aldehyde fixation and osmication and softening in glycerin, single myelinated fibers measuring 1 cm long are separated from each other and from their surrounding connective tissues utilizing fine needles under direct visualization with a dissecting microscope. Light microscopic

examination of the teased fibers demonstrates the relative positions of nodes of Ranvier, the length of internodes, integrity and caliber of the axons, and thickness of the overlying myelin, and thus can be helpful in discriminating primary axonopathies from primary demyelinating neuropathies (Fig. 12.9). The technique also allows for the demonstration of abnormalities in the thickness of the myelin as for example tomacula, or evidence of disintegration of the myelin/axon. Since the preparation

296

A.A. AMATO AND U. DE GIROLAMI

Fig. 12.9. Teased nerve fiber analysis of normal fiber (A), a thinly myelinated internode (B), and fiber with myelin ovoids indicating active axonal degeneration (C).

of individual teased nerve fibers is very time-consuming, the technique is often reserved for evaluation in specific problematic cases. Part of the tissue is frozen in isopentane cooled by liquid nitrogen or directly on the cryostat with appropriate embedding medium and transverse sections are cut on the cryostat. Direct immunofluorescence studies on frozen sections can be carried out using specific antibodies to identify abnormal deposits of immunoglobulins in endoneurium or in myelinated fibers. The last bit of tissue is quickly frozen in isopentane cooled by liquid nitrogen and stored in the ultra-low freezer for potential biochemical or genetic studies.

STRUCTURE OF NORMAL NERVE Peripheral nerves are composed of axons, Schwann cells, myelin sheaths, and supporting tissue. Individual nerve fibers are surrounded by endoneurial connective tissue and grouped into fascicles encased by perineurial

sheaths. All the fascicles within a nerve in turn are surrounded by epineurial connective tissue. A blood–nerve barrier is regulated between the perineurial cells and endoneurial capillaries branching from the vasa nervorum. The connective tissue sheathes that surround nerve roots, dorsal root ganglia, autonomic ganglia, and terminal twigs are different from those of peripheral nerves and so are the corresponding barrier permeabilities. Individual nerve fibers are embedded and in connective tissue compartments. Within each fascicle, myelinated and unmyelinated nerves and their supporting cells are surrounded by delicate connective tissue strands containing blood vessels, termed the endoneurium. Nerve fascicles are bounded by the perineurium, a complex structure with important physiologic barrier functions consisting of multiple overlapping concentric layers of specialized connective tissue cells called perineurial cells. The epineurium is the fibroadipose connective tissue which surrounds the fascicules of nerve and contains the blood vessels supplying the nerve.

NEUROMUSCULAR PATHOLOGY The organization of a myelinated peripheral nerve fiber is such that the axon emanating from the cell body of the neuron is surrounded by a myelin sheath. The myelin sheath is interrupted at regular intervals along its length. The space between two adjacent myelinated segments is the node of Ranvier. The myelinated segment of nerve between one node of Ranvier and the next is referred to as an internode. Normally, the length of an internode is fairly constant along the axon and is proportional to the diameter of the axon. Individual myelin internodes extend from just beyond the neuronal cell body to just before the axon terminal. Schwann cells are the supporting cells of individual myelin internodes. They appear in abundance within the endoneurial compartment, identifiable as cells with a pale nucleus and evenly dispersed chromatin. A Schwann cell can be distinguished with certainty from endoneurial fibroblasts on EM because it is surrounded by basement membrane. In longitudinal sections of nerve, elongated Schwann-cell nuclei are normally roughly equidistant from the two adjacent nodes of Ranvier, positioned toward the middle of the internode. In mature nerve, the cytoplasm, between the inner lamella of the myelin sheath and the axon, and between the outermost myelin lamella and the cell membrane, is sparse. EM examination demonstrates that Schwann-cell cytoplasm contains endoplasmic reticulum, a Golgi apparatus, mitochondria, sometimes a centriole, and complex multilamellar lipid membranous granules termed p granules or granules of Reich. The myelin sheath is made up of regularly arranged concentric lamellae with a 12- to 17-nm periodicity, forming major dense lines separated by electron-lucent zones in which one or two interperiod lines can be observed. Discontinuity in the compaction of the lamellae is seen at the Schmidt–Lanterman incisures. Here, invaginations of Schwann cell cytoplasm penetrate openings of the major dense lines. The specialized Schwann cell that supports unmyelinated fibers and does not form myelin is known as the Remak cells. In a given cross section a single Remak cell can surround and support multiple unmyelinated axons, and given the absence of nodes, they are evenly distributed along the length of the fiber. The ratio of unmyelinated nerve fibers to myelinated nerve fibers in the sural nerve is approximate 4 to 1. The normal diameter of myelinated fibers ranges from 2 to 12 mm in a bimodal distribution with peaks at 3–6 mm and 9–12 mm. In young adults, the average number of myelinated nerve fibers is 7000–10,000 fibers/mm2 of endoneurial area. The diameter of unmyelinated axons ranges from 0.2 to 2.5 mm in a unimodal distribution with a peak at 1.4–1.6 mm, while the density ranges from 20,000 to 35,000/mm2. Axonal caliber is related to,

297

among other things, the number of neurofilaments and neurotubules contained in the axon. There is a nearly linear relationship between diameter of the axon and thickness myelin lamellae. The normal “g” ratio, or ratio of axonal diameter/total diameter, is approximately 0.6 and can commonly be used to distinguish disorders that affect primarily the axon from those that are first directed toward the myelin sheath. Myelinated axons are surrounded by a 7–8 nm-thick membrane, the axolemma. The axolemma has similar ultrastructural characteristics as the cytoplasmic membranes of Schwann cells. This basal lamina passes from one Schwann cell to the next without interruption at the nodes of Ranvier. The cytoplasm within the axons, the axoplasm, contains longitudinally oriented mitochondria, smooth endoplasmic reticulum, multivesicular bodies, neurofilaments, and microtubules. Neurofilaments have a mean diameter of 10 nm, whereas microtubules contain a central lumen and have an external diameter of approximately 25 nm. The nerve–CSF barrier is formed by the tight junctions between the cells that form the outer layer of the arachnoid membrane. These cells fuse with the perineurium of the roots and cranial nerves as these leave the subarachnoid space.

REACTIONS TO INJURY Although disease processes affecting nerves have different pathogenic mechanisms, these lead to two principal reactions to injury: demyelination or axonal degeneration. Damage to Schwann cells or the myelin sheath itself can lead to demyelination. Because these diseases affect individual Schwann cells to varying degrees, the process is characteristically segmental along the length of the nerve. The disintegrating myelin is phagocytosed by Schwann cells and macrophages (Figs. 12.7B and C and 12.8A). Schwann cells are also stimulated to remyelinate the denuded axon. The myelin sheath of the newly remyelinated axons is disproportionally thinner in diameter and the internodes are shorter than normal. These features that are well seen with teased nerve preparations and can also appreciated on semithin sections (Fig. 12.7D) and on EM. With sequential episodes of demyelination and remyelination, concentric tiers of Schwann cell processes accumulate around the axons forming “onion bulbs” (Fig. 12.8B). Some disease processes are associated with inclusions within Schwann cells (e.g., metachromatic leukodystrophy and certain toxic neuropathies). Other abnormalities in the myelin sheath include tomacula (redundant folds of myelin characteristic of hereditary neuropathy with liability to pressure palsies) and widened periodicity of compacted myelin (seen in neuropathy associated with myelin-associated antibodies).

298

A.A. AMATO AND U. DE GIROLAMI

Primary damage to the axon may either be due to a discrete, localized event (trauma, ischemia, etc.) or be due to an underlying abnormality of the neuronal cell body or ganglion (neuronopathy) or its axon (axonopathy). These processes lead to axonal degeneration with secondary disintegration of its myelin sheath. If a nerve is transected, the distal portion of the nerve undergoes an acute disintegration (Wallerian degeneration) characterized by breakdown of the axon and its myelin sheath into fragments forming small oval compartments (i.e., myelin ovoids). These breakdown products undergo phagocytosis by macrophages and Schwann cells. Most neuronopathies or axonopathies evolve more slowly; therefore, evidence of active axon and myelin breakdown is scant because only a few fibers are degenerating at any given time. The proximal stumps of axons that have degenerated sprouts of new axons may attempt to grow along the course of the degenerated axon. Small clusters of these regenerated axons, which are small in diameter and thinly myelinated, can be recognized on cross section of semithin and EM sections. Also, as axonal transport of essential proteins and other substances synthesized in the perikaryon is often impaired in axonopathies, this leads to axonal atrophy that again is apparent on the semithin and EM sections (G ratio less than 0.6). In contrast, enlarged axons are seen in giant axonal neuropathy and hexacarbon toxicity. In addition, nerve biopsies can reveal evidence of disease processes like those found in other organ systems. Amyloid deposition around blood vessels or within the endoneurium can be seen in systemic amyloidosis or in a familial amyloidotic polyneuropathy (Fig. 12.10). In systemic or isolated peripheral nerve vasculitis, there is transmural infiltration of vessel walls by inflammatory cells associated with fibrinoid necrosis of the vessel walls (Fig. 12.11). Because nerve fibers course between

different fascicles along the length of the nerve and vasculitis can be patchy asymmetric loss of axons within and between fascicles is a characteristic finding of ischemic nerve injury. Infiltration of the nerve by neoplastic or inflammatory cells can also be recognized. Leprosy is one of the most common etiologies of polyneuropathy in the world. When granulomas or diffuse inflammation of the nerve are seen, a Fite stain can demonstrate the acid-fast bacilli, depending on the form of the disease.

Skin biopsy Skin biopsies are increasingly being performed to evaluate patients with peripheral neuropathy (Holland et al., 1997, 1998; McArthur et al., 1998; Herrmann et al., 1999; Periquet et al., 1999; Tobin et al., 1999; Smith et al., 2001; Amato and Oaklander, 2004; Wendelschafer-Crabb et al., 2006; Sommer and Lauria, 2007; Walk et al., 2007; Sommer, 2018; Nolano et al., 2020). These are most useful in patients with small fiber neuropathies in which other testing modalities provide normal or inconclusive results. Because nerve conduction studies only assess the conduction of large myelinated nerve fibers, patients with pure small fiber neuropathies will have normal nerve conduction studies. In at least a third of people with painful sensory neuropathies, intraepidermal nerve fibers density on skin biopsies may represent the only objective abnormality present following extensive evaluation. The rationale behind performing skin biopsies is to measure the density and assess the morphology of intraepidermal nerve fibers. These fibers represent the terminals of Ad and C nociceptors, and these may be decreased in patients with small fiber neuropathies in whom nerve conduction studies and routine nerve biopsies are often normal. Skin biopsies are relatively easy to

Fig. 12.10. AL amyloidosis. Paraffin sections of nerve stained with Congo red reveals bright red deposits surrounding small blood vessels in the epineurium (A), which turn apple-green under polarized light (B).

NEUROMUSCULAR PATHOLOGY

299

and then immunostaining protein gene product 9.5 (PGP 9.5) is applied to demonstrate the small intraepidermal fibers. Morphometric methods are used to assess the number and complexity of these nerves, through parameters such as the linear density (number of fibers per millimeter of biopsy) or total length of intraepidermal nerve fibers. The morphology of the intraepidermal nerve fibers can also be assessed. Axonal swellings may be an early marker of small fiber neuropathy and may be appreciated before a reduction in density. However, axonal swellings can be seen in normal individuals. Immunostaining for vasoactive intestinal polypeptide, substance P, or calcitonin gene-related proteins can be used to measure the density of sudomotor axons innervating sweat glands, piloerector nerves to hair follicles, and nerves to small arterioles.

FUTURE DIRECTION

Fig. 12.11. Vasculitis. Nerve biopsy in cross section reveals multiple occluded blood vessels and one with marked perivascular and transmural inflammation and neovascularization of an occluded lumen (A). A vessel cut longitudinally reveals medium sized vessel with transmural inflammation and fibrinoid necrosis (arrow) of vessel wall (B). Paraffin, H&E.

perform and are associated with a much lower morbidity than standard nerve biopsies. However, there are several drawbacks to the use of skin biopsies in these patients. Importantly, these usually may not add new information from other assessments of patients. That is, if a person complains of symmetric burning or tingling pain in the distal lower extremities, has normal strength and deep tendon reflexes, and has normal nerve conduction studies then he or she likely has a small fiber neuropathy. Skin biopsies are often not useful in identifying the etiology of the neuropathy. That said, assessing intraepidermal nerve fiber density and morphology may play a role in the future by defining the natural history of various neuropathies, monitoring response of the neuropathy to various therapies, and assessing for development of toxic neuropathies (e.g., during chemotherapy). Skin biopsies are usually done by performing a 3-mm punch biopsy of the skin under local anesthesia in the lower leg in an affected region. Other regions can be sampled to assess if there is a length-dependent loss of intraepidermal nerve fibers (e.g., in the dorsum of the foot, thigh, or forearm). The tissue is fixed in formalin,

The role of muscle and nerve biopsies are likely to be reduced with less invasive testing being more readily available and less expensive (ex. genetic testing). However, this will no obviate the need for biopsies as genetic testing at this time often is unremarkable or reveals variation of uncertain significance (VOUS). Muscle biopsies can be useful in such situations to assess if histopathological abnormalities support the features one would expect to see if these were pathogenic. Furthermore, muscle tissue can be used to assess RNA sequencing to assess if a particular sequence alteration leads to abnormal RNA splicing. Finally, we expect to see advances in in proteomic analysis of muscle tissue in very difficult cases in which there is no definite DNA or RNA abnormalities. There are many different subtypes of inflammatory myopathy associated with different types of autoantibodies and the histopathological and immunohistopathological features and genetic signatures may be distinct and lead to more focused immunopathies. Similar techniques may be employed as well in nerve biopsies.

CONCLUSION The majority of patients with myopathies and neuropathies and motor involvement can generally be diagnosed without invasive muscle or nerve biopsies. A biopsy should not be done because a patient has subjective muscle pain or weakness or unexplained numbness or tingling. There is still a utility of biopsy in patients with an unclear diagnosis, particularly if it will change management. Just doing a biopsy to find out if it is abnormal is not sufficient reason to do a biopsy. This needs to be explained to a patient before a biopsy is done. Advances in genetics will likely continue to reduce the need for a biopsy. However, this will not obviate the need as many

300

A.A. AMATO AND U. DE GIROLAMI

VOUS found on increasingly available genetic testing will lead to muscle biopsies to help determine if the mutation is pathogenic.

REFERENCES Amato AA, Oaklander AL (2004). Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 16-2004. A 76-year-old woman with numbness and pain in the feet and legs. N Engl J Med 350: 2181–2189. Amato AA, Russell J (2016). Neuromuscular disease, McGraw-Hill, New York. Banker BQ, Engel AG (2004). Basic reactions of muscle. In: AG Engel, C Franzini-Armstrong (Eds.), Myology, third edn. McGraw-Hill, New York. Brooke MH, Engel WK (1969a). The histographic analysis of human muscle biopsies with regard to fiber types. 1. Adult male and female. Neurology 19: 221–233. Brooke MH, Engel WK (1969b). The histographic analysis of human muscle biopsies with regard to fiber types. 2. Diseases of the upper and lower motor neuron. Neurology 19: 378–393. Brooke MH, Engel WK (1969c). The histographic analysis of human muscle biopsies with regard to fiber types. 3. Myotonias, myasthenia gravis, and hypokalemic periodic paralysis. Neurology 19: 469–477. Brooke MH, Engel WK (1969d). The histographic analysis of human muscle biopsies with regard to fiber types. 4. Children’s biopsies. Neurology 19: 591–605. Carpenter S, Karpati G (2001). Pathology of skeletal muscle, Oxford, New York. Chkheidze R, Pytel P (2020). What every neuropathologist needs to know: peripheral nerve biopsy. J Neuropathol Exp Neurol 79: 355–364. Cote AM, Jimenez L, Adelman LS et al. (1992). Needle muscle biopsy with the automatic biopsy instrument. Neurology 42: 2212–2213. de Girolami U, Frosch M, Amato AA (2003). Biopsy of nerve and muscle. In: M Samuels, S Feske (Eds.), Office practice of neurology, second edn. Harcourt Health Sciences, Philadelphia. Dyck PJ, Dyck PJB, Engelstad J (2005). Pathologic alterations of nerves. In: PJ Dyck, PK Thomas (Eds.), Peripheral neuropathy, fourth edn. W. B. Saunders, Philadelphia. Edwards RH, Round JM, Jones DA (1983). Needle biopsy of skeletal muscle: a review of 10 years experience. Muscle Nerve 6: 676–683. Engel AG (2004). The muscle biopsy. In: AG Engel, C Franzini-Armstrong (Eds.), Myology, third edn. McGraw-Hill, New York. Engel AG, Banker BQ (2004). Ultrastructural changes in diseased muscle. In: AG Engel, C Franzini-Armstrong (Eds.), Myology, third edn. McGraw-Hill, New York. Filosto M, Tonin P, Vattemi G et al. (2007). The role of muscle biopsy in investigating isolated muscle pain. Neurology 68: 181–186.

Gherardi G, Amato AA, Lidoy HG et al. (2019). Pathology of skeletal muscle. In: F Gary, C Duyckaerts, U De Girolami (Eds.), Escourolle and Poirier’s manual of basic neuropathology, sixth edn. Oxford University Press, New York. Heckmatt JZ, Moosa A, Hutson C et al. (1984). Diagnostic needle muscle biopsy. A practical and reliable alternative to open biopsy. Arch Dis Child 59: 528–532. Herrmann D, Griffin J, Hauer P et al. (1999). Epidermal nerve fiber density and sural nerve morphometry in peripheral neuropathies. Neurology 53: 1634. Holland NR, Stocks A, Hauer P et al. (1997). Intraepidermal nerve fiber density in patients with painful sensory neuropathy. Neurology 48: 708–711. Holland NR, Crawford TO, Hauer P et al. (1998). Small-fiber sensory neuropathies: clinical course and neuropathology of idiopathic cases. Ann Neurol 44: 47–59. Johnson MA, Polgar J, Weightman D et al. (1973). Data on the distribution of fibre types in thirty-six human muscles: an autopsy study. J Neurol Sci 18: 111–129. Magistris MR, Kohler A, Pizzolato G et al. (1998). Needle muscle biopsy in the investigation of neuromuscular disorders. Muscle Nerve 21: 194–200. McArthur JC, Stocks EA, Hauer P et al. (1998). Epidermal nerve fiber density: normative reference range and diagnostic efficiency. Arch Neurol 55: 1513–1520. Midroni G, Bilbao JM (1995). Biopsy of peripheral neuropathy, Butterworth-Heinemann, Boston. Nathani D, Spies J, Barnett MH et al. (2021). Nerve biopsy: current indications and decision tools. Muscle Nerve 64: 125–139. Nix JS, Moore SA (2020). What every neuropathologist needs to know: the muscle biopsy. J Neuropathol Exp Neurol 79: 719–733. Nolano M, Tozza S, Caporaso G et al. (2020). Contribution of skin biopsy in peripheral neuropathies. Brain Sci 10. Periquet MI, Novak V, Collins MP et al. (1999). Painful sensory neuropathy: prospective evaluation using skin biopsy. Neurology 53: 1641–1647. Richardson Jr EP, de Girolami U (1995). Pathology of the peripheral nerve, W. B. Saunders, Philadelphia. Smith AG, Ramachandran P, Tripp S et al. (2001). Epidermal nerve innervation in impaired glucose tolerance and diabetes-associated neuropathy. Neurology 57: 1701–1704. Sommer C (2018). Nerve and skin biopsy in neuropathies. Curr Opin Neurol 31: 534–540. Sommer C, Lauria G (2007). Skin biopsy in the management of peripheral neuropathy. Lancet Neurol 6: 632–642. Tobin K, Giuliani MJ, Lacomis D (1999). Comparison of different modalities for detection of small fiber neuropathy. Clin Neurophysiol 110: 1909–1912. Walk D, Wendelschafer-Crabb G, Davey C et al. (2007). Concordance between epidermal nerve fiber density and sensory examination in patients with symptoms of idiopathic small fiber neuropathy. J Neurol Sci 255: 23–26. Wendelschafer-Crabb G, Kennedy WR, Walk D (2006). Morphological features of nerves in skin biopsies. J Neurol Sci 242: 15–21.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00015-7 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 13

Electrophysiological assessment of peripheral and central autonomic disorders 1,2
JOEL GUTIERREZ * 1

Department of Clinical Neurophysiology, Cuban Institute of Neurology and Neurosurgery, Havana, Cuba 2

Department of Clinical Neurophysiology, Havana University of Medical Sciences, Havana, Cuba

Abstract The autonomic nervous system (ANS) coordinates multiple reflex actions which are essential for life. The tests employed to evaluate the ANS provide valuable information of the functional state of these reflex arcs. The ideal test should be simple to perform, noninvasive, reproducible, sensitive, specific, safe, and appropriate for longitudinal studies. The availability of computer-based techniques has facilitated the electrophysiological assessment of ANS-mediated reflexes. The information provided by autonomic testing must be analyzed in combination with the clinical history and physical examination of the patient, allowing for a hypothesis that can be tested. Properly performed and interpreted, ANS testing can be used to confirm the presence of an ANS disturbance and the involved functional pathways, as well as the extent, intensity, and site of injury. This chapter describes the most important electrophysiological tests used to evaluate the ANS control of cardiovascular reflexes and sweat gland activity.

INTRODUCTION The autonomic nervous system (ANS) regulates multiple reflex actions. Most of the tests employed to evaluate the ANS provide an overview of the functional state of these reflex arcs. The ideal test should be simple to perform, noninvasive, reproducible, sensitive, specific, safe, and appropriate for longitudinal studies. The availability of computer-based techniques has facilitated the electrophysiological assessment of the ANS (Low, 2003; Hilz and Dutsch, 2006). However, the information provided by autonomic testing is only useful when the indication is derived from proper prior analysis of the clinical history and physical examination of the patient, allowing for a hypothesis that can be tested. Properly performed and interpreted, ANS testing can be used to confirm the presence of an ANS disturbance and the involved functional pathways, as well as, the

extent, intensity, and site of injury (Appenzeller and Oribe, 1997; Low et al., 1998; Low, 2003).

HISTORICAL BACKGROUND Our current understanding of the ANS has been possible thanks to the contribution of many talented international researchers who, through several centuries, have dedicated efforts to study the anatomy and physiology of the nervous system. The idea that the body was divided into two systems, somatic, and autonomic comes originally from the ancient Greeks but the earliest documented reference about the ANS dates back to Galen (Oakes et al., 2016a). Based on animal dissections, Galen described the superior and inferior cervical ganglia, the semilunar (celiac) ganglia, the rami communicantes and the paravertebral chains with their central and peripheral extensions. He considered these were “tubes

*Correspondence to: Joel Guti
errez, Department of Clinical Neurophysiology, Cuban Institute of Neurology and Neurosurgery, Havana, Cuba.

302


J. GUTIERREZ

driving the animal spirit” to achieve “sympathy” or connection between the different parts of the body (Webber, 1978). Galen described the vagus and the sympathetic nerves and as a single functional unit, a mislead notion that was maintained by anatomists for several centuries. After Galen, there was a long pause of almost one millennium in the occurrence of new discoveries about the ANS until Vesalius, in the 16th century, made new descriptions of the sympathetic trunks and peripheral plexuses. Vesalius performed his own dissections and improved Galen descriptions but he perpetuated the belief that the sympathetic trunk was a branch of the
vagus nerve. It was Charles Etienne in 1545, and soon later Eustachius, who finally depicted the vagus and sympathetic nerves as independent structures (French, 1971; Ackerknecht, 1974; Appenzeller et al., 2022). Unfortunately, this important discovery was not published until 1714. Despite his important contributions, Eustachius persevered in the idea that the sympathetic nerve originated from the abducens nerve. It was Riolan who was the first anatomist to clearly discriminate between the upper and lower sympathetic ganglia and trace the sympathetic trunk (French, 1971). The next significant progress in the understanding of the ANS was done by Thomas Willis in 1664. Willis improved Galen’s anatomical description of the sympathetic chain which he named “intercostal nerves” and demonstrated its importance to modulated heart rate (HR) (Ackerknecht, 1974; Standring, 2016). Willis recaptured some of the Galen ideas about the ANS and he considered that the “sympathy” between organs could occur independently from our volition. It was Willis also who proposed the modern nomenclature of the cranial nerves and made a very accurate account of the vagus nerve topography (Oakes et al., 2016a). The term “great sympathetic nerve” was first used by Jacobus Winslow (1669–1760), a Danish professor settled in Paris, in 1732 (Todman, 2008). Winslow was also the first to describe the gray and white rami communicantes. Almost at the same time, Pourfour du Petit demonstrated that the carotid plexus originated from the sympathetic trunk and not from the cranial nerves as was believed by the contemporary anatomists (French, 1971). Unfortunately, that revolutionary finding was not accepted by the greater scientific community for some time (Oakes et al., 2016a). Whytt was another remarkable pioneer of ANS research in the 18th century. He was the first to describe the skeletal muscle tone, the reaction of pupils to light, and the responses of hollow muscles to mechanical distension (Clarke and Jacyna, 1988). He believed that involuntary motion was actually independent of the cerebellum, which went contrary to the prevailing belief at the time. The English anatomist Johnstone made also

important contributions related to the exact function of the autonomic ganglia which he considered as little brains filtering the communication between the viscera and the central nervous system (Clarke and Jacyna, 1988). The concepts of “animal life” and “organic life” were introduced by Bichat in 1802 (Oakes et al., 2016b). Bichat claimed there was much interdependence between both systems but the central nervous system was ultimately in command, however, he considered the rami communicantes were simple structural anastomoses joining the two systems without much functional consequence. He was also the first anatomist to associate ANS with metabolism (Clarke and Jacyna, 1988). The term “Vegetative Nervous System” was used initially by Reil in 1857 who was also a defender of Bichat’s ganglionic theory (Oakes et al., 2016b). In the late 19th century, Bernard’s observations on the vascular sympathetic innervation of the rabbit ear, demonstrated the existence of vascular sympathetic tone (Appenzeller et al., 2022). The most important highlights of ANS research in recent times were the experiments of Langley about the pharmacologic effects upon the ANS (Langley, 1901), the observations of the German microscopists Remak, M€uller, and Valentin who made the histological descriptions of white vs. gray rami communicantes, the experiments of Claude Bernard on the regulation of the chemical activity in the tissues (le milieu int
erieur), the finding of neurotransmitters and the discoveries of Brown-S
equard, Ernst Heinrich Weber, and Eduard Weber about the nervous system’s relationship with the circulatory system (Webber, 1978; Oakes et al., 2016b). The label “autonomic nervous system” was coined by Langley, a physiologist at the University of Cambridge, in 1898, who also proposed the existence of the sympathetic and parasympathetic classical ANS subdivisions (Oakes et al., 2016b). Each division of the ANS has distinct anatomical and functional features enabling their specific biological functions. Following is a brief description of these subdivisions needed to understand the design and interpretation of the functional tests of the ANS.

ANATOMY AND PHYSIOLOGY The ANS consists of a set of afferent pathways, a central nervous system (CNS) integrating complex in the brain and spinal cord, and two distinct efferent limbs, sympathetic and parasympathetic, each with preganglionic and postganglionic neurons (Karemaker, 2017). The parasympathetic division is topographically more restricted than the sympathetic division. The bodies of the preganglionic parasympathetic neurons are located in the brainstem nuclei of cranial nerve 3rd, 7th, 9th, and 10th,

PERIPHERAL AND CENTRAL AUTONOMIC DISORDERS and in the 2nd, 3rd, and 4th sacral segments of the spinal cord. Preganglionic fibers originate in the EdingerWestphal, upper and lower salivatory, ambiguous, and dorsal motor nuclei. Long, predominantly cholinergic parasympathetic preganglionic fibers travel relatively long distances to the sphenopalatine, submandibular, and otic cranial nerve ganglia located in the vicinity of the effector organs, which, in turn, originate short, predominantly cholinergic parasympathetic postganglionic fibers, which innervate the effector organs (Pincus and Magitsky, 1989). Vagal preganglionic fibers innervate all thoracic viscera and digestive tract up to the splenic flexure of the colon. The vagus and glossopharyngeal nerves contain afferent fibers which convey information to the CNS as to changes in blood pressure (BP) and HR essential in the integration of the baroreflex. Sacral segments provide parasympathetic innervation to the pelvic organs and the segments of the colon not innervated by the vagus nerves (Karemaker, 2017). The sympathetic nervous system consists of preganglionic sympathetic neurons located in the intermediolateral horn of the thoracic and lumbar spinal segments that elaborate short nerves which innervate postganglionic neurons located in the prevertebral, paravertebral and previsceral sympathetic ganglia, from whence long postganglionic axons innervate effector organs. Postganglionic axons are generally thin and unmyelinated, measuring 50 cm in length (Shields, 1993). Considering that such axons have a diameter of approximately 1.2 mm, the volume of these axons may reach about 565,000 mm3, in contrast to the volume of the soma of these neurons of only 14,000 mm3 (Wehrwein et al.,

303

2016). These morphological characteristics, together with its constant activity, make sympathetic fibers especially vulnerable to injury. Although parasympathetic and sympathetic effects have been traditionally regarded as antagonistic, it is now apparent that the interrelationship in more complex and context sensitive (Karemaker, 2017). For example, the specific effects of the autonomic innervation of a particular organ does not depend only on the type of autonomic organ receptor present, but the level of preactivation at the time of neurotransmitter stimulation (McCorry, 2007; Wehrwein et al., 2016). Activation of the sympathetic nervous system evokes a diffuse response preparing the body for situations requiring high metabolic demand and physical activity such as those of pupillary and bronchial dilation, increased HR and BP, decreased gastrointestinal motility, vasoconstriction, and increased glycogenolysis, but selectively activation can also occur (Navarro, 2002; McCorry, 2007). Whereas the parasympathetic nervous system regulates more restricted functions associated with resting activity, activation of this division promotes stages of energy conservation expressed as decreases in HR and BP, pupillary constriction and increased bronchial and gastrointestinal activity (Navarro, 2002; McCorry, 2007). Autonomic control is mainly based on autonomic reflexes (Shields, 1993). The baroreflex, the most important reflex for the control of rapid blood pressure oscillations, is a very illustrative example of how these reflexes work (Karemaker, 2017). The baroreflex is triggered by activation of stretch receptors located in the carotid sinus and the aortic arch. The afferent fibers from these receptors convey information along the glossopharyngeal and vagus nerves to the solitary tract nucleus of the brainstem (Wieling and Shepherd, 1992). The efferent arms of the baroreflex are the sympathetic and parasympathetic innervation of the heart and the sympathetic innervation of the smooth muscle of peripheral blood vessels (Wieling and Shepherd, 1992). The baroreflex controls the oscillations blood pressure within certain limits. For example, a sudden drop in blood pressure produces tachycardia and vasoconstriction, which helps to restore blood pressure to its normal limits (Shields, 1993; Karemaker, 2017). A detailed knowledge of ANS is essential for the design, implementation, and interpretation of ANS tests.

TESTS OF AUTONOMIC FUNCTION Although most tests are easy to perform, their interpretation is difficult due to the multiplicity of factors that can affect the responses and the complexity of the pathways and reflexes involved. Consequently, in clinical practice it is more reliable to perform a battery of ANS tests than a single test, the results of which will be complementary to

304


J. GUTIERREZ

each other, increasing clinically reliability (Hilz and Dutsch, 2006). Particular attention should be given to the influence of factors that may increase or decrease the sensitivity of the tests, as for example subjects who demonstrate completely normal vasoconstriction when tested under controlled laboratory conditions despite faulty vasoconstriction that leads to orthostatic hypotension under specific circumstances such as strenuous exercise, emotionally stressful situations and a hot environment (Low, 2003). It is advisable to perform the tests simulating factors that ordinarily trigger autonomic failure. Individual laboratories should have their own reference values corrected for the potential confounding among them, age, body mass index, laboratory conditions and the characteristics of the recording protocols such as the method of analysis, posture, test duration, order of application, levels of hydration, concurrent medication, prior consumption of caffeinated beverages, degree of physical activity and emotional state. Autonomic lesions can be underestimated both in the clinical and research setting due to their subclinical course, as well as, the mistaken attribution to a primary disturbance of the affected organ such as tachycardia erroneously attributed to heart failure instead of deficient cardiovagal control, and the relative insensitivity of vascular sympathetic testing (Low et al., 1998). The ANS, which regulates a wide range of biological functions, is amenable to standardized study of cardiovascular and sweat reflexes testing, with beat-to-beat heart rate variability (HRV) commonly employed to explore cardiovagal regulation, while variations in BP are used to assess the influence on sympathetic vasomotor sympathetic tone. Maneuvers that assess the integrity of cardiovascular reflexes can be performed simply by manual evaluation of the radial pulse and blood pressure taken with an ordinary sphygmomanometer, while more sophisticated and sensitive evaluation of these functions is required to record and mathematically analyze the HRV and beat-to-beat BP variations. The 1996 Task Force report of the American Academy of Neurology (AAN) evaluated the safety, reproducibility, ease of performance and sensitivity of available ANS tests (AAN committee, 1996). Those most useful to evaluate parasympathetic function included HRV during deep breathing with calculation of the expiratory/inspiratory ratio (E/I), Valsalva maneuver (VM) with calculation of the Valsalva ratio; HRV at rest and with active standing with calculation of the 30:15 ratio of the maximum RR interval (interval between two consecutive R waves) at 30 divided by the minimal interval at 15 s; while the recommendation for sympathetic function testing included BP response to the VM, during muscle contraction and active standing, sudomotor test of sympathetic skin response

(SSR), and quantitative sudomotor axon reflex test (QSART). While a wide variety of functional electrophysiological tests are available to study the ANS this chapter will emphasize in those with established practical value for the evaluation of autonomic cardiovascular and sudomotor reflexes.

Heart rate variability The autonomic regulation of HR depends upon the interactions between cardiovagal innervation, b-adrenergic sympathetic innervation, and the influence of circulating hormones. In physiologic conditions, the HR fluctuates according to the metabolic needs of the organism with a variation that is most evident during physical exertion and states of high emotional stress; however, HRV is a permanent phenomenon in normal subjects (Shields, 1993). The quantification of these variations is one of the most widely used tests to explore the ANS. Under physiological conditions, HR increases during inspiration and decreases during respiration termed respiratory sinus arrhythmia (RSA). Atropine and surgical destruction of the vagus nerves eliminates or reduces the RSA, suggesting that cardiovagal innervation is essential to its generation. The reflex arc of the RSA includes vagal afferent fibers from alveolar and thoracic aorta stretch receptors, carotid baroreceptors, dorsal motor ambiguous and vagus cranial nerve nuclei, and vagal efferent fibers to the heart (Wieling and Shepherd, 1992). The analysis of HRV requires the recording of an electrocardiogram (ECG) rhythm strip from lead II with clearly identifiable R waves. The intervals between consecutive R waves are manually or automatically measured producing a series of values of successive RR intervals. Most programs for the analysis of HRV plot RR intervals against time in their sequence of appearance (Fig. 13.1). Visual inspection of HRV curves gives an overview of the variability of RR intervals duration during the measured time period but HRV is better studied by mathematical methods, including analyses in time or spectral domains. Such studies, termed RR variability or RR intervalometry, are recorded at rest and with activating maneuvers including deep breathing at 6 cycles per second, VM, and active standing (Ewing et al., 1981).

Heart rate variability with deep breathing Patients are instructed to inhale and exhale deeply at a frequency of six cycles per minute (0.16 Hz). Each respiratory cycle lasts 10 s, 5 s of inspiration and 5 s of expiration. This frequency generates the most intense variations in the HRV. Some laboratories use visual signals on a monitor to guide the patient. It is advisable to

PERIPHERAL AND CENTRAL AUTONOMIC DISORDERS

305

Fig. 13.1. (A) Electrocardiogram and beat to beat blood pressure recordings. (B) Curves of RR intervals variability and blood pressure (systolic, mean, and diastolic) vs. time while the subject performs a Valsalva maneuver to activate autonomic cardiovascular reflexes.

record lead two of the resting ECG for 60 s before and after the maneuver. The commonest errors include completion of the breathing in less than 5 s and an inadequate respiratory effort. The commonest quantification of the change in HRV induced by deep breathing is the E/I ratio calculated by dividing the maximum and minimum RR intervals obtained during the deep breathing maneuver (Fig. 13.2), and measurement of the average of 3 maximal and minimal values obtained in 3 respiratory cycles, discarding the first cycle. Alternatively, the difference in the ratio of the longest and shortest intervals of 3 cycles with maximal variation can be computed, wherein a result greater than 15 beats per minute (bpm) can be considered normal and 10 bpm abnormal. Deep breathing responses are highly dependent upon patient age and the intensity of the respiratory cycles. Alteration of

HRV to deep breathing is considered a sensitive marker of impaired cardiovagal parasympathetic innervation with both afferent and efferent branches of this reflex mediated by the vagus nerves (Low, 1993). Given its simplicity of implementation and high sensitivity, it is widely employed in research investigations of parasympathetic ANS function, especially in prediction of the progression of diabetic autonomic neuropathy, wherein a prospective study of 373 insulin-dependent diabetic children without clinical manifestations of autonomic neuropathy determined that one-third had changes in HRV to deep breathing over a 7 year period (Stella et al., 2000). The deep breathing index has a higher sensitivity than the VM and the active standing indexes to identify parasympathetic ANS dysfunction (May and Arildsen, 2000).

306


J. GUTIERREZ

Fig. 13.2. (A) Effects of activating maneuvers on heart rate variability. (B) Calculation of deep breathing ratio. RR intervals are recorded while the subject is breathing at a frequency of 6 cycles per minute (5 s inspiration and 5 s expiration). The maxima and minima of the 3 widest respiratory cycles (arrows) are visually identified in the RR intervals curve. The deep breathing ratio is derived by dividing the RR maximum by the RR minimum for each cycle and averaging their results.

Heart rate variability with Valsalva maneuver The complex hemodynamic and autonomic changes induced by a VM depend on multiple interactions between blood pressure, venous return and the baroreflex, making the calculated VR sensitive to dysfunction of both the sympathetic and parasympathetic ANS divisions (Karemaker, 2017). To develop the VM maneuver subjects are instructed to blow vigorously into a mouthpiece or mask applied over the mouth, sustaining a pressure of 40 mmHg for 15 s while quantifying the expiratory force with a digital or aneroid manometer attached to the mask through a cannula. Subjects are

encouraged to watch the pressure gauge to monitor the intensity of the respiratory excursion which continues throughout the period of strain. The examiner informs the patient when to start and end the strain. It is important to ensure that the air is emanating actually from the lungs and not from the mouth alone as this will affect the desired effects of the VM on venous return. The ECG recording should commence 60 s before and last up to 60 s after the performance of the strain phase. The VM should be repeated several times until at least 2 trials yield reproducible HR and BP waveforms. Repeated maneuvers should be separated by intervals of 3 min to avoid overlapping the effects of adjacent strains. The VR, defined as the longest RR interval following

PERIPHERAL AND CENTRAL AUTONOMIC DISORDERS completion of the maneuver, divided by the shortest RR interval recorded during the strain phase (Fig. 13.2) is useful to evaluate the parasympathetic division, while cardiovagal innervation is more complex, since the resultant HR changes are secondary to alterations in systemic BP. Whereas both deep breathing and VR ratios explore HRV induced by cardiovagal innervation, these ratios have a different physiological basis, with the E/I ratio dependent solely upon afferent and efferent nerve fibers that course through the vagus nerves (Opfer-Gehrking and Low, 1993). In order to make a proper interpretation of the cardiovascular changes induced by a VM, it is necessary to evaluate both HR and beat-to-beat BP reflective of primary cardiovagal and a-adrenergic vascular impairment (Benarroch et al., 1991). The increased VR in patients with peripheral neuropathy results from concomitant impairment of vascular a-adrenergic and spares cardiac b-adrenergic autonomic innervation (Opfer-Gehrking and Low, 1993).

Heart rate variability with active standing The HR changes that occur during the first 30 s of active standing depend mainly upon cardiovagal regulation mediated by parasympathetic inhibition of the baroreflex with beat-to-beat recording showing a HR peak beginning between 3 and 12 s after standing, reaching a maximum value at 15 s, followed by a decrease in HR which starts at 20 s and reaches a maximum value at 30 s (Wehrwein et al., 2016). Therefore, the ratio of the maximum RR interval at 30 s divided by the minimum interval at 15 s, termed the 30:15 ratio is a reliable indicator of cardiovagal function and cardiac autonomic neuropathy. Patients are instructed to actively stand up being careful so as not to detach any recording electrodes or generate movement artifact. The determination of the 30:15 ratio requires recording of the initial 30–40 s period of active standing, however the test is usually extended for a longer time of 5, 15, or 30 min to explore long-term HR and BP responses generated by the postural change. As with the VM, the correct interpretation of HR changes induced by active standing requires the simultaneous recording of ECG and beat-to-beat BP. The Valsalva and 30:15 ratios, which are less sensitive markers of cardiovagal dysfunction than the deep breathing ratio, depend instead upon complex central and peripheral interactions rendering them less sensitive to identify mild autonomic abnormalities (Ewing et al., 1985). All these ratios are reproducible in healthy subjects but show a clear decline with age, as does the HRV which is sensitive to the presence of ectopic beats, technical artifacts, cardiac arrhythmias, inadequate identification of R waves, and wrong quantification of RR

307

intervals. The reliability of these indicators is severely limited in patients with HR > 100 bpm (Hilz and Dutsch, 2006).

Heart rate variability at rest The HRVat rest can be quantified in the time and spectral or frequency domains. Different periods of time could be analyzed: 1 min, 5 min, or 24 h. The variability could be quantified using simple indicators like the coefficient of variability which is defined as the count of the number or percentage of intervals that exceed certain duration or more complex indicators of variability like the standard deviation of RR intervals and the square of the differences between consecutive RR intervals are also employed (AAN committee, 1996). Spectral domain analysis, which quantifies the frequency and intensity of periodic changes of HRV in a period of time, show that rapid HRV oscillations depend upon the influence of breathing patterns and cardiac activity mediated by the parasympathetic system while slow changes in HRV are related to BP changes controlled by both parasympathetic and sympathetic influences. The ratio of low frequency (LF) and high frequency (HF) bands (LF/HF) is a useful indicator of the balance between the parasympathetic and sympathetic divisions (Niskanen et al., 2004). Therefore, a valuable application of the spectral analysis of HRV is the potential of estimating the relative contribution of the two ANS divisions in the control of HR more effectively than other methods of analysis. The spectral power of the HF band which has traditionally been accepted as a reliable indicator of cardiovagal activity is superior to that of the LF band that depends instead upon the combination of sympathetic and parasympathetic influences on the vascular and cardiac functions. The study of the balance between parasympathetic and sympathetic influences on the cardiovascular system is important because it has been extensively demonstrated that subjects with absolute or relative predominance of sympathetic activity are more likely to suffer myocardial infarction, cardiac arrhythmias, hypertension, sudden death, gout, obesity and premature aging than those with more balanced ANS function (Sica, 2000).

Beat-to-beat blood pressure and vascular sympathetic control The autonomic innervation of blood vessels and skin is mainly under the control of the sympathetic nervous system. The ANS is essential for the short-term regulation of BP including the fast compensation of transient changes that occur a few seconds after postural, hemodynamic and emotional changes (Wehrwein et al., 2016). Given the extended length of the sympathetic fibers throughout

308


J. GUTIERREZ

peripheral nerve trunks, it is likely that in the course of some neurological diseases, as for example lengthdependent neuropathy, peripheral sympathetic fibers are commonly injured earlier or more severely than parasympathetic fibers, the paths of which are usually much shorter (Guti
errez et al., 1998; Low et al., 2006). The recording of beat-to-beat BP changes is essential for evaluating the sympathetic control of peripheral blood vessels. Early investigations employing catheters inserted inside peripheral arteries provided very accurate data but were invasive and painful, and if continued in this manner, such analyses would have remained in research. Fortunately, noninvasive devices are available to record beat-to-beat BP for clinical use. There are two types of these devices: photoelectric cell–based sensors (Schutte et al., 2003; Maestri et al., 2005) and those that monitor the mechanical deformation of the arterial wall with a tonometric transducer (Zion et al., 2003). These transducers transform beat-to-beat BP oscillations into a curve as a function of time in which systolic peaks and diastolic valleys of each cardiac cycle amenable to identification and measurement (Fig. 13.3) (Langewouters et al., 1998).

Changes in beat-to-beat BP induced by Valsalva maneuver The evaluation of the changes in beat-to-beat BP induced by a Valsalva maneuver provides very valuable information about the functional state of peripheral vascular innervation (Denq et al., 1998). These changes are comprised of 4 distinctive phases (Fig.13.4). Phase I is associated with an increase in BP due to the mechanical compression motivated by the strain on thoracic great vessels. Phase I causes a transient bradycardia lasting 1–2 s. Phase IIA is characterized by reduced venous return and lowered cardiac output that follows compression of the great thoracic veins. Phase IIA is associated with a progressive decrease in BP lasting 5–7 s and baroreflex-mediated tachycardia that fails to reverse the fall in BP. Phase IIB is associated with progressive recovery of BP which lasts 5–7 s that results from the combined activation of the vasomotor and cardiovagalmediated tachycardia, initiated in phase IIA. Phase III is the mirror image of phase I wherein the brief drop in BP caused by mechanical decompression of thoracic great vessels, at the end of the strain phase, leads to

Fig. 13.3. Correlation between beat to beat blood pressure and muscle sympathetic nerve activity during a Valsalva maneuver. (A) Monitoring the duration and intensity of the respiratory effort. (B) Electrocardiogram. (C) Tonometric recordings of beat to beat blood pressure. (D) Microneurographic recordings of muscle nerve sympathetic activity from a peripheral nerve. Note that sympathetic nerve activity increases in response to blood pressure reduction in phase II and decreases following blood pressure recovery in phases IIB and IV (arrows).

PERIPHERAL AND CENTRAL AUTONOMIC DISORDERS

309

Fig. 13.4. Curves of RR interval duration (top tracing) and beat to beat systolic blood pressure (lower tracing) vs. time, recorded in a healthy subject while performing a standardized Valsalva maneuver. The heart rate and blood pressure changes associated to this maneuver are typically subdivided in four phases (roman numerals). Phases I and III are mainly due to mechanical compression of the large thoracic blood vessels.

progressive tachycardia. During phase IV the release of mechanical thoracic compression abruptly increases venous return and cardiac output, which in association with ongoing vasoconstriction, overshoots BP well above baseline values reaching a maximum in 5 s followed by a slow decline toward normal levels. The HR curve reveals a reflex bradycardia that reaches maximum in a few seconds after BP maximum and then falls to normal levels (Benarroch et al., 1991; Sandroni et al., 1991). Whereas BP recovery in phase II relies more upon increased vascular resistance mediated by a-adrenergic vasomotor fibers than increased HR mediated by cardiovagal parasympathetic inhibition (Sandroni et al., 1991), BP recovery during phase IV is a consequence of the combined effects of vascular a-adrenergic, cardiac b-adrenergic and cardiovagal functions. Mild a-adrenergic sympathetic disturbances yield delayed or impaired BP recovery during phase II with preserved phase IV. The differences between the physiological mechanisms underlying phases II and IV is shown by the administration of selective parasympathetic and sympathetic blocking medications. The administration of the a-adrenergic antagonist phentolamine produces a decrease in BP during phase II and an increase in BP during IV in normal subjects, while the b-adrenergic antagonist decreases the physiological increase in BP in phase IV without affecting BP in phase II, indicating that BP and HR

changes in in phase II are more dependent on a-adrenergic innervations, while those of phase IVare more reliant on b-adrenergic cardiac innervation. In general, a greater importance is attached to marked changes or absence of phase IV, and the reduction of BP recovery in phase IIA or its absence in phase IIB (Ferrer et al., 1991). The magnitude of quantifiable BP changes referenced to premaneuver baseline values, expressed as a percentage of the decrement of BP reached during phase IIA is a quantitative and standardized way of comparing to normal reference values. Such quantification may be more useful in detecting a-adrenergic abnormalities such as those associated with peripheral neuropathy, than other markers of the sympathetic ANS (Guti
errez et al., 2002). The BP changes that occur during the VM have been studied in patients with diabetic, alcoholic, demyelinating and nutritional neuropathies, and in those with neurally mediated syncope and orthostatic hypotension, in the early stages of which there may be delayed or ineffective recovery of BP during phase II, with other phases remaining normal (Fig. 13.5) (Ferrer et al., 1991; Guti
errez et al., 2002).

Changes in beat-to-beat BP induced by active and passive standing The sympathetic control of vascular tone is essential for maintaining BP during standing and the changes therein

310


J. GUTIERREZ

Fig. 13.5. Abnormalities in blood pressure change during the Valsalva maneuver in patients with autonomic disorders in the course of peripheral neuropathy. Blood pressure drop during phase IIA is deeper and more prolonged in patients with polyneuropathy than in normal controls. These patients have also a poor recovery of blood pressure in phase IIB (remains below baseline values). However, both groups have a similar blood pressure recovery in phase IV.

provide valuable information of the integrity of the baroreflex and other autonomic reflexes (Wehrwein et al., 2016). Standing and gravitational stress evokes an immediate displacement of 500–1000 mL of blood to the lower part of the body reducing BP and activating the baroreflex with resultant adjustments in vascular tone and HR. After several seconds the slight drop in BP generates a vagal inhibitory response and reflex tachycardia that reaches a peak at 15 s, which in association with progressive vasoconstriction, is followed by recovery in BP and reflex bradycardia that reaches a maximum at 30 s. Over the ensuing 1–2 min the BP remains steady in most subjects due to combined sympathetic and cardiovagal adjustments. Most of the ANS changes related to standing occur within the first 5 min while the long-term control of BP includes hormonal changes that influence vascular tone and fluid displacement (Wieling and Shepherd, 1992). While initial active standing in normal subjects leads to a slight decrease in BP and increase in HR of less than 30 bpm, in those with orthostatic hypotension, BP is severely reduced in the first few seconds and minutes after active standing. The criteria that define postural hypotension vary between laboratories but a symptomatic drop in systolic BP > 30 mmHg is conclusively abnormal. A severe drop in BP should lead to consideration of poor vascular sympathetic tone. A drop in BP may be associated with either tachycardia or bradycardia, depending upon the primary underlying cause of the hypotension. Some patients with chronic orthostatic hypotension experience persistent and pronounced tachycardia, a situation encountered in postural tachycardia syndrome

(POTS). Whereas normal subjects undergoing passive tilting of 60° to 90° can develop a slight transient drop in systolic, diastolic and mean BP that recovers in 1 min, and HR that increases from 10 to 20 bpm, those with significant adrenergic impairment have inadequate compensatory vasoconstriction leading to postural hypotension and a progressive fall in BP even during passive standing. The sensitivity of the tilt table test for confirming neurally mediated syncope varies from 40% to 60% with up to 50% of patients with a compatible clinical history demonstrating completely normal tilt table results. Accordingly, those with suspected neurally-mediate syncope that manifest normal HR and BP responses to passive standing should be studied for at least 30 to 45 min to increase the sensitivity of the study (Wieling and Shepherd, 1992).

SUDOMOTOR REFLEXES The regulation of body temperature is an important function of the ANS, and the control of sweating is essential for thermal homeostasis. Central and peripheral sudomotor sympathetic function is assessed by recording the responses of sweat glands to various types of stimuli. Such studies are especially useful in the early detection of small fiber neuropathy lesions (Low et al., 2006).

Thermoregulatory sweat test This test involves the application of a substance on the region of the body under examination capable of changing color in response to induced sweating. Sweating is stimulated when overall body temperature is raised after

PERIPHERAL AND CENTRAL AUTONOMIC DISORDERS placement in a room with elevated temperature or following exposure to a heating lamp to selectively stimulate a specific region of the body. The most commonly employed substances are starch iodide and corn starch with calcium carbonate, which change to a darker color upon contact with sweating. These studies are very useful to show the topographic pattern of sudomotor disturbances but are unable to differentiate central from peripheral autonomic disorders, and preganglionic or postganglionic sympathetic sudomotor disturbances (Fealey et al., 1989).

Sweat imprinting In this test, a silicone mold or other easily imprintable substances are applied over the body in the regions of postganglionic sympathetic-innervated sweat glands and activated by iontophoretically applied pilocarpine, leading to the accumulation of droplets of dermal sweat that becomes trapped in the mold, the volume of which can be ascertained (Kennedy and Navarro, 1989). The association of severe reduction in sweating with normal response to iontophoresis is consistent with a preganglionic or central autonomic injury.

Sympathetic skin response The SSR is a late electrical response of hairless skin characterized by a long reflex arc that involves central and peripheral ANS components, the former of which include synaptic connections from posterior hypothalamus to the brainstem reticular formation and spinal cord, and latter of which include pre-and postganglionic sympathetic sudomotor fibers and the sweat glands. Under physiological conditions of normal room temperature, the skin of the soles and palms is usually 10 to 25 mV more negative than that of other body regions, with spontaneous electrical oscillations. The SSR is generated in the deeper layers of the glabrous skin, due to the reflex activation of sweat glands, stimulated by acetylcholine secreting sudomotor sympathetic efferent fibers. The final morphology of the electrical responses recorded in the skin surface depends on the interaction between sweat glands and the surrounding skin tissue. Latency, amplitude, and morphology of the SSR are highly variable in the same subject during a given experimental session (Shahani et al., 1984). Central efferent fibers related to the SSR originate in the hypothalamus and travel caudally and ipsilaterally through the lateral columns of the spinal cord, synapsing upon preganglionic sympathetic neurons of the intermediate horn of the spinal cord. Preganglionic sympathetic fibers innervating the arms exit the spinal cord at the T4 to T7 spinal cord levels, while those innervating the legs depart from T10 to T12 segments. Postganglionic sympathetic fibers give

311

rise to sympathetic sudomotor fibers that innervate sweat glands of the hands and feet (Shields, 1993). The SSR responses are influenced by emotional, auditory and light stimuli, and in order to avoid the interference of these external factors, they should be performed in a quiet, sound-proof and temperature controlled room. Active electrodes are placed on hairless skin of the palms and soles and referenced to electrodes located on the hairy skin of the dorsum of the hands and feet. Electrophysiological activity is collected simultaneously from the arms and legs for 5–10 s after the stimulus (Fig. 13.6). Square pulses of electrical current lasting 0.2 ms, with an intensity of 10–30 mA, 2–3 fold the sensory threshold are delivered to the subject at irregular intervals of 30–60 s, with varying intensity, to any sensory nerve; however, the supraorbital, median, sural, and posterior tibial are routinely used. If stimulation of a particular nerve is not effective to generate responses, others can be tried to avoid the effect of focal lesions of the afferent reflex. Electrical stimuli that fail to evoke responses can be switched to auditory, emotional or light stimulation, as well as, brisk and deep inspiration to induce SSR. With an average conduction velocity of 1–2 m/s, conduction through type C postganglionic fibers and sweat gland activation account for more than 95% of the SSR latency (Shahani et al., 1984). The total amplitude of the SSR depends on the interaction of two components, the activation of the sweat gland itself and the influence of the surrounding skin tissue. Given its intrinsic vulnerability to emotional factors, the amplitude of the SSR shows 35%–45% variability, decreasing with age with up to one-half of normal subjects of age 60 years or older, demonstrating no recordable SSR. Accordingly, as a result of the expected physiological variability, some consider the SSR abnormal only when absent (Shahani et al., 1984). One advantage of the SSR is that it can be recorded with simple technology available in most clinical neurophysiology laboratories. The SSR has been used extensively in the study of diabetic, acute demyelinating, nutritional and toxic peripheral neuropathies, and in CNS disorders including Parkinson’s disease (PD), multiple system atrophy (MSA), multiple sclerosis (MS), and stroke. It is severely decreased or absent in patients with severe autonomic neuropathy, however, responses may remain within normal limits in up to 50% of patients of varying degrees of diabetic, uremic, amyloid, and alcoholic peripheral neuropathy, even in the presence of clinical dysautonomia. The responses have no predictable clinical-electrophysiological correlation (Guti
errez et al., 1998) and for uncertain reasons, absence of SSR should not be considered a unique result of sympathetic skin denervation because other factors can affect local skin conductance including trophic and callous skin

312


J. GUTIERREZ

Fig. 13.6. Sympathetic skin responses recorded in the four limbs. C1 and C5: Right palms, C2 and C6: Left palms, C3 and C7: Right plants, C4 and C8: Left plants. The onset latencies were approximately 1.5 s in the upper limbs and 2 s in the lower limbs (delay: 1 s).

changes. The diagnostic sensitivity of the SSR is higher in patients with PD, MSA, pure autonomic failure (PAF), and traumatic spinal cord injury, than in patients with peripheral neuropathy. Tetraplegic patients with spinal cord injuries above the T3 segment demonstrate absent SSR in the arms and legs (Arunodaya and Taly, 1995). An important factor in the interpretation of SSR is whether the causative lesion affects pre or postganglionic fibers. The most useful test to demonstrate injury to postganglionic sudomotor sympathetic fibers is the quantitative sudomotor axonal reflex test (QSART), which is performed by iontophoretically introducing pilocarpine into the dermis to directly stimulate the sweat glands. Action potentials of activated glands travel antidromically to the nearest axonal branching point where they are propagated to the other sweat glands innervated by the same postganglionic fiber, from which an electrical response can be recorded and quantified. Therefore, the functional integrity of the postganglionic axons is essential for the onset of normal QSART responses (Low, 2003). Concomitantly absent SSR and normal QSART responses suggest abnormality along preganglionic sudomotor fibers or of the CNS, a pattern which can be observed in those with MS, spinal cord injury, and stroke (Hilz and Dutsch, 2006). Whereas abnormality of SSR and QSART testing show a high correlation, the QSART is more sensitive than the SSR. The QSART can be evaluated also in proximal portions of the arms

and legs to prove the existence of a length-dependent pattern of abnormalities. In the interpretation of the QSART, it should be noted that a normal QSART does not exclude the existence of central or preganglionic disorders, because this test does not assess these levels of the sudomotor pathways. The function of postganglionic sudomotor fibers and sweat glands can be evaluated also by measuring the electrochemical skin conductance with the Sudoscan test (Casellini et al., 2013). This test is based on the principle of reverse iontophoresis of chloride ions through the skin. It is relatively new, noninvasive and easy-to-perform. Subjects are asked to place their hands and feet on stainless steel–based plate electrodes which deliver a low voltage (6 months of age (Mercuri et al., 2018) showed benefit of treatment in the primary endpoint of a least-squares mean increase in the primary endpoint of the baseline to month 15 score on the Hammersmith Functional Motor Scale-Expanded (HFMSE) compared to controls (57% vs 26%). The NUTURE trial (De Vivo et al., 2019), an ongoing, phase 2, open-label, single-arm, multinational study of intrathecal nusinersen treatment of 25 children who initiated treatment early in infancy prior to the onset of SMA, reported follow-up after 2.9 years. None of the 25 subjects (15 of whom had two SMN2 copies and 10 had three SMN2 copies) met the primary endpoints of death or respiratory interventions; moreover all 25 were able to sit without support, and 88% were walking independently or with assistance. The functional outcomes and motor milestones (including independent walking) of subjects with three SMN2 copies were generally better than those with two copies. The findings highlight the substantial benefit of early therapy and point to the value of early diagnosis. Onasemnogene The FDA approved the biologics license application to market onasemnogene abeparvovec-xioi or onasemnogene (Zolgensma™) in May 2019 via single intravenous dose injection of adeno-associated viral vector (AAV) serotype 9 carrying SMN complementary DNA encoding the missing SMN protein. The Gene Transfer Clinical Trial for Spinal Muscular Atrophy Type 1 (Mendell et al., 2017) was a phase 1 open-label, dose-escalation 2-year safety trial of intravenously injected low- and high-dose AVXS-101 (onasemnogene abeparvovecxioi) in conjunction with corticosteroids with a primary safety and secondary outcomes of time to death or the need for permanent ventilatory assistance. All 15 patients remained alive and were event-free at 20 months of age, as compared with a rate of survival of 8% in a historical cohort. In the high-dose cohort, a rapid increase from baseline in the score on the CHOP INTEND scale followed gene delivery, with an increase of 9.8 points at 1 month and 15.4 points at 3 months, as compared with a decline in this score in a historical cohort. Of the 12 patients who had received the high dose, all patients were alive and event-free at 20 months of age; and 11 sat unassisted, 9 rolled over, 11 fed orally and could speak, and 2 walked independently. Liver enzyme elevation commonly seen in AAV therapy resolved with prednisolone treatment; and there were no clinically symptomatic acute immune-mediated responses. The Gene Replacement Therapy Clinical Trial for Participants with Spinal Muscular Atrophy Type 1

320

D.S. YOUNGER

(STR1VE) completed enrollment of SMA type 1 subjects and 1 or 2 copies of SMN2 and age 20). The CMTNS correlates well with other measures of disability including ambulation index, self-assessment, hand function, 9-hole peg test, and neuropathy impairment score. The CMT symptom score (CMTSS) and CMT examination score (CMTES) are subscores of the CMTNS, calculated by the sum of the symptoms (CMTSS) or the sum of the symptoms plus the signs (CMTES). The overall neuropathy limitation score (ONLS) (Graham and Hughes, 2006), is a composite of two scores, one for upper limb function and the other for lower limb function based on a person’s symptoms and gait analysis. The ONLS was employed as a secondary outcome measure in several of the initial ascorbic acid in CMT1 trials and was shown to be insensitive to change over the duration of the trial. Despite its lack of sensitivity, the ONLS remains one of only a handful of outcome measures that have been approved by the FDA for clinical trials in CMT. Although the CMTNS proved to be a useful measure of CMT severity, after using this scale for several years

327

(Shy et al., 2007; Pareyson et al., 2011), it was evident that some of the items had a ceiling or floor effect. Implementing the CMT Neuropathy Score (CMTNS) and the Short Form (SF)-36 in patients with CMT1A, Tozza and colleagues (Tozza et al., 2018) found a deterioration of all clinical measures after age 50 that was greater than controls. Thus, following the 168th ENMC International Workshop on outcome measures and clinical trials in CMT (Reilly et al., 2006), it was decided to make some modifications to the CMTNS to reduce floor and ceiling effects and to standardize patient assessment, aiming to improve its sensitivity for detecting change over time. This resulted in the updated version of the CMTNS (CMTNS2). The CMT Examination Score version 2 (CMTESv2) and Rasch-modified version (CMTESv2-R) are subscores of the CMTNSv2 comprising seven items from the patients’ symptoms and examination findings and was used as a primary outcome measure for the natural history study of CMT2A (Pipis et al., 2020) and CMT type C (Pan et al., 2020). For CMT1A, the commonest subtype of CMT and for many of the other types of CMT, in which onset of the disease is typically in the first or second decades of life, monitoring of progression in childhood presents particular difficulties due to the inherent changes in an outcome measure as the child grows and develops. The CMTPedS score comprises functional outcome measures for pediatric CMT (Burns et al., 2012a). The Charcot–Marie–Tooth Functional Outcome Measure (CMT-FOM) is a performance-based measure assessing functional ability in adults with CMT which has been designed to incorporate many of the items from the CMTPedS (Eichinger et al., 2018). It utilizes a number of outcome measures including hand grip, ankle–foot dorsiflexion and plantarflexion strength, nine-hole peg test to assess functional dexterity, balance beam, timed walk, stair climb and a timed up and go. The CMT-FOM is able to detect impairment in patients with CMT compared to controls. Its future use in clinical trials depends on the outcome of active longitudinal studies examining the psychometric properties of the score and its responsiveness to change. In RCTs, meaningful and clinically important differences should reflect significant benefits to a patient’s QoL and thus should be considered in the construct of a valid primary endpoint. To ensure that a patient’s QoL can be directly measured, a CMT health index questionnaire was developed (Johnson et al., 2018) that contains 18 themes identifying factors related to CMT disease burden. The score has been shown to have high internal consistency and test retest reliability and is able to discriminate between patients in different disease states.

328

D.S. YOUNGER

Table 14.3 CMT neuropathy score. Score Parameter

0

Sensory None symptoms Motor Symptoms Legs None

1

2

3

4

Limited to toes

Extend up to and may include ankle

Extend up to and may include knee

Extends above knees

AFO on at least 1 leg or ankle support Unable to do buttons or zippers but can write Reduced up to and may include wrist/ankle Reduced at wrist/ankle

Cane, walker, ankle surgery Cannot write or use keyboard Reduced up to and may include elbow/knee Reduced at elbow/knee

Wheelchair most of the time Proximal arms

3 Foot dorsiflexion

3 Dorsiflexion and plantar flexion 6 mV 4.0–5.9 mV (>4 mV) (2.8–3.9) >9 mV 6.0–8.9 mV (>22 mV) (14.0–21.9)

3 Intrinsic or finger extensors 2.0–3.9 mV (1.2–2.7) 3.0–5.9 mV (7.0–13.9)

0.1–1.9 mV (0.1–1.1) 0.1–2.9 mV (0.1–6.9)

Reduced above elbow/knee Reduced above elbow/knee Proximal weakness Weak above elbow Absent (Absent) Absent (Absent)

Adapted from Barohn RJ, Herbelin L & Dimachkie MM (2022). Clinical parameters in neuromuscular disease. In: Younger DS (ed.) Motor Disorders. 4th ed. Lanham: Roman and Littlefield. Abbreviations: AFO, ankle-foot orthosis; CMAP, compound muscle potential; SNAP, sensory nerve action potential.

RANDOMIZED CONTROLLED TRIALS With the development of preclinical models of CMT, together with advances in Adeno-Associated Viral (AAV) gene replacement therapy and antisense oligonucleotide (ASO)/RNAI biochemistry (Zhao et al., 2018) and NGS panels, more patients with CMT are receiving a genetic result than ever before. Thurs, the field is at a critical juncture to proceed to RCTs and test potentially useful therapies (Rossor et al., 2020).

Ascorbic acid There are 5 published RCT comparing the effects of oral ascorbic acid (1–4 g) with placebo in CMT1A for 1 year that have collectively enrolled 80 children (Burns et al., 2009) and 571 adults (Micallef et al., 2009; Verhamme et al., 2009; Pareyson et al., 2011; Lewis et al., 2013). All four trials in adults used the CMTNS as a composite disease-specific score; with other end-point measures including the CMTES (clinical component of the CMTNS), nine-hole peg test, 10-m walk test, hand grip

force, three-point pinch force, SF-36 bodily pain and ascorbic acid concentration. The trial in children used the median motor NCV at conclusion as the primary end-point measure, with secondary outcomes of foot and hand strength, motor function, walking ability, and QoL. Two studies continued treatment for 12 months (Micallef et al., 2009; Verhamme et al., 2009), and two for up to 24 months (Pareyson et al., 2011; Lewis et al., 2013). There were no significant differences reported in the primary outcome measure impairment of the CMTNS or the Neuropathy Impairment Score (NIS) (Bril, 1999) at 12 months, nor in secondary end-point measures of change in disability or sensory impairment on a validated scale, or a change in amplitude of the CMAP or grip force or other measures of force, or QoL. However, one primary end-point measure, the CMTES, that represents the CMTNS without the electrophysiological parameters, showed a significant benefit for ascorbic acid in one trial (Micallef et al., 2009), as well as the secondary measures of the sensory nerve action potential (SNAP) and the nine-hole peg test were reported as reaching significance in adults. Moreover, the ascorbic acid-treated cohort in children

ON THE PATH TO EVIDENCE-BASED THERAPY IN NEUROMUSCULAR DISORDERS (Burns et al., 2009) had a small, nonsignificant increase in median motor NCV compared with the placebo group.

FUTURE DIRECTIONS It is fair to conclude that ascorbic acid does not provide significant benefit for CMT1A nor would differing results be expected with further adaptation of study design or longer time periods (Gess et al., 2015). The finding of improvement in the median motor NCV in CMT1A children over the 2-year follow-up period (Burns et al., 2009) likely indicates partial restoration of peripheral nerve myelin or the reduction in over-thick myelin (Manganelli et al., 2016). If so, motor NCV could represent, at least in children, a useful outcome measure in future clinical trials of CMT1A.

Chronic inflammatory demyelinating polyradiculoneuropathy BACKGROUND The first case of recurrent neuritis was published by Eichhorst (1890). A century later, Austin (1958) identified steroid responsiveness and recurrence and CSF protein elevation, and Dyck et al. (1975) described the clinicopathological findings of monophasic progression, steady deterioration, stepwise progression, and prototypical inflammation and demyelination that can be seen in cutaneous nerve biopsy specimens (Fig. 14.7). Dyck and Arnason (1984) named the disorder, which accounted for 21% of initially undiagnosed patients with peripheral neuropathy at the Mayo Clinic (Dyck et al., 1981), and 13% to 20% of cases of immune-mediated neuropathy referred to academic centers worldwide (Barohn, 1998).

329

CLINICAL PRESENTATION Typical CIDP begins with paresthesia and weakness in the legs, as well as difficulty walking. Typical CIDP occurs at any age, from infancy and childhood (Nevo et al., 1996; Simmons et al., 1997) but most commonly between 40 and 60 years. The disease course steadily progresses for more than 8 weeks from onset; however, up to 13% of patients rapidly progressed over 4 weeks (Ruts et al., 2005), leading to uncertainty in the diagnosis in a third (Ruts et al., 2010). There are no specific clinical features or laboratory tests that distinguish CIDP from acute inflammatory demyelinating polyradiculoneuropathy (AIDP) in the acute stage of the disease. Affected patients can experience relapses after initial improvement, however, without cranial nerve, respiratory and autonomic involvement (Stojkovic et al., 2003; Henderson et al., 2005; Yamamoto et al., 2005; Stamboulis et al., 2006). Weakness varies in severity but is symmetric and characteristically involves proximal and distal leg muscles. Sensory manifestations consist of numbness and tingling, but painful paresthesia is not uncommon especially in the thighs at onset. Tendon reflexes are usually depressed or absent by the time the patient see a physician attesting to the insidiously slow decline.

DIAGNOSIS AND TREATMENT GUIDELINES The diagnosis of CIDP rests upon a combination of clinical, electrodiagnostic, and laboratory features with attention to variants and exclusions of mimics (Eftimov et al., 2012) to avoid misdiagnoses. Formal criteria have been closely linked to the performance of electrodiagnostic studies recognizing that errors in performance and

Fig. 14.7. Chronic inflammatory demyelinating polyradiculoneuropathy. Cutaneous nerve biopsy shows a few mononuclear cells surrounding an endoneurial blood vessel (curved arrow). The number of large myelinated fiber is decreased. Some thinly myelinated fibers have excessive Schwann cell process proliferation for onion bulbs (arrowhead). Toluidine Blue stain. 40.

330

D.S. YOUNGER

interpretation lead to misdiagnosis (Allen et al., 2018), errors in the performance or interpretation of electrodiagnostic studies, or the misperception of treatment benefit (Allen and Lewis, 2015; Kaplan and Brannagan, 2017). The standardization of formal criteria and guidelines for CIDP has been a long road. The cardinal electrodiagnostic features of CIDP include slow motor and sensory nerve conduction velocities, prolonged distal latencies and F wave latencies, partial conduction block, and abnormal temporal dispersion in nerves of more than one limb. However, 120%a

>150%b

49 50 41 41

39.2 40.0 32.8 32.8

34.3 35.0 28.7 28.7

4.5 3.6 6.6 6.0

5.6 4.5 8.2 7.5

6.7 5.4 9.9 9.0

31.0 32.0 58.0 58.0

37.2 38.4 69.6 69.6

46.5 48.0 87.0 87.0

Adapted from Dimachkie MM & Barohn RJ (2022). Chronic inflammatory demyelinating polyradiculoneuropathy and related disorders. In: Younger DS (ed.) Motor Disorders. 4th ed. Lanham: Roman and Littlefield. a If Amp > 80% LLN b If Amp < 80% LLN If: Median CMAP LLN 4.5 mV; then 80% LLN ¼ 3.6 mV. Ulnar CMAP LLN 5.0 mV; then 80% LLN ¼ 4 mV. Peroneal CMAP LLN 2.0 mV; then 80% LLN ¼ 1.6 mV. Tibial CMAP LLN 4.0 mV; then 80% LLN ¼ 3.2 mV. Abbreviations: CMAP, compound muscle action potential; DL, distal latency, LLN, lower limit of normal; ms, millisecond; mV, millivolt; NCV, nerve conduction velocity; ULN, upper limit of normal.

Table 14.6 INCAT disability scale. Arm disability 0 ¼ No upper limb problems 1 ¼ Symptoms, in one or both arms, not affecting the ability to perform any of the following functions: doing all zips and buttons; washing or brushing hair; using a knife and fork together; and handling small coins 2 ¼ Symptoms, in one arm or both arms, affecting but not preventing any of the above-mentioned functions 3 ¼ Symptoms, in one arm or both arms, preventing one or two of the above-mentioned functions 4 ¼ Symptoms, in one arm or both arms, preventing three or all of the functions listed, but some purposeful movements still possible 5 ¼ Inability to use either arm for any purposeful movement Leg disability 0 ¼ Walking not affected 1 ¼ Walking affected, but walks independently outdoors 2 ¼ Usually uses unilateral support (stick, single crutch, one arm) to walk outdoors 3 ¼ Usually uses bilateral support (sticks, crutches, frame, two arms) to walk outdoors 4 ¼ Usually uses wheelchair to travel outdoors, but able to stand and walk a few steps with help 5 ¼ Restricted to wheelchair, unable to stand and walk a few steps with help Overall disability ¼ Sum of arm and leg disability Adapted from Barohn RJ, Herbelin L & Dimachkie MM (2022). Clinical parameters in neuromuscular disease. In: Younger DS (ed.) Motor Disorders. 4th ed. Lanham: Roman and Littlefield.

(18–29 years, 30–59 years, and >60 year) and duration of symptoms (6 months to 1.9 years, 2–3.9 years, and >4 years) and randomized to treatment or no treatment with prednisone begun at 120 mg every second day,

falling to 0 mg tapering on alternate days by 5 mg over 12 weeks. Muscle strength, cutaneous sensation, CSF protein, and NCS were not significantly different between treated and control groups. The prednisone group included 7 patients with a progressive and 7 with a recurrent course, while the untreated group included 12 patients with a progressive course and 2 with a recurrent course. Prednisone was shown to cause a small but significant improvement over no treatment in the neurological disability score (NDS) (Dyck et al., 1980), computer-assisted sensory vibratory thresholds in the left hand and foot; maximum expiratory and inspiratory (PEmax and PImax) using a pressure bugle, graded muscle strength, and motor and sensory NCS. No subset of patients was more likely than another to be responsive to prednisone; as were both groups with either progressive or relapsing course. The investigators were unable to show that change in NDS was related to either course (progressive or recurrent), duration, CSF findings, or initial NDS. Nor could the answer to the value of prednisone usage for >12 weeks when balanced against the risk of AEs including hyperglycemic hyperosmolar coma, cataracts of the eye, infection, and aseptic necrosis of the hips. The alternative to oral daily and oral corticosteroid regimens is pulsed treatment with oral or IV preparations. Van Schaik and colleagues (van Schaik et al., 2010) reported the findings of 40 patients with newly diagnosed definite or probable CIDP randomized to either pulse high-dose dexamethasone (24 subjects) or standard oral prednisolone (16 subjects), noting that 16 patients achieved the primary outcome measure of remission at 12 months (10 dexamethasone, 6 prednisolone) defined as improvement of at least three points on the Rivermead mobility index and improvement of at least one point on the INCAT disability scale although not substantially

334

D.S. YOUNGER

different, with an odds ratio [OR] of 1.2 (95% CI: 0.3–4.4). Eftimov and colleagues (Eftimov et al., 2012) on behalf of PREDICT, compared the outcome of 6 monthly pulses (40 mg/day for 4 days a month) of dexamethasone with daily oral prednisolone (60 mg, slowly tapered over 8 month) in a prospective cohort of 40 patients assessed using the INCAT disability scale and Rivermead Mobility Index. Cure (>5 years off treatment) or remission was achieved in 10 (26%) patients. One half of the patients experienced a relapse after a median treatment-free interval that was (insignificantly) longer for those receiving dexamethasone (17.5 months) than prednisolone (11 months) indicating a similar effect in improving disability. According to the new Task Force of the 2021 EFNS/PNS (Van den Bergh et al., 2021) the best corticosteroid regimen is presently unknown. Pulsed high-dose corticosteroids with oral dexamethasone or IV methylprednisolone (IVMP) can be considered an alternative to daily oral prednisone/prednisolone or dexamethasone both for induction and maintenance treatment. The former agents generally have fewer side-effects that may include osteoporosis, gastric ulceration, diabetes, cataracts, avascular necrosis of long bones, arterial hypertension, which outweigh the benefit from treatment in low disability disease. In practice, patients so treated should be carefully monitored for treatment response, which usually starts after several weeks or months with attempted reductions in the dose steroid dose to investigate whether the current high dose is still required and whether and when the patient is achieving remission. Immune globulin Hughes and colleagues on behalf of the ICE Study Group (Hughes et al., 2008) first evidenced the short-term and long-term efficacy and safety of IVIg in CIDP in a double-blind, placebo-controlled response-crossover RCT enrolling 117 subjects meeting INCAT criteria for treatment with IVIg or placebo for up to 24 weeks, of whom 54% and 21%, respectively, achieved the primary outcome measure of sustained improvement in in the adjusted INCAT disability score. Overall, 94% of patients responded to 2 g/kg induction treatment and two subsequent treatments of 1 g/kg at 3 weeks intervals. Data extracted from the ICE trial (Hughes et al., 2008) by Latov et al. (2010) suggested that IVIg administered as a 2-g/kg loading dose followed by at least 2 courses of 1-g/kg maintenance administered 3 weeks apart was necessary to achieve a maximal therapeutic response. The open PRIMA (Leger et al., 2013) and PRISM (Nobile-Orazio et al., 2020) trials indicated a treatment response might only be observed after three to five infusions of 1 g/kg every 3 weeks. Then as now, it is unknown

whether the objective response following one or several treatments is due to a delayed treatment response or to the requirement of a different treatment regimen. While clinical experience suggests that switching products may be helpful in relieving side-effects, an RCT comparing 5% freeze-dried and 10% liquid IVIg preparations showed no difference in treatment efficacy (Kuitwaard et al., 2010). There is support by the 2021 EFNS/PNS 2021 Guidelines (Van den Bergh et al., 2021) for subcutaneous Ig (SCIg) as maintenance but not induction therapy for CIDP according to several clinical trials. Markvardsen and colleagues (Markvardsen et al., 2013) on behalf of the Danish CIDP and Multifocal Motor Neuropathy (MMN) Study group treated 30 adults with CIDP randomized to SCIg or placebo administered two or three times weekly for 12 weeks and assessed by isokinetic strength testing, and ODSS, 40-m-walking test (40-MWT), nine-hole-peg test, NIS, MRC score, grip strength, standardized electrophysiological recordings from three nerves, and plasma IgG levels, noting treated patients showing a significant benefit of SCIg compared to placebo. A year later, CIDP and MMN Study investigators (Markvardsen et al., 2014) reported the findings of 17 responders to IVIg in the previous study of SCIg vs placebo (Markvardsen et al., 2013) who were followed on therapy and evaluated after 3, 6, and 12 months noting an overall increase in isokinetic dynamometry by 7.2% suggesting it be considered as an alternative to IVIg for long-term management in CIDP. More recently, Danish CIDP and MMN Study Group investigators (Markvardsen et al., 2017) investigated whether multiple SCIg infusions were as effective as conventional loading dose therapy with IVIg in CIDP noting that treatment-naïve patients with CIDP, short-lasting SCIg and IVIg improved motor performance to a similar degree but with earlier maximal benefit following IVIg than SCIg. Twenty patients were randomized to either SCIg (0.4 g/kg/week) for 5 weeks or IVIg (0.4 g/kg/day) for 5 days for 10 weeks (Period I) and then crossed over to the opposite arm of therapy for an additional 10 weeks (Period II) whereupon they were evaluated for the primary end-point measure of isokinetic muscle strength and function. The investigators noted a similar effect in increase in isokinetic strength of 7.4% for SCIg and 6.9% with IVIg; however, improvement peaked 2 weeks after IVIg and 5 weeks after SCIg. Van Schaik and colleagues (van Schaik et al., 2019) conducted a 4 year open-label prospective PATH study in which adult subjects with CIDP were initially started on 0.2 g/kg or on 0.4 g/kg weekly were switched to 0.2 g/kg weekly after 24 weeks. After this dose reduction, 51% of patients relapsed (defined as a deterioration by at least 1 point in the total adjusted INCAT score)

ON THE PATH TO EVIDENCE-BASED THERAPY IN NEUROMUSCULAR DISORDERS of whom 92% improved after reinitiation of the 0.4 g/kg dose. Overall, two-thirds of patients who completed the PATH study remained relapse-free on the 0.2 g/kg dose. Immunoglobulin vs corticosteroids The new Task Force Guidelines (Van den Bergh et al., 2021) provide support for either treatment modality for CIDP citing that IVIg is preferable in the short-term especially when there are relative contraindications for corticosteroids. Several trials have compared corticosteroids and IVIg as first-line therapy to ascertain differences in short- and long-term effectiveness. Corticosteroids may be preferable for long-term treatment effectiveness because of a possible higher rate and longer duration of remission, especially when IVIg is unaffordable or unavailable. In 2012, Nobile-Orazio and colleagues (NobileOrazio et al., 2012) on behalf of the IMC Trial Group, randomized 45 subjects with CIDP in a double-blind, placebo controlled RCT to monthly IVIg (24 subjects) and IVMP (21 subjects) for 6 months with a primary outcome of the difference in the number of patients discontinuing either therapy owing to inefficacy or intolerance. After therapy discontinuation, patients were followed up for 6 months to assess relapses. More patients stopped methylprednisolone (11 [52%] of 21) than IVIg (three [13%] of 24), generally due to lack of efficacy (8 in the IVMP group vs 3 in the IVIg group). After therapy discontinuation, more patients on IVIg worsened and required further therapy (8 [38%] of 21) than did those on IVMP (0 of 10). Clinical improvement after IVIg may be faster and more enduring than IVMP according to Nobile-Orazio and IMC Trial Group investigators (Nobile-Orazio et al., 2015) who retrospectively compared the proportion of patients who eventually worsened after discontinuing therapy, noting that 87.5% of the patients who improved after IVIg therapy (as primary or secondary therapy after failing to respond to IVMP) were still improved after a median follow-up period of 42 months, compared with 54% of those treated with IVMP. Adrichem et al. (2020) on behalf of the OPTIC study group, extended the findings of an uncontrolled pilot study evaluating remission, rate of improvement and safety in 20 treatment-naive patients with CIDP receiving induction treatment with 2 g/kg loading dose and 1 g/kg maintenance every 3 weeks followed by 3-weekly 1-g IVMP infusions for a total of 18 weeks. The investigators hypothesized that combining the two agents would lead to more frequent remissions compared with IVIg alone, while maintaining the fast efficacy of IVIg at onset as induction therapy. Short-term combined induction treatment with IVIg and IVMP-induced remission in 10 of the 17 (59%) patients who completed the

335

treatment schedule, and 13 (76%) who showed improvement at 18 weeks as defined by a minimal clinically important difference (MCID) on the I-ROTS Disability Scale and/or an increase of 8 kPa in grip strength. This led to the protocol of an international double-blind RCT of IVIg and IVMP by Optic trial investigators (Bus et al., 2021) enrolling three group of “probable” or “definite” CIDP according to the EFNS/PNS 2010 criteria (Joint Task Force of the EFNS and the PNS, 2010; Van den Bergh et al., 2010) designated (1) treatment naïve, (2) known untreated CIDP with a relapse after >1 year; and (3) those improved within 3 months of a single course of IVIg, and subsequently deteriorated at any interval without having received additional treatment. Patients will be randomized to receive 7 infusions of 1000 mg of IVMP (intervention) or placebo (comparator) in combination with a loading dose (2 g/kg divided over 2–5 days) and 6 maintenance treatments of 1 g/kg (given over 2 days) every 3 weeks for 18 weeks. The primary outcome will be the number of patients in remission at 52 weeks after start of treatment defined as sustained improvement without the need for further treatment. Improvement will be defined by at least the MCID on the I-RODS and/or improvement of one or more points on the INCAT-DS at 18 weeks (end of treatment) compared to baseline. The trial finished recruiting its target group of 96 subjects in January 2022 with an estimated completion date of December 2024. Plasma exchange According to Oaklander and colleagues (Oaklander et al., 2017), in patients with good vascular access, PE may be an acceptable option for induction therapy of CIDP based upon very low certainty evidence at a dose of 2/week for 3 weeks followed by 1/week for 3 weeks. For maintenance treatment, there are no longterm efficacy or safety studies. Dyck and colleagues first established the efficacy of PE in CIDP more than three decades ago, demonstrating statistically significant improvement in the NDS and summated CMAPs in a sham-controlled, double-blind, prospective trial of PE in patients with static or progressive CIDP (Dyck et al., 1986). The treatment effect was rapid and large with only 15 patients in each arm of the trial, and with treatment lasting for only 3 weeks, and they inferred that neurological deficits probably were mainly due to alleviation of motor nerve conduction blocks. However, the authors later realized that this likely implied a large biological effect and in a later review of the CIDP (Dyck et al., 1993), advocated that PE be given intermittently over long periods to derive maximal benefit. The following year, Dyck et al. (1994) conducted a randomized blinded clinical study of 20 patients with CIDP assigned

336

D.S. YOUNGER

to PE or IVIg for an initial 6 week period followed by washout and a second period of cross-over treatment for 6 weeks. Primary end-points assessed before and after treatment for 6 weeks likewise included the NDS, weakness subset of the NDS, and summated CMAPs of the ulnar, median and fibular nerves; and secondary endpoint measures of the summated SNAPs of the median and sural nerves and computer-assisted vibratory detection threshold of the great toe. In a discussion of the three proven therapies for CIDP, prednisone, PE or IVIg should be recommended as primary therapy, the authors cited, similar to the 2021 Task Force (Van den Bergh et al., 2021), that since IVIg and PE are probably equally efficacious, there are certain situations in which IVIg would be preferred over PE. The first is the patient in whom venous access for PE is a problem. The second is when apheresis equipment is not readily available. Even in situations in which both modalities of treatment are available, IVIg may be preferred because treatment is simpler, expensive equipment is not needed and it is less invasive. Being able to give IVIg in the home is a further advantage of this treatment.

MYASTHENIA GRAVIS Background Myasthenia gravis is probably the best understood autoimmune disorder. None other has captured the attention of so many generations of neurologists and neuroscientists. Younger and colleagues (Younger et al., 1997) reviewed the history of MG. The first description of a patient with MG appeared in 1644 in correspondence from colonial Jamestown, Virginia (Marsteller, 1988) and described clinically more than two centuries later clinically by Wilks (1877), Erb (1879), and Goldflam (1893). Jolly (1895) named the disease “myasthenia gravis” pseudoparalytica and at the turn of the century, Campbell and Bramwell (1900) reported 60 cases of so-called MG, followed decades later, by a description of the efficacy of physostigmine by Walker (1934). Dale et al. (1936) described the chemical nature of neuromuscular transmission at motor end-plates, while Harvey and Masland (1941) summarized the salient electrophysiologic features. Blalock (1944) and Keynes (1946) asserted the role of thymectomy in the treatment of MG that included complete removal of the gland, whether or not a tumor was suspected preoperatively. The autoimmune cause of MG was suggested by Simpson (1960) and Nastuk et al. (1960); however, the immunologic basis of MG awaited basic understanding of acetylcholine (ACh) release at motor end-plates, as described by Katz and Miledi (Katz and Miledi, 1967).

Nature provided two important clues to the characterization of the nicotinic ACh receptors (AChR): one was the neuromuscular toxin, alpha-bungarotoxin (BuTx) isolated from krait snakes (Chang and Lee, 1966; Changeux et al., 1970), and the other was the electric organ of Torpediniformes (Torpedo) eels (Miledi et al., 1971) that served as rich reservoirs of AChRs. A turning point came shortly thereafter when Patrick and Lindstrom (Lindstrom et al., 1976b) injected rabbits with the electric organ intending to make AChR antibodies, and to see if they blocked the function of AChRs in intact electric organ cells. The antibodies did block; however, the immunized rabbits became paralyzed and died. Contemporaneously, Fambrough and colleagues (Fambrough et al., 1973) applied botulinum toxin to motor point biopsies from patients with MG and found a marked reduction in the number of AChRs, averaging 20% of controls. Both human MG and experimental autoimmune myasthenia gravis (EAMG) (Lennon et al., 1975) resulted from the autoimmune attack against native AChRs. By 1980, Toyka, Lindstrom, Engel, Lennon, Richman and colleagues (Toyka et al., 1975; Engel et al., 1976; Lindstrom et al., 1976a; Richman et al., 1980) reproduced the essential clinical and morphologic correlates of human MG in animals by passive transfer of human MG serum and AChR-specific monoclonal antibodies. The integral elements of neuromuscular transmission assure the functional success of different motor neurons to generate action potentials (APs) with characteristic temporal patterns to subserve diverse motor functions in turn, ensuring that all of the fibers in each motor unit produce a contraction suited to a particular need and function. For example, at the level of the motor neuron, those innervating slow muscle fibers tend to be relatively small, have a low threshold and fire continuously at a low frequency (e.g., 20 Hz), whereas those innervating fast fibers are larger, have a higher threshold, and fire in brief high frequency (e.g., 100 Hz). An important feature of a given pattern of firing is that every impulse in the motor axon should reliably excite every muscle fiber in the motor unit. At the ultrastructural level, normal excitability depends upon a variety of factors which ensure the security of neuromuscular transmission. This involves the concerted actions of elements of the NMJ (or endplate) in particular AChRs depicted in Fig. 14.8, which depend upon the release of ACh contained in synaptic vesicles along specialized areas of the presynaptic membrane called active zones in association with calcium channels where vesicular release occurs. Each vesicle contains about 5000 molecules of ACh and there are a large number of synaptic vesicles in each bouton. The boutons cluster in rows along specialized regions of the presynaptic membrane that facilitate the secretion

ON THE PATH TO EVIDENCE-BASED THERAPY IN NEUROMUSCULAR DISORDERS

337

Fig. 14.8. Structure of the NMJ. (A) Diagram of the mouse NMJ showing three muscle fibers, two of which are innervated by branches of a single motor axon. The highly branched nerve terminals are each capped by several Schwann cells. (B) Human NMJ. The nerve terminal (green) branches extensively within the NMJ. The terminal boutons are aligned with regions of high AChR (red) density in the muscle fiber surface. Many nuclei (blue) from muscle and Schwann cells are seen. (C) Ion channels in the muscle fiber surface. AChR clusters (green) are surrounded by regions of high NaV1 (red) density. Regions of optical overlap appear yellow. (D) Mouse NMJ showing nerve, muscle and Schwann cells. The AChRs have been labeled with an electron-dense conjugate of aBgTx and appear as a black labeling at the crests of the postsynaptic folds. (E) Diagram of ion channel domains at level of the electron microscope. The AChRs at the crests between the folds are shown in green and the NaV1 channels in the depths of the folds in red. Reproduced from Slater CR (2008). Reliability of neuromuscular transmission and how it is maintained. Handb Clin Neurol 91: 27-101, with permission.

of ACh called active zones (AZs), where vesicles fuse and ACh is released by exocytosis. The postsynaptic membrane is distinguished by the presence of infoldings, which lie opposite the AZs. The AChR and NaV1.4 proteins occupy separate postsynaptic domains. The AChRs are concentrated at the crest of the folds, near the nerve, while the NaV1.4 channels are concentrated in the depths of the folds. Neuromuscular transmission involves the release of ACh under the action of voltage-gated calcium channels triggered by the incoming nerve impulse causing 20 to 200 of vesicles to release their contents. The high density of AChRs in the postsynaptic membrane of the muscle fiber where ligand-gated ion channels open briefly when bound by ACh, causes an influx of positive ions or endplate current (EPC) and a transient local

depolarization of the muscle membrane also known as the endplate potential (EPP). EPPs that summate rising to a peak of 25–44 mV form the resting potential of 75 mV in about 1 ms in turn leads to the opening of voltage-gated Nav1.4 channels, initiating the muscle action potential. These events are terminated by the action of acetylcholinesterase (AChE) associated with the synaptic basal lamina which splits ACh into acetate and choline, a function that is prevented by (ChE) inhibitors or anti-ChE medications to treat MG. An essential feature of the safety factor of NMJ transmission thus lies in the fact that the summed effect of released ACh into the synaptic cleft, which normally depolarizes the muscle fiber is more than is required to reach the threshold to evoke an AP despite intense physical activity when there

338

D.S. YOUNGER

is a temporary depletion of vesicles available for rapid release, resulting in the reduction in the amount of ACh released by individual nerve impulses (Wood and Slater, 2001). Much has also been learned about the role of thymusand bone-marrow-derived B-cells in the pathogenesis of MG since the early reports of altered cellular immunity in human MG (Richman et al., 1976) and EAMG (Lennon et al., 1976). ACh receptor antibodies are a product of plasma cells of the lymphocyte lineage (Storb, 1993). AChR antibody production starts with activation of T-cells in a trimolecular complex (TMC) composed of major histocompatibility class II molecules (MHC II), native AChR, and antigen-presenting cells (APCs) (Theofilopoulos et al., 1993). The initial events in the pathogenesis of myasthenia, i.e., a loss of selftolerance are not yet well understood, but several factors probably contribute to it, including autoreactive T-cells sensitive to native AChR antigens in thymus and blood that are usually quiescent and not deleted or anergized.

Clinical overview Autoimmune acquired MG is recognized by a pattern of weakness that has the features of fluctuation and variability over the course of a day or over months or years, leading to perceptible exacerbations and remissions. The distribution of weakness is characteristic, affecting ocular, facial, oropharyngeal, and limb muscles. Ocular muscles are usually involved in MG and are uniquely suited for direct observation of weakness. Asymmetrical weakness of several muscles in both eyes is typical, and the pattern of weakness resulting in fluctuating asymmetric ptosis and oculomotor function that spares the pupils does not usually fit lesions affecting one or more individual nerves or an intramedullary lesions. When ptosis is mild, frontalis muscle contraction may be a clue that eyelid elevation is abnormal. Several simple maneuvers accentuate the apparent facial and oropharyngeal muscle

involvement: prolonged upgaze accentuates ptosis, passively lifting the opposite lid results in the “curtainsign” (Fig. 14.9), vertical excursion of the eyes back to primary position leads to a quick “lid twitch,” attempting to smile results in a characteristic “sneer” (Fig. 14.10), and there is difficulty making the high-pitched “eeee” sound. Other features include excessive throat clearing, coughing, choking on mouth secretions and nasal regurgitation while eating. Any limb muscle can be weak, but neck flexors are generally worse than extensors, and every affected muscle may show characteristic “give-away” weakness. The diagnosis is confirmed by unequivocal and reproducible improvement after intravenous administration of edrophonium chloride, a rapidly acting anticholinesterase drug. Formal diagnosis is bolstered by finding a decremental response to repetitive nerve stimulation (RNS) or single-fiber electromyography (SFEMG); and in particular, the identification of binding, blocking, or modulating AChR antibodies in the serum (Lindstrom et al., 1976b) with postulated actions that include accelerated degradation, endocytosis, and crosslinking of receptors (Drachman et al., 1978), functional blockade (Drachman et al., 1982) and complement-mediated lysis of end-plates by the membrane attack complex leading to flattening and simplification of postsynaptic junctional folds (Engel and Arahata, 1987).

Nosology and classification It has been said that efforts to define a disease are attempts to understand the concept of the disorder. In no other motor diseases has this been more evident than in the NMJ disorders that span the simple spectrum of presynaptic, synaptic, and postsynaptic localization, reflecting the continued reappraisal of the AChR and its diverse genetic etiopathogenic factors. Congenital myasthenic syndromes (CMS) (Engel et al., 1997), Lambert-Eaton myasthenic syndrome (LEMS)

Fig. 14.9. Curtain sign. Passive elevation of each eyelid in turn unmasks or exaggerates ptosis in the contralateral lid. Reproduced from Sanders DB & Massey JM (2008). Clinical features of myasthenia gravis. Handb Clin Neurol 91: 229-252, with permission.

ON THE PATH TO EVIDENCE-BASED THERAPY IN NEUROMUSCULAR DISORDERS

Fig. 14.10. Sneer sign. During attempted smile, there is contraction of the median portion of the upper lip and horizontal contraction of the corners of the mouth without the natural upward curling. There is noticeable ptosis with elevation of the facial forehead muscles and upward movements of the eyebrows. Reproduced from Sanders DB & Massey JM (2008). Clinical features of myasthenia gravis. Handb Clin Neurol 91: 229-252, with permission.

(Lennon, 1997), thymomatous (Lovelace and Younger, 1997) and seronegative MG (Sanders et al., 1997), and the natural history of MG have posed challenges to nosology and classification. For example, although LEMS is just as presynaptic as many CMS (Engel, 2018), it is instead immunologically-mediated and distinguished by circulating P/Q-type calcium channelbinding antibodies and occasionally striational antibodies (StrAbs) with or without evidence of cancer. Coexistence of MG and LEMS has not convincingly been documented by electrophysiological criteria, but there is a one-way serological overlap between the two disorders in that about 13% of LEMS cases have AChR binding or StrAbs (Lennon, 1997). The exclusively presynaptic lesion found micro-electrophysiologically in biopsied nerve muscle preparations of several LEMS patients who were seropositive for AChR autoantibodies (both AChR-binding and AChR-modulating (Howard et al., 1987; Sano et al., 1992) implies that these antibodies are probably nonpathogenic, and are presumably directed against cytoplasmic, intramembranous or fetal-type muscle epitopes. The clinical features of seropositive and seronegative MG are similar, although the latter are more likely to have purely ocular myasthenia and milder disease. An autoimmune process underlies seronegative cases as shown by improvement after immunotherapy and thymectomy, and abnormal NMJ transmission that transfers to animals by injection of immunoglobulin (Ig) from seronegative patients, with direct blocking effects when applied in vitro to nervemuscle preparations (Yamamoto et al., 1991). Two antibodies can be found in seronegative cases. One is musclespecific receptor tyrosine kinase (MuSK) identified in up

339

to one-half of seronegative cases, with little or no overlap among those with antibodies to the ACh and MuSK receptors (Hoch et al., 2001; Liyanage et al., 2002) and a mechanism not yet well understood; however, advanced serological assay show anti-MuSK antibody binding to clusters of AChR on cell surfaces with activation of complement (Leite et al., 2008). A second antibody to low-density lipoprotein receptor-related protein 4 (LRP4) and agrin have been described (Yu et al., 2021). Agrin activates LRP4 to interact with MuSK, which, is critical for proper expression of AChR, allowing normal neuromuscular transmission. The usefulness of these antibodies in the diagnosis of MG in patients without AChR or MuSK antibodies, in so-called doubleseronegative MG, is still under investigation, yet only a small percentage of such patients have LRP4 and/or agrin antibodies (Rivner et al., 2020). The classification and nosology have been problematic in MG. While one could potentially apply the term myasthenia to all postsynaptic forms (when certain), including the clearly postsynaptic congenital forms (Engel, 2018) and reserve myasthenic for presynaptic or synaptic space disorders, over time the term myasthenia has been reserved instead for acquired immunologically-mediated postsynaptic disorders, whether generalized or ocular, adult, juvenile or neonatal and transient. Historically, there have been several proposed classifications; however, few institutions endorsed the same ones. Such early attempts emphasized duration of symptoms because it was believed that the disorder might be progressive (Osserman, 1971), transitioning from ocular to generalized disease and culminating in crisis. However, available natural history retrospective studies show a different pattern and outcome of MG. A half-century ago, patients had a 50:50 chance of surviving a myasthenic crisis (Rowland et al., 1956), which typically occurs in the first 2 years of illness yet is not a sign of disease progression. Successful treatment of MG has little impact on the crisis, which occurs in about 15% of cases at a frequency that is unchanged in several decades (Cohen and Younger, 1981) with a mortality that now comes close to nil with the modern intensive care unit (ICU) (Thomas et al., 1997).

Natural history studies Oosterhuis (1989) conducted an long-term follow-up study in 73 MG patients in Amsterdam between 1926 and 1965, treated mainly with cholinesterase inhibitors alone demonstrating the evolution of their clinical state. The maximum severity of the disease occurred during the first 7 years after onset (in 87%). Eighteen (29%) patients died, of whom eight had a thymoma. Spontaneous improvement or remission occurred at any time during

340

D.S. YOUNGER

the follow-up. At the end of the study (in 1985) 16 (22%) patients were in a complete clinical remission, 13 (18%) had improved considerably (3 with prednisone), 12 (16%) had improved moderately, 12 (16%) remained unchanged and two had deteriorated. Associated autoimmune diseases were diagnosed in 25% (n ¼ 58). The largest natural history study of 1976 MG patients treated between 1940 and 2000 in Baltimore, Maryland (first 500 cases), and the remainder in Brooklyn, New York was conducted by Grob et al. (2008) that included 246 with ocular MG and 1730 patients with generalized MG. The study found that in >90% of cases the distribution, severity, and course of the disease were in fact determined within the first 1 or 2 years, with subsequent improvement afterward. The initial and almost uniform occurrence of ptosis or diplopia, and its improvement after anti-ChE medication enabled early detection of whom 17% had symptoms ultimately restricted to the eyes for at least 2 years, and a 90% likelihood not to become more generalized. In about 40% of cases of generalized MG, whose symptoms became severe, up to one-half suffered transient respiratory involvement, also during the first 2 years. However, the natural course of MG was general improvement overall in 70% including remission in 13%, with 20% of patients remaining unchanged, and a stunning 4% of the patients who survived the first 2 years worse than the time of maximum weakness; with fatalities in the remaining 5% to 9%.

Clinical status and outcome measures The first Task Force of the Medical Scientific Advisory Board (MSAB) of the Myasthenia Gravis Foundation of America (MGFA) (Jaretzki et al., 2000) ostensibly to develop standardized classification and outcome measures pertaining to thymectomy trials in MG. However, it became quickly apparent that their efforts would apply to all therapeutic trials. Thus, their scope was expanded to the need for universally accepted clinical definition, classifications, grading systems, and methods of analysis guiding the performance of RCTs (Beggs and Kunkel, 1990; Schulz, 1997), and future guidelines to achieve the highest standards of evidence-based patient care (Miller et al., 1999). These recommendations for clinical research standards included the MGFA Clinical Classification (a tool that identifies subgroups of MG patients who share distinct clinical features), the Quantitative MG Scale (QMG-a quantitative score of disease severity), and the MGFA Post-Intervention Status (a system to classify clinical status after therapy). During the evaluation of therapeutic options, the Task Force recommended that assessment of the QoL be performed. The second task force of the MSAB of the

MGFA (Benatar et al., 2012) proposed the use of a quantitative measure, such as the MG-Composite score, weighted for clinical significance and incorporates patient reported outcomes. These and other primary and secondary endpoint measures listed below have been incorporated into prospective RCTs of therapy for MG.

MGFA CLINICAL CLASSIFICATION In developing the MGFA Clinical Classification, the Task Force recognized the importance of identifying subgroups of patients with MG sharing distinct clinical features or severity of disease that may indicate different prognoses or responses to therapy. It was not intended to be used as an outcome measure. Moreover, it deferred quantitative assessment of muscle weakness and response to therapy to the more precise quantitative MG score (QMG). Without necessarily replacing other classifications in use, the Task Force recommended that the most severely affected muscles be employed to define the patient’s Class and that the “maximum severity” designation be used to identify the most severe pretreatment clinical classification status. The “maximum severity” designation could be made historically and employed as a point of reference. It can be simply summarized as follows: Class I (ocular); Class II, “mild” (generalized) weakness affecting limb (IIa) and oropharyngeal muscles (IIb); Class III “moderate” generalized affecting limb (IIIa) and oropharyngeal or respiratory muscles (IIIb); Class IV “severe” affecting limb (IVa) and oropharyngeal or respiratory muscles (IVb); and Class V “crisis” defined by intubation). Ascertaining respiratory (muscle) involvement in MG relies upon pulmonary function test (PFT) measurements, especially vital capacity (VC) measured in liters, which is largely dependent upon inspiratory muscle function, measured as maximal inspiratory pressure (PImax) in cm of H2O, but also expiratory muscle pressure, reflected in the measured PEmax that is important in generating an effective cough to clear airway secretions and prevent aspiration (Younger et al., 1984).

QUANTITATIVE MG SCALE The QMG (Table 14.7) expanded and modified earlier scales that consisted of 8 items, each graded 0–3 with the score of 3 being the most severe. It was expanded to 13 items Barohn and coworkers (Barohn et al., 1998) replaced 3 subjective items in facial muscles, chewing and swallowing rendering it more objective, enabling it to reach the 95% confidence level in interrater reliability as a primary outcome measure, such that QMG scores did not differ by more than 2.63 units, translating to a required sample size of 17 patients in each treatment group allowing the detection of a significant

ON THE PATH TO EVIDENCE-BASED THERAPY IN NEUROMUSCULAR DISORDERS

341

Table 14.7 Quantitative MG scale.

Test items weakness(score)

None (0)

Mild (1)

1. Double vision on lateral gaze right 61 11–60 or left (circle one), seconds 2. Ptosis (upward gaze), seconds 61 11–60 3. Facial Muscles Normal lid Complete, weak, closure some resistance 4. Swallowing 4 oz./120 mL water Normal Minimal coughing or throat clearing 5. Speech following counting aloud None Dysarthria from 1 to 50 at #50 at #30–49 6. Right arm outstretched (90o sitting), seconds 240 90–239 7. Left arm outstretched (90o sitting), seconds 240 90–239 8. Vital Capacity (% predicted) mouthpiece or facemask 80% 65%–79% (circle one; best of 3) 9. Right hand grip: (best of 2) Male 45 15–44 (kgw) female 30 10–29 10. Left hand grip: (best of 2) Male 35 15–34 (kgw) female 25 10–24 11. Head, lifted (45o supine), seconds 120 30–119 12. Right leg outstretched 100 31–99 (45o supine), seconds 31–99 13. Left leg outstretched (45o supine), 100 seconds

Moderate (2)

Severe (3)

1–10

Spontaneous

1–10 Complete, without resistance

Spontaneous Incomplete

Severe coughing/ choking or nasal regurgitation Dysarthria at #10–29

Cannot swallow (test not attempted) Dysarthria at #9

10–89

0–9

10–89

0–9

50%–64%

C in subacute-onset, painless and progressive visual loss affecting McFarland et al. (2007) MT-ND6 and m.3460G>A both eyes. “LHON plus” refers to rare manifestations such as in MT-ND1 account for dystonia and cardiac pre-excitation >90% of cases) nDNA (OPA1 mutations, ad) Dominant Optic Atrophy. 20% patients develop other neurological Amati-Bonneau et al. (2005), Amati-Bonneau et al. features including PEO, myopathy, neuropathy, mild spasticity (2008), and Yu-Wai-Man et al. (2010) and Parkinsonism nDNA (TYMP mutations, ar) Mitochondrial Neuro-Gastro-Intestinal Encephalomyopathy. Severe Hirano et al. (1994), Nishino et al. (1999), and gut dysmotility, malnutrition, PEO and demyelinating Halter et al. (2015) neuropathy. Leukodystrophic changes (clinically asymptomatic) on MR imaging mtDNA (MT-ATP6 Neurogenic weakness, Ataxia and Retinitis Pigmentosa. Other Holt et al. (1990), Thorburn and Rahman (1993), variants, e.g., m.8993T>C, features including seizures and cognitive impairment Ng et al. (2019b), and Stendel et al. (2020) m.8993T>G and m.9176T>C) nDNA (POLG1 mutations, ad Encompasses two overlapping syndromic descriptions: SANDO Van Goethem et al. (2003), Luoma et al. (2004), and ar) (sensory ataxia, neuropathy, dysarthria and ophthalmoparesis) Cohen et al. (1993), and Hikmat et al. (2020) and MEMSA (myoclonic epilepsy, myopathy and sensory ataxia). Other features include PEO, cerebellar ataxia, Parkinsonism and premature ovarian failure mtDNA (single, large-scale PEO, myopathy and mild bulbar dysfunction Holt et al. (1988), Grady et al. (2014), and Mancuso mtDNA deletion, sporadic) et al. (2015) nDNA (genes involved in maintenance of mtDNA and associated with mtDNA depletion and multiple deletions) TWNK (ad) (previously known Late-onset PEO and mild myopathy. Autosomal recessive disease is Fratter et al. (2010) and Nikali et al. (2005) as PEO1) associated with infantile-onset spinocerebellar ataxia (IOSCA) RRM2B (ad) Autosomal dominant disease is characterized by late-onset PEO and Pitceathly et al. (2012), Bourdon et al. (2007), and mild myopathy. Autosomal recessive disease is associated with Lim et al. (1993) severe SNHL, myopathy, PEO, renal disease and gut dysmotility TK2 (ar) The severity of myopathy is determined by the disease onset. PEO, Saada et al. (2001), Garone et al. (2018), and facial weakness, respiratory muscle weakness and dysphagia Domínguez-González et al. (2019) SLC25A4 (ad) (previously Indolent PEO and myopathy Kaukonen et al. (2000) known as ANT1) SPG7 (ar) PEO, spastic paraparesis, cerebellar ataxia, and myopathy Pfeffer et al. (2014) and Hewamadduma et al. (2018)

Adapted from a previous publication Gorman GS, Chinnery PF, DiMauro S, et al. (2016). Mitochondrial diseases. Nat Rev Dis Primers 2: 16080. AD, autosomal dominant; AGK, Acylglycerol Kinase gene; ANT1, Adenine Nucleotide Translocator 1; AR, autosomal recessive; BCS1L, BCS1 Homolog, Ubiquinol-Cytochrome C Reductase Complex Chaperone gene; CSF, cerebrospinal fluid; FSGS, focal segmental glomerulosclerosis; MR, magnetic resonance; MT-ATP6, mitochondrially encoded ATP Synthase Membrane Subunit 6 gene; mtDNA, mitochondrial DNA; MT-ND1, mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 1 gene; MT-ND4, mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 4 gene; MT-ND6, mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 6 gene MT-TK, mitochondrially encoded tRNA lysine; MT-TL1, mitochondrially encoded tRNA leucine 1 gene; nDNA, nuclear DNA; OPA1, optic atrophy 1 gene; PEO, progressive external ophthalmoplegia; POLG1, Mitochondrial DNA Polymerase Gamma 1 gene; RRM2B, Ribonucleotide Reductase Regulatory TP53 Inducible Subunit M2B gene; SERAC1, Serine Active Site Containing 1 gene; SLC25A4, Solute Carrier Family 25 member 4 gene; SNHL, sensorineural hearing loss; SPG7, Spastic Paraplegia 7 gene; TAZ, Taffazin gene; TK2, Thymidine Kinase 2 gene; TWNK, Twinkle mtDNA Helicase gene; TYMP, Thymidine Phosphorylase gene.

566

Y.S. NG AND R. MCFARLAND

A key initial step in the investigation of a patient with suspected mitochondrial disease is the meticulous clinical assessment and review of family history. Further ancillary investigations such as laboratory tests and imaging studies are often necessary to help determine the overall phenotype and whether this is a recognizable syndrome, a nonspecific multisystem picture or a solitary organ involvement such as optic neuropathy. This chapter highlights the neurological features frequently encountered in the setting of both pediatric and adult neurology clinical practice and to provide an overview of mitochondrial genetics.

OVERVIEW OF MITOCHONDRIAL GENETICS Mitochondria are double membrane organelles present in all nucleated cells in the body. They are dynamic organelles undergoing fission and fusion depending on the metabolic state of the cell (Chapman et al., 2020). Mitochondria have multiple functions within cells and are involved in tricarboxylic acid (TCA) cycle, fatty acid oxidation, oxidative phosphorylation, urea cycle, gluconeogenesis, and ketogenesis. They also play an important role in several other important cellular processes including calcium homeostasis, amino acid and lipid metabolism, synthesis of iron–sulfur cluster, (nonshivering) thermogenesis, biosynthesis of hemes, and apoptosis (Ng et al., 2021c). Oxidative phosphorylation (OXPHOS) occurs in the inner mitochondrial membrane, where five multisubunit enzymatic complexes directly involved in OXPHOS are colocated, three of which pump protons into the intermembrane space (complexes I, III, and IV). This generates an electrochemical gradient across the near impermeable inner mitochondrial membrane, which then dissipates through complex V, also known as adenosine triphosphate (ATP) synthetase, to generate ATP through the phosphorylation of adenosine diphosphate (ADP). The complexes involved in OXPHOS include subunits encoded by both mitochondrial and nuclear genomes highlighting the complex genetics of mitochondria. Mitochondria are unique from the genetic perspective as they contain the extrachromosomal deoxyribonucleic acid (DNA) referred as mitochondrial DNA (mtDNA). MtDNA is a small (16.6 kb) circular double-stranded DNA molecule that contains 37 genes encoding 13 protein subunits of OXPHOS, 22 transfer ribonucleic acids (tRNAs), and two ribosomal ribonucleic acid (rRNAs) (Anderson et al., 1981). The 13 protein subunits consist of 7 subunits of complex I, 1 subunit of complex III, 3 subunits of complex IV, and 2 subunits of complex V. The rRNA molecules enable the intramitochondrial synthesis of these subunits. The remaining mitochondrial

proteins, including all the other subunits of OXPHOS, are encoded by the nuclear DNA and transported into mitochondria after synthesis in the cytosol. Mitochondrial DNA genetics is different from that of nuclear DNA in two main respects: it is maternally inherited and present in multicopy. Consequently, mtDNA diseases are only passed down the female line and, given the potential for thousands of copies of mtDNA within a single cell, it is possible to have both mutated and wild type molecules coexisting in the same cell—a situation known as heteroplasmy that is frequently associated with pathological variants of mtDNA. Heteroplasmy is often quoted as a percentage that describes the ratio of mutated to wild type copies of mtDNA present in a specific cell or tissue. Alternatively, all copies of mtDNA may be identical for a pathological variant or, more commonly, wild type mtDNA; in this circumstance, the term homoplasmy is used. Most heteroplasmic mtDNA defects are functionally recessive and thus high levels of mutated mtDNA are required before incurring a biochemical deficiency and the subsequent manifestation of disease. This disease threshold is known to be dependent on the specific mtDNA variant and tissue type, but other considerations include nuclear and environmental factors as well as mtDNA copy number (Grady et al., 2018; Bernardino Gomes et al., 2021). From a clinical perspective, it is important to recognize there is often a difference between the level of heteroplasmy observed in different tissues, with lower levels in mitotic tissues such as blood and much higher levels in postmitotic tissues, for example, muscle (Grady et al., 2018; Ng et al., 2018). This phenomenon has consequences for the choice of tissue for investigating mitochondrial disease, timing in relation to age and interpretation of the result—some clinically relevant pathogenic mtDNA variants, such as m.3243A>G, can be “lost” (undetectably low heteroplasmy) from the blood by early adult life (Grady et al., 2018). In being transmitted through the maternal germline mtDNA experiences significant fluctuations in copy number that create a “genetic bottleneck” (Stewart and Chinnery, 2015). In the presence of maternal heteroplasmy, this bottleneck can result in offspring with sometimes quite markedly divergent mtDNA heteroplasmy from that seen in their mother or siblings. This is important when considering reproductive options for women with pathogenic mtDNA variants. All female carriers of pathogenic mtDNA are at risk of transmitting the mtDNAvariant and thus mtDNA disease to their offspring. The risk very much depends on the mtDNA variant. For example, the majority of single, large-scale mtDNA deletions are sporadic so mothers with one affected child may well have other clinically unaffected children (Chinnery et al., 2004). For patients with the m.3243A>G variant, and many other pathogenic variants, virtually all mothers

MITOCHONDRIAL ENCEPHALOMYOPATHY transmit the pathogenic variant to their offspring, at varying levels of heteroplasmy (Pickett et al., 2019). For mothers with homoplasmic mtDNA defects, all offspring will inherit the pathogenic variant. There is no risk of paternal transmission of pathogenic mitochondrial DNA variants to their offspring (Taylor et al., 2003). In view of the multiple functions of mitochondria, the diseases that are traditionally termed mitochondrial are those in which defects of OXPHOS are the primary abnormality. There are approximately 1100 nuclear mitochondrial proteins, including the assembly factors of OXPHOS subunits, proteins involving in mitochondrial dynamics, and those required for the many other functions that mitochondria undertake (Rath et al., 2021). Nuclear mitochondrial proteins are also directly, and indirectly, involved in the replication and maintenance of mtDNA, and thus nuclear mitochondrial defects can lead to decrease in the mtDNA copy number (depletion) or multiple rearrangements (multiple deletions) of mtDNA (El-Hattab et al., 2017b). Clinically, defects of mtDNA and nuclear DNA can look very similar, but the consequences for disease progression, genetics counseling and reproductive options are different and so establishing a genetic diagnosis is crucial (Ng et al., 2021a).

NEUROLOGICAL MANIFESTATIONS OF MITOCHONDRIAL DISEASE Hypotonia and neurodevelopment Abnormal tone including hypotonia is a prominent and early feature of childhood-onset mitochondrial disease. Although typically evident during the first year of life, hypotonia may go undetected until the infant is examined by an experienced healthcare professional. In mitochondrial disease, weakness commonly accompanies hypotonia and together they contribute to a delay in the acquisition of developmental motor milestones. As with many other chronic pediatric neurological conditions, the hypotonia of mitochondrial disease is a consequence of CNS dysfunction. Moreover, in infants with the subacute necrotizing disorder, Leigh syndrome (LS) (Leigh, 1951), they may be initially floppy due to reduced muscle tone, but later develop superimposed spasticity and dystonia due to subsequent brainstem and basal ganglia involvement resulting in axial (spinal) hypotonia and limb hypertonia. Given the importance (and abundance) of mitochondria in both skeletal muscle and peripheral nerve tissue, peripheral hypotonia may be a contributory factor in other cases of early-onset mitochondrial disease, with only rare instances if ever, of “pure mitochondrial myopathy.” Children with central hypotonia can be distinguished by the delay in meeting their motor milestones that is

567

most often a consequence of accompanying muscle weakness. Developmental delay, and the loss of developmental milestones (developmental regression), may not be confined to impaired motor abilities, although these are often the most clinically apparent in prelingual infant. Cognitive, language and social domains of neurodevelopment may all be affected to varying degrees in childhoodonset mitochondrial diseases. Severe early-onset disorders associated with epileptic encephalopathy, such as occurs in arginyl (R) aminoacyl transfer RNA (tRNA) synthetase 2 (RARS2) gene-related mitochondrial disease (see below), allow little scope for neurodevelopment. More commonly, for example, in the case of many patients with deficiency of surfeit locus protein 1 (SURF1) (Tiranti et al., 1998; Zhu et al., 1998), development during the first year of life is considered within normal limits, however it is the loss of previously acquired (motor) skills during the second year of life, in the context of a child who is also failing to thrive, that prompts clinical assessment and investigation (Wedatilake et al., 2013). In patients with LS, the loss of neurodevelopmental skills can occur abruptly, sometimes in conjunction with signs of concomitant infection or metabolic crisis, as in those with Leigh syndrome. These skills may be regained, at least to some extent, as affected children enter a period of neurodevelopmental stagnation. Recurrent subacute episodes of neurodevelopmental regression, with or without recovery, often punctuate the natural history and overall disease burden accumulates with duration of illness.

Mitochondrial epilepsy of childhood Although epilepsy is considered a common complication of mitochondrial disease in both children and adults (Rahman, 2012; Lim and Thomas, 2020), there are few comprehensive epidemiological studies confirming the veracity and extent of this assumption (Whittaker et al., 2015). Nevertheless, epilepsy is recognized as a major, and in some cases, a syndrome defining, clinical feature of several forms of childhood-onset mitochondrial disease including Alpers–Huttenlocher syndrome, Leigh syndrome, Mitochondrial Encephalomyopathy Lactic Acidosis and Stroke-like episodes (MELAS), and pyruvate dehydrogenase complex (PDHc) deficiency (PDCD). The genetic etiology of mitochondrial epilepsy is diverse, with causative pathogenic variants in mitochondrial and nuclear genomes. These genetic disorders impair mtDNA maintenance (e.g., Polymerase gamma 1 gene [POLG1] (Hikmat et al., 2020), Ribonucleotide Reductase Regulatory TP53 Inducible Subunit M2B gene [RRM2B] (Bourdon et al., 2007), Mitochondrial Succinyl-CoA Ligase Subunit Beta gene [SUCLA2] (Elpeleg et al., 2005)), and mtDNA translation

568

Y.S. NG AND R. MCFARLAND

(e.g., genes encoding mitochondrial aminoacyl tRNA synthetases, such as Arginyl [RARS2], Phenylalanine [FARS2] and Isoleucine [IARS2]) (Diodato et al., 2014). They also cause structural abnormalities of mitochondrial respiratory chain complexes (e.g., variants in the Mitochondrially Encoded ATP Synthase Membrane Subunit 6 gene [MTATP6] (Ng et al., 2019b; Licchetta et al., 2021), Succinate Dehydrogenase Complex Flavoprotein Subunit A gene [SDHA] (Bourgeron et al., 1995), NADH:Ubiquinone Oxidoreductase Core Subunit V1 gene [NDUFV1] (Schuelke et al., 1999), NADH dehydrogenase iron–sulfur protein 4 gene [NDUFS4] (Budde et al., 2000)), adversely impact mitochondrial respiratory chain complex assembly (e.g., Transmembrane Protein 70 gene [TMEM70]—complex V (Honzik et al., 2012), BCS1 Homolog, UbiquinolCytochrome C Reductase Complex Chaperone gene [BCS1L]—complex III (Hikmat et al., 2021) and NADH: Ubiquinone Oxidoreductase Complex Assembly Factor 2 gene [NDUFAF2]—complex I (Nouws et al., 2012)), decrease biosynthesis of coenzyme Q10 (e.g., decaprenyl diphosphate synthase subunit 2 gene [PDSS2], Coenzyme Q8A gene [CoQ8A]) (Salviati et al., 1993; Sondheimer et al., 2017) and severely limit solute transport (e.g., Solute carrier family 25 member 22 gene [SLC25A22]). While impairment of these diverse processes could lead to a final common result of limited mitochondrial ATP supply, that does not adequately explain the diversity of seizure semiology in mitochondrial disease, where generalized tonic–clonic, focal, myoclonic, tonic, infantile spasms, atypical absence, and focal dyscognitive seizures are observed. Nonconvulsive status epilepticus and convulsive status epilepticus (focal and generalized) are also recognized complications of various forms of mitochondrial disease (see below). It is likely that multiple pathological mechanisms lead to such wide-ranging epileptic semiology and disruption of intracellular calcium storage and flux, impaired vesicular transport and release of neurotransmitters, vulnerability of specific inhibitory interneurons to mitochondrial dysfunction (Smith et al., 2022) and reactive oxygen species (ROS) damage to ion channels in the neuronal membrane (Zsurka and Kunz, 2015). Infantile spasms (IS) are an age-dependent response of the immature brain to a variety of pathological insults and thus a form of epileptic seizure rarely seen outside the first year of life (Wilmshurst et al., 2017). IS are often associated with an electroencephalographic (EEG) appearance of hypsarrhythmia, when the electro-clinical syndrome is known as West Syndrome. It is a particularly common problem in young children with a PDHc deficiency, a multiple enzyme platform located on the inner mitochondrial membrane that is key to carbohydrate metabolism, converting pyruvate to acetyl CoA

(and carbon dioxide), which then enters the TCA cycle. Other features associated with PDHc deficiency include an elevated lactate:pyruvate ratio (>20) in blood and evidence of corpus callosal dysgenesis on cranial magnetic resonance imaging (MRI) (Barnerias et al., 2010; Patel et al., 2012). However, IS is not limited to PDHc deficiency and can be observed in a range of childhood onset mitochondrial diseases such as those caused by mitochondrial aminoacyl tRNA synthetase deficiencies (e.g., Arginyl—RARS2 (Ngoh et al., 2016), Asparaginyl—NARS2 (Mizuguchi et al., 2017), and Phenylalanine—FARS2 (Almalki et al., 2014; Raviglione et al., 2016)), pathogenic variants in mtDNA (e.g., m.13513G>A (Monlleo-Neila et al., 2013)). While conventional treatment for IS would involve a combination of steroid hormone and Vigabatrin, many children with PDCD respond well to a ketogenic diet (KD) (Sofou et al., 2017) and the recent identification of decanoic acid (a principal component of the KD) as an effective anticonvulsant, holds the promise of a more tolerable form of treatment (Wright et al., 2020).

Stroke-like episodes A mitochondrial stroke-like episode is defined as a subacute, evolving brain syndrome driven by seizure activity in genetically confirmed mitochondrial disease (Ng et al., 2019a). Stroke-like episodes are commonly characterized by severe headache, nausea and vomiting, encephalopathy, unilateral visual disturbances, focalonset seizures, and/or neuropsychiatric symptoms prior to the development of any focal neurological deficits such as visual field defect (hemianopia and cortical blindness), aphasia and apraxia. Nonconvulsive seizures arising from temporal and parietal lobe lesions can manifest with new-onset psychosis, aggression and confusion (Kaufman et al., 2010). Refractory, focal-onset status epilepticus is almost invariably evident in patients with POLG-related mitochondrial disease whereas it is present in less than a third of patients with mtDNArelated MELAS syndrome (Li et al., 2021; Ng et al., 2021b). It is rare that stroke-like episode manifests with an acute-onset profound hemiparesis (Ng et al., 2019a). Although stroke-like episodes were originally described in individuals G variant in the transfer RNA, mitochondria, leucine 1 (MT-TL1) gene accounting for up to two-thirds of the cases (Ng et al., 2021b); pathogenic variants in other mtDNA genes (Ng et al., 2018; Seed et al., 2022) and in the nuclear genes such as POLG and

MITOCHONDRIAL ENCEPHALOMYOPATHY

569

Fig. 23.1. Examples of neuroimaging findings in mitochondrial diseases. (A) Evolution of stroke-like lesions in a young woman with the m.3243A>G-related MELAS (Mitochondrial Encephalomyopathy, Lactic Acidosis and Stroke-like episodes) syndrome. Magnetic resonance fluid-attenuated inversion recovery (FLAIR) sequence images showed extensive signal abnormalities involving right occipital, temporal, parietal (not shown) lobes crossing vascular territories and bilateral insular cortices on admission (A(i–ii)); these changes were associated with restricted diffusion and mixed apparent diffusion coefficient (ADC) map. The signal changes became more confluent with new thalamic lesion a week later (A(iii–iv)). The signal abnormalities resolved in most areas, corresponding to the clinical improvement, with evidence of atrophy in the temporal-occipital region 5 weeks after the first scan (A(v–vi)). (B) Marked cerebral atrophy following recurrent stroke-like episodes in a man with MELAS syndrome in his 50s. (C) Marked cerebellar atrophy in a young adult patient with m.8993T>G-related Leigh syndrome. (D) Symmetrical leukodystrophic changes in a case of MNGIE (Mitochondrial Neuro-Gastro-Intestinal Encephalomyopathy). (E) Neuroimaging changes of a 4-year-old child with surfeit locus protein 1 (SURF1)-related Leigh syndrome. FLAIR signal abnormalities identified in (E(i)) medulla and cerebellar deep nuclei, (E(ii)) pons, (E(iii)) midbrain, (E(iv)) thalami, (E(v)) globus pallidus and putamen, (E(vi)) caudate nuclei. Figures adapted from a previous publication Lim AZ, Ng YS, Blain A, et al. (2022). Natural History of Leigh Syndrome: A Study of Disease Burden and Progression. Ann Neurol 91: 117-130.

CoQ8A (also known as the aarF-domain-containing kinase 3 gene, [ADCK3]) can cause stroke-like episodes (Hikmat et al., 2016; Ng et al., 2019a; Hikmat et al., 2020). Stroke-like lesions do not typically occur in specific vascular territories, but involve cerebral cortex and juxtacortical white matter on computed tomography (CT) and MRI (Fig. 23.1). Stroke-like lesions show restricted diffusion with variable apparent diffusion coefficient (ADC) map changes (Ng et al., 2021b), depending on the time interval between the onset of symptoms and scan (Tzoulis and Bindoff, 2009; Kim et al., 2011). The evolution of stroke-like lesions on serial MRI scans frequently corresponds to the clinical status, for example, the stroke-like lesions increase in size and appear to be “spreading to the adjacent cortical areas” as the patient’s neurological status deteriorates due to ongoing seizure activities and encephalopathy (Iizuka et al., 2002, 2003; Tzoulis et al., 2010). These signal changes may resolve

completely or partially when the patient shows clinical recovery from the acute episode if appropriate and timely treatment has been instigated. Some signal abnormalities resolve completely, however, most severe lesions develop into cortical laminar necrosis, gliosis and significant brain volume loss (Ng et al., 2021b), potentially leading to progressive physical decline and development of dementia. The pathogenesis and pathophysiology of stroke-like episodes have been debated over the last few decades (Lax et al., 2016a; Ng et al., 2019a; Tetsuka et al., 2021). The “mitochondrial angiopathic theory” as proposed is upon the identification of abnormal accumulation of mitochondria within the smooth muscle layer and endothelium of small arteries and arterioles, which show strong reactivity to succinate dehydrogenase (Ohama et al., 1987; Hasegawa et al., 1991). It has been hypothesized that the abnormal mitochondrial proliferation within the vessel walls results in nitric oxide

570

Y.S. NG AND R. MCFARLAND

deficiency (Koenig et al., 2016; El-Hattab et al., 2017a), low plasma arginine (Koga et al., 2002, 2005) and citrulline (Naini et al., 2005) levels, leading to the loss of cerebrovascular autoregulation and vasodilatation, consequently causing tissue ischemia and development of stroke-like lesions. However, the microangiopathy theory has not been supported by several radiological and neuropathological findings. First, there is no evidence of arterial thrombosis based on both angiographic and perfusion studies; increased perfusion to the acute stroke-like lesions have been reported. Second, there is no evidence of microvascular occlusion on postmortem brain studies (Tzoulis and Bindoff, 2012; Lax et al., 2012b; Tzoulis et al., 2017). Third, neurons and astrocytes in the cerebral cortex have more severe mitochondrial respiratory chain defects than the endothelium and smooth muscle layers of the microvasculature (Ng et al., 2021b). On the other hand, there is compelling evidence to support that stroke-like episodes are predominantly driven by neuronal hyperexcitability. First, focal-onset seizures are common in stroke-like episodes (Hirano et al., 1992; Tzoulis and Bindoff, 2012; Ng et al., 2021b). Second, stroke-like lesions identified on neuroimaging are likely seizure-related changes which often colocalize with focal epileptiform discharges detected on EEGs (Iizuka et al., 2002, 2003; Ng et al., 2021b). Third, postmortem brain studies have revealed selective vulnerabilities of inhibitory interneurons and Purkinje neurons to mitochondrial respiratory chain defects, and these findings provide the basis of neuronal hyperexcitability in these patients manifesting with MELAS and Alpers–Huttenlocher syndromes (Lax et al., 2016b; Hayhurst et al., 2019; Smith et al., 2022).

Myoclonus Myoclonus is an involuntary movement characterized by sudden, brief, jerky, shock-like movements involving the limbs, face, and trunk, without loss of consciousness (Shibasaki and Hallett, 2005). Cortical, generalized/ multifocal myoclonus is most commonly associated with a classic mitochondrial syndrome called myoclonic epilepsy with ragged-red fibers (MERRF) syndrome (Fukuhara et al., 1980), which is most frequently caused by the m.8344A>G variant (Wallace et al., 1988b). In addition to photosensitive cortical myoclonus and generalized epilepsy, MERRF syndrome is associated with other neurological findings including progressive cerebellar ataxia, proximal myopathy, cognitive impairment, sensorineural hearing loss and optic neuropathy (Mancuso et al., 2013; Altmann et al., 2016). Multiple lipomatosis especially in the neck and upper trunk is a characteristic feature of MERRF syndrome, but it is

rarely identified in other forms of mitochondrial disease (Ekbom, 1975; Musumeci et al., 2019). While cortical myoclonus is assumed to be the most prevalent form of myoclonus in mitochondrial disease, there have been limited studies applying electrophysiological techniques to investigate subcortical and spinal myoclonus in these patients. Apart from MERRF syndrome, myoclonus has also been reported in several other mitochondrial diseases such as POLG-related Alpers’ syndrome, Leigh syndrome and MELAS syndrome (Mancuso et al., 2014; Lamperti and Zeviani, 2016; Ghaoui and Sue, 2018). Overall, myoclonus is uncommon in both pediatric and adult mitochondrial diseases, that it only accounts for 4% and 3.6% of Italian pediatric (Ticci et al., 2021) and adult (Montano et al., 2022) cohort studies, respectively.

Movement disorders A broad spectrum of other abnormal motor manifestations may occur in mitochondrial diseases including dystonia, Parkinsonism, and ataxia, with marked heterogeneity, even in patients with the same genetic mutation (Ghaoui and Sue, 2018).

Dystonia Symmetrical bilateral basal ganglia involvement, often in association with symmetric brainstem lesions, is a hallmark of LS, the archetypal childhood onset mitochondrial disorder first described by Denis Leigh (Leigh, 1951). The etiology of LS is diverse, with numerous pathogenic variants in mtDNA, some of which have been more commonly associated with other classic mitochondrial syndromes (McFarland et al., 2007), and in excess of 75 different nuclear genetic variants causing this progressive neurodegenerative condition (Rahman and Thorburn, 1993–2023; Lake et al., 2016). Abnormalities of posture, increased limb tone and spasms are prominent features of LS and these features of dystonia are usually persistent, generalized and accompanied by axial hypotonia. Bulbar involvement with dysphagia and dysphonia is a common clinical consequence in LS and often necessitates insertion of a percutaneous gastrostomy tube and use of electronic communication aids. Dystonia and other movement disorders appear to be a significant contributor to impaired quality of life in children with LS (Ticci et al., 2021; Lim et al., 2022). Management of dystonia with conventional therapies, such as baclofen, trihexyphenidyl, benzodiazepines, levodopa, botulinum toxin, and deep brain stimulation (MartinezRamirez et al., 2015), is variably effective and patients often require multiple therapeutic approaches (Ticci et al., 2021).

MITOCHONDRIAL ENCEPHALOMYOPATHY

Parkinsonism The association of Parkinsonism, chronic progressive external ophthalmoplegia (CPEO) and dominant POLG disease was described in early 2004 (Luoma et al., 2004). These patients developed unilateral signs of Parkinsonism at the onset, their dopamine active transporter (DaT) scans demonstrated reduced dopamine uptake in the basal ganglia, and clinically they showed good response to levodopa treatment. Subsequently, compound heterozygous POLG mutations were also identified in two sisters who developed Parkinsonism in their early 20s and peripheral neuropathy (Davidzon et al., 2006). In a large, UK national cohort study, Parkinsonism accounted for 43% of all extrapyramidal movement disorders in patients with mitochondrial disease, making it the most common extrapyramidal movement disorder identified in this population (Martikainen et al., 2016). Parkinsonism has been reported as the second most common movement disorder in an Italian cohort of adult mitochondrial disease, with the mean age of onset was 62.7 11.2 years (Montano et al., 2022). Importantly, 70% of patients with the mitochondrial Parkinsonism were dopamine responsive. However, the mechanistic link between POLG deficiency and Parkinsonism is not straightforward. In a Norwegian study of both postmortem samples and living patients with POLG mutations (Tzoulis et al., 2013), the features of Parkinsonism were not clinically evident despite marked neuronal loss in the substantia nigra and reduced dopamine uptake detected on the DaT scans. More recently, other nuclear genes involving in mitochondrial DNA maintenance such as the optic atrophy 1 gene [OPA1] (Carelli et al., 2015) and the twinkle mtDNA helicase gene [TWNK] (Percetti et al., 2022) have also been observed in association with Parkinsonism.

Ataxia Ataxia is the most common movement disorder in both pediatric (50%) (Ticci et al., 2021) and adult (60%) (Montano et al., 2022) patients with mitochondrial disease. Ataxia can be a prominent manifestation in certain types of mitochondrial disease such as POLGrelated mitochondrial disease and neurogenic weakness, ataxia and retinitis pigmentosa (NARP) syndrome. However, ataxia more commonly features as part of a multisystem disorder, for example, Kearns-Sayre syndrome, MERRF and MELAS syndromes (Ghaoui and Sue, 2018). According to an Italian retrospective cohort study, ataxia due to cerebellar degeneration was identified in 64% of the patients, followed by spinocerebellar (22%) and sensory ataxia (14%) (Montano et al., 2022). Some patients develop additional movement disorders such as dystonia and Parkinsonism as their disease evolve, in the pediatric and adult cohorts, respectively.

571

Mixed cerebellar and sensory ataxia are frequently observed in patients harboring pathogenic variants in POLG (Rahman and Copeland, 2019) and MT-ATP6 (Ng et al., 2019b). The clinical signs of a cerebellar syndrome in mitochondrial disease are similar in cerebellar ataxia of other etiologies; however, eye signs such as square-wave jerk, nonfatiguable nystagmus, and hypo-/ hyper-metric saccadic movements are not always apparent especially among patients who have extra-ocular muscle weakness due to CPEO. The progressive ataxic symptoms usually lead to greater physical disabilities such as falls and loss of ambulation, dysarthria and dysphagia, than the myopathic symptoms. Brain MRI findings of patients with mitochondrial ataxia often include nonspecific volumetric loss of cerebellum, and symmetrical signal abnormalities in dentate nuclei are occasionally evident in patients with POLG mutations (Tzoulis et al., 2006). Purkinje neuronal loss and cerebellar atrophy due to mitochondrial respiratory chain deficiency has been documented in the neuropathology studies of mitochondrial disease (Lax et al., 2012a). Nerve conduction studies (NCS) usually show changes of axonal neuropathy and/or neuronopathy; and neuropathological studies have revealed striking neuronal cell loss from the dorsal root ganglia and atrophy of posterior columns in POLG cases (Lax et al., 2012c), resembling findings identified in Friedreich ataxia. Mutations in the spastic paraplegia type 7 gene [SPG7], which encodes the mitochondrial metalloprotease paraplegin, were originally reported to cause autosomal recessive (AR) hereditary spastic paraplegia type 7 with abnormal muscle biopsy findings showing ragged red fibers and cytochrome c oxidase (COX) deficiency (Casari et al., 1998). The phenotypic spectrum of SPG7 mutations has been expanded in the subsequent studies to include spastic ataxia (Hewamadduma et al., 2018), CPEO with multiple mtDNA deletions (Pfeffer et al., 2014) and late-onset cerebellar syndrome (Pfeffer et al., 2015). It has been suggested mutations in SPG7 are one of the most common causes of AR cerebellar ataxia, with an estimated prevalence of 0.72 per 100,000 in North East England (Pfeffer et al., 2014). Coenzyme Q10 or ubiquinone is one of the electron transporters in the oxidative phosphorylation system that shuttles electrons between complexes I, II and III. There are at least 12 nuclear-encoded genes required for the synthesis of coenzyme Q10 in mitochondria and mutations in these genes lead to primary coenzyme Q10 deficiency. Cerebellar ataxia is a prominent finding in PDSS2, coenzyme Q6 gene [CoQ6] and CoQ8A gene defects; however, broader phenotypic spectrum of primary coenzyme Q10 deficiency such as stroke-like episodes, seizures, neurodevelopmental delay, neuromuscular weakness, cardiomyopathy, and nephrotic syndrome has been

572

Y.S. NG AND R. MCFARLAND

reported (Salviati et al., 1993; Acosta et al., 2016). The clinical spectrum of biallelic CoQ8A variants has been better delineated by a multicenter, retrospective study showing the age of disease onset was 90% or near the homoplasmic state in cases of Leigh syndrome (Ng et al., 2019b); although m.8993T>G does

574

Y.S. NG AND R. MCFARLAND

appear to have a lower threshold for causing severe disease when compared with other MT-ATP6 variants. Several cohort studies have shown that common neurological features of MT-ATP6 disease include ataxia, peripheral neuropathy, cognitive dysfunction including learning difficulties, and seizures (Ng et al., 2019b; Stendel et al., 2020). Retinitis pigmentosa is a relatively infrequent finding in 10% to 30% of cases. Indeed, most patients with MT-ATP6 disease do not manifest with full NARP syndrome (Ng et al., 2019b; Stendel et al., 2020). Nuclear genetic defects involving the maintenance and replication of mitochondrial DNA, such as POLG (see section “Ataxia”), mitochondrial inner membrane protein 17 gene [MPV17] (Karadimas et al., 2006) and thymidine phosphorylase gene [TYMP] (Nishino et al., 1999) are associated with prominent neuropathy. Patients with recessive MPV17 disease typically present with a neonatal/infantile onset hepatocerebral syndrome characterized by progressive liver failure (>90%) [for which transplant is often indicated], neurodevelopmental delay (>80%), hypotonia (>70%), and sensorimotor neuropathy (20%) (El-Hattab et al., 2018). Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is an ultra-rare disease (estimated prevalence A (mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 1 gene [MT-ND1]), m.11778G>A (mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 4 gene [MT-ND4]) and m.14484T>C (mitochondrially encoded NADH:Ubiquinone Oxidoreductase Core Subunit 6 gene [MT-ND6]), account for >90% of LHON cases; with m.11778G>A variant being the most prevalent worldwide. The penetrance of LHON is different between the sexes, with risks for male and female carriers estimated to be 50% and 10%–30%, respectively. However, a more recent Australian study suggested that the overall risk of visual loss among the mutation carriers is much lower, although it remains true that men have a higher risk than women (17.5% vs 5.4%) (Lopez Sanchez et al., 2021). In addition, a later disease onset has also been observed in women than men although most cases develop visual loss by the age of 50 years. Smoking and excess alcohol are purported environmental risk factors for the disease manifestation (Carelli et al., 2016). Better education regarding these risk factors in LHON families, together with a general improvement in public smoking habits may be behind the decreased risk of blindness observed in the Australian cohort study. Also, and rather interestingly, estrogen has been demonstrated to have a protective effect in LHON patient cell line studies, which may explain a reduced penetrance in female carriers (Giordano et al., 2011). Spontaneous recovery of visual function occurs in patients with LHON and the best prognosis is associated with the m.14484T>C variant, where improvement of visual acuity has been documented in 70% of the cases (Riordan-Eva et al., 1995). In contrast, less than 15% of patients who were older than 15 years old with the m.11778G>A variant showed meaningful visual recovery (Newman et al., 2020; Yu-Wai-Man et al., 2022). A small number of cases have been described to exhibit LHON plus phenotype, characterized by generalized dystonia and other movement disorders (Nikoskelainen et al., 1995), and abnormal MRI head scan findings reminiscent of Leigh-like syndrome (McFarland et al., 2007). The association of optic neuropathy and multiple sclerosis in carriers of LHON variants is referred to as Harding’s disease (Harding et al., 1992). However, it remains controversial as to whether Harding’s disease represents a phenotypic expression of pathogenic mtDNA variants (Matthews et al., 2014) or is incidental association between two different conditions (Pfeffer et al., 2013).

DOMINANT OPTIC ATROPHY (DOA) About 60% of autosomal dominant (AD) optic atrophy cases are due to heterozygous variants of OPA1, which

MITOCHONDRIAL ENCEPHALOMYOPATHY have an estimated prevalence of 1 in 25,000 (Yu-WaiMan and Chinnery, 2013). It is characterized by a slowly progressive, childhood-onset optic neuropathy with moderate to severe loss of visual acuity. Overall, the severity of visual impairment is milder in DOA cases when compared to LHON, although spontaneous recovery of visual function is not observed (Newman et al., 2022). Around 20% patients with OPA1 pathogenic variants develop extra neurologic manifestations, e.g., bilateral sensorineural hearing loss, CPEO, peripheral neuropathy, spasticity, ataxia and Parkinsonism (Yu-Wai-Man et al., 2010; Carelli et al., 2015).

DIAGNOSTIC INVESTIGATION Making a diagnosis of mitochondrial disease relies on a meticulous clinical history-taking, physical examination and analysis of the family pedigree. Clinical investigations such as blood or cerebrospinal fluid (CSF) lactate may be elevated and helpfully signpost the diagnosis of mitochondrial disease, but unfortunately the latter cannot be excluded on the basis of normal lactate results. Mild-moderate elevation of the serum CK is also supportive, but not by any means definitive, though grossly elevated CK in the absence of rhabdomyolysis would be most unusual for mitochondrial disease. More specialized bedside tests, such as serum fibroblast growth factor 21 (FGF21) levels, can be helpful in distinguishing myopathy manifesting mitochondrial disease from other causes of muscle weakness (Suomalainen et al., 2011), but is of no help in diagnosing mitochondrial disease where CNS, PNS, cardiac, liver, eye or renal involvement are primary features (Lin et al., 2020). Neuroradiological investigations, specifically MRI and magnetic resonance spectroscopy (MRS), can be helpful in identifying the subacute onset nonvascular territory T2 hyperintense lesions of stroke-like episodes or brainstem lesions of Leigh syndrome (Tschampa et al., 2013; Bindu et al., 2015; Bonfante et al., 2016). Other central nervous system features of mitochondrial disease, that might not be associated with an acute/subacute presentation, can also be identified, such as leukodystrophy (e.g., Aspartyl (D)aminoacyl trNA synthetase gene [DARS2], NADH:Ubiquinone Oxidoreductase Core Subunit S2 gene [NDUFS2], NDUFV1), symmetrical basal ganglia involvement (LS/MELAS), corpus callosal dysplasia (PDCD) or elevated lactate peak on MRS (Bricout et al., 2014). Global cerebral and cerebellar atrophy are nonspecific common neuroimaging findings identified in both pediatric- and adult-onset mitochondrial diseases. The role of electroencephalography in diagnosis of mitochondrial epilepsy and encephalopathy is discussed elsewhere in this chapter, and while it undoubtedly has a

575

central role in the diagnostic work-up, many of the features observed on EEG are not unique to mitochondrial disease. However, POLG-related epilepsy and Alpers’ syndrome in particular, is one of the few causes of Rhythmic High Amplitude Delta with superimposed Spikes (RHADS) and should raise clinical suspicion of this diagnosis (Anagnostou et al., 2016). Nonconvulsive status epilepticus should be considered as a potential cause for confusion/encephalopathy when patients present with a stroke-like episode and EEG is integral to the diagnostic work-up of such patients (Ng et al., 2019a). As for specialized diagnostic testing, there has been a transformation in the approach to this over the last 10 years since the widespread introduction of relatively inexpensive next-generation sequencing (NGS) technology. The “gold standard” of skeletal muscle biopsy, which stood front and central in mitochondrial diagnostic algorithms around the world (Taylor and Turnbull, 2005) has largely been replaced by whole-exome (WES) or whole-genome sequencing (WGS), the latter often being able to read the mitochondrial genome too (Stenton and Prokisch, 2020; Thompson et al., 2020; Alston et al., 2021). An in-depth discussion of the merits of NGS technologies is beyond the scope of this chapter but DNA derived from patient peripheral blood leukocytes is now the tissue of choice in investigating mitochondrial disease for both nuclear and mitochondrial genomes. One note of caution however, some pathogenic mtDNA variants are restricted to skeletal muscle and not detectable at all in blood (Blakely et al., 2005; Hardy et al., 2016) while others (e.g., m.3243A>G) are “lost” from blood as the patient ages. The latter results from the predictable rate of decline of heteroplasmy in blood, but can be corrected for mathematically (Grady et al., 2018; Franco et al., 2022). There remains a role for skeletal muscle biopsy in patients when WES/WGS are negative (Ng et al., 2021a), yet there is still a strong clinical suspicion of mitochondrial disease; however the best NGS pipelines are not achieving diagnostic hit rates of more than 60% to 70% (Taylor et al., 2014; Stenton and Prokisch, 2020). Skeletal muscle biopsy allows histochemical analysis by sequential COX and succinate dehydrogenase (SDH) assays (Taylor et al., 2004), or more recently multiplex immunofluorescent analysis (Rocha et al., 2015). Isolation of a mitochondrial fraction from skeletal muscle allows direct assessment of individual mitochondrial respiratory chain complexes (Frazier et al., 2020). The skeletal muscle is also a source of DNA for mitochondrial genome sequencing and is also useful in functional validation of suspected pathogenic variants of mtDNA using techniques such as Northern and Western blotting (Zierz et al., 2019; Ng et al., 2020). Skin biopsy, followed by fibroblast culture, also proves extremely helpful in functional validation of

576

Y.S. NG AND R. MCFARLAND

possible pathogenic variants though, as low energy demanding cells, skin fibroblasts may not exhibit florid functional consequences of disruptions to OXPHOS caused by the pathogenic variant (Thompson et al., 2020). Metabolomics, proteomics, lipidomics, and transcriptomics (Ren et al., 2018; Frazier et al., 2021; Yepez et al., 2022) complement the current genomic approach and are all likely to become established in the diagnostic algorithm for mitochondrial disease (Alston et al., 2021). This multiomic diagnostic approach will not only help resolve the 30% to 40% of patients with a suspected genetic diagnosis who are at present undiagnosed, but will also help inform our understanding of the basic biology, function and pathological processes that affect mitochondria and hopefully identify potential targets for treatment.

MANAGEMENT Acute neurological presentations Stroke-like episodes are neurological emergencies in patients with MELAS syndrome and the European-based consensus guidance emphasizes that instigation of intravenous antiepileptic drug treatment should be prioritized early in the clinical course of the event (Ng et al., 2019a). The diagnosis of nonconvulsive status epilepticus requires a high index of clinical suspicion and should be confirmed with an EEG study. Patients presenting with the POLG-related seizures are at risk of developing supra-refractory status epilepticus with a very poor prognosis (Anagnostou et al., 2016; Ng et al., 2021b) although an early escalation to general anesthetic agents may modify the survival probability and disease trajectory (Hikmat et al., 2017). Sodium valproate is an absolute contraindication in patients with POLG-related disease due to the risk of precipitating fatal liver failure (Stewart et al., 2010). A systematic review has clearly demonstrated there is very poor evidence proving the efficacy of L-arginine in both acute and prophylactic settings (Stefanetti et al., 2022). There are some potential safety concerns regarding use of L-arginine including precipitation of metabolic/lactic acidosis and renal failure. Over 50% of patients presenting with stroke-like episodes develop intestinal pseudo-obstruction and active medical management is required to circumvent aspiration pneumonia (Ng et al., 2016). Children and young adults with LS are at risk of developing episodic, subacute brainstem dysfunction, often precipitated by inter-current febrile illness or physical stress (Sofou et al., 2014; Ng et al., 2019b). Prompt recognition is crucial and critical care support for low conscious level, dysphagia and central hypoventilation might be required for some cases. The disease burden and prognosis of LS is variable and in part, is determined

by the genotype, as shown by several cohort studies (Sofou et al., 2018; Ogawa et al., 2020; Lim et al., 2022). Idebenone, a synthetic short-chain benzoquinone, has become the first licensed treatment for patients with LHON in the European Union since 2015 following the publication of a randomized controlled study (RHODOS) (Klopstock et al., 2011) and data derived subsequently from an expanded access program (Newman et al., 2022). Approximately 50% of patients receiving idebenone treatment for 18 to 24 months showed clinically relevant recovery, defined as an improvement from off-chart to on-chart by at least one full line (5 letters on ETDRS chart), or an improvement in an on-chart best corrected visual acuity by at least 2 lines (10 letters) according to the most recent real-world data (Catarino et al., 2020).

Chronic neurological presentations Progressive eyelid ptosis can result in restriction of the visual field, as well as a social stigma in patients living with CPEO, however the performance of oculoplastic surgery can lead to significant functional and esthetic improvements (Sebastiá et al., 2015; Eshaghi et al., 2021). The symptomatic management of various chronic neurologic complications in mitochondrial diseases, such as dystonia, Parkinsonism and neuropathy, are similar to other neuromuscular disorders. There is no specific safety concern about common pharmacological agents used in these settings (De Vries et al., 2020). Muscle fatigue and exercise intolerance are common and debilitating symptoms experienced by many patients, irrespective of the underlying genotypes (Mancuso et al., 2012; Gorman et al., 2015b). Patients and their caregivers should be advised of the importance in pacing their physical activities and maintaining hydration. They should be encouraged to exercise within their limits as aerobic training is safe and beneficial for the overall cardiorespiratory fitness in mitochondrial myopathy (Bates et al., 2013; Jeppesen, 2020; Stefanetti et al., 2020). Patients benefit from periodic physiotherapy and occupational therapy assessments, and access to aids such as cutlery aids and wheelchairs as the needs arise. Regular evaluation of respiratory function is indicated in cases which significant muscle weakness are frequently present, for example, in patients with TK2—(Garone et al., 2018), RRM2B—(Lim et al., 1993), and m.8344A>Grelated (Catteruccia et al., 2015) mitochondrial diseases.

Other systemic involvements Given the multisystem nature of many mitochondrial diseases, regular surveillance for common complications such as sensorineural deafness, cardiac involvement, gut dysmotility and renal disease should be coordinated with different medical specialities and best practice

MITOCHONDRIAL ENCEPHALOMYOPATHY guidelines are available (Parikh et al., 2017; WCMR, 2022). Some patients with profound deafness can benefit from cochlear implant if the hearing function is not corrected by the use of hearing aids (Zia et al., 2021). A minority of patients develop end-organ failure (e.g., heart, kidney, and liver) and organ transplantation should be considered in selected cases as the survival has been shown to be comparable to the other patients without mitochondrial disease (Parikh et al., 2016; Shimura et al., 2020; Weiner et al., 2020; Nishida et al., 2022). There are no controlled clinical trials to support routine dietary supplements in patients with mitochondrial disease (Pfeffer et al., 2012). There are specific mitochondrial diseases such as primary coenzyme Q10 deficiency, primary disorders of vitamin cofactor metabolism (e.g., thiamine, biotin, and riboflavin) and multiple acyl-CoA dehydrogenase deficiency that are likely to benefit from the supplementation (Distelmaier et al., 2017).

CONCLUSION Mitochondrial diseases are individually rare clinical conditions that together are the largest single group of inherited neurometabolic conditions. They encompass a wide range of pathologies involving different organ systems, but share a common etiology of mitochondrial dysfunction. The CNS and skeletal muscle are prominently involved in many mitochondrial diseases, manifesting as encephalomyopathy, and exhibit a spectrum of associated clinical features including epilepsy, visual impairment, deafness, neurodevelopmental regression/ delay, neuropathy, weakness, fatigue, movement disorders, and stroke-like episodes. More broadly, perturbation of mitochondrial function may play a significant role in a number of more common neurodegenerative disorders including Parkinson disease and the dementias. While much has been learned about mitochondrial disease in the last decade, particularly since the advent of NGS technology, there is still much to be learned. The validated multiomic tools now at our disposal will be crucial not only in improving diagnosis of mitochondrial disease but in better understanding the pathological processes at play and factors that influence disease expression. Creating better disease models as a result of this knowledge will help identify life-changing therapy for these most devastating of diseases (Russell et al., 2020).

ABBREVIATION AD, autosomal dominant; ADC, apparent diffusion coefficient; ADCK3, aarF domain containing kinase 3; ADP, adenosine diphosphate; AGK, acylglycerol kinase; ANT1, Adenine Nucleotide Translocator 1; AR,

577

autosomal recessive; ATP, adenosine triphosphate; BCS1L, BCS1 Homologubiquinol-cytochrome C reductase complex chaperone; CMT, Charcot-Marie-Tooth; CNS, central nervous system; CoQ8A, coenzyme Q8A; COX, cytochrome c oxidase; CPEO, chronic progressive external ophthalmoplegia; CSF, cerebrospinal fluid; CT, computed tomography; Dat, dopamine active transporter; DNA, deoxyribonucleic acid; EEG, electroencephalogram; FSGS, focal segmental glomerulosclerosis; GDAP1, Ganglioside-induced differentiation-associated protein; LHON, Leber hereditary optic neuropathy; LS, Leigh syndrome; LARS2, leucyl-tRNA synthetase 2, mitochondrial; MEERF, myoclonic epilepsy with ragged-red fibers; MELAS, mitochondrial encephalomyopathy lactic acidosis and stroke-like episodes; MFN2, Mitofusin-2; MNGIE, mitochondrial neurogastrointestinal encephalopathy; MPV17, mitochondrial inner membrane protein MPV17; MRI, magnetic resonance imaging; MRS, magnetic resonance spectroscopy; MT-ATP6, mitochondrially encoded ATP synthase membrane subunit 6; mtDNA, mitochondrial DNA; MT-ND1, mitochondrially encoded NADH:Ubiquinone oxidoreductase core subunit 1; MT-ND4, mitochondrially encoded NADH:Ubiquinone oxidoreductase core subunit 4; MT-ND6, mitochondrially encoded NADH:Ubiquinone oxidoreductase core subunit 6; MT-TK, mitochondrially encoded tRNA lysine; MT-TL1, mitochondrially encoded tRNA leucine 1; NARP, neuropathy, ataxi, retinitis pigmentosa; nDNA, nuclear DNA; NDUFAF2, NADH:ubiquinone oxidoreductase complex assembly factor 2; NDUFV1, NADH: ubiquinone oxidoreductase core subunit V1; NDUFS4, NADH dehydrogenase [ubiquinone] iron–sulfur protein 4, mitochondrial; NGS, next-generation sequencing; OPA1, optic atrophy 1; OXPHOS, oxidative phosphorylation; PDCD, pyruvate dehydrogenase complex deficiency; PDSS2, Decaprenyl-diphosphate synthase subunit 2; PEO, progressive external ophthalmoplegia; PNS, peripheral nervous system; POLG1, mitochondrial DNA polymerase gamma 1; RARS2, arginyl-tRNA synthetase 2, mitochondria; RNA, ribonucleic acid; ROS, reactive oxygen species; RRM2B, ribonucleotide reductase regulatory TP53 inducible subunit M2B; SDH, succinate dehydrogenase; SERAC1, serine active site containing 1; SLC25A4, solute carrier family 25 member 4; SLC25AA22, solute carrier family 25 member 22; SNHL, sensorineural hearing loss; SPG7, spastic paraplegia 7; SUCLA2, Succinyl-CoA ligase [ADPforming] subunit beta, mitochondrial; SURF1, surfeit locus protein 1; TAZ, Taffazin; TCA, tricarboxylic acid; TK2, thymidine kinase 2; TMEM70, Transmembrane protein 70; tRNA, transfer RNA; TWNK, twinkle mtDNA helicase; TYMP, thymidine phosphorylase; UK, United Kingdom; WES, whole-exome sequencing; WGS, whole-genome sequencing.

578

Y.S. NG AND R. MCFARLAND

REFERENCES Acosta MJ, Vazquez Fonseca L, Desbats MA et al. (2016). Coenzyme Q biosynthesis in health and disease. Biochim Biophys Acta - Bioenerg 1857: 1079–1085. Almalki A, Alston CL, Parker A et al. (2014). Mutation of the human mitochondrial phenylalanine-tRNA synthetase causes infantile-onset epilepsy and cytochrome c oxidase deficiency. Biochim Biophys Acta 1842: 56–64. Alston CL, Stenton SL, Hudson G et al. (2021). The genetics of mitochondrial disease: dissecting mitochondrial pathology using multi-omic pipelines. J Pathol 254: 430–442. https:// doi.org/10.1002/path.5641. PMID: 33586140; PMCID: PMC8600955. Altmann J, Buchner B, Nadaj-Pakleza A et al. (2016). Expanded phenotypic spectrum of the m.8344A>G “MERRF” mutation: data from the German mitoNET registry. J Neurol 263: 961–972. Amati-Bonneau P, Guichet A, Olichon A et al. (2005). OPA1 R445H mutation in optic atrophy associated with sensorineural deafness. Ann Neurol 58: 958–963. Amati-Bonneau P, Valentino ML, Reynier P et al. (2008). OPA1 mutations induce mitochondrial DNA instability and optic atrophy ‘plus’ phenotypes. Brain 131: 338–351. Anagnostou ME, Ng YS, Taylor RW et al. (2016). Epilepsy due to mutations in the mitochondrial polymerase gamma (POLG) gene: a clinical and molecular genetic review. Epilepsia 57: 1531–1545. Anderson S, Bankier AT, Barrel BG et al. (1981). Sequence and organization of the human mitochondrial genome. Nature 290: 457–465. Barnerias C, Saudubray J-M, Touati G et al. (2010). Pyruvate dehydrogenase complex deficiency: four neurological phenotypes with differing pathogenesis. Dev Med Child Neurol 52: e1–e9. Barth PG, Wanders RJ, Vreken P et al. (1999). X-linked cardioskeletal myopathy and neutropenia (Barth syndrome) (MIM 302060). J Inherit Metab Dis 22: 555–567. Bates MGD, Newman JH, Jakovljevic DG et al. (2013). Defining cardiac adaptations and safety of endurance training in patients with m.3243A>G-related mitochondrial disease. Int J Cardiol 168: 3599–3608. Bernardino Gomes TM, Ng YS, Pickett SJ et al. (2021). Mitochondrial DNA disorders: from pathogenic variants to preventing transmission. Hum Mol Genet 30: R245–R253. Bindu PS, Arvinda H, Taly AB et al. (2015). Magnetic resonance imaging correlates of genetically characterized patients with mitochondrial disorders: a study from south India. Mitochondrion 25: 6–16. Blakely EL, Mitchell AL, Fisher N et al. (2005). A mitochondrial cytochrome b mutation causing severe respiratory chain enzyme deficiency in humans and yeast. FEBS J 272: 3583–3592. Bonfante E, Koenig MK, Adejumo RB et al. (2016). The neuroimaging of Leigh syndrome: case series and review of the literature. Pediatr Radiol 46: 443–451. Bourdon A, Minai L, Serre V et al. (2007). Mutation of RRM2B, encoding p53-controlled ribonucleotide reductase (p53R2), causes severe mitochondrial DNA depletion. Nat Genet 39: 776–780.

Bourgeron T, Rustin P, Chretien D et al. (1995). Mutation of a nuclear succinate dehydrogenase gene results in mitochondrial respiratory chain deficiency. Nat Genet 11: 144–149. Braz LP, Ng YS, Gorman GS et al. (2019). Neuromuscular junction abnormalities in mitochondrial disease. An observational cohort study. Neurol Clin Pract. https://doi.org/ 10.1212/CPJ.0000000000000795. Bricout M, Grevent D, Lebre AS et al. (2014). Brain imaging in mitochondrial respiratory chain deficiency: combination of brain MRI features as a useful tool for genotype/phenotype correlations. J Med Genet 51: 429–435. Broomfield A, Sweeney MG, Woodward CE et al. (2015). Paediatric single mitochondrial DNA deletion disorders: an overlapping spectrum of disease. J Inherit Metab Dis 38: 445–457. Budde SM, van den Heuvel LP, Janssen AJ et al. (2000). Combined enzymatic complex I and III deficiency associated with mutations in the nuclear encoded NDUFS4 gene. Biochem Biophys Res Commun 275: 63–68. Carelli V, Musumeci O, Caporali L et al. (2015). Syndromic parkinsonism and dementia associated with OPA1 missense mutations. Ann Neurol 78: 21–38. Carelli V, d’Adamo P, Valentino ML et al. (2016). Parsing the differences in affected with LHON: genetic versus environmental triggers of disease conversion. Brain 139: e17. Casari G, De Fusco M, Ciarmatori S et al. (1998). Spastic paraplegia and OXPHOS impairment caused by mutations in paraplegin, a nuclear-encoded mitochondrial metalloprotease. Cell 93: 973–983. Catarino CB, von Livonius B, Priglinger C et al. (2020). Realworld clinical experience with idebenone in the treatment of leber hereditary optic neuropathy. J Neuroophthalmol 40: 558–565. Catteruccia M, Sauchelli D, Della Marca G et al. (2015). “Myo-cardiomyopathy” is commonly associated with the A8344G “MERRF” mutation. J Neurol 262: 701–710. https://doi.org/10.1007/s00415-014-7632-0. PMID: 25559684. Chapman J, Ng YS, Nicholls TJ (2020). The maintenance of mitochondrial DNA integrity and dynamics by mitochondrial membranes. Life (Basel) 10. Chinnery PF, DiMauro S, Shanske S et al. (2004). Risk of developing a mitochondrial DNA deletion disorder. The Lancet 364: 592–596. Ciafaloni E, Santorelli FM, Shanske S et al. (1993). Maternally inherited Leigh syndrome. J Pediatr 122: 419–422. Clarke NF, Waddell LB, Cooper ST et al. (2010). Recessive mutations in RYR1 are a common cause of congenital fiber type disproportion. Hum Mutat 31: E1544–E1550. Clarke SLN, Bowron A, Gonzalez IL et al. (2013). Barth syndrome. Orphanet J Rare Dis 8: 23. Cohen BH, Chinnery PF, Copeland WC (1993). POLGRelated Disorders. In: RA Pagon, MP Adam, HH Ardinger et al. (Eds.), GeneReviews(R). University of Washington, Seattle, Seattle (WA). University of Washington, Seattle. All rights reserved. Davidzon G, Greene P, Mancuso M et al. (2006). Early-onset familial parkinsonism due to POLG mutations. Ann Neurol 59: 859–862.

MITOCHONDRIAL ENCEPHALOMYOPATHY de Laat P, Zweers HE, Knuijt S et al. (2015). Dysphagia, malnutrition and gastrointestinal problems in patients with mitochondrial disease caused by the m3243A>G mutation. Neth J Med 73: 30–36. De Vries MC, Brown DA, Allen ME et al. (2020). Safety of drug use in patients with a primary mitochondrial disease: an international Delphi-based consensus. J Inherit Metab Dis 43: 800–818. Diodato D, Ghezzi D, Tiranti V (2014). The mitochondrial aminoacyl tRNA synthetases: genes and syndromes. Int J Cell Biol 2014: 787956. Distelmaier F, Haack TB, Wortmann SB et al. (2017). Treatable mitochondrial diseases: cofactor metabolism and beyond. Brain 140: e11. Domı´nguez-Gonza´lez C, Herna´ndez-Laı´n A, Rivas E et al. (2019). Late-onset thymidine kinase 2 deficiency: a review of 18 cases. Orphanet J Rare Dis 14: 100. Ekbom K (1975). Hereditary ataxia, photomyoclonus, skeletal deformities and lipoma. Acta Neurol Scand 51: 393–404. El-Hattab AW, Almannai M, Scaglia F (2017a). Arginine and citrulline for the treatment of MELAS syndrome. J Inborn Errors Metab Screen 5. https://doi.org/10.1177/ 2326409817697399. El-Hattab AW, Craigen WJ, Scaglia F (2017b). Mitochondrial DNA maintenance defects. Biochim Biophys Acta Mol Basis Dis 1863: 1539–1555. El-Hattab AW, Wang J, Dai H et al. (2018). MPV17-related mitochondrial DNA maintenance defect: New cases and review of clinical, biochemical, and molecular aspects. Hum Mutat 39: 461–470. Elpeleg O, Miller C, Hershkovitz E et al. (2005). Deficiency of the ADP-forming succinyl-CoA synthase activity is associated with encephalomyopathy and mitochondrial DNA depletion. Am J Hum Genet 76: 1081–1086. Emmanuele V, Lopez LC, Berardo A et al. (2012). Heterogeneity of coenzyme Q10 deficiency: patient study and literature review. Arch Neurol 69: 978–983. Engelsen BA, Tzoulis C, Karlsen B et al. (2008). POLG1 mutations cause a syndromic epilepsy with occipital lobe predilection. Brain 131: 818–828. Eshaghi M, Arabi A, Eshaghi S (2021). Surgical management of ptosis in chronic progressive external ophthalmoplegia. Eur J Ophthalmol 31: 2064–2068. Feely SM, Laura M, Siskind CE et al. (2011). MFN2 mutations cause severe phenotypes in most patients with CMT2A. Neurology 76: 1690–1696. Franco M, Pickett SJ, Fleischmann Z et al. (2022). Dynamics of the most common pathogenic mtDNA variant m.3243A > G demonstrate frequency-dependency in blood and positive selection in the germline. Hum Mol Genet 31: 4075–4086. Fratter C, Gorman GS, Stewart JD et al. (2010). The clinical, histochemical, and molecular spectrum of PEO1 (Twinkle)-linked adPEO. Neurology 74: 1619–1626. Frazier AE, Vincent AE, Turnbull DM et al. (2020). Assessment of mitochondrial respiratory chain enzymes in cells and tissues. Methods Cell Biol 155: 121–156. Frazier AE, Compton AG, Kishita Y et al. (2021). Fatal perinatal mitochondrial cardiac failure caused by recurrent

579

de novo duplications in the ATAD3 locus. Med (N Y) 2: 49–73. Fukuhara N, Tokiguchi S, Shirakawa K et al. (1980). Myoclonus epilepsy associated with ragged-red fibres (mitochondrial abnormalities): disease entity or a syndrome? Light-and electron-microscopic studies of two cases and review of literature. J Neurol Sci 47: 117–133. Garone C, Taylor RW, Nascimento A et al. (2018). Retrospective natural history of thymidine kinase 2 deficiency. J Med Genet 55: 515–521. Ghaoui R, Sue CM (2018). Movement disorders in mitochondrial disease. J Neurol 265: 1230–1240. Giordano C, Montopoli M, Perli E et al. (2011). Oestrogens ameliorate mitochondrial dysfunction in Leber’s hereditary optic neuropathy. Brain 134: 220–234. Gorman GS, Blakely EL, Hornig-Do HT et al. (2015a). Novel MTND1 mutations cause isolated exercise intolerance, complex I deficiency and increased assembly factor expression. Clin Sci (Lond) 128: 895–904. Gorman GS, Elson JL, Newman J et al. (2015b). Perceived fatigue is highly prevalent and debilitating in patients with mitochondrial disease. Neuromuscul Disord 25: 563–566. Gorman GS, Schaefer AM, Ng Y et al. (2015c). Prevalence of nuclear and mitochondrial DNA mutations related to adult mitochondrial disease. Ann Neurol 77: 753–759. Gorman GS, Chinnery PF, DiMauro S et al. (2016). Mitochondrial diseases. Nat Rev Dis Primers 2: 16080. Goto Y, Nonaka I, Horai S (1990). A mutation in the tRNA(Leu)(UUR) gene associated with the MELAS subgroup of mitochondrial encephalomyopathies. Nature 348: 651–653. Grady JP, Campbell G, Ratnaike T et al. (2014). Disease progression in patients with single, large-scale mitochondrial DNA deletions. Brain 137: 323–334. Grady JP, Pickett SJ, Ng YS et al. (2018). mtDNA heteroplasmy level and copy number indicate disease burden in m.3243A>G mitochondrial disease. EMBO Mol Med 10. Haghighi A, Haack TB, Atiq M et al. (2014). Sengers syndrome: six novel AGK mutations in seven new families and review of the phenotypic and mutational spectrum of 29 patients. Orphanet J Rare Dis 9: 1–12. Halter JP, Michael W, Schupbach M et al. (2015). Allogeneic haematopoietic stem cell transplantation for mitochondrial neurogastrointestinal encephalomyopathy. Brain 138: 2847–2858. Hammans SM (2020). Mitochondrial neurogastrointestinal encephalopathy disease (MNGIE). Pract Neurol 21: 43–47. Harding AE, Sweeney MG, Miller DH et al. (1992). Occurrence of a multiple sclerosis-like illness in women who have a Leber’s hereditary optic neuropathy mitochondrial DNA mutation. Brain 115: 979–989. Hardy SA, Blakely EL, Purvis AI et al. (2016). Pathogenic mtDNA mutations causing mitochondrial myopathy: The need for muscle biopsy. Neurol Genet 2. Hasegawa H, Matsuoka T, Goto Y et al. (1991). Strongly succinate dehydrogenase-reactive blood vessels in muscles from patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes. Ann Neurol 29: 601–605.

580

Y.S. NG AND R. MCFARLAND

Hayhurst H, Anagnostou ME, Bogle HJ et al. (2019). Dissecting the neuronal vulnerability underpinning Alpers’ syndrome: a clinical and neuropathological study. Brain Pathol 29: 97–113. Hewamadduma CA, Hoggard N, O’Malley R et al. (2018). Novel genotype-phenotype and MRI correlations in a large cohort of patients with SPG7 mutations. Neurol Genet 4: e279. Hikmat O, Tzoulis C, Knappskog PM et al. (2016). ADCK3 mutations with epilepsy, stroke-like episodes and ataxia: a POLG mimic? Eur J Neurol 23: 1188–1194. Hikmat O, Eichele T, Tzoulis C et al. (2017). Understanding the epilepsy in POLG related disease. Int J Mol Sci 18. Hikmat O, Naess K, Engvall M et al. (2020). Simplifying the clinical classification of polymerase gamma (POLG) disease based on age of onset; studies using a cohort of 155 cases. J Inherit Metab Dis 43: 726–736. Hikmat O, Isohanni P, Keshavan N et al. (2021). Expanding the phenotypic spectrum of BCS1L-related mitochondrial disease. Ann Clin Transl Neurol 8: 2155–2165. Hirano M, Ricci E, Koenigsberger MR et al. (1992). Melas: an original case and clinical criteria for diagnosis. Neuromuscul Disord 2: 125–135. Hirano M, Silvestri G, Blake DM et al. (1994). Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE): clinical, biochemical, and genetic features of an autosomal recessive mitochondrial disorder. Neurology 44: 721–727. Holt IJ, Harding AE, Morgan-Hughes JA (1988). Deletions of muscle mitochondrial DNA in patients with mitochondrial myopathies. Nature 331: 717–719. Holt IJ, Harding AE, Petty RK et al. (1990). A new mitochondrial disease associated with mitochondrial DNA heteroplasmy. Am J Hum Genet 46: 428–433. Honzik T, Tesarova M, Magner M et al. (2012). Neonatal onset of Mitochondrial disorders in 129 patients: clinical and laboratory characteristics and a new approach to diagnosis. J Inherit Metab Dis 35: 749–759. Iizuka T, Sakai F, Suzuki N et al. (2002). Neuronal hyperexcitability in stroke-like episodes of MELAS syndrome. Neurology 59: 816–824. Iizuka T, Sakai F, Kan S et al. (2003). Slowly progressive spread of the stroke-like lesions in MELAS. Neurology 61: 1238–1244. Jeppesen TD (2020). Aerobic exercise training in patients with mtDNA-related mitochondrial myopathy. Front Physiol 11: 349. Jeppesen TD, Madsen KL, Poulsen NS et al. (2021). Exercise testing, physical training and fatigue in patients with mitochondrial myopathy related to mtDNA mutations. J Clin Med 10: 1796. Karadimas CL, Vu TH, Holve SA et al. (2006). Navajo neurohepatopathy is caused by a mutation in the MPV17 gene. Am J Hum Genet 79: 544–548. Kaufman KR, Zuber N, Rueda-Lara MA et al. (2010). MELAS with recurrent complex partial seizures, nonconvulsive status epilepticus, psychosis, and behavioral disturbances: case analysis with literature review. Epilepsy Behav 18: 494–497.

Kaukonen J, Juselius JK, Tiranti V et al. (2000). Role of adenine nucleotide translocator 1 in mtDNA maintenance. Science 289: 782–785. Kearns TP, Sayre GP (1958). Retinitis pigmentosa, external ophthalmophegia, and complete heart block: unusual syndrome with histologic study in one of two cases. AMA Arch Ophthalmol 60: 280–289. Kim JH, Lim MK, Jeon TY et al. (2011). Diffusion and perfusion characteristics of MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episode) in thirteen patients. Korean J Radiol 12: 15–24. Klopstock T, Yu-Wai-Man P, Dimitriadis K et al. (2011). A randomized placebo-controlled trial of idebenone in Leber’s hereditary optic neuropathy. Brain 134: 2677–2686. Koenig MK, Emrick L, Karaa A et al. (2016). Recommendations for the management of strokelike episodes in patients with mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes. JAMA Neurol 73: 591–594. Koga Y, Ishibashi M, Ueki I et al. (2002). Effects of L-arginine on the acute phase of strokes in three patients with MELAS. Neurology 58: 827–828. Koga Y, Akita Y, Nishioka J et al. (2005). L-arginine improves the symptoms of strokelike episodes in MELAS. Neurology 64: 710–712. Lake NJ, Compton AG, Rahman S et al. (2016). Leigh syndrome: one disorder, more than 75 monogenic causes. Ann Neurol 79: 190–203. Lamperti C, Zeviani M (2016). Myoclonus epilepsy in mitochondrial disorders. Epileptic Disord 18: 94–102. Lax NZ, Hepplewhite PD, Reeve AK et al. (2012a). Cerebellar ataxia in patients with mitochondrial DNA disease: a molecular clinicopathological study. J Neuropathol Exp Neurol 71: 148–161. Lax NZ, Pienaar IS, Reeve AK et al. (2012b). Microangiopathy in the cerebellum of patients with mitochondrial DNA disease. Brain 135: 1736–1750. Lax NZ, Whittaker RG, Hepplewhite PD et al. (2012c). Sensory neuronopathy in patients harbouring recessive polymerase mutations. Brain 135: 62–71. Lax NZ, Gorman GS, Turnbull DM (2016a). Invited review: central nervous system involvement in mitochondrial disease. Neuropathol Appl Neurobiol 43: 102–118. Lax NZ, Grady J, Laude A et al. (2016b). Extensive respiratory chain defects in inhibitory interneurones in patients with mitochondrial disease. Neuropathol Appl Neurobiol 42: 180–193. Leigh D (1951). Subacute necrotizing encephalomyelopathy in an infant. J Neurol Neurosurg Psychiatry 14: 216–221. Li J, Zhang W, Cui Z et al. (2021). Epilepsy associated with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes. Front Neurol 12: 675816. Licchetta L, Ferri L, La Morgia C et al. (2021). Epilepsy in MT-ATP6-related mils/NARP: correlation of elettroclinical features with heteroplasmy. Ann Clin Transl Neurol 8: 704–710.

MITOCHONDRIAL ENCEPHALOMYOPATHY Lim A, Thomas RH (2020). The mitochondrial epilepsies. Eur J Paediatr Neurol 24: 47–52. Lim AZ, McFarland R, Taylor RW et al. (1993). RRM2B Mitochondrial DNA Maintenance Defects. In: MP Adam, DB Everman, GM Mirzaa et al. (Eds.), GeneReviews(®). University of Washington, Seattle, Seattle (WA) Copyright © 1993-2022, University of Washington, Seattle. GeneReviews is a registered trademark of the University of Washington, Seattle. All rights reserved. Lim AZ, Ng YS, Blain A et al. (2022). Natural history of Leigh syndrome: a study of disease burden and progression. Ann Neurol 91: 117–130. Lin Y, Ji K, Ma X et al. (2020). Accuracy of FGF-21 and GDF-15 for the diagnosis of mitochondrial disorders: a meta-analysis. Ann Clin Transl Neurol 7: 1204–1213. Lopez Sanchez MIG, Kearns LS, Staffieri SE et al. (2021). Establishing risk of vision loss in Leber hereditary optic neuropathy. Am J Hum Genet 108: 2159–2170. L€ uhl S, Bode H, Schl€otzer W et al. (2016). Novel homozygous RARS2 mutation in two siblings without pontocerebellar hypoplasia - further expansion of the phenotypic spectrum. Orphanet J Rare Dis 11: 140. https:// doi.org/10.1186/s13023-016-0525-9. PMID: 27769281; PMCID: PMC5073905. Luoma P, Melberg A, Rinne JO et al. (2004). Parkinsonism, premature menopause, and mitochondrial DNA polymerase gamma mutations: clinical and molecular genetic study. Lancet 364: 875–882. Maas RR, Iwanicka-Pronicka K, Kalkan Ucar S et al. (2017). Progressive deafness–dystonia due to SERAC1 mutations: A study of 67 cases. Ann Neurol 82: 1004–1015. Man PYW, Turnbull DM, Chinnery PF (2002). Leber hereditary optic neuropathy. J Med Genet 39: 162–169. Mancuso M, Angelini C, Bertini E et al. (2012). Fatigue and exercise intolerance in mitochondrial diseases. Literature revision and experience of the Italian network of mitochondrial diseases. Neuromuscul Disord 22 Suppl 3: S226–S229. Mancuso M, Orsucci D, Angelini C et al. (2013). Phenotypic heterogeneity of the 8344A>G mtDNA “MERRF” mutation. Neurology 80: 2049–2054. Mancuso M, Orsucci D, Angelini C et al. (2014). Myoclonus in mitochondrial disorders. Mov Disord 29: 722–728. Mancuso M, Orsucci D, Angelini C et al. (2015). Redefining phenotypes associated with mitochondrial DNA single deletion. J Neurol 262: 1301–1309. Martikainen MH, Ng YS, Gorman GS et al. (2016). Clinical, genetic, and radiological features of extrapyramidal movement disorders in mitochondrial disease. JAMA Neurol 73: 668–674. Martinez-Ramirez D, Hack N, Vasquez ML et al. (2015). Deep brain stimulation in a case of mitochondrial disease. Mov Disord Clin Pract 3: 139–145. https://doi.org/10.1002/ mdc3.12241. PMID: 30713906; PMCID: PMC6353366. Mascialino B, Leinonen M, Meier T (2012). Meta-analysis of the prevalence of Leber hereditary optic neuropathy mtDNA mutations in Europe. Eur J Ophthalmol 22: 461–465.

581

Matthews L, Enzinger C, Fazekas F et al. (2014). MRI in Leber’s hereditary optic neuropathy: the relationship to multiple sclerosis. J Neurol Neurosurg Psychiatry 86: 537–542. Mayr JA, Haack TB, Graf E et al. (2012). Lack of the mitochondrial protein acylglycerol kinase causes Sengers syndrome. Am J Hum Genet 90: 314–320. McFarland R, Chinnery PF, Blakely EL et al. (2007). Homoplasmy, heteroplasmy, and mitochondrial dystonia. Neurology 69: 911–916. McFarland R, Taylor RW, Turnbull DM (2010). A neurological perspective on mitochondrial disease. Lancet Neurol 9: 829–840. Mizuguchi T, Nakashima M, Kato M et al. (2017). PARS2 and NARS2 mutations in infantile-onset neurodegenerative disorder. J Hum Genet 62: 525–529. Monlleo-Neila L, Toro MD, Bornstein B et al. (2013). Leigh syndrome and the mitochondrial m.13513G>A mutation: expanding the clinical spectrum. J Child Neurol 28: 1531–1534. Montano V, Orsucci D, Carelli V et al. (2022). Adult-onset mitochondrial movement disorders: a national picture from the Italian Network. J Neurol 269: 1413–1421. Musumeci O, Barca E, Lamperti C et al. (2019). Lipomatosis incidence and characteristics in an Italian cohort of mitochondrial patients. Front Neurol 10: 160. Naini A, Kaufmann P, Shanske S et al. (2005). Hypocitrullinemia in patients with MELAS: an insight into the “MELAS paradox”. J Neurol Sci 229-230: 187–193. Naviaux RK, Nyhan WL, Barshop BA et al. (1999). Mitochondrial DNA polymerase gamma deficiency and mtDNA depletion in a child with Alpers’ syndrome. Ann Neurol 45: 54–58. Nesbitt V, Pitceathly RDS, Turnbull DM et al. (2013). The UK MRC Mitochondrial Disease Patient Cohort Study: clinical phenotypes associated with the m.3243A>G mutation— implications for diagnosis and management. J Neurol Neurosurg Psychiatry 84: 936–938. Newman NJ, Carelli V, Taiel M et al. (2020). Visual outcomes in leber hereditary optic neuropathy patients with the m.11778G>A (MTND4) mitochondrial DNA mutation. J Neuroophthalmol 40: 547–557. Newman NJ, Yu-Wai-Man P, Biousse V et al. (2022). Understanding the molecular basis and pathogenesis of hereditary optic neuropathies: towards improved diagnosis and management. Lancet Neurol 22: 172–188. Ng YS, Feeney C, Schaefer AM et al. (2016). Pseudoobstruction, stroke, and mitochondrial dysfunction: a lethal combination. Ann Neurol 80: 686–692. Ng YS, Lax NZ, Maddison P et al. (2018). MT-ND5 mutation exhibits highly variable neurological manifestations at low mutant load. EBioMedicine 30: 86–93. Ng YS, Bindoff LA, Gorman GS et al. (2019a). Consensusbased statements for the management of mitochondrial stroke-like episodes. Wellcome Open Res 4: 201. Ng YS, Martikainen MH, Gorman GS et al. (2019b). Pathogenic variants in MT-ATP6: a United Kingdombased mitochondrial disease cohort study. Ann Neurol 86: 310–315.

582

Y.S. NG AND R. MCFARLAND

Ng YS, Thompson K, Loher D et al. (2020). Novel MT-ND gene variants causing adult-onset mitochondrial disease and isolated Complex I deficiency. Front Genet 11: 24. Ng YS, Bindoff LA, Gorman GS et al. (2021a). Mitochondrial disease in adults: recent advances and future promise. Lancet Neurol 20: 573–584. Ng YS, Lax NZ, Blain AP et al. (2021b). Forecasting strokelike episodes and outcomes in mitochondrial disease. Brain. Ng YS, Lim AZ, Panagiotou G et al. (2021c). Endocrine manifestations and new developments in mitochondrial disease. Endocr Rev. Ngoh A, Bras J, Guerreiro R et al. (2016). RARS2 mutations in a sibship with infantile spasms. Epilepsia 57: e97–e102. Nikali K, Suomalainen A, Saharinen J et al. (2005). Infantile onset spinocerebellar ataxia is caused by recessive mutations in mitochondrial proteins Twinkle and Twinky. Hum Mol Genet 14: 2981–2990. Nikoskelainen EK, Marttila RJ, Huoponen K et al. (1995). Leber’s “plus”: neurological abnormalities in patients with Leber’s hereditary optic neuropathy. J Neurol Neurosurg Psychiatry 59: 160–164. Nishida H, Nawano T, Fukuhara H et al. (2022). Outcomes of living kidney transplantation for mitochondrial disease patients: a case series. Transplant Proc 54: 267–271. Nishino I, Spinazzola A, Hirano M (1999). Thymidine phosphorylase gene mutations in MNGIE, a human mitochondrial disorder. Science 283: 689–692. Nouws J, Nijtmans LG, Smeitink JA et al. (2012). Assembly factors as a new class of disease genes for mitochondrial complex I deficiency: cause, pathology and treatment options. Brain 135: 12–22. Ogawa E, Fushimi T, Ogawa-Tominaga M et al. (2020). Mortality of Japanese patients with Leigh syndrome: effects of age at onset and genetic diagnosis. J Inherit Metab Dis 43: 819–826. Ohama E, Ohara S, Ikuta F et al. (1987). Mitochondrial angiopathy in cerebral blood vessels of mitochondrial encephalomyopathy. Acta Neuropathol 74: 226–233. Orsucci D, Angelini C, Bertini E et al. (2017). Revisiting mitochondrial ocular myopathies: a study from the Italian Network. J Neurol 264: 1777–1784. Pareyson D, Piscosquito G, Moroni I et al. (2013). Peripheral neuropathy in mitochondrial disorders. Lancet Neurol 12: 1011–1024. Parikh S, Karaa A, Goldstein A et al. (2016). Solid organ transplantation in primary mitochondrial disease: proceed with caution. Mol Genet Metab 118: 178–184. Parikh S, Goldstein A, Karaa A et al. (2017). Patient care standards for primary mitochondrial disease: a consensus statement from the Mitochondrial Medicine Society. Genet Med 19. Parikh S, Karaa A, Goldstein A et al. (2019). Diagnosis of ‘possible’ mitochondrial disease: an existential crisis. J Med Genet 56: 123–130. Patel KP, O’Brien TW, Subramony SH et al. (2012). The spectrum of pyruvate dehydrogenase complex deficiency: clinical, biochemical and genetic features in 371 patients. Mol Genet Metab 105: 34–43.

Pavlakis SG, Phillips PC, DiMauro S et al. (1984). Mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes: a distinctive clinical syndrome. Ann Neurol 16: 481–488. Pearson HA, Lobel JS, Kocoshis SA et al. (1979). A new syndrome of refractory sideroblastic anemia with vacuolization of marrow precursors and exocrine pancreatic dysfunction. J Pediatr 95: 976–984. Percetti M, Franco G, Monfrini E et al. (2022). TWNK in Parkinson’s disease: a movement disorder and mitochondrial disease center perspective study. Mov Disord 37: 1938–1943. Petty RK, Harding AE, Morgan-Hughes JA (1986). The clinical features of mitochondrial myopathy. Brain 109: 915–938. Pfeffer G, Majamaa K, Turnbull DM et al. (2012). Treatment for mitochondrial disorders. Cochrane Database Syst Rev 4: Cd004426. Pfeffer G, Burke A, Yu-Wai-Man P et al. (2013). Clinical features of MS associated with Leber hereditary optic neuropathy mtDNA mutations. Neurology 81: 2073–2081. Pfeffer G, Gorman GS, Griffin H et al. (2014). Mutations in the SPG7 gene cause chronic progressive external ophthalmoplegia through disordered mitochondrial DNA maintenance. Brain 137: 1323–1336. Pfeffer G, Pyle A, Griffin H et al. (2015). SPG7 mutations are a common cause of undiagnosed ataxia. Neurology 84: 1174–1176. Pickett SJ, Grady JP, Ng YS et al. (2018). Phenotypic heterogeneity in m.3243A>G mitochondrial disease: the role of nuclear factors. Ann Clin Transl Neurol 5: 333–345. Pickett SJ, Blain A, Ng YS et al. (2019). Mitochondrial donation—which women could benefit? N Engl J Med 380: 1971–1972. Pitceathly RDS, Smith C, Fratter C et al. (2012). Adults with RRM2B-related mitochondrial disease have distinct clinical and molecular characteristics. Brain 135: 3392–3403. Rahman S (2012). Mitochondrial disease and epilepsy. Dev Med Child Neurol 54: 397–406. Rahman S, Copeland WC (2019). POLG-related disorders and their neurological manifestations. Nat Rev Neurol 15: 40–52. Rahman S, Blok RB, Dahl HH et al. (1996). Leigh syndrome: clinical features and biochemical and DNA abnormalities. Ann Neurol 39: 343–351. Rahman S, Thorburn D (1993–2023). Nuclear gene-encoded Leigh syndrome spectrum overview. 2015 Oct 1 [Updated 2020 Jul 16]. In: MP Adam, DB Everman, GM Mirzaa (Eds.), GeneReviews® [Internet]. University of Washington, Seattle. Available from: https://www. ncbi.nlm.nih.gov/sites/books/NBK320989/. Rath S, Sharma R, Gupta R et al. (2021). MitoCarta3.0: an updated mitochondrial proteome now with sub-organelle localization and pathway annotations. Nucleic Acids Res 49: D1541–d1547. Raviglione F, Conte G, Ghezzi D et al. (2016). Clinical findings in a patient with FARS2 mutations and early-infantileencephalopathy with epilepsy. Am J Med Genet A 170: 3004–3007.

MITOCHONDRIAL ENCEPHALOMYOPATHY Ren JL, Zhang AH, Kong L et al. (2018). Advances in mass spectrometry-based metabolomics for investigation of metabolites. RSC Adv 8: 22335–22350. Riordan-Eva P, Sanders MD, Govan GG et al. (1995). The clinical features of Leber’s hereditary optic neuropathy defined by the presence of a pathogenic mitochondrial DNA mutation. Brain 118: 319–337. Rocha MC, Grady JP, Grunewald A et al. (2015). A novel immunofluorescent assay to investigate oxidative phosphorylation deficiency in mitochondrial myopathy: understanding mechanisms and improving diagnosis. Sci Rep 5: 15037. Rodrı´guez-Lo´pez C, Garcı´a-Ca´rdaba LM, Bla´zquez A et al. (2020). Clinical, pathological and genetic spectrum in 89 cases of mitochondrial progressive external ophthalmoplegia. J Med Genet 57: 643–646. Rotig A, Colonna M, Bonnefont JP et al. (1989). Mitochondrial DNA deletion in Pearson’s marrow/pancreas syndrome. Lancet 1: 902–903. Russell OM, Gorman GS, Lightowlers RN et al. (2020). Mitochondrial diseases: hope for the future. Cell 181: 168–188. Saada A, Shaag A, Mandel H et al. (2001). Mutant mitochondrial thymidine kinase in mitochondrial DNA depletion myopathy. Nat Genet 29: 342–344. Sakiyama Y, Okamoto Y, Higuchi I et al. (2011). A new phenotype of mitochondrial disease characterized by familial late-onset predominant axial myopathy and encephalopathy. Acta Neuropathol 121: 775–783. Salviati L, Trevisson E, Doimo M et al. (1993). Primary Coenzyme Q(10) Deficiency. In: MP Adam, DB Everman, GM Mirzaa et al. (Eds.), GeneReviews(®). University of Washington, Seattle, Seattle (WA). Copyright © 19932022, University of Washington, Seattle. GeneReviews is a registered trademark of the University of Washington, Seattle. All rights reserved. Schuelke M, Smeitink J, Mariman E et al. (1999). Mutant NDUFV1 subunit of mitochondrial complex I causes leukodystrophy and myoclonic epilepsy. Nat Genet 21: 260–261. Sebastia´ R, Fallico E, Fallico M et al. (2015). Bilateral lid/brow elevation procedure for severe ptosis in Kearns-Sayre syndrome, a mitochondrial cytopathy. Clin Ophthalmol 9: 25–31. Seed LM, Dean A, Krishnakumar D et al. (2022). Molecular and neurological features of MELAS syndrome in paediatric patients: a case series and review of the literature. Mol Genet. Genomic Med e1955. Serrano M, Garcia-Silva MT, Martin-Hernandez E et al. (2010). Kearns-Sayre syndrome: cerebral folate deficiency, MRI findings and new cerebrospinal fluid biochemical features. Mitochondrion 10: 429–432. Shibasaki H, Hallett M (2005). Electrophysiological studies of myoclonus. Muscle Nerve 31: 157–174. Shimura M, Kuranobu N, Ogawa-Tominaga M et al. (2020). Clinical and molecular basis of hepatocerebral mitochondrial DNA depletion syndrome in Japan: evaluation of outcomes after liver transplantation. Orphanet J Rare Dis 15: 169.

583

Shoffner JM, Lott MT, Lezza AM et al. (1990). Myoclonic epilepsy and ragged-red fiber disease (MERRF) is associated with a mitochondrial DNA tRNA(Lys) mutation. Cell 61: 931–937. Smith LA, Erskine D, Blain A et al. (2022). Delineating selective vulnerability of inhibitory interneurons in Alpers’ syndrome. Neuropathol Appl Neurobiol e12833. Sofou K, De Coo IF, Isohanni P et al. (2014). A multicenter study on Leigh syndrome: disease course and predictors of survival. Orphanet J Rare Dis 9: 52. Sofou K, Dahlin M, Hallb€ o€ ok T et al. (2017). Ketogenic diet in pyruvate dehydrogenase complex deficiency: short- and long-term outcomes. J Inherit Metab Dis 40: 237–245. Sofou K, de Coo IFM, Ostergaard E et al. (2018). Phenotypegenotype correlations in Leigh syndrome: new insights from a multicentre study of 96 patients. J Med Genet 55: 21–27. Sommerville EW, Chinnery PF, Gorman GS et al. (2014). Adult-onset Mendelian PEO associated with mitochondrial disease. J Neuromuscul Dis 1: 119–133. Sommerville EW, Ng YS, Alston CL et al. (2017). Clinical features, molecular heterogeneity, and prognostic implications in YARS2-related mitochondrial myopathy. JAMA Neurol 74: 686–694. Sondheimer N, Hewson S, Cameron JM et al. (2017). Novel recessive mutations in COQ4 cause severe infantile cardiomyopathy and encephalopathy associated with CoQ(10) deficiency. Mol Genet Metab Rep 12: 23–27. Stefanetti RJ, Blain A, Jimenez-Moreno C et al. (2020). Measuring the effects of exercise in neuromuscular disorders: a systematic review and meta-analyses. Wellcome Open Res 5: 84. Stefanetti R, Ng Y, Errington L et al. (2022). L-arginine in mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes: a systematic review. Neurology. Stendel C, Neuhofer C, Floride E et al. (2020). Delineating MT-ATP6-associated disease: from isolated neuropathy to early onset neurodegeneration. Neurol Genet 6: e393. Stenton SL, Prokisch H (2020). Genetics of mitochondrial diseases: identifying mutations to help diagnosis. EBioMedicine 56: 102784. Stewart JB, Chinnery PF (2015). The dynamics of mitochondrial DNA heteroplasmy: implications for human health and disease. Nat Rev Genet 16: 530–542. Stewart JD, Horvath R, Baruffini E et al. (2010). Polymerase gGene POLG determines the risk of sodium valproateinduced liver toxicity. Hepatology 52: 1791–1796. Suomalainen A, Elo JM, Pietil€ainen KH et al. (2011). FGF-21 as a biomarker for muscle-manifesting mitochondrial respiratory chain deficiencies: a diagnostic study. Lancet Neurol 10: 806–818. Taivassalo T, Dysgaard Jensen T, Kennaway N et al. (2003). The spectrum of exercise tolerance in mitochondrial myopathies: a study of 40 patients. Brain 126: 413–423. Tatuch Y, Christodoulou J, Feigenbaum A et al. (1992). Heteroplasmic mtDNA mutation (T——G) at 8993 can cause Leigh disease when the percentage of abnormal mtDNA is high. Am J Hum Genet 50: 852–858.

584

Y.S. NG AND R. MCFARLAND

Taylor RW, Turnbull DM (2005). Mitochondrial DNA mutations in human disease. Nat Rev Genet 6: 389–402. Taylor RW, McDonnell MT, Blakely EL et al. (2003). Genotypes from patients indicate no paternal mitochondrial DNA contribution. Ann Neurol 54: 521–524. Taylor RW, Schaefer AM, Barron MJ et al. (2004). The diagnosis of mitochondrial muscle disease. Neuromuscul Disord 14: 237–245. Taylor RW, Pyle A, Griffin H et al. (2014). Use of wholeexome sequencing to determine the genetic basis of multiple mitochondrial respiratory chain complex deficiencies. JAMA 312: 68–77. Tetsuka S, Ogawa T, Hashimoto R et al. (2021). Clinical features, pathogenesis, and management of stroke-like episodes due to MELAS. Metab Brain Dis 36: 2181–2193. Thompson K, Collier JJ, Glasgow RIC et al. (2020). Recent advances in understanding the molecular genetic basis of mitochondrial disease. J Inherit Metab Dis 43: 36–50. Thorburn DR, Rahman S (1993). Mitochondrial DNAAssociated Leigh Syndrome and NARP. In: RA Pagon, MP Adam, HH Ardinger et al. (Eds.), GeneReviews(R). University of Washington, Seattle, Seattle (WA). Ticci C, Orsucci D, Ardissone A et al. (2021). movement disorders in children with a mitochondrial disease: a cross-sectional survey from the nationwide Italian collaborative network of mitochondrial diseases. J Clin Med 10. Tiranti V, Hoertnagel K, Carrozzo R et al. (1998). Mutations of SURF-1 in Leigh disease associated with cytochrome c oxidase deficiency. Am J Hum Genet 63: 1609–1621. Trasch€utz A, Schirinzi T, Laugwitz L et al. (2020). Clinicogenetic, imaging and molecular delineation of COQ8AAtaxia: a multicenter study of 59 patients. Ann Neurol 88: 251–263. Tschampa HJ, Urbach H, Greschus S et al. (2013). Neuroimaging characteristics in mitochondrial encephalopathies associated with the m.3243A>G MTTL1 mutation. J Neurol 260: 1071–1080. Tzoulis C, Bindoff LA (2009). Serial diffusion imaging in a case of mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes. Stroke 40: e15–e17. Tzoulis C, Bindoff LA (2012). Acute mitochondrial encephalopathy reflects neuronal energy failure irrespective of which genome the genetic defect affects. Brain 135: 3627–3634. Tzoulis C, Engelsen BA, Telstad W et al. (2006). The spectrum of clinical disease caused by the A467T and W748S POLG mutations: a study of 26 cases. Brain 129: 1685–1692. Tzoulis C, Neckelmann G, Mørk SJ et al. (2010). Localized cerebral energy failure in DNA polymerase gamma-associated encephalopathy syndromes. Brain 133: 1428–1437. Tzoulis C, Tran GT, Schwarzlmuller T et al. (2013). Severe nigrostriatal degeneration without clinical parkinsonism in patients with polymerase gamma mutations. Brain 136: 2393–2404. Tzoulis C, Henriksen E, Miletic H et al. (2017). No evidence of ischemia in stroke-like lesions of mitochondrial POLG encephalopathy. Mitochondrion 32: 10–15.

van den Ouweland JM, Lemkes HH, Ruitenbeek W et al. (1992). Mutation in mitochondrial tRNA(Leu)(UUR) gene in a large pedigree with maternally transmitted type II diabetes mellitus and deafness. Nat Genet 1: 368–371. Van Goethem G, Martin JJ, Dermaut B et al. (2003). Recessive POLG mutations presenting with sensory and ataxic neuropathy in compound heterozygote patients with progressive external ophthalmoplegia. Neuromuscul Disord 13: 133–142. Visap€a€a I, Fellman V, Vesa J et al. (2002). GRACILE syndrome, a lethal metabolic disorder with iron overload, is caused by a point mutation in BCS1L. Am J Hum Genet 71: 863–876. Wallace DC, Singh G, Lott MT et al. (1988a). Mitochondrial DNA mutation associated with Leber’s hereditary optic neuropathy. Science 242: 1427–1430. Wallace DC, Zheng XX, Lott MT et al. (1988b). Familial mitochondrial encephalomyopathy (MERRF): genetic, pathophysiological, and biochemical characterization of a mitochondrial DNA disease. Cell 55: 601–610. WCMR (2022). Wellcome Centre for Mitochondrial Research, UK. https://www.newcastle-mitochondria.com/ guidelines/ (accessed on 4th Nov 2022). Wedatilake Y, Brown RM, McFarland R et al. (2013). SURF1 deficiency: a multi-centre natural history study. Orphanet J Rare Dis 8: 96. Weiner JG, Lambert AN, Thurm C et al. (2020). Heart transplantation in children with mitochondrial disease. J Pediatr 217: 46–51. Whittaker RG, Devine HE, Gorman GS et al. (2015). Epilepsy in adults with mitochondrial disease: a cohort study. Ann Neurol 78: 949–957. Wilmshurst JM, Ibekwe RC, O’Callaghan FJK (2017). Epileptic spasms—175 years on: trying to teach an old dog new tricks. Seizure 44: 81–86. Wortmann SB, Vaz FM, Gardeitchik T et al. (2012). Mutations in the phospholipid remodeling gene SERAC1 impair mitochondrial function and intracellular cholesterol trafficking and cause dystonia and deafness. Nat Genet 44: 797–802. Wright SK, Wilson MA, Walsh R et al. (2020). Abolishing spontaneous epileptiform activity in human brain tissue through AMPA receptor inhibition. Ann Clin Transl Neurol 7: 883–890. Yepez VA, Gusic M, Kopajtich R et al. (2022). Clinical implementation of RNA sequencing for Mendelian disease diagnostics. Genome Med 14: 38. Yoshimi A, Ishikawa K, Niemeyer C et al. (2022). Pearson syndrome: a multisystem mitochondrial disease with bone marrow failure. Orphanet J Rare Dis 17: 379. Yu-Wai-Man P, Chinnery PF (2013). Dominant optic atrophy: novel OPA1 mutations and revised prevalence estimates. Ophthalmology 120: 1712. Yu-Wai-Man P, Griffiths PG, Gorman GS et al. (2010). Multisystem neurological disease is common in patients with OPA1 mutations. Brain 133: 771–786.

MITOCHONDRIAL ENCEPHALOMYOPATHY Yu-Wai-Man P, Newman NJ, Carelli V et al. (2022). Natural history of patients with Leber hereditary optic neuropathyresults from the REALITY study. Eye (Lond) 36: 818–826. Zhu Z, Yao J, Johns T et al. (1998). SURF1, encoding a factor involved in the biogenesis of cytochrome c oxidase, is mutated in Leigh syndrome. Nat Genet 20: 337–343. Zia N, Nikookam Y, Muzaffar J et al. (2021). Cochlear implantation outcomes in patients with mitochondrial hearing loss: a systematic review and narrative synthesis. J Int Adv Otol 17: 72–80.

585

Zierz CM, Baty K, Blakely EL et al. (2019). A novel pathogenic variant in MT-CO2 causes an isolated mitochondrial complex iv deficiency and late-onset cerebellar ataxia. J Clin Med 8: 789. Zsurka G, Kunz WS (2015). Mitochondrial dysfunction and seizures: the neuronal energy crisis. Lancet Neurol 14: 956–966. Z€ uchner S, Mersiyanova IV, Muglia M et al. (2004). Mutations in the mitochondrial GTPase mitofusin 2 cause CharcotMarie-Tooth neuropathy type 2A. Nat Genet 36: 449–451.

This page intentionally left blank

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00004-2 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 24

Autoimmune polyneuropathies RYAN NAUM1 AND KELLY GRAHAM GWATHMEY2* 1

Department of Neurology, Virginia Commonwealth University, Richmond, VA, United States

2

Neuromuscular Division, Department of Neurology, Virginia Commonwealth University, Richmond, VA, United States

Abstract The autoimmune peripheral neuropathies with prominent motor manifestations are a diverse collection of unusual peripheral neuropathies that are appreciated in vast clinical settings. This chapter highlights the most common immune-mediated, motor predominant neuropathies excluding acute, and chronic inflammatory demyelinating polyradiculoneuropathy (AIDP and CIDP, respectively). Other acquired demyelinating neuropathies such as distal CIDP and multifocal motor neuropathy will be covered. Additionally, the radiculoplexus neuropathies, resulting from microvasculitis-induced injury to nerve roots, plexuses, and nerves, including diabetic and nondiabetic lumbosacral radiculoplexus neuropathy and neuralgic amyotrophy (i.e., Parsonage–Turner syndrome), will be included. Finally, the motor predominant peripheral neuropathies encountered in association with rheumatological disease, particularly Sj€ogren’s syndrome and rheumatoid arthritis, are covered. Early recognition of these distinct motor predominant autoimmune neuropathies and initiation of immunomodulatory and immunosuppressant treatment likely result in improved outcomes.

INTRODUCTION In this chapter, the initial section will address the acquired demyelinating neuropathies and those associated with monoclonal gammopathies, exclusive of chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), and acute inflammatory demyelinating polyradiculoneuropathy (AIDP). The topics of CIDP and AIDP are covered in Chapter 26. Following the acquired demyelinating neuropathies, “Section 2: Radiculoplexus Neuropathies” comprises the radiculoplexus neuropathies including the lumbosacral radiculoplexus neuropathies (LRPN), neuralgic amyotrophy, and the variants. Finally, in “Section 3: Motor Neuropathies in the Setting of Rheumatological Disease,” the neuropathies associated with rheumatological disease will be covered. As the emphasis of this chapter on the motor manifestations of neuropathies and considering most overlap neuropathies have a sensorypredominant presentation, the motor presentations will be primarily highlighted.

SECTION 1 ACQUIRED DEMYELINATING NEUROPATHIES Distal CIDP (distal acquired demyelinating symmetric neuropathy) Distal acquired demyelinating symmetric (DADS) is a CIDP variant characterized by symmetrical, distal lower extremity sensory or sensorimotor deficits with dramatically prolonged distal motor latencies on electrodiagnostic studies (Katz et al., 2000). In the recently updated European Academy of Neurology/Peripheral Nerve Society guideline on diagnosis and treatment of chronic inflammatory demyelinating polyradiculoneuropathy, DADS has been classified as the CIDP variant, distal CIDP (Van den Bergh et al., 2021). Approximately two-thirds of patients with this phenotype will have an IgM paraproteinemia with frequent association with myelin associated glycoprotein (MAG) antibodies (Maisonobe et al., 1996; Larue et al., 2011; Lunn and Nobile-Orazio, 2016).

*Correspondence to: Kelly Graham Gwathmey, MD, 11958 West Broad Street, Henrico, VA 23233, United States. Tel: +1-804-3604669, Fax: +1-804-3646521, E-mail: [email protected]

588

R. NAUM AND K.G. GWATHMEY

MAG is a Schwann cell–based glycoprotein that acts as a mediator of axonal cytoskeleton integrity (Albany, 2009). Monoclonal IgM antibodies target the basement membrane and myelin components, causing a widening of the myelin lamellae and demyelination, leading to a peripheral neuropathy (Ritz et al., 1999; H€anggi et al., 2021). Initially, DADS neuropathy was classified as either DADS-M (with an associated monoclonal gammopathy, typically IgM-kappa) or DADS-I (idiopathic) (Katz et al., 2000). More recently, it has been proposed to divide these disorders into DADS-I, DADS-M (with IgM paraproteinemia and no MAG antibodies) and anti-MAG neuropathy (which may or may not follow the classic clinical phenotype) (Menon et al., 2021). It has an overall prevalence of approximately 1 in 100,000 people, with a 2.7:1 male to female ratio (H€anggi et al., 2021).

CLINICAL PRESENTATION Distal CIDP is a chronic inflammatory neuropathy characterized by significant sensory loss in the distal upper and lower extremities as well as sensory ataxia, paresthesia, neuropathic pain, cramping, and tremor (Pedersen et al., 1997; Delmont et al., 2017; Fatehi et al., 2017; Rajabally et al., 2018; Svahn et al., 2018; H€anggi et al., 2021; Van den Bergh et al., 2021). When weakness occurs, it is usually distally accentuated in the lower limbs, more than upper limbs. This disorder is found more commonly in men in their 6th to 7th decades of life (Svahn et al., 2018; Katz et al., 2000). Rarely patients with anti-MAG antibodies may also present with acute or chronic sensorimotor polyradiculoneuropathies (
12%) or asymmetric/ multifocal neuropathy (
3% collectively), though these presentations are atypical (Svahn et al., 2018).

DIAGNOSTIC EVALUATION Approximately 50% of patients with DADS phenotype will have detectable anti-MAG antibody titers at diagnosis with approximately 70% of patients also having an IgM monoclonal gammopathy of undetermined significance, typically IgM-kappa subtype (Svahn et al., 2018). There is evidence to suggest that higher anti-MAG titers correlate with therapeutic response and are the characteristic anti-MAG neuropathy phenotype (Latov, 2021). Whereas, mild elevation of anti-MAG antibodies may be appreciated in those with CIDP. Nerve conduction studies will demonstrate a distal demyelinating pattern, including prolonged distal motor latency and decreased terminal latency index (distal nerve conduction distance/[proximal motor conduction velocity  distal motor latency])  0.25, a severe diminution of sensory nerve action potentials (SNAPs), and

absence of conduction block (Khadilkar et al., 2018a; Capasso et al., 2002). The 2021 European Academy of Neurology/Peripheral Nerve Society guideline on diagnosis and treatment of chronic inflammatory demyelinating polyradiculoneuropathy criteria states that distal CIDP motor conduction criteria must be fulfilled in at least two upper limb nerves to confirm the clinical diagnosis of distal CIDP and the distal negative peak CMAP amplitude should be at least 1 mV (Van den Bergh et al., 2021). If criteria are fulfilled in two lower limb nerves, but not upper limb nerves, or if criteria fulfilled in only one upper limb nerve, then maximum diagnostic certainty is possible distal CIDP. Sensory conduction abnormalities must be present in two nerves. EMG findings will typically be consistent with axonal loss or reinnervation of motor units potentials (Fatehi et al., 2017). Nerve biopsy is not required for diagnosis, but if performed, should show classic widening of myelin lamellae, as well as deposits of monoclonal IgM on the myelin sheath, which pathologically discriminates antiMAG from CIDP (Ritz et al., 1999; Fatehi et al., 2017).

TREATMENT At the time of this writing, there is no FDA approved treatments for anti-MAG neuropathy. The mainstay of therapy is directed at reducing the antibody concentration and depleting monoclonal B-cells. Intravenous immune globulins (IVIg), therapeutic plasma exchange, and various biologic drugs such as rituximab and obinutuzumab have been used in an attempt to treat anti-MAG neuropathy (H€anggi et al., 2021). High doses of rituximab have shown promise as a possible treatment by reducing B cell clones and reducing serum concentrations of IgM (Renaud et al., 2006). Two large, controlled studies have been performed. One demonstrated that 30% of rituximab-randomized patients improved by >1 point on the Inflammatory Neuropathy Cause and Treatment (INCAT) leg disability score compared with 0% placebo-treated controls and 53.8% improved walking and daily activities compared to 0% placebo-treated controls (Dalakas et al., 2009). A second, larger controlled study of 54 subjects did not demonstrate any difference on sensory scales between rituximab and placebo-treated patients (Leger et al., 2013). Some studies have shown that about 30% of patients experience stabilization of disease in 7–12 months after rituximab treatment, though the overall reported efficacy of the drug is mixed (Delmont et al., 2017; Svahn et al., 2018). It has been suggested that anti-MAG patients with proximal lower extremity weakness and subacute evolution may be most responsive to treatment with rituximab (Gazzola et al., 2017). There is evidence to suggest that a

AUTOIMMUNE POLYNEUROPATHIES sustained reduction of anti-MAG antibody titers of at least 50% compared to pretreatment titers or levels could be an indicator for therapeutic response and reduction in neuropathic symptoms (H€anggi et al., 2021). It should be noted that those with DADS-I, or distal CIDP without anti-MAG antibodies, may be treated as if with typical CIDP with considerably better response to treatments with IVIg and corticosteroids. In a small series of non-anti-MAG DADS patients with M protein, the treatment response to immunomodulatory therapies was similar to CIDP (Larue et al., 2011). For those patients with monoclonal gammopathies, continued surveillance with serum protein electrophoresis (SPEP) and serum immunofixation (SIFE) is necessary given the 1% chance per year of malignant transformation (Menon et al., 2021).

PROGNOSIS Generally, patients remain functional since motor function is relatively spared or confined to the distal extremities. Patients tend to remain ambulatory throughout most of the disease course.

Monoclonal gammopathy of undetermined significance (paraproteinemic neuropathy) Immunoglobulins, which are secreted by plasma cells, are comprised of two heavy chains and two light chains. Each plasma cell generates a specific immunoglobulin and a single type of heavy chain with a unique antigen-binding site. If a single plasma cell clone proliferates, and there is secretion of a monoclonal immunoglobulin (M protein or M spike), this is considered a monoclonal gammopathy. The primary monoclonal gammopathies that are encountered clinically are IgG, IgA, and IgM with either kappa or lambda light chains. Importantly, monoclonal gammopathies may be light chain only. Monoclonal gammopathies of undetermined significance (MGUS) are classified under the spectrum of clonal plasma cell disorders that range from a benign process to a malignant lymphoplasmacytic disorder such as multiple myeloma or Waldenstr€ om macroglobulinemia. When monoclonal gammopathies result in high levels of plasma cells, the monoclonal proteins may cause organ damage. If not, the “benign” monoclonal gammopathy is referred to as MGUS. The definition of MGUS requires serum monoclonal protein level less than 3 g/dL, less than 10% plasma cells on bone marrow biopsy, and less than 500 mg M protein in urine per 24 h. MGUS is relatively common with an incidence of 3.2% in individuals older than 50 years of age and greater than 5% after 70 years of age (Kyle et al., 2006; Hoffman et al., 2015; Hanewinckel et al., 2016). Monoclonal gammopathy of

589

undetermined significance is commonly associated with a distal and symmetric polyneuropathy of sensory predominance but may also be associated with CIDP (see Chapter 26), distal CIDP (see preceding section), and polyneuropathy, organomegaly, endocrinopathy, monoclonal plasma cell disorder, and skin changes (POEMS, not covered here) (Callaghan et al., 2015). The pathophysiology revolves around the concept of interaction of antibodies produced by clonal proliferation of plasma cells with specific antigenic targets on peripheral nerves. Antibodies are directed against peripheral nerve glycolipids and glycoproteins like myelin-associated glycoprotein (MAG), cross-reactive glycolipid sulfoglucuronyl paragloboside (SGOG), or gangliosides (GM1, GM2). IgM antibodies tend to be associated with demyelinating neuropathy, whereas IgG and IgA antibodies tend to cause axonal damage (Khadilkar et al., 2018b).

CLINICAL PRESENTATION IgM is the most common monoclonal gammopathy subtype encountered in patients with peripheral neuropathy, and IgG is the most common in the general population. IgM is also the only MGUS subtype that has been definitively associated with peripheral neuropathy without an underlying hematologic malignancy or amyloidosis (Naddaf and Mauermann, 2020). In one study, nearly 60% of MGUS-associated neuropathies were associated with an IgM monoclonal gammopathy, 30% IgG, and 10% IgA (Yeung et al., 1991). Most patients with monoclonal gammopathies do not have evidence of any overt malignancy. Only 3%–5% of patients with peripheral neuropathy have a monoclonal gammopathy and approximately 30% of patients with IgM MGUS may have a peripheral neuropathy (Nobile-Orazio et al., 1992; Chaudhry et al., 2017). Many patients may have idiopathic neuropathy, and the identification of the M protein is purely incidental (Chaudhry et al., 2017). Though monoclonal gammopathies may be associated with neuropathies in the setting CIDP, POEMS, chronic ataxic neuropathy ophthalmoplegia M protein, agglutination, disialosyl antibodies syndrome (CANOMAD), Waldenstr€om, amyloidosis, and multiple myeloma, the primary focus of this chapter is autoimmune neuropathies, therefore MGUS-peripheral neuropathy will be covered, and not malignancy-associated neuropathies. IgM MGUS-associated peripheral neuropathy patients present with a distal CIDP presentation, described in detail above. These patients present with progressive, distal lower extremity sensory loss with sensory ataxia, and occasionally tremor and distal weakness. If a patient develops a small fiber predominant peripheral neuropathy, without obvious large fiber involvement on

590

R. NAUM AND K.G. GWATHMEY

exam and without the characteristic demyelinating features on electrodiagnostic studies, the IgM MGUS may be incidental (Naddaf and Mauermann, 2020). Should a patient with small fiber-predominant peripheral neuropathy develop rapid progression or significant dysautonomia, further evaluation for amyloidosis is indicated. IgA and IgG MGUS are not clearly associated with peripheral neuropathy apart from in the context of multiple myeloma, osteosclerotic myeloma, or amyloidosis. Therefore, in patients with IgG or IgA monoclonal gammopathy and peripheral neuropathy, the association is likely coincidental though non-IgM monoclonal proteins have been reported in the full spectrum of peripheral neuropathy phenotypes including lengthdependent sensorimotor polyneuropathy and CIDP (Di Troia et al., 1999).

DIAGNOSTIC EVALUATION OF MONOCLONAL GAMMOPATHY

Evaluation for a monoclonal protein in a patient with a peripheral neuropathy should include SPEP, SIFE and quantification of free light chains with a sensitivity of 97% (Dispenzieri et al., 2009). Serum immunofixation or SPEP in MGUS patients will demonstrate elevated IgM kappa or IgG/IgA kappa/lambda levels of less than 3 g/dL, less than 10% plasma cells in bone marrow, and less than 500 mg of M protein in 24-h urine sample. Patients should lack evidence of end organ damage as expressed by the acronym “CRAB” (hypercalcemia, renal failure, anemia, or bone lesions). Complete blood counts, creatinine, calcium, and skeletal survey will usually be within normal limits. Approximately 50% of patients with IgM MGUS will be positive for antiMAG antibody (Khadilkar et al., 2018b). Once a monoclonal gammopathy has been identified, the patient should be referred to a hematologist or an internist for further investigation to determine the need for long-term monitoring as the lifelong risk of progression to lymphoplasmacytic malignancy is 1% per year (Kyle et al., 2018). The three main risk factors for progression into a lymphoplasmacytic malignancy are IgG subtype, M protein greater than or equal to 1.5 g/dL, and abnormal serum free light chain ratio (Kyle et al., 2011, 2018). The presence of any of these factors warrants further testing with bone marrow biopsy as well as skeletal survey in patients with IgG and IgA subtypes. All patients with MGUS should have repeat evaluation with CBC, SPEP, free light chains, calcium, and creatinine levels measured 6 months after diagnosis and on a yearly basis thereafter (Naddaf and Mauermann, 2020). Unfortunately, there are no specific tests that can distinguish between a causal relationship of the

monoclonal gammopathy and the peripheral neuropathy (Chaudhry et al., 2017).

DIAGNOSTIC EVALUATION OF PERIPHERAL NEUROPATHY

Please see the distal CIDP section for description of the electrodiagnostic features of DADS-M. Whereas those with DADS-M have demyelinating features, with significant involvement of the terminal nerves, those with IgG monoclonal gammopathies more commonly have axonal features, though in some classic CIDP and motor neuropathy with conduction block has been reported (Suarez and Kelly, 1993).

TREATMENT When considering treatment, it is necessary to recall that often M proteins and peripheral neuropathy association is coincidental (Chaudhry et al., 2017). In the setting of non-IgM MGUS and peripheral neuropathy, there is very scant evidence on treatment efficacy. Before embarking on treatment, the physician must be confident that the neuropathy is secondary to the peripheral neuropathy (see Fig. 24.1). In those with non-IgM M proteins, that have a peripheral neuropathy that does not resemble CIDP, then the risk of harm with therapy may outweigh any benefit (Chaudhry et al., 2017). Numerous immunosuppressive therapies have been investigated and reported in MGUS polyneuropathies. In terms of IgM monoclonal gammopathy-associated peripheral neuropathy, several studies have looked at the use of IVIg with mixed results (Leger et al., 1994; Dalakas et al., 1996; Ellie et al., 1996; Mariette et al., 1997; Giancarlo et al., 2002). In general, these studies may support modest, short-term benefit with IVIg. For those with an IgG or IgA associated neuropathy, IVIg has not been well studied. One retrospective study of 20 IgG peripheral neuropathy patients treated with IVIg, demonstrated benefit in 40% of patients, most of whom were felt to have demyelinating neuropathies (Gorson et al., 2002). Plasma exchange may have benefit in those with IgG or IgA gammopathies compared with IgM gammopathy based off of an older study (Dyck et al., 1991). There is some literature in support of rituximab in certain patients with DADS-M (see above). Neuropathic pain should also be treated with pregabalin, amitriptyline, gabapentin, or duloxetine.

PROGNOSIS Long-term follow-up is necessary as there is a possibility of malignant transformation in a portion of patients with MGUS. Approximately 1% of MGUS patients annually

AUTOIMMUNE POLYNEUROPATHIES

591

Fig. 24.1. Approach to Management of Monoclonal Gammopathy Associated Peripheral Neuropathy. *High probability of monoclonal gammopathy associated peripheral neuropathy after excluding other causes of peripheral neuropathy, POEMS syndrome, and AL amyloid neuropathy. Neuropathy considered severe and/or progressive enough to warrant intervention. †Unlikely that M protein is causally related to neuropathy; most associations are likely coincidental. DADS, distal acquired, demyelinating symmetric neuropathy; IVIG, intravenous immune globulin; CIDP, chronic inflammatory demyelinating polyneuropathy. (Reproduced with permission from Chaudhry HM, Mauermann ML, Rajkumar SV. (2017). Monoclonal gammopathy-associated peripheral neuropathy: diagnosis and management. Mayo Clin Proc 92: 838–850.)

transform to multiple myeloma or other malignant B-cell disorders (Kyle et al., 2018). Patients with no high-risk factors have a risk of progression into lymphoplasmacytic malignancy of about 7% over 20 years (Kyle et al., 2004, 2018; Go and Vincent Rajkumar, 2018). IgG and IgA MGUS can progress into multiple myeloma or osteosclerotic myeloma, and light chain MGUS can progress into light chain myeloma (Naddaf and Mauermann, 2020). All MGUS subtypes have the potential to progress into amyloidosis.

Multifocal motor neuropathy Multifocal motor neuropathy (MMN) is a chronic, progressive, autoimmune motor neuropathy which is characterized by asymmetrical weakness and electrophysiological evidence of partial motor conduction block (Yeh et al., 2020). A rare neuropathy, MMN has an estimated prevalence of 0.29–0.7 per 100,000 people (Matsui et al., 2013; Mahdi-Rogers and Hughes, 2014; Lefter et al., 2017). There are still considerable unknowns when it comes to the pathophysiology of the disease,

diagnostic criteria and optimal treatment approach (Yeh et al., 2020). Multifocal motor neuropathy is not simply a demyelinating neuropathy. There are likely multiple events driven by antibody mediated damage at the node of Ranvier and paranodal region that result in the characteristic conduction block (Franssen, 2014; Uncini and Kuwabara, 2015; Beadon et al., 2018; Fehmi et al., 2018). The disruption of saltatory conduction may stem from many different insults to the nodal and paranodal region including nodal lengthening, voltage gated sodium channel disruption, disorganized polarization of the axolemma, myelin detachment at the paranode (Franssen and Straver, 2014; Uncini and Kuwabara, 2015; Beadon et al., 2018). Complement activation may also play a role in the pathophysiology of the disease (Harschnitz et al., 2016).

CLINICAL PRESENTATION Multifocal motor neuropathy more commonly impacts males with a male-to-female ratio of 2.7:1 and average

592

R. NAUM AND K.G. GWATHMEY

age of onset 40 years old (Yeh et al., 2020). Patients experience progressive multifocal involvement of at least two nerves, in the absence of significant sensory involvement. Often MMN presents initially as distal upper limb weakness with relative sparing of flinger flexors, but one-third will first experience foot drop (Cats et al., 2010). Patients will have progression of their symptoms to affect other limbs. Cramps and fasciculations are very common (Beadon et al., 2018; Yeh et al., 2020). Though it can easily be mistaken for ALS given the weakness, atrophy, and fasciculations, MMN patients lack upper motor neuron signs, have peripheral nerve distribution weakness, rather than weakness in a myotomal pattern, and bulbar and respiratory muscles should be spared. Cranial mononeuropathies, however, are rarely reported. Cold aggravation of weakness is common and reported in 83% in one series (Straver et al., 2011). It is hypothesized that cold paresis in MMN does not reflect demyelination only, but that it may indicate the existence of inflammatory nerve lesions resulting in depolarized axons that are barely able to conduct at normal temperatures and fail at lower temperatures. Reflexes are typically diminished in the affected limbs, but may be normal or even brisk (Oshima et al., 2002). Sensory involvement should not occur but minor sensory symptoms have been reported in 20% (Nobile-Orazio and Gallia, 2013). Diagnostic criteria (Table 24.1) was published by the European Federation of Neurological Societies/Peripheral Nerve Society in 2010 (Joint Task Force of the EFNS and the PNS, 2010a).

DIAGNOSTIC EVALUATION Nerve conduction studies will show partial motor conduction block at noncompressible sites most commonly in the median and ulnar nerves (Taylor et al., 2000). Please reference Table 24.2 for the electrophysiological criteria for conduction block. Electrophysiologic studies will show other features of demyelination such as prolonged distal latencies, temporal dispersion, slow conduction velocity, and/or delayed or absent F-waves. Though partial motor conduction block is considered the electrophysiologic hallmark of MMN, when impacting the proximal sites, may be difficult to demonstrate. The Revised EFNS/PNS 2010 guidelines also provide four supportive criteria: (1) elevated IgM anti-GM1 antibodies; (2) increased cerebrospinal fluid (CSF) protein; (3) MRI demonstrating increased signal intensity on T2-weighted imaging of the brachial plexus; (4) objective clinical improvement following IVIg (Joint Task Force of the EFNS and the PNS, 2010b). Motor nerve axolemma at the node of Ranvier and Schwann cells harbor the ganglioside GM1. IgM antibodies directed toward GM1 are present in 40%–60%

Table 24.1 Clinical criteria for multifocal motor neuropathy Core criteria (both must be present)

1. Slowly progressive or stepwise progressive, focal, asymmetrica limb weakness, that is, motor involvement in the motor nerve distribution of at least two nerves, for more than 1 month.b If symptoms and signs are present only in the distribution of one nerve only a possible diagnosis can be made. 2. No objective sensory abnormalities except for minor vibration sense abnormalities in the lower limbsc

Supportive clinical criteria

1. Predominant upper limb involvementd 2. Decreased or absent tendon reflexes in the affected limbe 3. Absence of cranial nerve involvementf 4. Cramps and fasciculations in the affected limb 5. Response in terms of disability or muscle strength to immunomodulatory treatment

Exclusion criteria

1. Upper motor neuron signs 2. Marked bulbar involvement 3. Sensory impairment more marked than minor vibration loss in the lower limbs 4. Diffuse symmetric weakness during the initial weeks

(Reproduced with permission from: Joint Task Force of the EFNS and the PNS (2010). European Federation of Neurological Societies/ Peripheral Nerve Society guideline on management of multifocal motor neuropathy. Report of a joint task force of the European Federation of Neurological Societies and the Peripheral Nerve Society—first revision. J Peripher Nerv Syst 15: 295–301.) a Asymmetric ¼ a difference of 1 MRC grade if strength is MRC >3 and 2 MRC grades if strength is MRC  3. b Usually more than 6 months. c Sensory signs and symptoms may develop over the course of MMN. d At onset, predominantly lower limb involvement account for nearly 10% of the cases. e Slightly increased tendon reflexes, in particular in the affected arm, have been reported and do not exclude the diagnosis of MMN provided criterion 8 is met. f Twelfth nerve palsy has been reported.

of MMN cases and are thought to be produced by a limited number of B cell clones (Taylor et al., 2000; Cats et al., 2015; Vlam et al., 2015; Beadon et al., 2018). The sensitivity is rather low and these antibodies may be seen in healthy individuals, ALS, and other neuropathies.

AUTOIMMUNE POLYNEUROPATHIES Table 24.2 Electrophysiological criteria for conduction block 1. Definite conduction block

2. Probable motor conduction block

3. Normal sensory conductions

Negative peak CMAP area reduction on proximal vs. distal stimulation of at least 50% whatever the nerve segment length (median, ulnar, and peroneal). Negative peak CMAP amplitude on stimulation of the distal part of the segment with motor CB must be >20% of the lower limit of normal and >1 mV and increase of proximal to distal negative peak CMAP duration must be 30% Negative peak CMAP area reduction of at least 30% over a long segment (e.g., wrist to elbow or elbow to axilla) of an upper limb nerve with increase of proximal to distal negative peak CMAP duration 30% OR Negative peak CMAP area reduction of at least 50% (same as definite) with an increase of proximal to distal negative peak CMAP duration >30% In the upper limb segments with CB

(Reproduced with permission from: Joint Task Force of the EFNS and the PNS (2010). European Federation of Neurological Societies/ Peripheral Nerve Society guideline on management of multifocal motor neuropathy. Report of a joint task force of the European Federation of Neurological Societies and the Peripheral Nerve Society–first revision. J Peripher Nerv Syst 15: 295–301.) CB, conduction block; CMAP, compound muscle action potential. *Evidence for CB must be found at sites distinct from common entrapment or compression syndromes.

If elevated titers and the appropriate context, the specificity may exceed 90% (Taylor et al., 1996). Testing of GM1/ galactocerebroside (GM1/GalC) may increase the positivity rate to 70% (Nobile-Orazio et al., 2014; Delmont et al., 2015). Rarer antibodies such as GM2, GD1A, asialo-GM1 are uncommonly detected (Beadon et al., 2018). MRI imaging is not required but may show increased signal intensity on T2-weighted imaging associated with a diffuse nerve swelling of affected nerves. MRI of the cervical roots and plexus may demonstrate abnormalities in 35%–50% (Goedee et al., 2017; Jongbloed et al., 2017). Similar findings can be appreciated in other inflammatory neuropathies such as CIDP. In some cases, high-resolution ultrasound (HRUS) has been shown to be more sensitive to MRI (Jongbloed et al., 2016). Cross-sectional area values of the median nerve in the

593

forearm and the ulnar nerve distal to the sulcus and tibial nerve have been shown to be significantly enlarged in patients with MMN (Kerasnoudis et al., 2014). Nerve biopsies are not routinely done in evaluation of MMN, as cutaneous sensory nerves which are typically sampled, would be unrevealing. If a lumbar puncture is to be performed, CSF studies are expected to demonstrate a normal protein level.

TREATMENT Numerous immunomodulatory treatments have been used with poor response such as steroids and plasma exchange, which may even worsen the disease. IVIg remains the standard of care treatment. There have been five different randomized clinical trials demonstrating benefit of IVIg for MMN (Azulay et al., 1994; Van Den Berg et al., 1995; Federico et al., 2000; Leger et al., 2001; Hahn et al., 2013). The largest trial, involved 33 subjects in a randomized, crossover design, comparing IVIg and placebo (Hahn et al., 2013). The primary outcome was grip strength which demonstrated a decline of 31.38% in placebo arm and increase of 3.75% in the treatment group (P ¼ 0.005). Placebo-treated patients also demonstrated worsening disability for the upper limb on the Guys’ Neurologic Disability Score (GDNS) (P ¼ 0.021). The optimal dose and frequency of IVIg infusions remains unknown. Despite long term treatment, strength may decline as a result of ongoing axonal loss (Van Den Berg-Vos et al., 2002). The earlier the treatment, the better, though response may diminish over time (Cats et al., 2010). Continued assessment with objective markers of disease activity (e.g., grip strength) is recommended (Kumar et al., 2017). High dose IVIg (>2 g/kg/6 weeks) had been reported to be effective in MMN patients who otherwise did not respond to standard dose (Kapoor et al., 2022). Subcutaneous immune globulin treatment is likely a good alternative to IVIg and allows for uniform serum levels of IgG (Harbo et al., 2009; Al-Zuhairy et al., 2019a, b; Gentile et al., 2021). Rituximab was studied in a small (n ¼ 6) prospective study in MMN and only one patient was able to reduce IVIg dose (Chaudhry and Cornblath, 2010). Medical Research Council (MRC) sum score worsened after 12 months by average 1.17 3.37 points. Adjunctive treatment with mycophenolate mofetil (1000 mg twice daily) in a randomized controlled trial of 28 patients was well-tolerated and safe, but did not allow lowering of the IVIg dose (Piepers et al., 2007). Cyclophosphamide is occasionally needed for IVIg-refractory MMN. There have been reports of high dose IV cyclophosphamide (50 mg/kg  4 days) resulting in dramatic clinical improvement (Brannagan et al., 2006).

594

R. NAUM AND K.G. GWATHMEY

PROGNOSIS Early IVIg may delay axonal damage and resultant permanent deficits. A recent study of 100 MMN patients, 60 of which were studied over time, suggested that lower strength and absence of reflexes early in the disease course resulted in more progressive disease (Herraets et al., 2020). Despite maintenance therapy, patients gradually developed worsening of muscle strength, vibratory sensation, and deterioration of reflexes. Delay in start of therapy likely leads to poorer long-term outcome without any influence of disease duration (Al-Zuhairy et al., 2019a, b) Many patients may have moderate to severe impairment affecting the ankle dexterity and stability.

SECTION 2: RADICULOPLEXUS NEUROPATHIES Lumbosacral radiculoplexus neuropathy In 1890, a German neuropathologist, Bruns, first described diabetic lumbosacral radiculoplexus neuropathy (DLRPN). Sixty years later, Garland then coined the term “diabetic amyotrophy” though this syndrome has also been designated the eponymous “Bruns-Garland syndrome” (Llewelyn and Llewelyn, 2019). There is also a nondiabetic form (NDLRPN) and for the purposes of this article both will be considered under the term of lumbosacral radiculoplexus neuropathy (LRPN). Surprisingly common, the incidence of LRPN in Olmsted County, Minnesota was reported to be 4.16/100,000/year, a 3-fold increased incidence compared with CIDP in the same population, making it the most common inflammatory neuropathy (Pinto et al., 2021). Lumbosacral radiculoplexus neuropathy is considered a variant of nonsystemic vasculitic neuropathy with the pathophysiology thought to be the result of microvasculitis-induced ischemia of the nerves and upregulation of inflammatory mediators (Said et al., 1994; Dyck and Norell, 1999; Dyck et al., 2001; Collins and Hadden, 2017). The role diabetes has in triggering the autoimmune attack is unclear and initially LRPN was felt to occur solely in diabetics (Pinto et al., 2022). Once nondiabetic patients were described, the term nondiabetic LRPN (NDLRPN) was coined (Dyck et al., 2001). In Olmstead County, Minnesota, a significantly higher occurrence of diabetes was reported in LRPN patients compared to age and sex-matched controls (Ng et al., 2019). In this series, diabetic patients had an odds ratio (OR) of 7.91 for developing LRPN compared with nondiabetics. LRPN tends to target those with type 2 diabetes mellitus. In a recent retrospective epidemiological study of 59 LRPN patients compared with 177 age and sex-matched

controls, LRPN patients had hypertension, diabetes, obesity, dyslipidemia, stroke, autoimmune disorders, and dementia more commonly (Pinto et al., 2022). Stroke and dyslipidemia were more common in the NDLRPN patients. On multivariate logistic regression analysis, diabetes was the greatest risk factor followed by comorbid autoimmune disorders, stroke and body mass index (BMI). Patients often do not have diabetic retinopathy and nephropathy, which are more commonly associated with diabetic peripheral neuropathy (Dyck and Windebank, 2002).

CLINICAL PRESENTATION Lumbosacral radiculoplexus neuropathy typically starts unilaterally in a proximal lower extremity (less commonly distal lower extremity) with pain, followed by sensory disturbance and ultimately weakness and atrophy (Glenn and Jabari, 2020). It will spread, over time, to affect the unaffected thigh or leg as well as spread contralaterally and occasionally impacts the upper limbs and thoracic roots (Barohn et al., 1991; Said et al., 2003; Massie et al., 2012; Ng et al., 2019). Lumbosacral radiculoplexus neuropathy, as the name implies, targets the lumbosacral nerve roots, lumbosacral plexus, as well as the peripheral nerves. Characteristically patients lose weight, up to 20–30kg around the time of symptom onset (Llewelyn and Llewelyn, 2019).

CLINICAL VARIANTS Two clinical variants of LRPN have recently been described and warrant mention. Diabetic cervical radiculoplexus neuropathy shares many clinical and pathological alterations with DLRPN, suggesting these two entities could be categorized under diabetic radiculoplexus neuropathies (Massie et al., 2012). In a large series of 85 patients from the Mayo Clinic, pain was the most common initial symptom, and weakness was nearly universal (84/85) at time of presentation. The upper, middle and lower portions of the brachial plexus were equally affected with some patients having a pan-plexopathy. Over one-half of patients had an additional body region affected including contralateral cervical, lumbosacral and thoracic regions. Electrodiagnostic studies, cerebrospinal fluid protein and MRI all demonstrated similar features to DLRPN (see below). Painless diabetic motor neuropathy, has been reported as a likely variant of DLRPN (Garces-Sanchez et al., 2011). These patients develop a painless, motor predominant neuropathy over weeks to months with bilateral foot drop that progressed to proximal lower extremity muscles and less commonly upper extremities. Weight loss was similarly quite common. The impairment can

AUTOIMMUNE POLYNEUROPATHIES be severe with half patient requiring a wheelchair at presentation. Most had sensory symptoms (22/23) and many had autonomic dysfunction (9/23). In contrast to DLRPN, the distal lower extremities were more severely affected. Electrodiagnostic features resembled DLRPN and the pathological hallmarks of DLRPN were appreciated including ischemic injury, perineural thickening, injury neuroma, inflammation, neovascularization, and universally microvasculitis. The authors emphasized that the electrodiagnostic studies and pathological features differed greatly from CIDP.

DIAGNOSTIC EVALUATION It is recommended patients with suspected LRPN have complete blood counts, blood urea nitrogen, electrolytes, liver enzymes, glycosylated hemoglobin, antinuclear antibody, serum C reactive protein, erythrocyte sedimentation rate, and serum calcium (Llewelyn and Llewelyn, 2019). Cerebrospinal fluid is usually not necessary, but if obtained may show elevated proteins without cells and negative oligoclonal bands (Dyck and Windebank, 2002; Llewelyn and Llewelyn, 2019). On electrodiagnostic studies, the nerve conduction studies will demonstrate reduced motor and sensory potential amplitudes signifying axonal loss (Dyck and Norell, 1999; Dyck et al., 2001). The low amplitude sensory responses suggests a postganglionic process (plexus or individual peripheral nerves) contrasting it from a preganglionic, pure nerve root localization. On EMG, there is active denervation, and chronic neurogenic changes in muscles supplied by multiple roots and nerves. Often the electrophysiological abnormalities are more widespread than the clinical involvement. Often patients with LRPN will have a lumbosacral spine MRI as part of the diagnostic work up. Common MRI findings include increased T2-weighted signal, enhancement of the roots, plexus, and nerves, as well as T2 hyperintensity of the affected muscles though this is not appreciated in all patients (Filosto et al., 2013; Pinto et al., 2021). Root and peripheral nerve enlargement may be seen (Hlis et al., 2019), but is not specific to DLRPN and may be seen in radiculitis, and demyelinating neuropathies (Glenn and Jabari, 2020). Lumbosacral radiculoplexus neuropathy pathology demonstrates axonal degeneration, nerve ischemia, inflammation, and microvasculitic changes with upregulated inflammatory mediators (Said et al., 1994; Llewelyn et al., 1998; Dyck and Norell, 1999; Dyck et al., 2000; Kelkar et al., 2000; Kawamura et al., 2008). Nerve biopsy is usually unnecessary in straight forward cases, but may be needed if a clinical suspicion of an alternative diagnosis such as malignant infiltration exists (Glenn and Jabari, 2020).

595

TREATMENT Treatment approach of LRPN should include pain control, rehabilitation and physical therapy, and consideration of immunotherapy. As these patients may present with treatment refractory pain, standard neuropathic pain medications should be offered (tricyclic antidepressants, gabapentinoids, serotonin, and norepinephrine reuptake inhibitors (SNRIs), and may need to be escalated to include opiates. Though there are no published, randomized, placebocontrolled trials of immunomodulating and immunosuppressant treatments in LRPN, there are several intriguing case reports and case series. One placebo-controlled trial of intravenous methylprednisolone in DLRPN was published in abstract format in 2006 (Dyck et al., 2006). Seventy-five participants, mean age of 65.3 were included. Forty-nine subjects received methylprednisolone dosed 1 g, three times weekly, with decreasing dosage and frequency over 12 weeks and 26 subjects received placebo. Interval assessments occurred from baseline to week 104. Though the study’s primary endpoint of improvement in Neuropathy Impairment Score (lower limb) was not met, the treatment group had improvement in neuropathic symptoms. Other studies have described the benefit of corticosteroids in LRPN including prednisolone(Said et al., 1994; Krendel et al., 1995; Pascoe et al., 1997) and methylprednisolone (Kilfoyle et al., 2003). In a retrospective series of 62 LRPN patients, 16 received immunotherapy (IVMP 13/16, 1/16 prednisone and 2/16 IVIg) (Pinto et al., 2021). The Neuropathy Impairment Score and Modified Rankin Score were not different between those treated and those who were not. Two grams per kilogram of IVIg has been described to be beneficial in DLRPN patient who failed corticosteroid and analgesic treatment in a small study (Tamburin and Zanette, 2009). Patients reported improved pain, improved strength and walking distance. The benefit of IVIg has been described in other DLRPN studies (Krendel et al., 1995; Pascoe et al., 1997; Jaradeh et al., 1999). In NDLRPN, one open study of IV methylprednisolone in 11 patients has been published (Dyck and Norell, 2001). Intravenous methylprednisolone was dose as 1 g/week for 8–16 weeks with the median pretreatment symptom duration of 5 months. The Neuropathy Impairment Score improved in all 11 patients.

PROGNOSIS Lumbosacral radiculoplexus neuropathy is self-limiting, with slow and often incomplete recovery (Glenn and Jabari, 2020). In a large series of patients, the median time from neuropathy onset to beginning of recovery

596

R. NAUM AND K.G. GWATHMEY

was 2 months for pain and 3 months for weakness (Pinto et al., 2021). Of 59 patients, only two were symptom-free at follow up with normal exam (both with NDLRPN), 15.9% required wheelchair, 27.3% needed gait aids, and 56.8% were walking independently. Patients with LRPN had a 76% increased risk of death compared to age- and sex-matched controls (P ¼ 0.0164) (Pinto et al., 2021).

Neuralgic amyotrophy Neuralgic amyotrophy (NA, i.e., Parsonage–Turner syndrome or idiopathic brachial plexopathy) is a multifocal inflammatory motor-predominant neuropathy that targets primarily the upper extremities. Though hereditary forms of the disorder exist, this article will focus on the autoimmune presentation. Though NA was initially described in 1879 by Joffroy (1879), it was not until Parsonage and Turner provided a detailed report in 1948, that it gained its eponymous name (Parsonage and Aldren Turner, 1948). Idiopathic NA has an average age of onset around 40 years (van Alfen and van Engelen, 2006). Likely underrecognized, a recent prospective study found a one-year incidence rate of 1 per 1000 (Van Alfen et al., 2015) suggesting that it is actually quite common and as frequent as cervical radiculopathies (IJspeert et al., 2021). The male-to-female ratio is 2:1 (van Alfen and van Engelen, 2006; Collins et al., 2019). Similar to LRPN, NA, is suspected to immunemediated and the result of focal inflammatory infiltrates, release of inflammatory mediators and resultant ischemia of nerve fibers resulting in axonal loss and denervation (van Alfen, 2011). Many different factors have been reported to trigger NA including vaccinations, infections (including SARS CoV-2), immune-checkpoint inhibitors, surgery, pregnancy/childbirth, trauma (van Alfen and van Engelen, 2006; Alhammad et al., 2017; Collins et al., 2019; Porambo et al., 2019; Siepmann et al., 2020; Ismail et al., 2021). Hepatitis E virus is likely a potent trigger (Scanvion et al., 2017; Van Eijk et al., 2017; Ripellino et al., 2019). For an extensive list of antecedent infections that trigger NA, please refer to Table 24.2. Hepatitis E virus warrants particular attention as this virus may be responsible for over 10% of acute NA episodes (Van Eijk et al., 2014). HEV rarely causes hepatitis and those NA patients with or without ribavirin had no specific improvement (Van Eijk et al., 2017). Some experts view it as a self-limited variant of nonsystemic vasculitic neuropathy considering limited biopsy data supporting histopathological evidence of vasculitis (Collins et al., 2019). Additionally mechanical factors such as strenuous activity and local trauma may contribute (IJspeert et al., 2021). Mechanical strain on the brachial plexus is thought to result in disturbance of the blood nerve

barrier, resulting in susceptibility to autoimmune attack (Van Alfen et al., 2005). This is true for the brachial plexus and arm nerves that may routinely undergo large mechanical deformation (IJspeert et al., 2021). This is followed by activation of the immune system which results in an autoimmune response targeting the brachial plexus/arm nerves are affected by the leaky blood-nerve barrier (IJspeert et al., 2021).

CLINICAL PRESENTATION More than 95% of patients develop acute, severe, constant pain that will last an average of 20 days (van Alfen and van Engelen, 2006). Sixty-five percent of patients go on to develop musculoskeletal pain. Weakness then occurs on average 1–2 days later but may be delayed for up to 2–4 weeks. The disease targets motor predominant nerves and the “classic” presentation is present in 70% of patients and tends to target the suprascapular, superficial radial, anterior interosseous, and long thoracic nerves (IJspeert et al., 2021). Other less frequent presentations include lower trunk brachial plexus involvement, phrenic nerve involvement, a pure sensory form and recurrent laryngeal nerve form (IJspeert et al., 2021). Many isolated anterior interosseous and posterior interosseous nerve palsies can be considered part of the NA spectrum. Hepatitis E virus has also been described to cause severe, bilateral NA in middle-aged men with concomitant phrenic neuropathies (Scanvion et al., 2017). Even in idiopathic NA, recurrences may occur at a rate of 25% in the first 5–10 years, but are undeniably less common than in hereditary neuralgic amyotrophy (IJspeert et al., 2021). There is also a rare, new phenotype of patients presenting with chronic, progressive pain and axonal loss, which is steroid responsive (Lieba-Samal et al., 2018). Diabetes does not appear to be commonly associated with NA in contrast to the other localized vasculitic neuropathies (IJspeert et al., 2021). Pain is present in over 95% of patients (Van Eijk et al., 2016) (Table 24.3). On examination, the most commonly affected muscle groups are shoulder anteflexion, shoulder external rotation, and flexor pollicis longus (Van Alfen et al., 2015). Phrenic nerve involvement results in significant orthopnea and dyspnea on exertion. Chronically patients may have persistent upper extreme pain and weakness due to alteration of posture and scapular movements resulting in muscle strain, subpectoral impingement, and subacromial tendinopathy (Cup et al., 2013; Van Eijk et al., 2016).

DIAGNOSTIC EVALUATION Before the widespread adoption of imaging such as MRI and ultrasound to support a diagnosis of NA, the diagnosis was made based on clinical presentation (van Alfen and van Engelen, 2006; van Alfen, 2011).

AUTOIMMUNE POLYNEUROPATHIES Table 24.3 Antecedent infections associated with neuralgic amyotrophy Bacterial

Viral

Other

Escherichia Coli Staphylococcus aureus Neisseria gonorrhea Yersinia enterocolica

SARS-CoV2 Hepatitis E virus Hepatitis B virus Varicella zoster virus Herpes simplex virus Cytomegalovirus HIV

Aspergillus

Borrelia burgdorferi Streptococcus group A Mycoplasma pneumoniae Bartonella henselae Brucella Coxiella burnetti Chlamydophila pneumoniae Leptospirosis

West Nile virus Coxsackie B virus Parvovirus B19 Dengue virus

Modified from Table 1 in IJspeert J, Janssen RMJ, van Alfen N (2021). Neuralgic amyotrophy. Curr Opin Neurol 34: 605–612. https://doi.org/ 10.1097/WCO.0000000000000968. PMID: 34054111.

Electrodiagnostic testing may determine the specific nerves involved, but is limited as it may take up to 4 weeks to become abnormal, limiting its value in the acute setting (Feinberg, 2006). One should not rely on electrodiagnostic studies to confirm the diagnosis (Ijspeert et al., 2013; Van Eijk et al., 2016). Longitudinally, nearly 80% of patients will demonstrate electrodiagnostic evidence of initial recovery by 6 months with 31% demonstrating complete electrodiagnostic recovery by a mean of 1 year (Feinberg et al., 2017). Laboratory studies are generally unhelpful unless they point to a specific underlying infection (Van Eijk et al., 2016). High-resolution MRI and HRUS demonstrate both segmental swelling, torsion, and constriction of clinically affected nerves and roots in 75%–80% of NA patients (Arányi et al., 2015; Lieba-Samal et al., 2016; Sneag et al., 2018, 2020; van Rosmalen et al., 2019; IJspeert et al., 2021). These changes are often present in the distal nerves rather than the brachial plexus (Arányi et al., 2015, 2017). MRI and HRUS may be considered confirmatory testing. Affected nerves may demonstrate focal enlargement, inflammation and some have typical focal “hourglass” constrictions (Pan et al., 2014). Some authors have described a “bulls-eye sign” of the nerve identified proximal to sites of constriction and manifesting as peripheral signal hyperintensity and central hypointensity orthogonal to the long axis of the nerve (Sneag et al., 2017). For two commonly affected nerves, the anterior interosseous nerve (AIN) and posterior

597

interosseous nerve (PIN), specific constrictions have been reported. For the AIN, the constriction is found within the median nerve between 2.5 and 7 cm proximal to the medial epicondyle (Nagano et al., 1996). In the PIN, the constrictions are found between 0.2 and 5.2 cm proximal to the supinator arcade (Wu et al., 2014). With MRI findings demonstrating relatively normal brachial plexus imaging in the context of evidence of multiple mononeuropathies, NA may in fact not be a brachial plexopathy (Sneag et al., 2018).

TREATMENT Though there is no high quality evidence supporting the optimal treatment approach in NA (van Alfen et al., 2009a, b), one retrospective study suggested early oral prednisone use may shorten the duration of the pain and accelerate recovery in some (van Alfen and van Engelen, 2006). Treatment approach with corticosteroids and IVIg is primarily anecdotal (van Alfen and van Engelen, 2006; van Eijk et al., 2009, 2017; Moriguchi et al., 2011; Naito et al., 2012; Al-Ghamdi and Ghosh, 2018; Ripellino et al., 2019). Patients with NA are typically managed with pain management and physical therapy to address the weakness (Van Eijk et al., 2016). It has been proposed that some patients are nearly pain free within 24–48 h of starting treatment if they will respond. Treatment started more than 2 weeks after symptom onset is not expected to have a significant effect (IJspeert et al., 2021). Pain control can be addressed with nonsteroid antiinflammatory drugs and most patients will have a pain numerical rating score 7 (IJspeert et al., 2021). Patients with phrenic mononeuropathy may need noninvasive nighttime positive pressure ventilation and occasionally may need a diaphragmatic plication if they do not respond spontaneously (Van Alfen et al., 2018; Farr et al., 2020). Surgical intervention can be pursued in the appropriate setting in select patients (Arányi et al., 2015; ArÁnyi et al., 2017). In a recently reported large cohort of 31 patients with surgical intervention, the majority of nerves addressed with surgical treatment demonstrated excellent regeneration (80.6% M4) (Pan et al., 2014). One series of 41 patients with spontaneous PIN palsy with hourglass constrictions received conservative and surgical treatment (Wu et al., 2014). Of the 24 who were surgically treated after 3 months of conservative treatment, 20 had a good recovery (83.3%). Neurolysis was helpful in those with mild to moderate constrictions. Most with severe constrictions underwent neurorrhaphy or grafting with positive results in 12/14 (85.7%). A study of 20 patients with spontaneous nerve palsy and hourglass constrictions was published in 2019 and all 16 patients who had long-term follow-up showed good recovery (Wang et al., 2019).

598

R. NAUM AND K.G. GWATHMEY

A surgical treatment algorithm for NA has been published (Gstoettner et al., 2020). The emphasis of this algorithm is utilizing high-resolution imaging, conservative management for at least 3 months to include possible use of oral corticosteroids, and pain management. Those without obvious constrictions following 3 months warrant conservative treatment. Those with severe constrictions and rotational phenomena may need surgical intervention prior to the 3-month interval. When there is near complete paralysis after 6 months, surgical neurolysis is indicated within 6–12 months and up to 90% of patients improve (Gstoettner et al., 2020; Krishnan et al., 2021). If neurolysis isn’t an option, then nerve transfer or secondary surgery using tendon transfers are an option (Jones and Machado, 2011; Midha and Grochmal, 2019).

PROGNOSIS Most patient with NA are left with residual pain and fatigue that impacts their daily activities (van Eijk et al., 2009; van Alfen et al., 2009a, b; Cup et al., 2013). In a large study of 89 patients followed long-term, about ¼–⅓ of patients reported significant long-term fatigue and pain, and ½–⅓ reported daily impairments (van Alfen et al., 2009a, b). In another study including 49 patients followed at least 3 years, only 4% reported full recovery, 50% had chronic pain, and 85%–90% had residual weakness (van Alfen and van Engelen, 2006). Interestingly there was no correlation between pain and fatigue with residual weakness. Thus, it is important to incorporate outpatient rehabilitation treatment. It is recommended that behavioral change techniques such as motivational interviewing also be employed (Nijs et al., 2020). Unfortunately, despite this, up to 30% of NA patient will have residual motor deficits (van Alfen and van Engelen, 2006).

SECTION 3: MOTOR NEUROPATHIES IN THE SETTING OF RHEUMATOLOGICAL DISEASE In addition to the connective tissue and solid organ manifestations of rheumatologic diseases, the peripheral nervous system can also be targeted. The autoimmune neuropathies that result are characterized by their unusual presentations. These rare peripheral neuropathies are often distinguished by acute or subacute onset, non–lengthdependent symptoms, and refractoriness to treatment. Considering these overlap peripheral neuropathies tend to be heavily weighted toward peripheral sensory nerve and dorsal root ganglion involvement, and this Handbook of Clinical Neurology volume focuses specifically on motor system disorders, this chapter will emphasize the overlap neuropathies with motor manifestations. Sj€ ogren’s syndrome and rheumatoid arthritis-associated peripheral neuropathies will be highlighted, with several additional,

less common overlap neuropathies included in Table 24.4. The sensory peripheral neuropathies will be briefly mentioned for completeness-sake. Often the neurological symptoms predate the diagnosis of a rheumatological disease emphasizing the importance of testing for rheumatological disorders in patients with progressive neuropathies especially with unusual clinical features (Gwathmey and Satkowiak, 2021). It should also be highlighted that patients’ rheumatological disorders who present purely with muscle weakness are more likely to have a concomitant myopathy, than a motor neuropathy. This underscores the importance of electrodiagnostic testing, and if indicated tissue biopsy. Also one must not forget, that patients with rheumatological disorders may in fact have a peripheral neuropathy resulting from an alternative etiology (Paik et al., 2016; Gwathmey and Satkowiak, 2021). As such testing for alternative causes such as diabetes, SPEP, SIFE, B12, methylmalonic acid or homocysteine is always indicated.

Sj€ ogren’s syndrome peripheral neuropathy CLINICAL PRESENTATION Sj€ogren’s syndrome results in inflammation of the salivary and lacrimal glands causing sicca symptoms including xerophthalmia (dry eyes) and xerostomia (dry mouth). Sj€ogren’s syndrome impacts the peripheral nervous system in up to 60%, though estimates vary widely (Margaretten, 2017). Peripheral nervous system involvement was recently reported however to just be 3.7% of an Italian primary Sj€ogren’s syndrome (pSS) cohort of 1695 patients (Cafaro et al., 2021). Sj€ogren’s syndrome is classically associated with sensory polyneuropathies including most commonly small fiber neuropathy, sensory neuronopathies (dorsal root ganglionopathies), vasculitic neuropathies, and pure sensory trigeminal neuropathies. The sensorimotor and motor neuropathies are equally as diverse. Distal, symmetrical sensorimotor neuropathies may be seen, as well as demyelinating polyneuropathies, polyradiculoneuropathies, and rarely motor neuron disease (Turner et al., 2013; Tanaka et al., 2016; Tominaga et al., 2016; Zahlane et al., 2016; Gwathmey and Satkowiak, 2021). In one series of 44 pSS patients with severe neuropathies and associated weakness, only 48% were seropositive for anti-SSA antibodies (Seeliger et al., 2019). Eighty-four percent had symmetrical weakness, and importantly, most had their neuropathy symptoms precede the pSS diagnosis. Eleven percent had pure motor symptoms, and more than 50% met the EFNS/PNS criteria for CIDP. A recent series of 54 CIDP patients (fulfilling EFNS/PNS criteria) with Sj€ogren’s syndrome (fulfilling ACR/EULAR classification criteria), was published

AUTOIMMUNE POLYNEUROPATHIES

599

Table 24.4 Motor peripheral neuropathies in rheumatological disease Rheumatological disease

Peripheral neuropathy with motor involvement

Systemic lupus erythematosus

Clinical features

Treatment

Length-dependent sensorimotor polyneuropathy

● Sensory or sensorimotor

Corticosteroids, cyclophosphamide, azathioprine, IVIg, mycophenolate mofetil (Toledano et al., 2017; Bortoluzzi et al., 2019; Fargetti et al., 2019)

Chronic inflammatory demyelinating polyradiculoneuropathy

● Very rare ● May present with quadriparesis,

Vasculitic neuropathy

● Multifocal, asymmetric, mixed

polyneuropathies are the most common (Florica et al., 2011; Fargetti et al., 2019) ● In one series over ½ had asymmetrical presentation and over 1/3 had lower extremity weakness (Florica et al., 2011) ● 1/3 may have upper extremity involvement (Fargetti et al., 2019)

paraparesis, generalized areflexia or hyporeflexia, and ataxic gait (Julio et al., 2021) sensorimotor mononeuropathies (Rivière et al., 2017) ● Affects common fibular nerve ● Less commonly involves tibial nerve and upper extremities

Scleroderma

Sarcoidosis

Length-dependent sensorimotor polyneuropathy

● More typically associated with

Polyradiculoneuropathy

● Will impact thoracic and lumbar

● Painful sensorimotor

mononeuropathies ● Noncaseating granulomas on nerve and/or muscle Length-dependent sensorimotor polyneuropathy

Corticosteroids, IV cyclophosphamide, IVIg (Rivière et al., 2017; Toledano et al., 2017; Bortoluzzi et al., 2019) Unknown

small and large fiber sensory neuropathies (Paik et al., 2016) ● A large series of 9506 scleroderma patients, demonstrated 14.25% had peripheral sensorimotor polyneuropathy (Amaral et al., 2013) roots ● May be asymptomatic ● May arise from hematogenous or gravitational nerve root sleeve seeding (Koffman et al., 1999)

Vasculitic neuropathy

Corticosteroids or IVIg (Julio et al., 2021)

● Noncaseating granulomas on

nerve and/or muscle

Corticosteroids, methotrexate, azathioprine (Koffman et al., 1999; Kaiboriboon et al., 2005)

Corticosteroids (Said et al., 2002; Zivkovic and Lacomis, 2004)

Corticosteroids (Zivkovic and Lacomis, 2004)

600

R. NAUM AND K.G. GWATHMEY

(Seeliger et al., 2021). Compared to 100 patients meeting EFNS/PNS criteria for CIDP without pSS, those with both pSS and CIDP were more commonly women (52% vs. 28%) and had cranial nerve impairment (39% vs. 14%). Otherwise the two groups were similar in terms of initial symptoms, presentation, disability, CSF, laboratory and electrophysiological findings. Interestingly, there have been several case reports of a motor neuron disease-like presentation (i.e., pseudoamyotrophic lateral sclerosis) overlapping with pSS in the literature (Katz et al., 1999; Lafitte et al., 2001; Mori et al., 2005; Zahlane et al., 2016; Margaretten, 2017). These patients commonly have additional features, such as extraglandular involvement, the classic sicca symptoms, sensory involvement, cognitive impairment, and positive serologies. Considering these patients may stabilize with corticosteroids and azathioprine, this overlap syndrome should not go unrecognized.

DIAGNOSTIC EVALUATION Patients presenting with sicca syndrome and symptoms consistent with a sensory neuropathy should have an antinuclear antibody (ANA), anti-SSA antibody, and anti-SSB antibody titers drawn. These antibodies will occur in approximately 60% of patients and can serve as additional useful, but nonspecific, serologic markers to help guide diagnosis (Fox, n.d.) Negative serology does not preclude the diagnosis of pSS in a patient with sicca symptoms and neurologic involvement. Evaluation of Sj€ ogren’s syndrome should still be performed even in the absence of sicca syndrome if suspicion is high, as the neuropathy may precede sicca syndrome (Pavlakis et al., 2012; Seeliger et al., 2019). Should serologies be negative, a minor salivary gland biopsy is recommended in patients with clinical syndromes strongly suggestive of Sj€ ogren’s syndrome. The widely accepted 2016 ACREULAR Classification Criteria for Primary Sj€ ogren’s syndrome calculates the weighted sum of 5 items: antiSSA(Ro) antibodies and focal lymphocytic sialadenitis with a focus score 1 foci/mm2 (3 points each); an abnormal ocular staining score 5 (or van Bijsterveld score 4), a Schirmer test 5 mm/5 min, and an unstimulated salivary flow rate  0.1 mL/min (1 point each) (Shiboski et al., 2017). Individuals who have the clinical features of pSS, who have a total score 4 for these items, meet the criteria for pSS. Electrodiagnostic testing is performed in most patients with pSS and peripheral neuropathy. Considering the wide spectrum of peripheral nervous system involvement, it is not surprising that the electrophysiological features are equally as diverse. Most frequently electrodiagnostic testing demonstrates a symmetric axonal sensorimotor polyneuropathy.

TREATMENT Management of pSS involves treatment of the sicca symptoms via topical and systemic medications to improve glandular function and stimulate glandular secretion. There is no clear optimal management approach to the peripheral nervous system complications as no randomized controlled trials have been conducted to support the use of any specific medication. It is suggested that severe disabling neuropathies, such as vasculitic neuropathies and those resembling CIDP, as a result of pSS, should be treated with immunosuppression. Several case series have shown benefit with IVIg, steroids, rituximab, or mycophenolate mofetil in sensory or sensorimotor axonal polyneuropathies (Levy et al., 2005; Morozumi et al., 2009; Pavlakis et al., 2012; Berkowitz and Samuels, 2014; Seeliger et al., 2019; Gwathmey and Satkowiak, 2021). Demyelinating neuropathy can be treated similarly to CIDP with steroids and IVIg. Symptomatic treatments for associated neuropathic pain include topical agents such as lidocaine or capsaicin, or oral agents such as gabapentin, pregabalin, or duloxetine. Traditional therapies for peripheral sensory neuropathies, such as tricyclic agents, may not be tolerated at therapeutic doses due to their anticholinergic side effects, exacerbating sicca symptoms (Fox, n.d.; Pavlakis et al., 2012).

PROGNOSIS Response to therapy is variable. Symptoms are frequently mild but can be particularly severe or disabling (Fox, n.d.). Without formal treatment algorithms and randomized, placebo-controlled trials, most patients (apart from those with mild-to-moderate small fiber or sensorimotor polyneuropathy) are subjected to immunosuppressant and immunomodulatory treatments. As we lack long-term natural history studies in these overlap neuropathies, the prognosis for these different forms of Sj€ogren’s peripheral neuropathy remains uncertain.

Rheumatoid arthritis peripheral neuropathy CLINICAL PRESENTATION Rheumatoid arthritis (RA) is a chronic systemic inflammatory disorder primarily affecting the articular structures. The typical presentation is bilaterally symmetric polyarthritis mainly affecting the proximal interphalangeal and metacarpophalangeal joints. It can involve any small or large joint in the upper and lower extremities but spares the vertebral column, except in the cervical spine. Systemic manifestations result from release of inflammatory mediators from inflamed synovial tissues can cause constitutional symptoms, circulating immune

AUTOIMMUNE POLYNEUROPATHIES complexes can give rise to systemic vasculitis, and rheumatoid nodules and inflammation can involve extraarticular structures including nervous system structures. Peripheral neuropathy is common, with over 50% of patients having electrodiagnostic evidence, though only one-quarter had neuropathic symptoms or clinical signs of neuropathy in one series (Agarwal et al., 2008). These overlap neuropathies are often subclinical, and similar to Sj€ ogren’s, tend to be heavily weighted to sensory or sensorimotor axonopathies (Agarwal et al., 2008; Biswas et al., 2011). Perhaps most importantly, rheumatoid arthritis can result in motor predominant vasculitic neuropathy. Rheumatoid vasculitis is increasingly rare considering our increasingly effective disease modifying therapies as it has classically been considered a late presentation of severe seropositive RA (Ntatsaki et al., 2014). Peripheral neuropathy due to vasculitis is explained by immune complex-mediated damage of the vessel wall or myelinated nerves. Between 30% and 54% of patients with rheumatoid vasculitis will develop vasculitic neuropathy (Puechal et al., 1995; Makol et al., 2014). Similar to other vasculitic neuropathies these patients present with acute or subacute, multiple painful sensorimotor mononeuropathies (Burns et al., 2007). Skin changes are a frequent accompaniment (Puechal et al., 1995).

TREATMENT The treatment approach to sensory and sensorimotor polyneuropathy in RA is unknown (Gwathmey and Satkowiak, 2021). In cases of rheumatoid vasculitic neuropathy, glucocorticoids and cyclophosphamide have been used (Puechal et al., 1995; Genta et al., 2006). Rituximab has been used less commonly (DeQuattro and Imboden, 2017).

PROGNOSIS As mentioned above, the distal sensory and sensorimotor polyneuropathies are often mild or even subclinical. In contrast, the mortality of rheumatoid vasculitis is still considered relatively high at 25% over 5 years (Makol et al., 2014). There are several other rheumatological disorders associated with autoimmune peripheral neuropathies with motor involvement. Please reference Table 24.4 for additional etiologies, presentations, and treatment of these overlap neuropathies.

EVALUATION In 2010 the American College of Rheumatology/ European League Against Rheumatism classification criteria for rheumatoid arthritis was published (Aletaha et al., 2010). Patients reaching a score of 6/10 points are classified as definite RA. Point values are assigned for number of small and large joints affected, serological status (rheumatoid factor and anticitrullinated protein antibody positivity), elevation of acute-phase reactants (CRP/ESR), and duration of symptoms. Laboratory markers in those with rheumatoid vasculitis may demonstrate elevated erythrocyte sedimentation rate, eosinophilia, perinuclear antineutrophil cytoplasmic antibodies (p-ANCA), and hypocomplementemia (DeQuattro and Imboden, 2017). As mentioned above, electrodiagnostic evidence of a mild length-dependent sensory or sensorimotor polyneuropathy is commonly encountered, often in otherwise asymptomatic RA patients. In those with suspected rheumatoid vasculitis and resultant rheumatoid vasculitic neuropathy, electrodiagnostic play a particularly important role. They may demonstrate either multiple mononeuropathies, an asymmetrical mixed sensory and motor axonopathy or occasionally a length-dependent mixed sensory and motor polyneuropathy. They can serve to guide biopsy of a cutaneous nerve.

601

Biopsy of a cutaneous nerve may demonstrate inflammation and vasa nervorum damage resulting in nerve ischemia resembling polyarteritis nodosa–associated vasculitic neuropathy (DeQuattro and Imboden, 2017).

CONCLUSION The autoimmune, motor-predominant neuropathies are diverse in their clinical presentation and associated conditions. Given their immune-mediated etiologies, it is not surprising that many of these conditions present acute or subacutely, with asymmetrical, non–length-dependent presentations, that may respond to immunomodulatory and immunosuppressant treatments. Unfortunately, due to rarity of these conditions, large, randomized, placebocontrolled trials are lacking. Resultantly, clinicians are left without formal guidance. It must be emphasized, however, that early identification of the wide array of motor predominant autoimmune neuropathies and initiation of treatment aimed at stopping the autoimmune attack, likely results in improved outcomes.

REFERENCES Agarwal V et al. (2008). A clinical, electrophysiological, and pathological study of neuropathy in rheumatoid arthritis. Clin Rheumatol 27: 841–844. https://doi.org/10.1007/ s10067-007-0804-x. Albany C (2009). Anti-myelin-associated glycoprotein peripheral neuropathy as the only presentation of low grade lymphoma: a case report. Cases J 2. https://doi.org/ 10.4076/1757-1626-2-6370.

602

R. NAUM AND K.G. GWATHMEY

Aletaha D et al. (2010). 2010 rheumatoid arthritis classification criteria: an American College of Rheumatology/ European league against rheumatism collaborative initiative. Arthritis Rheum 62: 2569–2581. https://doi.org/ 10.1002/art.27584. Al-Ghamdi F, Ghosh PS (2018). Neuralgic amyotrophy in children. Muscle Nerve 57: 932–936. https://doi.org/ 10.1002/MUS.26060. Alhammad RM et al. (2017). Brachial plexus neuritis associated with anti-programmed cell death-1 antibodies: report of 2 cases. Mayo Clin Proc 1: 192–197. https://doi.org/ 10.1016/j.mayocpiqo.2017.07.004. Al-Zuhairy A et al. (2019a). A population-based and crosssectional study of the long-term prognosis in multifocal motor neuropathy. J Peripher Nerv Syst 24: 64–71. https://doi.org/10.1111/JNS.12311. Al-Zuhairy A et al. (2019b). Randomized trial of facilitated subcutaneous immunoglobulin in multifocal motor neuropathy. Eur J Neurol 26: 1289–e82. https://doi.org/10.1111/ ene.13978. Amaral TN et al. (2013). Neurologic involvement in scleroderma: a systematic review. Semin Arthritis Rheum 43: 335–347. https://doi.org/10.1016/j.semarthrit.2013.05.002. Ara´nyi Z et al. (2015). Ultrasonographic identification of nerve pathology in neuralgic amyotrophy: enlargement, constriction, fascicular entwinement, and torsion. Muscle Nerve 52: 503–511. https://doi.org/10.1002/MUS.24615. ´ nyi Z et al. (2017). Ultrasonography in neuralgic amyotroArA phy: sensitivity, spectrum of findings, and clinical correlations. Muscle Nerve 56: 1054–1062. https://doi.org/ 10.1002/MUS.25708. Azulay JP et al. (1994). Intravenous immunoglobulin treatment in patients with motor neuron syndromes associated with anti-GM1 antibodies: a double-blind, placebo-controlled study. Neurology 44: 429–432. https://doi.org/10.1212/ WNL.44.3_PART_1.429. Barohn RJ et al. (1991). The Bruns-Garland syndrome (diabetic amyotrophy). Revisited 100 years later. Arch Neurol 48: 1130–1135. https://doi.org/10.1001/ARCHNEUR.1991. 00530230038018. Beadon K, Guimara˜es-Costa R, Leger J-M (2018). Multifocal motor neuropathy. Curr Opin Neurol 31: 559–564. https:// doi.org/10.1097/WCO.0000000000000605. Berkowitz AL, Samuels MA (2014). The neurology of Sjogren’s syndrome and the rheumatology of peripheral neuropathy and myelitis. Pract Neurol 14: 14–22. https:// doi.org/10.1136/practneurol-2013-000651. Biswas M et al. (2011). Prevalence, types, clinical associations, and determinants of peripheral neuropathy in rheumatoid patients. Ann Indian Acad Neurol 14: 194–197. https://doi.org/10.4103/0972-2327.85893. Bortoluzzi A et al. (2019). Peripheral nervous system involvement in systemic lupus erythematosus: a retrospective study on prevalence, associated factors and outcome. Lupus 28: 465–474. https://doi.org/10.1177/0961203319828499. Brannagan TH, Alaedini A, Gladstone DE (2006). High-dose cyclophosphamide without stem cell rescue for refractory multifocal motor neuropathy. Muscle Nerve 34: 246–250. https://doi.org/10.1002/mus.20524.

Burns TM, Schaublin G, Dyck PJB (2007). Vasculitic neuropathies. Neurol Clin 25: 89–113. https://doi.org/10.1016/j. ncl.2006.11.002. Cafaro G et al. (2021). Peripheral nervous system involvement in Sj€ ogren’s syndrome: analysis of a cohort from the Italian research group on Sj€ ogren’s syndrome. Front Immunol 12. https://doi.org/10.3389/FIMMU.2021.615656. Callaghan BC, Price RS, Feldman EL (2015). Distal symmetric polyneuropathy: a review. JAMA 314: 2172–2181. https://doi.org/10.1001/JAMA.2015.13611. Capasso M et al. (2002). Can electrophysiology differentiate polyneuropathy with anti-MAG/SGPG antibodies from chronic inflammatory demyelinating polyneuropathy? Clin Neurophysiol 113: 346–353. https://doi.org/10.1016/ s1388-2457(02)00011-1. Cats EA et al. (2010). Multifocal motor neuropathy: association of anti-GM1 IgM antibodies with clinical features. Neurology 75: 1961–1967. https://doi.org/10.1212/WNL. 0B013E3181FF94C2. Cats EA et al. (2015). Clonality of anti-GM1 IgM antibodies in multifocal motor neuropathy and the Guillain-Barre syndrome. J Neurol Neurosurg Psychiatry 86: 502–504. https://doi.org/10.1136/JNNP-2014-308118. Chaudhry V, Cornblath DR (2010). An open-label trial of rituximab (Rituxan®) in multifocal motor neuropathy. J Peripher Nerv Syst 15: 196–201. https://doi.org/ 10.1111/J.1529-8027.2010.00270.X. Chaudhry HM, Mauermann ML, Rajkumar SV (2017). Monoclonal gammopathy-associated peripheral neuropathy: diagnosis and management. Mayo Clin Proc 92: 838–850. https://doi.org/10.1016/J.MAYOCP.2017.02.003. Collins MP, Hadden RD (2017). The nonsystemic vasculitic neuropathies. Nat Rev Neurol 13: 302–316. https://doi. org/10.1038/nrneurol.2017.42. Collins MP, Dyck PJB, Hadden RDM (2019). Update on classification, epidemiology, clinical phenotype and imaging of the nonsystemic vasculitic neuropathies. Curr Opin Neurol 32: 684–695. https://doi.org/10.1097/WCO. 0000000000000727. Cup EH et al. (2013). Residual complaints after neuralgic amyotrophy. Arch Phys Med Rehabil 94: 67–73. https:// doi.org/10.1016/J.APMR.2012.07.014. Dalakas MC et al. (1996). A controlled study of intravenous immunoglobulin in demyelinating neuropathy with IgM gammopathy. Ann Neurol 40: 792–795. https://doi.org/ 10.1002/ANA.410400516. Dalakas MC et al. (2009). Placebo-controlled trial of rituximab in IgM anti-myelin-associated glycoprotein antibody demyelinating neuropathy. Ann Neurol 65: 286–293. https://doi.org/10.1002/ana.21577. Delmont E et al. (2015). Improving the detection of IgM antibodies against glycolipids complexes of GM1 and Galactocerebroside in multifocal motor neuropathy using glycoarray and ELISA assays. J Neuroimmunol 278: 159–161. https://doi.org/10.1016/J.JNEUROIM.2014.11.001. Delmont E et al. (2017). Determinants of health-related quality of life in anti-MAG neuropathy: a cross-sectional multicentre European study. J Peripher Nerv Syst 22: 27–33. https:// doi.org/10.1111/JNS.12197.

AUTOIMMUNE POLYNEUROPATHIES DeQuattro K, Imboden JB (2017). Neurologic manifestations of rheumatoid arthritis. Rheum Dis Clin N Am 43: 561–571. https://doi.org/10.1016/j.rdc.2017.06.005. Di Troia A et al. (1999). Clinical features and anti-neural reactivity in neuropathy associated with IgG monoclonal gammopathy of undetermined significance. J Neurol Sci 164: 64–71. https://doi.org/10.1016/S0022-510X(99) 00049-0. Dispenzieri A et al. (2009). International myeloma working group guidelines for serum-free light chain analysis in multiple myeloma and related disorders. Leukemia 23: 215–224. https://doi.org/10.1038/LEU.2008.307. Dyck PJ, Norell JE (1999). Microvasculitis and ischemia in diabetic lumbosacral radiculoplexus neuropathy. Neurology 53: 2113–2121. Available at:. http://www.ncbi.nlm.nih.gov/ pubmed/10599791. (Accessed: 27 May 2013). Dyck PJ, Norell JE (2001). Methylprednisolone may improve lumbosacral radiculoplexus neuropathy. Can J Neurol Sci 28: 224–227. Available at: http://www.ncbi.nlm.nih.gov/ pubmed/11513340. (Accessed: 9 April 2013). Dyck PJB, Windebank AJ (2002). Diabetic and nondiabetic lumbosacral radiculoplexus neuropathies: new insights into pathophysiology and treatment. Muscle Nerve25: 477–491. https://doi.org/10.1002/mus.10080. Dyck PJ et al. (1991). Plasma exchange in polyneuropathy associated with monoclonal gammopathy of undetermined significance. N Engl J Med 325: 1482–1486. https://doi. org/10.1056/NEJM199111213252105. Dyck PJ, Engelstad J, Norell J (2000). Microvasculitis in nondiabetic lumbosacral radiculoplexus neuropathy (LSRPN): similarity to the diabetic variety (DLSRPN). J Neuropathol Exp Neurol 59: 525–538. Available at: http://www.ncbi. nlm.nih.gov/pubmed/10850865. (Accessed: 2 May 2013). Dyck PJ, Norell JE, Dyck PJ (2001). Non-diabetic lumbosacral radiculoplexus neuropathy: natural history, outcome and comparison with the diabetic variety. Brain 124: 1197–1207. Available at: http://www.ncbi.nlm.nih. gov/pubmed/11353735. Dyck PJB, O’Brien P, Bosch P et al. (2006). The multi-center double-blind controlled trial of IV methylprednisolone in diabetic lumbosacral radiculoplexus neuropathy [abstract]. Neurology 66: p. A191. Ellie E et al. (1996). Neuropathy associated with “benign” anti-myelin-associated glycoprotein IgM gammopathy: clinical, immunological, neurophysiological pathological findings and response to treatment in 33 cases. J Neurol 243: 34–43. https://doi.org/10.1007/BF00878529. Fargetti S et al. (2019). Short- and long-term outcome of systemic lupus erythematosus peripheral neuropathy: bimodal pattern of onset and treatment response. J Clin Rheumatol. https://doi.org/10.1097/RHU.0000000000001201. Farr E, D’Andrea D, Franz CK (2020). Phrenic nerve involvement in neuralgic amyotrophy (Parsonage-Turner syndrome). Sleep Med Clin 15: 539–543. https://doi.org/ 10.1016/j.jsmc.2020.08.002. Fatehi F et al. (2017). Motor unit number index (MUNIX) in patients with anti-MAG neuropathy. Clin Neurophysiol 128: 1264–1269. https://doi.org/10.1016/J. CLINPH.2017.04.022.

603

Federico P et al. (2000). Multifocal motor neuropathy improved by IVIg: randomized, double-blind, placebocontrolled study. Neurology 55: 1256–1262. https://doi. org/10.1212/WNL.55.9.1256. Fehmi J et al. (2018). Nodes, paranodes and neuropathies. J Neurol Neurosurg Psychiatry 89: 61–71. https://doi.org/ 10.1136/JNNP-2016-315480. Feinberg J (2006). EMG: myths and facts. HSS J 2: 19–21. https://doi.org/10.1007/S11420-005-0124-0. Feinberg JH et al. (2017). The electrodiagnostic natural history of parsonage–turner syndrome. Muscle Nerve 56: 737–743. https://doi.org/10.1002/mus.25558. Filosto M et al. (2013). MR neurography in diagnosing nondiabetic lumbosacral radiculoplexus neuropathy. J Neuroimaging 23: 543–544. https://doi.org/10.1111/ JON.12039. Florica B et al. (2011). Peripheral neuropathy in patients with systemic lupus erythematosus. Semin Arthritis Rheum 41: 203–211. https://doi.org/10.1016/j.semarthrit.2011.04.001. Fox, R. I. (n.d.) Sj€ ogren’s syndrome. Lancet, 366, 321–31. https://doi.org/10.1016/S0140-6736(05)66990-5. Franssen H (2014). The node of Ranvier in multifocal motor neuropathy. J Clin Immunol 34. https://doi.org/10.1007/ S10875-014-0023-6. Franssen H, Straver DCG (2014). Pathophysiology of immune-mediated demyelinating neuropathies—part II: neurology. Muscle Nerve 49: 4–20. https://doi.org/ 10.1002/mus.24068. Garces-Sanchez M et al. (2011). Painless diabetic motor neuropathy: a variant of diabetic lumbosacral radiculoplexus neuropathy? Ann Neurol 69: 1043–1054. https://doi.org/ 10.1002/ana.22334. Gazzola S et al. (2017). Predictive factors of efficacy of rituximab in patients with anti-MAG neuropathy. J Neurol Sci 377: 144–148. https://doi.org/10.1016/j.jns.2017.04.015. Genta MS, Genta RM, Gabay C (2006). Systemic rheumatoid vasculitis: a review. Semin Arthritis Rheum 36: 88–98. https://doi.org/10.1016/j.semarthrit.2006.04.006. Gentile L et al. (2021). Long-term treatment with subcutaneous immunoglobulin in multifocal motor neuropathy. Sci Rep 11. https://doi.org/10.1038/S41598-02188711-9. Giancarlo C et al. (2002). A randomised controlled trial of intravenous immunoglobulin in IgM paraprotein associated demyelinating neuropathy. J Neurol 249: 1370–1377. https://doi.org/10.1007/S00415-002-0808-Z. Glenn MD, Jabari D (2020). Diabetic lumbosacral Radiculoplexus neuropathy (diabetic amyotrophy). Neurol Clin 38: 553–564. https://doi.org/10.1016/J.NCL.2020. 03.010. Go RS, Vincent Rajkumar S (2018). How I manage monoclonal gammopathy of undetermined significance. Blood 131: 163–173. https://doi.org/10.1182/BLOOD-2017-09807560. Goedee HS et al. (2017). A comparative study of brachial plexus sonography and magnetic resonance imaging in chronic inflammatory demyelinating neuropathy and multifocal motor neuropathy. Eur J Neurol 24: 1307–1313. https://doi.org/10.1111/ENE.13380.

604

R. NAUM AND K.G. GWATHMEY

Gorson KC et al. (2002). Efficacy of intravenous immunoglobulin in patients with IgG monoclonal gammopathy and polyneuropathy. Arch Neurol 59: 766–772. https://doi. org/10.1001/ARCHNEUR.59.5.766. Gstoettner C et al. (2020). Neuralgic amyotrophy: a paradigm shift in diagnosis and treatment. J Neurol Neurosurg Psychiatry 91: 879–888. https://doi.org/10.1136/JNNP2020-323164. Gwathmey KG, Satkowiak K (2021). Peripheral nervous system manifestations of rheumatological diseases. J Neurol Sci 424. https://doi.org/10.1016/J.JNS.2021.117421. Hahn AF et al. (2013). A controlled trial of intravenous immunoglobulin in multifocal motor neuropathy. J Peripher Nerv Syst 18: 321–330. https://doi.org/10.1111/JNS5.12046. Hanewinckel R et al. (2016). Prevalence of polyneuropathy in the general middle-aged and elderly population. Neurology 87: 1892–1898. https://doi.org/10.1212/WNL. 0000000000003293. H€anggi P et al. (2021). Decrease in serum anti-MAG autoantibodies is associated with therapy response in patients with anti-MAG neuropathy: retrospective study. Neurol Neuroimmunol NeuroInflammation 9. https://doi. org/10.1212/NXI.0000000000001109. Harbo T et al. (2009). Subcutaneous versus intravenous immunoglobulin in multifocal motor neuropathy: a randomized, single-blinded cross-over trial. Eur J Neurol 16: 631–638. https://doi.org/10.1111/j.1468-1331.2009. 02568.x. Harschnitz O et al. (2016). Autoantibody pathogenicity in a multifocal motor neuropathy induced pluripotent stem cell-derived model. Ann Neurol 80: 71–88. https://doi. org/10.1002/ana.24680. Herraets I et al. (2020). Clinical outcomes in multifocal motor neuropathy: A combined cross-sectional and follow-up study. Neurology 95: e1979–e1987. https://doi.org/ 10.1212/WNL.0000000000010538. Hlis R et al. (2019). Quantitative assessment of diabetic amyotrophy using magnetic resonance neurography—a casecontrol analysis. Eur Radiol 29: 5910–5919. https://doi. org/10.1007/S00330-019-06162-3. Hoffman EM et al. (2015). Impairments and comorbidities of polyneuropathy revealed by population-based analyses. Neurology 84: 1644–1651. https://doi.org/10.1212/WNL. 0000000000001492. Ijspeert J et al. (2013). Efficacy of a combined physical and occupational therapy intervention in patients with subacute neuralgic amyotrophy: a pilot study. NeuroRehabilitation 33: 657–665. https://doi.org/10.3233/NRE-130993. IJspeert J, Janssen RMJ, van Alfen N (2021). Neuralgic amyotrophy. Curr Opin Neurol 34: 605–612. https://doi.org/ 10.1097/WCO.0000000000000968. Ismail II et al. (2021). Neuralgic amyotrophy associated with COVID-19 infection: a case report and review of the literature. Neurol Sci 42: 2161–2165. https://doi.org/ 10.1007/S10072-021-05197-Z. Jaradeh SS, Prieto TE, Lobeck LJ (1999). Progressive polyradiculoneuropathy in diabetes: correlation of variables and clinical outcome after immunotherapy. J Neurol Neurosurg Psychiatry 67: 607–612. https://doi. org/10.1136/JNNP.67.5.607.

Joffroy A (1879). De la nevrite parenchymateuse spontanee, generalisee ou partielle. Arch Physiol 6: 172–198. Joint Task Force of the EFNS and the PNS (2010a). European Federation of Neurological Societies/peripheral nerve society guideline on management of multifocal motor neuropathy. Report of a joint task force of the European Federation of Neurological Societies and the peripheral nerve society— first revision. J Peripher Nerv Syst 15: 295–301. https://doi. org/10.1111/j.1529-8027.2010.00290.x. Joint Task Force of the EFNS and the PNS (2010b). European Federation of Neurological Societies/peripheral nerve society guideline on management of paraproteinemic demyelinating neuropathies. Report of a joint task force of the European Federation of Neurological Societies and the peripheral nerve society. J Peripher Nerv Syst 15: 185–195. https://doi.org/ 10.1111/j.1529-8027.2010.00278.x. Jones NF, Machado GR (2011). Tendon transfers for radial, median, and ulnar nerve injuries: current surgical techniques. Clin Plast Surg 38: 621–642. https://doi.org/ 10.1016/J.CPS.2011.07.002. Jongbloed BA et al. (2016). Comparative study of peripheral nerve Mri and ultrasound in multifocal motor neuropathy and amyotrophic lateral sclerosis. Muscle Nerve 54: 1133–1135. https://doi.org/10.1002/mus.25391. Jongbloed BA et al. (2017). Brachial plexus magnetic resonance imaging differentiates between inflammatory neuropathies and does not predict disease course. Brain Behav 7: e00632. https://doi.org/10.1002/brb3.632. Julio PR et al. (2021). Chronic inflammatory demyelinating polyradiculoneuropathy associated with systemic lupus erythematosus. Semin Arthritis Rheum 51: 158–165. https://doi.org/10.1016/J.SEMARTHRIT.2020.09.018. Kaiboriboon K, Olsen TJ, Hayat GR (2005). Cauda equina and conus medullaris syndrome in sarcoidosis. Neurology 11: 179–183. https://doi.org/10.1097/01.nrl.0000159983. 19068.21. Kapoor M et al. (2022). Dramatic clinical response to ultrahigh dose IVIg in otherwise treatment resistant inflammatory neuropathies. Int J Neurosci 132: 352–361. https://doi. org/10.1080/00207454.2020.1815733. Katz JS, Houroupian D, Ross MA (1999). Multisystem neuronal involvement and sicca complex: broadening the spectrum of complications. Muscle Nerve 22: 404–407. https://doi.org/10.1002/(sici)1097-4598(199903)22:33.0.co;2-e. Katz JS et al. (2000). Distal acquired demyelinating symmetric neuropathy. Neurology 54: 615–620. Available at: http:// www.ncbi.nlm.nih.gov/pubmed/10680792. Kawamura N et al. (2008). Inflammatory mediators in diabetic and non-diabetic lumbosacral radiculoplexus neuropathy. Acta Neuropathol 115: 231–239. https://doi.org/10.1007/ S00401-007-0326-2. Kelkar P, Masood M, Parry GJ (2000). Distinctive pathologic findings in proximal diabetic neuropathy (diabetic amyotrophy). Neurology 55: 83–88. https://doi.org/10.1212/ WNL.55.1.83. Kerasnoudis A et al. (2014). Sarcoid neuropathy: correlation of nerve ultrasound, electrophysiological and clinical findings. J Neurol Sci 347: 129–136. https://doi.org/10.1016/ j.jns.2014.09.033.

AUTOIMMUNE POLYNEUROPATHIES Khadilkar SV, Yadav RS, Patel BA (2018a). Distal acquired demyelinating symmetric neuropathy. Neuromuscul Disord 515–518. https://doi.org/10.1007/978-981-105361-0_45. Khadilkar SV, Yadav RS, Patel BA (2018b). Neuromuscular disorders: a comprehensive review with illustrative cases, 1st edn. Springer. Kilfoyle D, Kelkar P, Parry GJ (2003). Pulsed methylprednisolone is a safe and effective treatment for diabetic amyotrophy. J Clin Neuromuscul Dis 4: 168–170. Available at: http:// www.ncbi.nlm.nih.gov/pubmed/19078710. (Accessed: 9 April 2013). Koffman B et al. (1999). Polyradiculopathy in sarcoidosis. Muscle Nerve 22: 608–613. https://doi.org/10.1002/ (sici)1097-4598(199905)22:53.0.co;2-l. Krendel DA, Costigan DA, Hopkins LC (1995). Successful treatment of neuropathies in patients with diabetes mellitus. Arch Neurol 52: 1053–1061. Available at: http:// www.ncbi.nlm.nih.gov/pubmed/7487556. (Accessed: 9 April 2013). Krishnan KR et al. (2021). Outcomes of microneurolysis of hourglass constrictions in chronic neuralgic amyotrophy. J Hand Surg [Am] 46: 43–53. https://doi.org/10.1016/J. JHSA.2020.07.015. Kumar A, Patwa HS, Nowak RJ (2017). Immunoglobulin therapy in the treatment of multifocal motor neuropathy. J Neurol Sci 375: 190–197. https://doi.org/10.1016/J. JNS.2017.01.061. Kyle RA et al. (2004). Incidence of multiple myeloma in Olmsted County, Minnesota: trend over 6 decades. Cancer 101: 2667–2674. https://doi.org/10.1002/CNCR.20652. Kyle RA et al. (2006). Prevalence of monoclonal gammopathy of undetermined significance. N Engl J Med 354: 1362–1369. https://doi.org/10.1056/NEJMOA054494. Kyle RA, Buadi F, Vincent Rajkumar S (2011). Management of monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM). Oncology (Williston Park) 25: Accessed: 11 June 2022. Available at https://pubmed-ncbi-nlm-nih-gov.proxy.library. vcu.edu/21888255/. Kyle RA et al. (2018). Long-term follow-up of monoclonal gammopathy of undetermined significance. N Engl J Med 378: 241–249. https://doi.org/10.1056/NEJMOA1709974. Lafitte C et al. (2001). Neurological complications of primary Sj€ogren’s syndrome. J Neurol 248: 577–584. https://doi. org/10.1007/s004150170135. Larue S et al. (2011). Non-anti-MAG DADS neuropathy as a variant of CIDP: clinical, electrophysiological, laboratory features and response to treatment in 10 cases. Eur J Neurol 18: 899–905. https://doi.org/10.1111/j.14681331.2010.03312.x. Latov N (2021). Antibody testing in neuropathy associated with anti-myelin-associated glycoprotein antibodies: where we are after 40 years. Curr Opin Neurol 34: 625–630. https:// doi.org/10.1097/WCO.0000000000000975. Lefter S, Hardiman O, Ryan AM (2017). A population-based epidemiologic study of adult neuromuscular disease in the Republic of Ireland. Neurology 88: 304–313. https://doi. org/10.1212/WNL.0000000000003504.

605

Leger JM et al. (1994). Human immunoglobulin treatment of multifocal motor neuropathy and polyneuropathy associated with monoclonal gammopathy. J Neurol Neurosurg Psychiatry 57: 46–49. https://doi.org/10.1136/JNNP.57. SUPPL.46. Leger JM et al. (2001). Intravenous immunoglobulin therapy in multifocal motor neuropathy: a double-blind, placebocontrolled study. Brain 124: 145–153. https://doi.org/ 10.1093/BRAIN/124.1.145. Leger J-M et al. (2013). Placebo-controlled trial of rituximab in IgM anti-myelin-associated glycoprotein neuropathy. Neurology 80: 2217–2225. https://doi.org/10.1212/WNL. 0b013e318296e92b. Levy Y et al. (2005). Response of vasculitic peripheral neuropathy to intravenous immunoglobulin. Ann N Y Acad Sci 1051: 779–786. https://doi.org/10.1196/annals.1361.121. Lieba-Samal D et al. (2016). Neuroimaging of classic neuralgic amyotrophy. Muscle Nerve 54: 1079–1085. https://doi. org/10.1002/MUS.25147. Lieba-Samal D et al. (2018). Extremely painful multifocal acquired predominant axonal sensorimotor neuropathy of the upper limb. J Ultrasound Med 37: 1565–1574. https:// doi.org/10.1002/JUM.14492. Llewelyn D, Llewelyn JG (2019). Diabetic amyotrophy: a painful radiculoplexus neuropathy. Pract Neurol 19: 164–167. https://doi.org/10.1136/practneurol-2018-002105. Llewelyn JG, Thomas PK, King RHM (1998). Epineurial microvasculitis in proximal diabetic neuropathy. J Neurol 245: 159–165. https://doi.org/10.1007/ S004150050197. Lunn MPT, Nobile-Orazio E (2016). Immunotherapy for IgM anti-myelin-associated glycoprotein paraprotein-associated peripheral neuropathies. Cochrane Database Syst Rev 10. https://doi.org/10.1002/14651858.CD002827.PUB4. Mahdi-Rogers M, Hughes RAC (2014). Epidemiology of chronic inflammatory neuropathies in Southeast England. Eur J Neurol 21: 28–33. https://doi.org/10.1111/ ene.12190. Maisonobe T et al. (1996). Chronic dysimmune demyelinating polyneuropathy: a clinical and electrophysiological study of 93 patients. J Neurol Neurosurg Psychiatry 61: 36–42. https://doi.org/10.1136/JNNP.61.1.36. Makol A et al. (2014). Vasculitis associated with rheumatoid arthritis: a case-control study. Rheumatology (Oxford) 53: 890–899. https://doi.org/10.1093/rheumatology/ket475. Margaretten M (2017). Neurologic manifestations of primary Sj€ ogren syndrome. In: Rheumatic disease clinics of North America, W.B. Saunders 519–529. https://doi.org/ 10.1016/j.rdc.2017.06.002. Mariette X et al. (1997). A randomised clinical trial comparing interferon-alpha and intravenous immunoglobulin in polyneuropathy associated with monoclonal IgM. The IgM-associated polyneuropathy study group. J Neurol Neurosurg Psychiatry 63: 28–34. https://doi.org/10.1136/ JNNP.63.1.28. Massie R et al. (2012). Diabetic cervical radiculoplexus neuropathy: a distinct syndrome expanding the spectrum of diabetic radiculoplexus neuropathies. Brain 135: 3074–3088. https:// doi.org/10.1093/brain/aws244.

606

R. NAUM AND K.G. GWATHMEY

Matsui N, Miyashiro A, Kaji R (2013). Update of multifocal motor neuropathy. Nippon Rinsho 71: 861–864. Available at: https://pubmed-ncbi-nlm-nih-gov.proxy.library.vcu.edu/ 23777095/. (Accessed: 11 June 2022). Menon D, Katzberg HD, Bril V (2021). Treatment approaches for atypical CIDP. Front Neurol 12: 319. https://doi.org/ 10.3389/FNEUR.2021.653734/BIBTEX. Midha R, Grochmal J (2019). Surgery for nerve injury: current and future perspectives. J Neurosurg 130: 675–685. https:// doi.org/10.3171/2018.11.JNS181520. Mori K et al. (2005). The wide spectrum of clinical manifestations in Sj€ogren’s syndrome-associated neuropathy. Brain 128: 2518–2534. https://doi.org/10.1093/brain/awh605. Moriguchi K et al. (2011). Four cases of anti-ganglioside antibody-positive neuralgic amyotrophy with good response to intravenous immunoglobulin infusion therapy. J Neuroimmunol 238: 107–109. https://doi.org/10.1016/J. JNEUROIM.2011.08.005. Morozumi S et al. (2009). Intravenous immunoglobulin treatment for painful sensory neuropathy associated with Sj€ ogren’s syndrome. J Neurol Sci 279: 57–61. https://doi. org/10.1016/j.jns.2008.12.018. Naddaf E, Mauermann ML (2020). Peripheral neuropathies associated with monoclonal gammopathies. Continuum (Minneapolis, Minn) 26: 1369–1383. https://doi.org/ 10.1212/CON.0000000000000919. Nagano A et al. (1996). Spontaneous anterior interosseous nerve palsy with hourglass-like fascicular constriction within the main trunk of the median nerve. J Hand Surg [Am] 21: 266–270. https://doi.org/10.1016/S0363-5023 (96)80114-6. Naito KS et al. (2012). Intravenous immunoglobulin (IVIg) with methylprednisolone pulse therapy for motor impairment of neuralgic amyotrophy: clinical observations in 10 cases. Intern Med 51: 1493–1500. https://doi.org/ 10.2169/INTERNALMEDICINE.51.7049. Ng PS et al. (2019). Lumbosacral radiculoplexus neuropathy: incidence and the association with diabetes mellitus. Neurology 92: E1188–E1194. https://doi.org/10.1212/ WNL.0000000000007020. Nijs J et al. (2020). Integrating motivational interviewing in pain neuroscience education for people with chronic pain: a practical guide for clinicians. Phys Ther 100: 846–859. https://doi.org/10.1093/PTJ/PZAA021. Nobile-Orazio E, Gallia F (2013). Multifocal motor neuropathy: current therapies and novel strategies. Drugs 73: 397–406. https://doi.org/10.1007/s40265-013-0029-z. Nobile-Orazio E et al. (1992). Peripheral neuropathy in monoclonal gammopathy of undetermined significance: prevalence and immunopathogenetic studies. Acta Neurol Scand 85: 383–390. https://doi.org/10.1111/J.1600-0404. 1992.TB06033.X. Nobile-Orazio E et al. (2014). Sensitivity and predictive value of anti-GM1/galactocerebroside IgM antibodies in multifocal motor neuropathy. J Neurol Neurosurg Psychiatry 85: 754–758. https://doi.org/10.1136/jnnp2013-305755. Ntatsaki E et al. (2014). Systemic rheumatoid vasculitis in the era of modern immunosuppressive therapy. Rheumatology

(Oxford) 53: 145–152. https://doi.org/10.1093/rheumatology/ket326. Oshima Y et al. (2002). Central motor conduction in patients with anti-ganglioside antibody associated neuropathy syndromes and hyperreflexia. J Neurol Neurosurg Psychiatry 73: 568–573. https://doi.org/10.1136/JNNP. 73.5.568. Paik JJ et al. (2016). Symptomatic and electrodiagnostic features of peripheral neuropathy in scleroderma. Arthritis Care Res 68: 1150–1157. https://doi.org/10.1002/ACR.22818. Pan Y et al. (2014). Hourglass-like constrictions of peripheral nerve in the upper extremity: a clinical review and pathological study. Neurosurgery 75: 10–22. https://doi.org/ 10.1227/NEU.0000000000000350. Parsonage MJ, Aldren Turner JW (1948). Neuralgic amyotrophy; the shoulder-girdle syndrome. Lancet 1: 973–978. https://doi.org/10.1016/S0140-6736(48)90611-4. Pascoe MK et al. (1997). Subacute diabetic proximal neuropathy. Mayo Clinic proceedings. Mayo Clinic 72: 1123–1132. https://doi.org/10.1016/S0025-6196(11)63674-4. Pavlakis PP et al. (2012). Peripheral neuropathies in Sj€ ogren’s syndrome: a critical update on clinical features and pathogenetic mechanisms. J Autoimmun 39: 27–33. https://doi. org/10.1016/j.jaut.2012.01.003. Pedersen SF et al. (1997). Physiological tremor analysis of patients with anti-myelin-associated glycoprotein associated neuropathy and tremor. Muscle Nerve 20: 38–44. Available at: http://www.ncbi.nlm.nih.gov/pubmed/ 8995581. Piepers S et al. (2007). Mycophenolate mofetil as adjunctive therapy for MMN patients: a randomized, controlled trial. Brain 130: 2004–2010. https://doi.org/10.1093/BRAIN/ AWM144. Pinto MV et al. (2021). Lumbosacral radiculoplexus neuropathy: neurologic outcomes and survival in a populationbased study. Neurology 96: e2098–e2108. https://doi.org/ 10.1212/WNL.0000000000011799. Pinto MV et al. (2022). Risk factors for lumbosacral radiculoplexus neuropathy. Muscle Nerve 65: 593–598. https://doi. org/10.1002/mus.27484. Porambo ME et al. (2019). Nivolumab-induced neuralgic amyotrophy with hourglass-like constriction of the anterior interosseous nerve. Muscle Nerve 59: E40–E42. https://doi. org/10.1002/MUS.26454. Puechal X et al. (1995). Peripheral neuropathy with necrotizing vasculitis in rheumatoid arthritis. A clinicopathologic and prognostic study of thirty-two patients. Arthritis Rheum 38: 1618–1629. Available at: http://www.ncbi. nlm.nih.gov/pubmed/7488283 (Accessed: 8 April 2013). Rajabally YA et al. (2018). Prevalence, correlates and impact of pain and cramps in anti-MAG neuropathy: a multicentre European study. Eur J Neurol 25: 135–141. https://doi.org/ 10.1111/ENE.13459. Renaud S et al. (2006). High-dose rituximab and anti-MAGassociated polyneuropathy. Neurology 66: 742–744. https://doi.org/10.1212/01.wnl.0000201193.00382.b3. Ripellino P et al. (2019). Neurologic complications of acute hepatitis E virus infection. Neurol Neuroimmunol NeuroInflammation 7. https://doi.org/10.1212/NXI. 0000000000000643.

AUTOIMMUNE POLYNEUROPATHIES Ritz MF et al. (1999). Anti-MAG IgM penetration into myelinated fibers correlates with the extent of myelin widening. Muscle Nerve 22: 1030–1037. https://doi.org/ 10.1002/(sici)1097-4598(199908)22:83.0.co;2-h. Rivie`re E et al. (2017). Clinicopathological features of multiple mononeuropathy associated with systemic lupus erythematosus: a multicenter study. J Neurol 264: 1218–1226. https://doi.org/10.1007/s00415-017-8519-7. Said G et al. (1994). Nerve biopsy findings in different patterns of proximal diabetic neuropathy. Ann Neurol 35: 559–569. https://doi.org/10.1002/ANA.410350509. Said G et al. (2002). Nerve granulomas and vasculitis in sarcoid peripheral neuropathy. A clinicopathological study of 11 patients. Brain 125: 264–275. https://doi.org/10.1093/ brain/awf027. Said G et al. (2003). Inflammatory vasculopathy in multifocal diabetic neuropathy. Brain 126: 376–385. https://doi.org/ 10.1093/BRAIN/AWG029. Scanvion Q et al. (2017). Neuralgic amyotrophy triggered by hepatitis E virus: a particular phenotype. J Neurol 264: 770–780. https://doi.org/10.1007/S00415-017-8433-Z. Seeliger T et al. (2019). Neuro-Sj€ogren: peripheral neuropathy with limb weakness in Sj€ogren’s syndrome. Front Immunol 10: 1600. https://doi.org/10.3389/fimmu.2019.01600. Seeliger T et al. (2021). CIDP associated with Sj€ogren’s syndrome. J Neurol 268: 2908–2912. https://doi.org/10.1007/ S00415-021-10459-Z. Shiboski CH et al. (2017). 2016 ACR-EULAR classification criteria for primary Sj€ogren’s syndrome: a consensus and data-driven methodology involving three international patient cohorts. Arthritis Rheum 69: 35. https://doi.org/ 10.1002/ART.39859. Siepmann T et al. (2020). Neuralgic amyotrophy following infection with SARS-CoV-2. Muscle Nerve 62: E68–E70. https://doi.org/10.1002/MUS.27035. Sneag DB et al. (2017). MRI Bullseye sign: an indicator of peripheral nerve constriction in parsonage-turner syndrome. Muscle Nerve 56: 99–106. https://doi.org/10.1002/ mus.25480. Sneag DB et al. (2018). Brachial plexitis or neuritis? MRI features of lesion distribution in Parsonage-Turner syndrome. Muscle Nerve 58: 359–366. https://doi.org/ 10.1002/MUS.26108. Sneag DB et al. (2020). Fascicular constrictions above elbow typify anterior interosseous nerve syndrome. Muscle Nerve 61: 301–310. https://doi.org/10.1002/MUS.26768. Straver DCG et al. (2011). Activity-dependent conduction block in multifocal motor neuropathy. Muscle Nerve 43: 31–36. https://doi.org/10.1002/MUS.21843. Suarez GA, Kelly JJ (1993). Polyneuropathy associated with monoclonal gammopathy of undetermined significance: further evidence that IgM-MGUS neuropathies are different than IgG-MGUS. Neurology 43: 1304–1308. https:// doi.org/10.1212/WNL.43.7.1304. Svahn J et al. (2018). Anti-MAG antibodies in 202 patients: clinicopathological and therapeutic features. J Neurol Neurosurg Psychiatry 89: 499–505. https://doi. org/10.1136/jnnp-2017-316715.

607

Tamburin S, Zanette G (2009). Intravenous immunoglobulin for the treatment of diabetic lumbosacral radiculoplexus neuropathy. Pain Med 10: 1476–1480. https://doi.org/ 10.1111/j.1526-4637.2009.00704.x. Tanaka K et al. (2016). Acute motor-dominant polyneuropathy as Guillain-Barre syndrome and multiple mononeuropathies in a patient with Sj€ ogren’s syndrome. Intern Med 55: 2717–2722. https://doi.org/10.2169/ INTERNALMEDICINE.55.6881. Taylor BV, Gross L, Windebank AJ (1996). The sensitivity and specificity of anti-GM1 antibody testing. Neurology 47: 951–955. https://doi.org/10.1212/WNL.47.4.951. Taylor BV et al. (2000). Natural history of 46 patients with multifocal motor neuropathy with conduction block. Muscle Nerve 23: 900–908. https://doi.org/ 10.1002/(sici)1097-4598(200006)23:63.0. co;2-y. Toledano P et al. (2017). Peripheral nervous system involvement in systemic lupus erythematosus: prevalence, clinical and immunological characteristics, treatment and outcome of a large cohort from a single centre. Autoimmun Rev 750–755. https://doi.org/10.1016/j.autrev.2017.05.011. Tominaga N et al. (2016). Autoimmune brachial amyotrophic diplegia and sensory neuronopathy with Sj€ ogren’s syndrome: a case report. J Neurol Sci 368: 1–3. https://doi. org/10.1016/J.JNS.2016.06.053. Turner MR et al. (2013). Autoimmune disease preceding amyotrophic lateral sclerosis: an epidemiologic study. Neurology 81: 1222–1225. https://doi.org/10.1212/WNL. 0B013E3182A6CC13. Uncini A, Kuwabara S (2015). Nodopathies of the peripheral nerve: an emerging concept. J Neurol Neurosurg Psychiatry 86: 1186–1195. https://doi.org/10.1136/JNNP2014-310097. van Alfen N (2011). Clinical and pathophysiological concepts of neuralgic amyotrophy. Nat Rev Neurol 7: 315–322. https://doi.org/10.1038/nrneurol.2011.62. van Alfen N, van Engelen BGM (2006). The clinical spectrum of neuralgic amyotrophy in 246 cases. Brain 129: 438–450. https://doi.org/10.1093/brain/awh722. Van Alfen N et al. (2005). Histology of hereditary neuralgic amyotrophy. J Neurol Neurosurg Psychiatry 76: 445–447. https://doi.org/10.1136/JNNP.2004.044370. van Alfen N, van Engelen BGM, Hughes RAC (2009a). Treatment for idiopathic and hereditary neuralgic amyotrophy (brachial neuritis). Cochrane Database Syst Rev (Online) 3. https://doi.org/10.1002/14651858.CD006976. pub2. p. CD006976. van Alfen N, van der Werf SP, van Engelen BG (2009b). Longterm pain, fatigue, and impairment in neuralgic amyotrophy. Arch Phys Med Rehabil 90: 435–439. https://doi. org/10.1016/J.APMR.2008.08.216. Van Alfen N et al. (2015). Incidence of neuralgic amyotrophy (Parsonage Turner syndrome) in a primary care setting–a prospective cohort study. PLoS One 10. https://doi.org/ 10.1371/JOURNAL.PONE.0128361. Van Alfen N et al. (2018). Phrenic neuropathy and diaphragm dysfunction in neuralgic amyotrophy. Neurology 91: e843–e849. https://doi.org/10.1212/WNL.0000000000006076.

608

R. NAUM AND K.G. GWATHMEY

Van Den Berg LH et al. (1995). Treatment of multifocal motor neuropathy with high dose intravenous immunoglobulins: a double blind, placebo controlled study. J Neurol Neurosurg Psychiatry 59: 248–252. https://doi. org/10.1136/JNNP.59.3.248. Van den Bergh PYK et al. (2021). European academy of neurology/peripheral nerve society guideline on diagnosis and treatment of chronic inflammatory demyelinating polyradiculoneuropathy: report of a joint task forcesecond revision. J Peripher Nerv Syst 26: 242–268. https://doi.org/10.1111/JNS.12455. Van Den Berg-Vos RM et al. (2002). Multifocal motor neuropathy: long-term clinical and electrophysiological assessment of intravenous immunoglobulin maintenance treatment. Brain 125: 1875–1886. https://doi.org/ 10.1093/BRAIN/AWF193. van Eijk JJJ et al. (2009). Evaluation of prednisolone treatment in the acute phase of neuralgic amyotrophy: an observational study. J Neurol Neurosurg Psychiatry 80: 1120–1124. https://doi.org/10.1136/jnnp.2008. 163386. Van Eijk JJJ et al. (2014). Neuralgic amyotrophy and hepatitis E virus infection. Neurology 82: 498–503. https://doi.org/ 10.1212/WNL.0000000000000112. Van Eijk JJJ, Groothuis JT, Van Alfen N (2016). Neuralgic amyotrophy: an update on diagnosis, pathophysiology, and treatment. Muscle Nerve 53: 337–350. https://doi. org/10.1002/MUS.25008. Van Eijk JJJ et al. (2017). Clinical phenotype and outcome of hepatitis E virus-associated neuralgic amyotrophy. Neurology 89: 909–917. https://doi.org/10.1212/WNL. 0000000000004297.

van Rosmalen M et al. (2019). Ultrasound of peripheral nerves in neuralgic amyotrophy. Muscle Nerve 59: 55–59. https:// doi.org/10.1002/MUS.26322. Vlam L et al. (2015). Association of IgM monoclonal gammopathy with progressive muscular atrophy and multifocal motor neuropathy: a case-control study. J Neurol 262: 666–673. https://doi.org/10.1007/S00415-014-7612-4. Wang Y et al. (2019). Spontaneous peripheral nerve palsy with hourglass-like fascicular constriction in the upper extremity. J Neurosurg 131: 1876–1886. https://doi.org/ 10.3171/2018.8.JNS18419. Wu P et al. (2014). Surgical and conservative treatments of complete spontaneous posterior interosseous nerve palsy with hourglass-like fascicular constrictions: a retrospective study of 41 cases. Neurosurgery 75: 250–257. https://doi. org/10.1227/NEU.0000000000000424. Yeh WZ et al. (2020). Multifocal motor neuropathy: controversies and priorities. J Neurol Neurosurg Psychiatry 91: 140–148. https://doi.org/10.1136/jnnp-2019-321532. Yeung KB et al. (1991). The clinical spectrum of peripheral neuropathies associated with benign monoclonal IgM, IgG and IgA paraproteinaemia. Comparative clinical, immunological and nerve biopsy findings. J Neurol 238: 383–391. Available at: http://www.ncbi.nlm.nih.gov/ pubmed/1660064. (Accessed: 20 July 2013). Zahlane S et al. (2016). Anterior horn syndrome: a rare manifestation of primary Sj€ ogren’s syndrome. Joint Bone Spine 83: 448–450. https://doi.org/10.1016/j.jbspin.2016. 02.018. Zivkovic SA, Lacomis D (2004). Sarcoid neuropathy: case report and review of the literature. J Clin Neuromuscul Dis 5: 184–189. https://doi.org/10.1097/01. cnd.0000126502.72766.0e.

Handbook of Clinical Neurology, Vol. 195 (3rd series) Motor System Disorders, Part I: Normal Physiology and Function and Neuromuscular Disorders D.S. Younger, Editor https://doi.org/10.1016/B978-0-323-98818-6.00009-1 Copyright © 2023 Elsevier B.V. All rights reserved

Chapter 25

Hereditary neuropathy CHIARA PISCIOTTA1* AND MICHAEL E. SHY2 1

Department of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy 2

Department of Neurology, University of Iowa Hospitals and Clinics, Iowa City, IA, United States

Abstract The hereditary neuropathies, collectively referred as Charcot–Marie–Tooth disease (CMT) and related disorders, are heterogeneous genetic peripheral nerve disorders that collectively comprise the commonest inherited neurological disease with an estimated prevalence of 1:2500 individuals. The field of hereditary neuropathies has made significant progress in recent years with respect to both gene discovery and treatment as a result of next-generation sequencing (NGS) approach. These investigations which have identified over 100 causative genes and new mutations have made the classification of CMT even more challenging. Despite so many different mutated genes, the majority of CMT forms share a similar clinical phenotype, and due to this phenotypic homogeneity, genetic testing in CMT is increasingly being performed through the use of NGS panels. The majority of patients still have a mutation in one the four most common genes (PMP22 duplication—CMT1A, MPZ—CMT1B, GJB1—CMTX1, and MFN2— CMT2A). This chapter focuses primarily on these four forms and their potential therapeutic approaches.

INTRODUCTION The hereditary neuropathies, more commonly referred to as Charcot–Marie–Tooth disease (CMT) and related disorders, are heterogeneous genetic peripheral nerve disorders that collectively comprises the commonest inherited neurological disorders with an estimated prevalence of 1:2500 individuals. Historically, the classification of CMT has been based on the mode of inheritance and the primary pathology observed in nerve, as reflected in nerve conduction studies (NCS). CMT1 for demyelinating forms (with slowed nerve conduction velocities, NCV) and autosomal dominant (AD) inheritance; CMT4 for autosomal recessive (AR) demyelinating types; and CMT2 for primary axonal neuropathies and AD or AR transmission (AR-CMT2). There are intermediate forms of CMT with nerve conduction velocities (NCVs) falling in between CMT1 and CMT2. Pure motor forms of CMT are labeled distal Hereditary Motor Neuropathies (dHMN), and pure or predominantly sensory neuropathies are grouped under

the term of Hereditary Sensory and Autonomic Neuropathies (HSAN) (Pisciotta and Shy, 2018). The field of hereditary neuropathies has made significant progress in recent years with respect to both gene discovery and treatment with next-generation sequencing (NGS), which is becoming more accessible, and detecting over 100 causative genes so far and new mutations. This makes the classification of CMT more challenging which is gradually evolving to incorporate the specific gene that is mutated. Despite so many different mutated genes, the majority of CMT forms share a similar clinical phenotype and a precise diagnosis at the bedside can be difficult. Common clinical manifestations are foot deformities, including pes cavus, pes planus, or hammertoes, length-dependent muscle weakness and atrophy, and length-dependent sensory loss. Deep tendon reflexes are typically diminished or absent; however, they can also be preserved or rarely even brisk in select forms of CMT when the causal gene is also

*Correspondence to: Chiara Pisciotta, MD, PhD, Department of Clinical Neurosciences, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy. Tel: +39-02-23943001, Fax: +39-02-23942293, E-mail: [email protected]

610

C. PISCIOTTA AND M.E. SHY

expressed in CNS neurons or glia (Pisciotta and Shy, 2018). Due to this phenotypic homogeneity, genetic testing in CMT is increasingly being performed through the use of NGS panels containing multiple disease relevant genes, rather than through sequential gene testing. However, despite the expediency and reliability of genetic testing, electrodiagnostic studies remain a valuable resource for clinical trials and practice, especially when valid functional parameters are needed to measure its progression. However, it should be kept in mind that about 90% of patients that achieve a definite genetic diagnosis still have a mutation in one the four most common genes known to underlie CMT, namely, PMP22 (peripheral myelin protein 22) (duplication—CMT1A), MPZ (myelin protein zero) (CMT1B), GJB1 (gap junction protein beta 1) (CMTX1), and MFN2 (mitofusin 2) (CMT2A) (Saporta et al., 2011; Murphy et al., 2012; Fridman et al., 2015). This chapter focuses primarily focus on these four forms. In addition, we report on the recent discovery of recessive mutations in sorbitol dehydrogenase gene (SORD) emerging as one of the most common and potentially treatable causes of axonal CMT (CMT2) (Cortese et al., 2020). Moreover, looking at other genetic neuromuscular disorders as familial transthyretin amyloidosis (hATTR) with polyneuropathy and spinal muscular atrophy (SMA), where rapid successful therapeutic developments and achievements occurred in recent years, it is likely that we are facing remarkable advances in CMT treatment in the near future. Ideally, the aim is to treat the disease before it causes significant axonal degeneration and fiber loss, which are difficult to revert. As the vast majority of CMT types have childhood onset, the ultimate aim is to treat children as soon as they are diagnosed with the disease. Thus far, most clinical studies targeted adult patients for practical reasons and regulatory aspects, but future studies should be addressed mainly to children. We will discuss the potential therapeutic option for the most common forms of CMT and the current management of the disease.

CMT1A CMT1A is the most common type of CMT accounting for almost 80% of cases of demyelinating CMT. It results from a duplication of the short arm of chromosome 17p containing the PMP22 gene. A deletion of the same region of chromosome 17p leads to hereditary neuropathy with liability to pressure palsies (HNPP) (Pisciotta and Shy, 2018). PMP22 encodes peripheral myelin protein 22, a 22 kDa hydrophobic transmembrane glycoprotein that accounts for approximately 2%–5% of peripheral nerve myelin protein. PMP22 is expressed in myelinating Schwann cells and is critical for both the synthesis and

maintenance of myelin, though the exact mechanisms by which the protein functions remain not well known (Garbay et al., 2000). CMT1A has a de novo mutation rate of 10%, and patients therefore may report the absence of relevant family history (Blair et al., 1996). Searching for the PMP22 duplication is typically the first genetic test performed in a patient with uniform demyelinating CMT. Patients affected by CMT1A usually have the “classic CMT phenotype” with disease onset in the first two decades of life. They complain of walking difficulties, distal weakness associated with wasting, sensory loss and foot deformities. Patients progress very slowly and they often require ankle-foot-orthotics (AFOs) (Kramarz and Rossor, 2022). The involvement of small unmyelinated fibers, confirmed by several studies, explain the high prevalence of neuropathic pain observed in some patients (Nolano et al., 2015; Peretti et al., 2022). Inter- and intrafamilial variability is frequently observed likely related to the influence of genetic modifiers, as well as other epigenetic or environmental factors (Tao et al., 2019). The largest sample of electrophysiological data obtained so far from CMT1A patients confirmed the uniform slowing of NCV in the demyelinating range and the absence of acquired demyelinating features (i.e., partial conduction block) and that secondary axonal degeneration reflected clinical impairment. The authors concluded that CMT1A was mainly a dysmyelinating rather than a demyelinating neuropathy because of the uniform reduction of NCV present from the first years of life (Manganelli et al., 2016). Moreover, dermal myelinated nerve fibers in CMT1A have also revealed uniformly shortened internodes, potentially related to a developmental defect (Manganelli et al., 2015). The possibility of a primary developmental defect in myelin formation in CMT1A strengthens the need to treat the disease in the early phase. Reducing PMP22 gene expression would be an obvious therapeutic strategy for CMT1A. Recent studies have demonstrated the ability to measure elevated PMP22 mRNA (messenger RNA) from skin biopsies in both patients and mouse models suggesting that this technology may be useful in demonstrating target identification in clinical trials (Nobbio et al., 2014; Svaren et al., 2019). One of the first compounds to be tested in humans was the ascorbic acid, which was hypothesized to reduce PMP22 expression through a cAMP-mediated mechanism, but the studies failed to demonstrate efficacy (Pareyson et al., 2011). However, these trials demonstrated the need to develop additional clinical outcome assessments and biomarkers, as more sensitive methods to detect disease progression. A number of these are in place in adults such as the CMT Functional Outcome Measure (CMT-FOM) (Eichinger et al., 2018), the patient reported CMT-Health Index (CMT-HI) (Johnson et al., 2018), magnetic resonance imaging (MRI) of

HEREDITARY NEUROPATHY intramuscular fat accumulation in the legs (Morrow et al., 2018; Bas et al., 2020) and plasma biomarkers including neurofilament light (NfL) (Sandelius et al., 2018; Millere et al., 2021; Rossor et al., 2022) and Transmembrane Serine Protease 5 (TMPRSS5) (Wang et al., 2020). Outcome assessment in children now includes the CMT Pediatric Scale (CMTPedS) (Burns et al., 2012) and the CMT Infant and Toddler Scale (CMTInfS) (Mandarakas et al., 2018), and the patient reported CMT pediatric quality of life scale (pCMT-QOL) (Ramchandren et al., 2021). A recent study in CMT1A utilizes PXT3003, which combines three drugs at low doses: baclofen, sorbitol, and naltrexone; it is predicted to lower PMP22 levels and its efficacy in a preclinical and phase II study in CMT1A lead to a double-blind phase III study (Prukop et al., 2020; Attarian et al., 2021). Unfortunately, the phase III was interrupted by the unblinding of the high-dose group because of lack of stability of the formulation (Attarian et al., 2021); a novel international phase III trial required by the FDA has started. The recent success of genetic therapies, including antisense oligonucleotides (ASOs) and small-interfering RNA (siRNA)-based treatments, in other neuromuscular disorders has greatly aroused enthusiasm in the CMT field, promoting the hope for a disease modifying treatment for CMT, particularly since CMT1A is a gene dosage disorder that would seem suitable for a genetic approach aiming to reduce total PMP22 transcript levels. These studies have demonstrated efficacy in rodent models of CMT1A, though there are challenges that need to be addressed for human trials. There are potential concerns about lowering PMP22 levels to much such that HNPP might be induced. Second there are challenges to reaching sufficient number of Schwann cells with these agents which would need to broach the blood nerve barrier and reach the endoneurium to reach Schwann cells. Intrathecal studies in rodents have been encouraging in this respect. Approaches in rodents to date have included: (a) subcutaneous ASOs (Zhao et al., 2018); (b) subcutaneous siRNA conjugated to squalene nanoparticles (Boutary et al., 2021); (c) intranerve injection of small hairpin RNA (shRNAs) with an adeno-associated viral serotype 9 (AAV2/9) vector (Gautier et al., 2021); and (d) injection into the sciatic nerve of liposomeencapsulated single guide RNA (sgRNA) aimed at deleting the PMP22 TATA-Box by CRISPR-Cas9 editing (Lee et al., 2020). Normalization of PMP22 levels and improvement in locomotor, electrophysiological, and pathological parameters in mice have been obtained with these approaches. An additional study provided preclinical data demonstrating efficacy of adeno-associated virus (AAV)-mediated neurotrophin 3 (NT-3) (AAV1-NT-3) gene therapy in a mouse model of CMT1A (Sahenk et al., 2005). Previously, a pilot study in eight patients

611

with CMT1A undergoing subcutaneous injection of NT-3 resulted in an increase in myelinated fiber density at nerve biopsy, a reduction in the neurologic impairment score, and improved sensory modalities as compared with controls (Sahenk et al., 2005). Therefore, an openlabel study has been designed to deliver ascending doses of the AAV1-NT-3 gene intramuscularly in both legs (in medial and lateral heads of triceps and in tibialis anterior muscles) in 3 young CMT1A subjects (ClinicalTrials. gov NCT03520751).

CMT1B CMT1B, caused by mutations in the MPZ gene, accounts for 5% of all of CMT and 10% of all demyelinating forms of CMT. MPZ, also termed P0, is the major protein in peripheral nerve myelin and a member of the immunoglobulin (Ig) supergene family. The protein plays an important role both in the formation of myelin and in the maintenance of myelin homeostasis and stability. MPZ localizes to compact myelin, where it is responsible for maintaining both the major dense line and intraperiod line (Shy et al., 2004). MPZ-related neuropathies are genetically heterogeneous, with over 200 different disease-causing mutations identified to date. MPZ phenotypes are being classified by the patient’s age at onset, the primary nerve pathology, and the specific genetic mutation. Genotype– phenotype correlation studies have identified three distinct types of MPZ-neuropathy, including an infantile form (clinical presentation prior to 3 years of age with severely slowed motor conduction velocities (NCV; 9.5 ms, fibular >8.8 ms, tibial >9.2 ms (for LFF of 2 Hz; normals for higher LFF available in (Van den Bergh et al., 2021) Abnormal temporal dispersion: increase in CMAP duration of >30% from distal to proximal stimulation at noncompressible sites. The tibial nerve is usually excluded due to a high rate of technical factors at the proximal stimulation site, or required to be at least 100% increase in duration. Conduction block: a drop in CMAP amplitude from distal to proximal stimulation with a corresponding reduction in CMAP area at noncompressible sites. For median and ulnar nerves, CMAP amplitude and area drop by 50%. For all other nerves, CMAP amplitude and area drop by 30%. Distal CMAP must be 20% LLN. The tibial nerve is excluded due to a high rate of technical factors at the proximal stimulation site.

Cross-sectional area median nerve >10 mm2 at forearm, >13 mm2 upper arm Cross-sectional area > 9 mm2 interscalene (trunks) Cross-sectional area > 12 mm2 for nerve roots

Adapted from Van den Bergh P, Pieter A, Hadden R et al. (2021). European academy of neurology/peripheral nerve society guideline on diagnosis and treatment of chronic inflammatory demyelinating polyradiculoneuropathy: report of a joint task force-second revision. Eur J Neurol 28: 3556–3583. Abbreviations: CMAP, compound muscle action potential; Hz, hertz; LFF, low frequency filter; LLN, lower limit of normal; ms, milliseconds; ULN, upper limit of normal.

Table 26.6 Electrodiagnostic criteria for CIDP. Typical, distal and multifocal CIDP At least two motor nerves must have demyelinating range abnormalities that meet motor nerve conduction criteria (see above) Sensory abnormalities in at least two nerves (prolonged distal latency, reduced SNAP amplitude, slow conduction velocity) Motor CIDP Same as for typical CIDP minus the sensory requirements Sensory CIDP Sensory nerve CV 80% LLN or < 70% LLN for SNAP 100 mg/dL) (45%) and normal glucose overall in 77% (103/135). There

662

D.S. YOUNGER

were 51 cases of GANS in association with other conditions, including 11 cases of TA or generalized GCA; 12 cases of HZV, 9 with lymphoproliferative tumors (7 alone, and 2 with HZV), 6 cases of sarcoidosis, 10 cases of amyloid angiopathy, 1 case of SLE with HZV, and 2 cases of GANS with human immunodeficiency virus (HIV) without acquired immunodeficiency syndrome (AIDS). The neurologic disorder associated with temporal or GCA differed in the predilection for large intracranial vessels, and the relentless progression despite corticosteroids. The neurologic presentation of contralateral hemiparesis associated with HZV typically followed appearance of an ophthalmicus (V1) rash by 2–3 weeks with skin lesions and vasculitic involvement of the ipsilatera1 carotid, middle or anterior cerebral artery in 9 cases; in addition to 4 cases of nonophthalmicus or spinal radicular dermatomal lesions, and 2 with disseminated HZV. Cases of lymphoma-associated disease had a subacute or chronic neurologic illness, with 4 cases manifesting preceding lymphoma for up to 3 years, and 3 with occult lymphoma detected only after diagnosis of granulomatous angiitis (Fig. 28.2). One patient with a cavernous sinus syndrome had biopsyproven granulomatous angiitis and occult lymphoma of the midbrain at autopsy. A second patient had successfully treated malignant lymphoma of the parotid gland 2 years before the onset of severe back pain and headache, mental change, focal cerebral and long tract signs, coma, and death in 2 days. At autopsy there was evidence of granulomatous angiitis affecting spinal and cerebral leptomeningeal vessels, and a malignant lymphoma of the parotid gland. A third patient had headaches and

Fig. 28.2. Central nervous system vasculitis. The media and adventitia of a small leptomeningeal artery is almost completely replaced by multinucleated giant cells (arrowheads). There is intimal proliferation with obliteration of the vascular lumen and a dense perivascular mononuclear inflammatory infiltrate (hematoxylin and eosin; original magnification, x250). Reproduced with permission of the publisher (Younger DS (2015). Adult and childhood vasculitis of the nervous system. In: Younger DS (Ed.), Motor disorders. 3rd edn. David S Younger MD PC, New York).

progressive spastic paraparesis that prompted brain biopsy and discovery of granulomatous angiitis; re-evaluation of a submandibular mass showed Hodgkin’s lymphoma. Treatment of the tumor with combination chemotherapy resulted in remission of the cancer and stabilization of the neurologic disorder. The neurologic disease in the six patients with sarcoidosis was essentially similar to other cases of GANS, in that two patients with known systemic disease were being treated with CS when neurologic symptoms emerged. Asymptomatic systemic involvement was present in 3 of 4 others diagnosed at postmortem examination. The 10 patients with amyloid angiopathy were clinically and pathologically inseparable from other cases of granulomatous angiitis, Headache, mental change, gait difficulty, and focal cerebral signs were present at onset in seven patients (70%) and progressive stupor, coma, and death followed in five patients. All so studied at postmortem examination showed diffuse granulomatous vasculitis. These cases are of considerable interest because it is not yet known whether the combination of amyloid angiopathy and granulomatous angiitis is more than a fortuitous association. The pathologic heterogeneity of GANS was exemplified by the predilection of lesions for vessels of variable caliber, regardless of the presenting clinical syndrome or associated disorder. Of 50 cases studied postmortem, 6 had predominant involvement of small arteries and veins, 11 involved small and large vessels together, 28 cases involved small- and medium-sized arteries; and 5 involved large cerebral vessel alone. Isolated microscopic foci of vascular inflammation noted in heart, lungs, and kidney specimens at general autopsy in 19 cases, was considered insufficient evidence for systemic vasculitis. Two factors emerged that changed modern concepts of the disorder. First, patients with beading on cerebral angiography were diagnosed with benign angiopathy of the CNS (BACNS) (Calabrese et al., 1993) including many who may have been diagnosed with PACNS and so treated. Second, there was increasing recognition of serious side effects in up to 40% of patients treated with oral CYC for GPA (despite its lack of inferiority to parenteral cyclophosphamide), that waned enthusiasm for the selection of patients with PACNS for empiric highdose prednisone and long-term maintenance cytotoxic therapy. Table 28.2 shows the survival among comparable immunosuppressive regimens for histologicallyproven GANS in treated and untreated historical controls (Younger et al., 1997). Among 30 such cases diagnosed by meningeal and brain biopsy, 28 were treated with CS alone (11 patients) or with oral CYC (16 patients) or azathioprine (1 patient), and followed for up to a year of whom 18 (64%) improved, 7 (25%) were unchanged, and 3 (11%) died (with roughly equally satisfactory

ADULT AND CHILDHOOD VASCULITIS Table 28.2 Outcome of 54 cases of granulomatous angiitis Outcome

CS

CS + CYC

CS + AZA

None

Total

Improved Same Dieda

8/0a 3/0 0/6

9/0 4/0 3/1

1/0 0 0

0 1/0 1/17

18/0 8/0 4/24

a

Patients diagnosed antemortem in the numerator, and postmortem in the denominator Abbreviations: CS, corticosteroids; CYC, cyclophosphamide; AZA, azathioprine

outcomes after treatment with CS with or without cyclophosphamide). However, 3 patients diagnosed antemortem died while taking CS and cyclophosphamide; and 2 suffered serious sequelae of the therapy including fatal lymphoma, immunosuppression and opportunistic infection, or pneumonia and leukopenia. Comparatively, among 24 patients diagnosed postmortem, 7 (29%) received treatment with CS alone (6 patients) or with CYC (1 patient) died, as did 17 (71%) who were untreated. Thus, 17 of 18 (94%) who received no treatment died, indicating that without therapy the disease was usually fatal. Treatment with CS alone or in combination with CYC was associated with a considerable reduction in mortality; 24 of 34 (70%) so treated survived, either improved (50%) or clinically unchanged. In this historical survey, there was no appreciable benefit in the addition of cyclophosphamide, however the numbers were small, unmatched for age, disease activity, or other factors, and follow-up was nonuniform. The authors suggested that CYC be reserved for histologically-confirmed cases of PCNSV and GANS, especially those who continue to progress or fail to improve on CS alone, and who can be monitored closely for serious medication side effects.

Theme 9: The rationale approach to diagnosis and management of adult and childhood CNS vasculitis requires the cooperation of stroke neurology and neuroradiology The neuroradiological approach to CNS vasculitis has evolved over the past several decades commensurate with improvements and the increased availability of highly precise methods to image the brain and the cerebral vasculature (Wynne et al., 1997) and understanding of the vascular inflammatory process. The pathophysiology of cerebrovascular injury in the course of CNS vasculitis is similar to other vascular beds with some differences. As in other vasculitides, mural changes lead to vessel stenosis or occlusion and endothelial inflammation promotes intraluminal coagulation and

663

thrombosis (Giannini et al., 2012). Perivascular inflammatory changes and edema also contribute to the pathologic picture. Arterial and venous components may be involved separately or together and the dural sinuses may be affected. The underlying generalized inflammatory processes can be associated with a secondary encephalitis or myelitis. Traditional radiologic methodologies do not adequately recognize the major categories of LVV, MVV, and SVV as in the systemic circulation. According to K€uker (2007), vasculitic involvement of the internal carotid (ICA), M1 and A1 segments of the middle (MCA) and anterior cerebral arteries (ACA), intracranial vertebral and basilar arteries, and P1 segment of the posterior cerebral artery (PCA), easily appreciated as large vessels on noninvasive imaging employing MRA and CTA (Fig. 28.3A–E), would nonetheless be equivalent to medium vessels by 2012 Revised CHCC nomenclature (Jennette et al., 2013). By contrast, arterial vessels distal to the MCA bifurcation, as well as the anterior (AComm) and posterior communicating (PComm) arteries, still considered MVV by the same CHCC criteria, would not be well visualized alongside intracranial LVV by MRA or CTA, and require CA to assess luminal irregularities. Finally, the smallest muscular arteries and arterioles within the brain parenchyma as well as the capillaries and proximal venules, all considered small vessels by their lumen size corresponding to a caliber of 200–500 mm or less (Kaufmann and Kallmes, 2008; Hajj-Ali and Calabrese, 2013), are all well beneath the resolution of invasive and noninvasive neuroimaging, and instead require tissue biopsy to diagnose vasculitic involvement. K€uker (2007) describes three steps in the diagnostic evaluation of CNS vasculitis beginning with the demonstration of brain lesions by T2- and diffusion and perfusion-weighted MRI, followed by the delineation of underlying vascular pathology by 1.5 Tesla (T) MRA to study the entire course of the carotid and vertebral arteries, as well as the circle of Willis. Time-of-flight (TOF) MRA sequences improves spatial resolution to detect subtle stenosis and mural thickness in basal brain arteries using MRA source images; however, hrMRI will discern mural enhancement. Conventional angiography with digital subtraction (DSA) evaluates both medium- and small brain vessels and the status of cerebral hemodynamics. Gomes (2010) divides available neuroimaging studies into three groups including those that investigate brain parenchyma, vessel lumen or the vessel wall. Parenchymal findings, while least specific, are necessary to detect the presence of the disease state and to follow progression and remission status. Vessel lumen abnormalities, while highly suggestive of vasculitis in LVV, are generally considered nonspecific in intracranial MVV, and insensitive in SVV.

Fig. 28.3. (A–E). Primary angiitis of the central nervous system. (A) Noncontrast CT (top) demonstrates multifocal regions of low attenuation. Those in the right frontal subcortical white matter and left basal ganglia (black arrows) are sharply defined, without mass effect and likely reflect old infarctions. Both the cortex and underlying white matter of the right occipital lobe are involved as is the right splenium of the corpus callosum (white arrows). In these locations the margins are more ill-defined and there is subtle mass effect characterized by sulcal and ventricular effacement, suggesting acute ischemia in the right posterior cerebral artery territory. MRI FLAIR imaging (middle) demonstrates central low and peripheral high signal intensity within the frontal and periventricular white matter lesions (black arrows) consistent with chronic encephalomalacia from old infarctions. The FLAIR hyperintense signal within the right occipital lobe is more confluent and extends to the posterior temporal lobe and splenium, involving both cortex and white matter (white arrows) and better delineates the extent of the acute infarct. DWI (bottom) demonstrates restricted diffusion consistent with acute ischemia. (B) T1-weighted imaging pre- and postgadolinium demonstrates extensive leptomeningeal enhancement along the cortical surface of the posterior temporal and occipital lobes. (C) CTA demonstrates multifocal vascular narrowing within several branches of the MCA (white arrows) with intervening regions of normal appearing vasculature. At the bottom of the image vascular narrowing within the posterior cerebral artery (not marked) is present. (D and E) CA reveals completely normal extracranial vasculature. The anterior cerebral (black arrowheads), middle cerebral (black arrows) and posterior cerebral artery (black outlined arrows) demonstrate mild to severe short segment stenoses. Abbreviations: CA, catheter angiography; CT, computed tomography; MRI, magnetic resonance imaging; FLAIR, fluid attenuation inversion recovery; CTA, computed tomographic angiography; DWI, diffusion-weighted imaging; MCA, middle cerebral artery. Images courtesy of Adam Davis, MD and Tibor Bescke, MD.

ADULT AND CHILDHOOD VASCULITIS

MRI IMAGING The MRI findings of CNS vasculitis have been previously described (Zuccoli et al., 2011), the commonest of which are T2/fluid attenuation inversion recovery (FLAIR) hyperintense lesions secondary to ischemia distributed throughout subcortical and deep white matter, the deep grey nuclei and the cortices. The MCA territory is the commonest involved in CNS vasculitis (Pomper et al., 1999; Wasserman et al., 2001). Diffusion-weighted imaging (DWI) helps to distinguish acute, subacute and chronic ischemia and is thus mandatory. Lesions are frequently bilateral and of differing ages. Involvement of multiple vascular territories or lesions within a frankly nonvascular territorial distribution may be clues to the diagnosis of CNS vasculitis although they can also be seen with nonvasculitic thrombophilic and cardiogenic emboli that produce ischemia. Acute ischemic lesions are present in up to one-half of patients with PACNS (Salvarani et al., 2007) however, subcortical white matter changes may also be found, or the only finding in symptomatic patients (Salvarani et al., 2007; Neel and Pagnoux, 2009), and unlikely due to atherosclerotic hypertensive disease especially in young patients, or difficult to distinguish from CNS demyelinating disease. Parenchymal and subarachnoid hemorrhages may be a presenting or associated clinical and radiographic feature of CNS vasculitis (Spitzer et al., 2005) although they occur less commonly than ischemic lesions (Salvarani et al., 2012) and there is uncertainty regarding microscopic hemorrhages in CNS vasculitis. T2-weighted gradient-echo MRI, which depicts chronic blood or hemosiderin products as regions with marked signal intensity loss (susceptibility effect) is useful in demonstrating multiple silent petechial hemorrhages scattered throughout the cerebral hemispheres in cortical– subcortical regions (Ay et al., 2002). Neither large nor small silent cortical hemorrhages were found among 25 patients with intracranial vasculitis employing T2-weighed MRI (Kuker et al., 2008). Leptomeningeal enhancement on MRI is noted in up to 10% of cases and may be the only clue of the vasculitic nature of neurocognitive disturbance especially when brain MRA and CA are normal in cases of small vessel PCNSV (Salvarani et al., 2012). Delayed perfusion and reduced cerebral blood volume may be seen on brain CT and MRI in patients with cerebral vasculitis; and magnetic resonance spectroscopy (MRS) cam reveal elevated glutamate and glutamine levels with reduced N-acetyl aspartate (NAA) levels with absence of the choline peak in cerebral vasculitis (Muccio et al., 2013; Park et al., 2014).

MAGNETIC RESONANCE ANGIOGRAPHY This noninvasive nonionizing indirect vessel wall imaging modality does not require intravenous contrast

665

administration for the assessment of intracranial stenoses and vascular occlusions in suspected CNS vasculitis. MRA may overestimate vascular stenosis secondary to diminished signal intensity from vessel tortuosity and slow flow. The more subtle finding of intracranial vessel irregularity is difficult to assess due to lower spatial resolution. Among nine arteries from 14 young patients with clinical and radiological suspicion of cerebral vasculitis, the sensitivity for detecting a stenosis by three-dimension (3D) TOF MRA or DSA varied from 62% to 79% for MRA, and 76% to 94% for DSA. The specificity for detecting a stenosis varied from 83% to 87% for MRA and from 83% to 97% for DSA. Using the criterion of >2 stenoses in 2 or more separate vascular distributions as a true positive criterion for cerebral vasculitis, the false positive rates for MRA and DSAwere comparable (Demaerel et al., 2004). When more than two stenoses are noted on MRA, DSA is unlikely to add further diagnostic precision in a given patient with suspected cerebral vasculitis, but yet might be useful when MRA was normal or disclosed 80% of cases received CS with CYC without subgroup analyses. It is of further interest

that 2 patients among de Boysson et al. (2017) with SV-PACNS did not receive induction treatment with CS. The present series analyses had several weaknesses. The numbers were small, cases were retrospective, unmatched for age, disease activity or other variables, and follow-up was not uniform, and summary data from case series varied in detail. Five literature cases were excluded from this analysis including 4 angiography-negative biopsy-proven cases of cPACNS (Benseler et al., 2005) because of involvement of histologic involvement of small and mediumsized vessels; and one recent case of biopsy-proven small-vessel cPACNS that did not undergo contemporaneous cerebral vascular imaging (Denny and Das, 2019).

Theme 10: The COVID-19 pandemic provided new insights into postinfectious autoimmunity CNS vasculitis The earliest reports of clusters of patients with pneumonia of unknown origin linked to exposure at a seafood and wet animal market in Wuhan (Hubei Province, China) (Zhu et al., 2020) was rapidly identified as a new beta coronavirus named Severe or novel Acute Respiratory Syndrome-Coronavirus-2 (SARS-nCoV-2 or SARS-CoV-2). These single-stranded ribonucleic acid (RNA) enveloped viruses have the largest known genome, ranging from 26.2 to 31.7 kilobases that encodes an important spike (S) glycoprotein that mediates viral entry and determines the range of potential host cell tropism and disease pathogenesis, hence it has been a major source of vaccine interest. Six coronavirus species cause human diseases (Su et al., 2016) associated with the common cold symptoms and two others, SARS-CoV-1, the causal agent of the SARS outbreaks in 2002 and 2003 of Guangdong Province, China (Zhong et al., 2003), and the Middle East Respiratory Syndrome or MERSCoV, responsible for outbreaks in 2012 (Zaki et al., 2012) are zoonotic beta coronaviruses and linked to fatal illness (Cui et al., 2019). SARS-Cov-1 and SARS-Cov-2 use angiotensin-converting enzyme 2 (ACE 2) receptor binding site to infect ciliated bronchial epithelial cells and type II pneumocystis, which explains the affinity of pulmonary involvement. Moreover, with five of the seven human coronavirus’ isolated in this century, they have assumed an important place in the 21st century. The novel SARS-CoV-2 originated in bats and reached humans via badgers, Himalayan palm civets and raccoon dogs, showing a similar capacity to infect humans, first by jumping across species from bat reservoirs. The MERS-CoV a decade earlier also originated in bats utilizing camels as intermediate hosts to human. There are animal models that convincingly demonstrate

ADULT AND CHILDHOOD VASCULITIS the capacity of coronaviruses to enter the CNS across the BBB. Older immunodeficient BALB/c mice exhibit a clinical syndrome, with increasing age as a risk factor (10). Transgenic K18-hACE2 mice infected with SARS-CoV (McCray et al., 2007) invoke infiltration of macrophages and lymphocytes to the lungs and a local release of pro-inflammatory cytokines that contribute to the SARS-CoV-2 cytokine storm. All individuals are generally susceptible to Covid-19. A multicenter retrospective Cox-proportional-hazards regression analysis of 147 critically-ill Chinese patients with COVID-19 (Xu et al., 2020) revealed that age >65years and thrombocytopenia at intensive care unit (ICU) admission, with acute respiratory distress syndrome (ARDS) and acute kidney injury (AKI) independently predicted a higher 60-day mortality. Epidemiological data reflect lower susceptibility and milder disease severity compared to adults, however, the large proportion of asymptomatic children makes epidemic surveillance more difficult, and contributes to greater spread of infection in the population.

COVID MULTISYSTEM INFLAMMATORY SYNDROME IN CHILDREN

One particularly severe childhood affliction, multisystem inflammatory syndrome in children (MIS-C) was noted early in the pandemic when incident cases of fever and mucocutaneous manifestations resembling KD (Baker et al., 2009), a rare vasculitis of childhood that causes coronary-artery aneurysms, emerged in Europe (Whittaker et al., 2020). Two contemporaneous reports in the New England Journal of Medicine described the epidemiology and clinical features of the US disorder (Dufort et al., 2020; Feldstein et al., 2020). With only about 6800 CDC-documented cases of MIS-C worldwide the incidence is vastly lower than SARS-CoV-2 for individuals 2) organ involvement (cardiac, renal, respiratory, hematologic, gastrointestinal, dermatologic or neurological); and no alternative plausible diagnoses; and positive for current or recent SARS-CoV-2 infection by real-time polymerase chain reaction (RT-PCR), serology, or antigen test; or suspected COVID-19 exposure within the 4 weeks prior to the onset of symptoms. Epidemiologic trends of MIS-C suggest that younger children are more likely to present with KD-like features, while older children are more likely to develop myocarditis and shock. The ACR has issued two versions of guidance on the management of MIS-C (Henderson et al., 2020) focusing on diagnosis and management, which includes CS and IVIg for first line management similar to KD, and the recombinant interleukin-1 (IL-1) receptor antagonist Anakinra for refractory cases. With just 59 fatalities from MIS-C, there is a paucity of neuropathology. Case 5 in the autopsy series of Duarte-Neto et al. (2021) described a postinfectious pathophysiology involving neurotropism reminiscent of CNS vasculitis associated with preceding varicellazoster virus infection (Wells et al., 2018). An 8-yearold girl had high fevers and headache for 5 days leading

672

D.S. YOUNGER

to altered mental state and status epilepticus treated with anticonvulsants and severe sedation requiring intubation and ventilatory support. There were ground glass lesions on chest CT, and by day 14 of hospitalization she was found to have left middle cerebral artery (MCA) spasm with on transcranial Doppler; by day 27, she developed circulatory shock and expired. Nasopharyngeal RT-PCR collected during hospitalization was negative and there was no history of SARS-Cov-2 exposure, however SARS-Cov-2 antigens were isolated in cerebral endothelial cells and in the perivascular astroglial cells by immunohistochemistry. There was in addition a thrombus in a medium sized artery, sparse small areas of ground-glass pulmonary infiltrates, with alveolar cell and cardiac endothelial expression of SARS-CoV-2 N antigens without signs of systemic vasculitis.

COVID-CHILDHOOD CNS VASCULITIS Pathologically-confirmed CNS vasculitis in association with SARS-CoV-2 infection without MIS-C was described by Poisson et al. (2022) in an 8-year-old girl with 5 days of fever, cough and headache followed 2 weeks later by hemiparesis prompting two open biopsies without contemporaneous cerebral angiography of an enhancing frontal brain lesion suspicious for tumor that showed transmural lymphohistiocytic and T-cell inflammation of parenchymal small-vessels. Despite sequential treatment with cyclophosphamide, IVIg, CS, rituximab and infliximab, she died 93 days after admission. Postmortem neuropathological examination confirmed the diagnosis of small-vessel vasculitis in the vicinity of a large area of brain infarction, histopathology similar to the earlier tissues specimens. Cerebrospinal fluid from hospital day 2 returned strongly positive for SARS-CoV-2 IgM.

ADULT CNS VASCULOPATHY The postmortem systemic findings of fatal COVID-19 illness have been described in hundreds of cases, which is miniscule in relation to the number of confirmed cases and reported deaths in the United States and worldwide. What was initially thought to be a self-limited disease almost exclusively involving the lungs now is being recognized as one that involves multiple organ systems including the brain. Younger (2021) summarized the neuropathological findings of the first 50 cases of COVID-19 illness to December 1, 2020 with detailed brain findings (Reichard et al., 2020; von Weyhern et al., 2020; Solomon et al., 2020; Al-Dalahmah et al., 2020; Kantonen et al., 2020; Bryce et al., 2020). Older age, male gender, increased serum cytokine and procoagulation markers, and critical care hospitalization for 10 days prior to death characterized

the cohort. Serum cytokine and procoagulant were consistently elevated in those so studied. The vast majority of patients were critically ill and managed in an ICU where the immediate causes of death was generally ascribed to cardiopulmonary failure. However, hypoxic–ischemic changes and neuronal loss was only seen in 25 (50%) of cases. Acute ischemic or hemorrhagic infarcts or petechial hemorrhages were noted in 9 (18%) of cases. There were no signs of vasculitis in any of the cases, however, 12 (24%) showed focal or diffuse cortical and brain leptomeningeal inflammation, characterized mainly as T-cell–mediated based upon flow cytometry, and 6 cases (12%) (von Weyhern et al., 2020) presented with histopathological feature of encephalitis including 6 with localized perivascular and interstitial infiltrates with neuronal cell loss and axonal degeneration involving brainstem nuclei and tracts without territorial infarctions, or evidence of virus infiltration. Two cases (4%) showed inflammatory T-cell infiltrates and clusters of macrophages and axonal injury tracking along cerebral vessels resembling acute disseminated encephalomyelitis (ADEM). Younger’s analysis (Younger, 2021) of critically ill COVID-19 cases reveal several important findings and implications. First, CNS vasculitis was not encountered and was not a cause of fatal COVID-19 neurological illness in this series however, the finding of focal or diffuse perivascular and leptomeningeal inflammation in 36% suggested an important contribution of cerebral vasculopathy. With an additional 16% of cases manifesting indolent brainstem encephalitis, this suggests the need for a high index of suspicion in patients presenting with altered sensorium. Second, hypoxia-ischemia did not account for all relevant neuropathological features of severe COVID-19. Third, patients presenting with elevated levels of circulating interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-a, suggests activation of innate and adaptive immunity indicative of a cytokine storm. Together with increased serum D-dimer and markers of hypercoagulability in 42% of cases, affected patients were at risk for thrombotic and hemorrhagic parenchymal tissue infarction so noted in nine (18%) of cases. There were several limitations of this small cohort analysis of literature cases. First, case series were often small and unselected often with missing demographic data and causes of death. Second, there were often contradictory conclusions about the significance of inflammatory vascular brain changes; moreover, there were all critically ill patients and there were no comparisons to control patients with sepsis. Third, it was uncertain whether negative in-situ SARS-CoV-2 RNA PCR results in those so studied makes a secondary inflammatory immune mechanisms of injury more likely. An updated clinicopathologic analysis of 141 COVID-19 cases comprising 91 additional cases,

ADULT AND CHILDHOOD VASCULITIS including 31 cases (Schaller et al., 2020; Puelles et al., 2020) initially excluded from the series of Younger (2021) for lack of description neuropathology, and 60 additional histopathologically-documented cases (Matschke et al., 2020; Hanley et al., 2020; Deigendesch et al., 2020) extending the period of observation to January 1, 2021 showed four notable findings. The first was the number of positive cases with SARS-CoV-2 genomes by PCR testing, accounting for 13 (48%) of 27 examined brains in the study by Matschke et al. (2020); in 4 (80%) of 5 brain tissue specimens studied by Hanley et al. (2020); and in 8 (38%) of 21 brain specimens examined by Puelles and coinvestigators (2020). Remarkably, SARS-CoV-2 presence did not correlate with the severity of neuropathological findings, thus it remains unclear whether a comparably low viral genome levels detectable by qRT-PCR in brain tissue could be blood-derived. A second finding was the increase in leptomeningeal and interstitial brainstem inflammation characterized in the study as cytotoxic T-cells in 34 (79%) cases according to Matschke et al. (2020), coinciding with the localization of SARS-CoV-2 viral proteins in cranial nerves and interstitial areas of the lower brainstem. Notwithstanding, there were no examples of frank CNS vasculitis. The detection of SARS-CoV-2 RNA specifically in olfactory bulb neurons and glial cells in 4 (57%) of 7 patients in another study cohort (Deigendesch et al., 2020), but not in any other brain regions, which lends support to a route of viral entry via the olfactory system and the importance of anosmia as an early clinical sign of COVID-19. As activated microglia localize to the olfactory bulb and medulla oblongata in COVID-19 brain tissues, both may be points of viral entry to the CNS. Especially considering the capability of SARS-CoV-2 to infect human gastrointestinal enterocytes as well as pneumocytes, it bears consideration whether the vagus nerve derived from the medulla could be another route of entry to the brain. A third findings was the detection of microglial activation and sparse perivascular and leptomeningeal T-cell infiltrates in COVID-19 brains, as well as controls with sepsis or systemic inflammation according in a small series in which authors (Deigendesch et al., 2020) suggest may represent a histopathological correlate of critical illness-related encephalopathy rather than a diseasespecific finding. Fourth, Matschke et al. (2020) who were interested in the neuronal cell types prone to SARS-CoV-2 infection screening gene expression datasets for signatures related to viral entry and persistence, noted high expression of ACE2 in oligodendrocytes and increased expression of transmembrane serine proteases 2 and 4 (TMPRSS2 and TMPRSS4) in neurons that respectively encode proteins implicated in host viral entry (ACE2) and pruning

673

of the viral-decorating spikes (TMPRSS2). Ramani et al. (2020) who employed a brain organoid model to examine whether SARS-CoV-2 directly targeted neurons and could lead to productive infection and neurotoxicity, compared cells from mock organoids that displayed a healthy nucleus labelled with 40 ,6-diamidino-2-phenylindole (DAPI) compared to SARS-CoV-2 exposed organoids that displayed increased terminal deoxynucleotidyl transferase dUTP nick end labeling) staining (TUNEL) detecting DNA breaks formed when DNA fragmentation occurs in the last phase of apoptosis. While most of the SARS-CoV-2-positive cells were TUNEL-positive, some were caspase-positive displaying pT231 Tau localization at the cell soma not observed in mock organoids. pT231-tau is highly neurotoxic and acts as an early driver of tauopathy in neurodegenerative diseases such as Alzheimer disease. In so much as Tau abnormalities in SARS-CoV-2 positive neurons could result from infection, it could also result from triggering of a cascade of downstream effects that results in immuneinflammation, neuronal stress, and direct neurotoxicity, all of which warrant future investigations.

Theme 11: Nonsystemic peripheral nerve vasculitides pose a challenge to diagnosis and treatment BACKGROUND The very concept of NPNV, which presumes that the necrotizing vasculitis process could be widespread within the PNS and not elsewhere in the body has frequently been called into question for several reasons. First, the reports of silent lesions in medium-sized muscular arteries in patients with clinically isolated vasculitic neuropathy make them anomalous examples of systemic vasculitis. As an example, patient 1 in the series of pathologically confirmed cases of PAN by Kernohan and Woltman (1938) was a 54-year-old man suffered for 5 years with progressive generalized painful peripheral neuropathy that was so severe before death that he was nearly paralyzed and unable to speak or swallow. Postmortem examination showed PAN limited to the nerve trunks of the arms and legs with otherwise normal brain, cranial nerves, and spinal cord tissues that was free of vasculitis. Examination of all other organs failed to reveal a single vascular lesion, except one small artery in the capsule of the prostate gland. Torvik and Berntzen (1968) described a 76-year-old woman with diffuse fever, pain, and central scotoma of the eye that improved with CS. Biopsy of the temporal artery and pectoralis muscle disclosed necrotizing arteries of small arteries and arterioles in small adventitial vessels of the temporal artery without frank temporal arteritis. However, postmortem examination showed evidence of healed vasculitis in numerous small arteries and arterioles of muscle

674

D.S. YOUNGER

and nerve tissue measuring 50 to 200 mm in diameter without vasculitis in visceral organ or the CNS. Second, vasculitis in muscle tissue is included in the definition of NSVN (Collins et al., 2010) suggesting instead the more appropriate term of peripheral nervous system vasculitis (PNSV) than NSPNV. Third, the varied long-term follow-up in cases series of NSPNV ranging from 6 months to 22 years (Dyck et al., 1987) that likely impacts a bias of ascertainment. Fourth, the notion that discovering vasculitis in a cutaneous nerve or muscle tissue specimen aids in the diagnosis of systemic vasculitides, particularly when no other site of vasculitis can be found. The FVSG database used nerve and muscle tissue biopsy to establish systemic vasculitis in a retrospective study cohort of PAN in the absence or presence of symptomatic peripheral neuropathy (Pagnoux et al., 2010). Of 129 patients who underwent nerve biopsy, including 108 with peripheral neuropathy and 21 without peripheral neuropathy, vasculitic lesions were noted respectively in 83% and 81% of patients compared to muscle biopsy which showed vasculitis respectively in 68% and 60%. Lacking a clear relationship to PNV, the 2010 guidelines (Collins et al., 2010) restricts the inclusion of cases associated with insulin dependent or T1D and noninsulin dependent or T2D, to avoid sources of ascertainment bias due to their frequent association with neuropathy. However, beyond the metabolic factors involved in the etiopathogenesis of the commonest form of neuropathy associated with either form of diabetes, namely distal symmetrical polyneuropathy (DSPN) (Kazamel et al., 2021), there are historical precedents for attributing associating systemic autoimmune and neuropathic ischemic aspects of diabetes with inflammatory changes in MNM and various forms of proximal diabetic neuropathy (PDN) notably the radiculoplexus neuropathies.

ISCHEMIC INFLAMMATORY AND AUTOIMMUNE FACTORS T1D is a T-cell–mediated autoimmune disease with an onset believed to be triggered by unknown environmental factors acting on a predisposing genetic background. The pancreas of newly diagnosed T1D shows inflammation in the region of insulin producing b-cells (Hanninen et al., 1992). The possibility that the onset of T1D might be triggered by a postinfectious autoimmune mechanism in genetically predisposed individuals due to molecular mimicry was investigated in two T1D children who died prematurely from brain swelling as a complication of ketoacidosis. Their autopsy showed evidence of islet infiltrating T-(IIT) cells with selective expression of a membrane-bound superantigen and selective expansion of a T-cell receptor (TCR) variable segment of the

b-chain (Vb7) indicating integrated bacterial or viral genes (Conrad et al., 1994). It was thought that if the initial exposure to a superantigen occurred at a very early age, potentially autoreactive T-cell clones might be inactivated. Whereas if exposure occured many years later, many different T-cell clones expressing the same T-cell receptor VΒ might be simultaneously activated, which in a genetically predisposed individual, could initiate a process that eventually results in the destruction of b-cells. Moreover, superantigens and peptidoglycans in AAV could induce skewing of T-cell responses toward pathogenic IL-17-producing T-helper cells (Th17), the overproduction of which could aggravate inflammatory responses and contribute to the production of autoantibodies as well as to granuloma formation and tissue injury (Abdulahad et al., 2009). Moreover, the expression of the variable region genes of the b-chain has been analyzed to study the involvement of peripheral blood T-cells in systemic vasculitis (Simpson et al., 1995). The genetic risk of T1D is strongly linked to HLA class II DR3 and DR4 haplotypes, with the highest risk in those with the DR3/DR4 genotype. The importance of HLA genes to T1D risk highlights the role of the adaptive immune system in the development of autoimmunity. Up to 90% of patients harbor one or more antibodies, most notably antibodies against cytoplasmic antigens such as ICA 69 and glutamic acid decarboxylase (GAD65) (Giorda et al., 1991; Atkinson and Maclaren, 1994) which may become accessible to T- and B-lymphocytes as a consequence of initial islet cell damage. The stimulation of self-antigen-selected T-cell clones and self-antigen-specific B-cell responses may augment the destruction of pancreatic b-cells. Finally, T1D occurs with increased frequency in association with other autoimmune disorders, including Grave disease, pernicious anemia, Hashimoto thyroiditis, myasthenia gravis, antiphospholipid antibody syndrome, and Addison disease. The views supporting an ischemic pathogenesis of diabetes stemmed from observations of diseased blood vessels and nerve trunks removed at autopsy or from amputated limbs (Pryce, 1887, 1893), however, those patients often had long-standing diabetes that tended to increase the likelihood of arteriosclerosis. Decades later, systematic studies of peripheral nerve microvessels stained by the periodic acid-Schiff method (Fagerberg, 1959) showed thickening of the walls and narrowing of endoneurial microvessels and nutrient artery lumina that later proved to be due to duplication of basal lamina, loss and degeneration of endothelia and pericytes, increased cellular debris, endothelial fenestration and dysjunction, altogether with the collective features of diabetic microangiopathy (Giannini and Dyck, 1994). Over the years there accumulated increasing support for the

ADULT AND CHILDHOOD VASCULITIS contribution of ischemic immunologic mechanisms in the pathogenesis of diabetic neuropathy, especially in patients with inordinately severe PDN, MNM, and DSPN. Early investigators (Woltman and Wilder, 1929; Dolman, 1963; Olsson et al., 1968; Duchen et al., 1980) noted the presence of inflammation in diabetic nerves, but their significance was not appreciated, partly because routine histology with hematoxylin and eosin probably under-estimated the number of infiltrating cells, and it was not known whether the observed infiltrates exceeded the expected number of cells in normal nerves. Raff et al. (1968) observed the ischemic consequences of inflammation of peripheral nerve microvessels with a diameter 95% specificity for systemic vasculitic neuropathy, namely ANCA and an erythrocyte sedimentation rate (ESR) >100 mm/h.

676

D.S. YOUNGER

The following year, Younger (2011) affirmed the existence of DLSRPN in a living case in whom superficial fibular sensory nerve biopsy tissue showed MV. However, the patient died of aspiration pneumonia several weeks after treatment with IVIg, CYC and methylprednisolone and postmortem examination showed no evidence of systemic or PNV. Sections of extradural lumbar plexus, sciatic, and femoral nerve tissue showed epineurial PV infiltration of adjacent endoneurium. A variant of DLSRPN, painless diabetic motor neuropathy (Garces-Sanchez et al., 2011) also due to ischemic injury and MV, differs from typical DLPRN being more insidious and symmetrical with slower evolution which may explain the lack of pain. The clinical parameters of diabetic cervical radiculoplexus neuropathy have recently been analyzed (Massie et al., 2012). It typically presents in the seventh decade in T2D with focal arm weakness followed by pain and numbness attributed to nearly equal involvement of the upper, middle and lower brachial plexus segments, individually or in combination, resulting at time in pan-plexopathy. Over one-half of cases showed at least one other body region affected (contralateral or thoracolumbar) and recurrence is not unusual. Nerve biopsies from the arm or leg showed ischemic injury with axonal degeneration, multifocal fiber loss, focal perineurial thickening and epineurial and perivascular inflammation, with hemosiderin deposition, vessel wall inflammation and MV. Sharing many of the clinical and pathological features of DLRPN, these two conditions are best categorized together within the same clinicopathologic spectrum.

CLASSIFICATION OF PERIPHERAL NERVE VASCULITIS Although the Revised 2012 Chapel Hill Consensus Conference (CHCC) (Jennette et al., 2013) does not directly consider NSVN, according to CHCC nosology, it probably should be recognized as a single-organ vasculitis (SOV) of the peripheral nervous system (PNS), analogous to PCNSV. In keeping with the CHCC nomenclature (Jennette et al., 2013), based upon the caliber of the vessels involved, NSVN is uncommon in LVVs involving the aorta, with the exception of its major branches and analogous veins; and more prevalent in MVVs such as PAN with a predilection for visceral arteries, or in SVVassociated with AAVand immune complex diseases, where it involves intraparenchymal arteries, arterioles, capillaries, veins and venules. Gwathmey and colleagues (2019) favor a simplified binary classification of PNV separating cases into nerve large arteriole vasculitis and nerve MV, based upon the caliber size of the affected vessels for prognostication and treatment approaches. The former category, which

includes most patients with MVV and SVV affects vessels ranging from 75 to 300 mm in diameter, with involvement of small arteries, large arterioles and variably smaller vessels; while nerve MV includes DLRPN, LRPN, DCRPN and PDMN, involving small arterioles, endoneurial microvessels, capillaries and venules all measuring 100 mg/dL with pronounced pleocytosis especially with an excess of lymphocytes is a promising sign of CNS vasculitis in both children and adults. The fluid should be sampled for an extensive panel of constituents with an extra tube placed in a negative 80 °C freezer for future analysis or to replace specimens lost in transport. In general, the neuroradiologic evaluation of patients will be guided by the expertise and technical advances available at the institution where care is being rendered,

Table 28.4 Laboratory evaluation of systemic and nervous system vasculitides Studies in blood, urine and body fluids CBC, chemistry panel, ANA, ANCA by IIF; ANCA ELISA serology specific for PR3 and MPO [in those with IIF ANCA seropositivity, other cytoplasmic fluorescence, and ANA that results in homogeneous or peripheral nuclear fluorescence]; ESR, CK, T- and B-cell subset panel, circulating IC, acute and convalescent viral, retroviral, bacterial, fungal, TB, syphilis and Lyme serology; quantitative immunoglobulins, IFE, C1q, complement proteins, RF, cryoglobulins, anticardiolipin and aPL, LAC, dsDNA antibodies, and appropriate HLA haplotypes; urinalysis for spot and 24 h collection for chemical and cellular microscopic analysis; bronchoscopy [in those with lung lesions] for lavage; lumbar CSF analysis for protein, glucose, cell count, IgG level, oligoclonal bands, cytology, VDRL, bacterial gram stain and culture; India ink, cryptococcal antigen and fungal culture; acid-fast and TB culture; viral encephalitis panel for realtime PCR analysis of DNA and RNA viruses; paired analysis of serum and CSF for autoantibody-mediated encephalopathy, and Lyme index with Borrelia burgdorferi and HIV serology. Radiological studies Screening color Doppler ultrasonography of the temporal arteries and great vessels, 3-T brain and spinal cord MRI and high field MRA or CTA and DSA of the brain and other vascular beds and major vessels; 18FDG body PET-CT, nuclear medicine cerebral perfusion with SPECT. Histopathological studies Bronchoscopy or needle tissue biopsy of lung lesion and endoscopic biopsy of kidney tissue; USG-guided temporal artery biopsy; open sural nerve and soleus muscle or superficial fibular nerve and peroneus muscle tissue biopsies for epineurial and epimysial vasculitic foci; en bloc open or sterotactic-guided biopsy of cortical brain and leptomeningeal tissues for vasculitic foci in arteries and veins, and 3 mm punch skin biopsy for ENF density and histology employing PLP 9.5, with IF of vessel walls and microscopic analysis for leukocytoclasia. Abbreviations: ANA, antinuclear antibody; ANCA, antineutrophil cytoplasmic antibody; aPL, antiphospholipid; CBC, complete blood count; CK, creatine kinase; CSF, cerebrospinal fluid; CT and CTA, computed tomography and angiography; DNA and RNA, deoxyribonucleic and ribonucleic acid; ds, double-stranded; DSA, digital subtraction angiography; ELISA, enzyme-linked immunosorbent assay; ENF, epidermal nerve fiber; ESR, erythrocyte sedimentation rate; 18FDG body PET-CT, Fluorodeoxyglucose positron emission tomography fused with CT; IC, HIV1, human immunodeficiency virus type 1; HLA, human leukocyte antigen; Ig, immunoglobulin; immune complexes; IFE, immunofixation electrophoresis; IIF, indirect immunofluorescence; dsDNA, LAC, lupus anticoagulant; MRI and MRA, magnetic resonance imaging and angiography; MPO, myeloperoxidase; PCR, polymerase chain reaction; PLP9.5, protein gene product 9.5; PR3, proteinase 3; RBC and WBC, red and white blood cells; RF, rheumatoid factor; SPECT, single photon emission CT; T and B, thymus and bone marrow derived cells; TB, tuberculosis; USG, ultrasonography; VDRL, Venereal Disease Research Laboratory.

ADULT AND CHILDHOOD VASCULITIS but will likely necessitate high resolution (hr) 3-T brain and spinal cord MRI and MRA or CTA before proceeding to CA to respectively view parenchymal tissues, vessel walls and luminal changes. Mural changes with vessel wall thickening correlate with early inflammation while vascular luminal stenosis and aneurysm formation often indicates late vasculitic complications. 18Fluorodeoxyglucose positron emission tomography (PET) detects metabolically active inflammatory cells infiltrating vessel walls in large vessels. However, the extent of disease and monitoring of luminal changes is best appreciated by employing a combination of PET and DSA techniques. Likewise, the combination of nuclear medicine cerebral perfusion with single photon emission-CT (SPECT) and PET fused with MRI (PET/MRI) provides a unique estimation of perfusion across the blood–brain barrier (BBB) in relation to cerebral regional metabolism.

Tissue biopsy studies in the vasculitides

679

obtained (Basu et al., 2010). Histologically-proven PAN was ascertained in 70% of patients by nerve, muscle or skin tissue biopsy in one large cohort (Pagnoux et al., 2010). Of 129 patients underwent nerve biopsy including 108 with peripheral neuropathy and 21 without peripheral neuropathy, vasculitic lesions were noted respectively in 83% and 81% of patients compared to muscle biopsy which showed vasculitis respectively in 68% and 60%. Angiography showed renal and gastrointestinal microaneurysms or stenosis respectively in 66% and 57% of patients. While PAN frequently involves the gastrointestinal tract especially the small intestine, biopsy of visually apparent lesions on luminal exam rarely provide histopathologic confirmation of vasculitis (Pagnoux et al., 2005). Renal biopsy may be negative even in patients with frank organ involvement due to sampling error. Unlike AAV, where the histopathologic renal lesion is the kidney glomerulus, microaneurysm formation due to PAN increases the risk of bleeding associated with renal biopsy.

AAV

TEMPORAL ARTERY BIOPSY

Tissue biopsy is important in defining the type, extent and character of the inflammatory process and categorizing the underlying systemic vasculitic disorder. In AAV, histologic evidence of necrotizing SVV with accompanying granulomatous inflammation noted in GPA can be used to support the diagnosis in conjunction with clinical and serologic information. Although nasal, sinus and upper airway involvement in GPA may be noted overall in 92% of patients at onset of disease (Hoffman et al., 1992), tissue biopsy of these regions is associated with vasculitis and necrosis in only 23% of upper airway biopsy specimens. The small amount of tissue available in biopsy specimens from areas of the head and neck make it surprisingly difficult or impossible to identify the classical pathological features of GPA. By comparison, renal biopsy in patients with GPA show varying degrees of segmental necrotizing glomerulonephritis in 80% of tissue (Hoffman et al., 1992). In the absence of renal involvement, targeted biopsy of radiographically abnormal lung parenchyma via a thoracoscopic or open lung biopsy technique shows granulomatous changes in 22% and capillaritis in 31% of those so studied (Hoffman et al., 1992) compared to the efficacy of transbronchial biopsy in establishing the diagnosis of pulmonary vasculitis in 10% of suspected tissues (Schnabel et al., 1997).

Histopathologic evidence of vascular wall infiltration by giant cells at the junction of the intima and media, leading to thrombosis, intimal hyperplasia, and fibrosis (Salvarani et al., 2002) is the gold standard method of GCA diagnosis. Accordingly, temporal artery biopsy is recommended for all patients with suspected GCA (Mukhtyar et al., 2009). The sensitivity of temporal artery biopsy in GCA has been estimated to be 85% (Younge et al., 2004). Evidence in support of unilateral as compared to bilateral temporal artery biopsy is lacking, leaving it up to the judgment of the treating physician. Temporal artery ultrasonography can be used to guide the surgeon to the arterial segment with the clearest halo sign in performance of temporal artery biopsy (Blockmans et al., 2009), however the discontinuous nature of inflammation in GCA leading to skip lesions may still lead to unforeseen false-negative results when the tissue specimen is of insufficient length and multiple sections are not examined. Certain clinical features, such as the presence of jaw claudication can increase the positivity predictive value of a temporal artery biopsy. Arteritis can be observed histologically even after several weeks of high dose corticosteroid therapy (Achkar et al., 1994). Notwithstanding, the heightened risk of ischemic complications culminating in irreversible visual loss in early untreated GCA has led to the recommendations for prompt administration of high-dose corticosteroid therapy while awaiting tissue biopsy results.

PAN The finding of focal, segmental necrotizing vasculitis in medium-sized arteries confirms the diagnosis of PAN in suspected patients. Angiographic demonstration of microaneurysm formation is an acceptable surrogate to the histopathologic confirmation when tissue cannot be readily

BRAIN AND MENINGEAL BIOPSY No typically abnormal neurological symptom, sign, or laboratory test establishes the diagnosis of CNS

680

D.S. YOUNGER

vasculitis in a living patient except a “positive” brain biopsy showing pathologic evidence of cerebral blood vessel inflammation. Biopsy specimens should be collected prior to treatment with CS and other immunosuppressive or biological agents whenever possible to avoid changes in the histopathology. Both lesional and nonlesional tissue specimens can yield a successful diagnosis, however a lesional specimen is preferred. Tissue samples of cortical gray matter, subcortical white matter tissue and overlying meninges should be removed en bloc prior to electrocautery and ideally measure 1 cm.3

MUSCLE AND NERVE AND EPIDERMAL NERVE FIBER ANALYSIS

Skin biopsy can be performed for IF analysis of vessel walls to search for microscopic analysis for leukocytoclasia, Ig and complement deposition. Two 3-mm punch skin biopsies of along the distal calf and proximal thigh fixed in antiprotein-gene-product 9.5 (PGP 9.5) solution will discern epidermal nerve fiber depletion useful in the designation of small and large fiber neuropathy, and in the selection of patient who should undergo surgical biopsy of the sural nerve or branch of the superficial fibular sensory nerve and corresponding soleus and peroneus brevis muscle to search for vasculitic lesions and confirm the neuropathic or myopathic changes found on electrodiagnostic studies.

TREATMENT Most experts concur with the following three principles to guide management of patients. First, vasculitis it is a potentially serious disorder with a propensity for permanent disability owing to tissue ischemia and infarction. Second, undiagnosed and untreated, the outcome of vasculitis is potentially fatal. Third, a favorable response to an empiric course of immunosuppressive and immunomodulating therapy should not be considered a substitute for the histolopathologic confirmation of vasculitis. Physicians treating vasculitides must choose the sequence and combination of available immunosuppressant and immunomodulating therapies to induce and sustain remission and treat relapses, recognizing the possible beneficial and adverse effects. Treatment options for the different categories of vasculitis are summarized in Table 28.5.

Corticosteroids Corticosteroids are frequently used alone or in combination and are the single most commonly used therapeutic agent for the treatment of systemic vasculitides. The clinical benefit of CS stems from their inhibitory properties on inflammatory and immune responses. Their broad

Table 28.5 Recommendations for the treatment of vasculitis in the literature Large vessel vasculitis GCA: EULAR (Hellmich et al., 2020), CS (Chandran et al., 2015; Yates et al., 2016), AZA (Boureau et al., 2016), MTX (Spiera et al., 2001; Jover et al., 2001; Hoffman et al., 2002), infliximab (Hoffman et al., 2007), or MMF (Sciascia et al., 2012) TAK: EULAR (Hellmich et al., 2020), CS (Shirai et al., 2021), AZA (Ohigashi et al., 2012), RTX (Pazzola et al., 2018), infliximab (Campochiaro et al., 2021), anti–TNF-a, anti–IL-6 (Regola et al., 2022), and tocilizumab (Salvarani and Hatemi, 2019; Nakaoka et al., 2020) Medium vessel vasculitis PAN: FVSG (Terrier et al., 2020), CS and CYC (Guillevin et al., 2003), MMF (Erden et al., 2017; Brogan et al., 2021), infliximab (Ginsberg et al., 2019), PE (Regent et al., 2020). KD: SHARE (de Graeff et al., 2019), ASA + IVIg (Durongpisitkul et al., 2003), CS (Newburger and Fulton, 2007; Furukawa et al., 2008) Small vessel vasculitides AAV GPA and MPA: EULAR (Yates et al., 2016), CS (Pagnoux and Guillevin, 2015), Induction with CS + CYC (Hoffman et al., 1992), CS + RTX (Stone et al., 2010; Jones et al., 2010; Jayne, 2013; Charles and Guillevin, 2013), CS + MMF (Silva et al., 2010), RTX (Cartin-Ceba et al., 2012; Smith et al., 2012; Specks et al., 2013), or MTX (De Groot et al., 2005), maintenance with AZA or MTX (Pagnoux et al., 2008; Gopaluni et al., 2017), and AZA or MMF (Hiemstra et al., 2010), PE (Jayne et al., 2007; Walsh et al., 2013; Regent et al., 2020), IVIg (Jayne et al., 2000) EGPA: EGPA Consensus Task Force (Groh et al., 2015), CS (Raffray and Guillevin, 2020), mepolizumab (Bel et al., 2014; Ortega et al., 2014) Childhood AAV (Yates et al., 2016; Calatroni et al., 2021), Remission induction: CS, CYC, MTX or RTX, and remission maintenance AZA MMF MTX CYC (Morishita et al., 2017) Immune-complex vasculitis CryoVas: (Perez-Alamino and Espinoza, 2014), RTX (De Vita et al., 2012; Sneller et al., 2012; Lesniak et al., 2021), RTX +belimumab (Saadoun et al., 2021) MC: (Ferri et al., 2011; Giuggioli et al., 2017), French (Terrier et al., 2015) IgAV: SHARE (Ozen et al., 2019), CS (Weiss et al., 2007), RTX (Hernández-Rodríguez et al., 2020), MMF (Ren et al., 2012; Nikibakhsh et al., 2014) Hypocomplementic-C1q: FVSG (Jachiet et al., 2015), omalizumab (Navarro-Navarro et al., 2020) Variable vessel vasculitis Cogan: CS (Gluth et al., 2006) Behc¸et: CS (Noel et al., 2014), MM (Shugaiv et al., 2011), colchicine or anti-TNFa (Nanthapisal et al., 2016; Hatemi et al., 2015)

ADULT AND CHILDHOOD VASCULITIS Table 28.5 Continued Single organ vasculitis Adult PACNS: CS alone or with CYC for induction of remission (de Boysson et al., 2014; Salvarani et al., 2015a, b; de Boysson et al., 2018; Schuster et al., 2019; Salvarani et al., 2020), CS-sparing with CYC, RTX, MTX, AZA (Schuster et al., 2019) cPACNS SV/AN: CS with CYC induction followed by CS taper and AZA or MMF maintenance (Hutchinson et al., 2010) LV: CS induction with ASA/clopidogrel followed by CS taper and MMF maintenance (Walsh et al., 2017; Beelen et al., 2019) APNP/APP: CS with CYC induction and acute AC followed by CS taper, APT, MMF (Beelen et al., 2019) Abbreviations: AAV, antineutrophil cytoplasmic antibody-associated vasculitis; AC, anticoagulation; AN, angiography-negative; ASA, aspirin; APNP, angiography-positive nonprogressive; APP, angiographypositive progressive; APT, antiplatelet therapy; AZA, azathioprine; BD, Behc¸et disease; CS, corticosteroids, CryoVas, cryoglobulinemia vasculitis; CYC, cyclophosphamide; EGPA, eosinophilic granulomatosis with polyangiitis; EULAR, European Alliance of Associations for Rheumatology; French Vasculitis Study Group; GCA, giant cell arteritis; GPA, granulomatosis with polyangiitis; HCV, hepatitis C virus; IgAV, IgA vasculitis; INF, interferon; IL, interleukin; IVIg, intravenous immune globulin; KD, Kawasaki disease; LV, large-vessel; MC, mixed cryoglobulinemia; MTX, methotrexate; MPA, microscopic polyangiitis; MMF, mycophenolate mofetil; MTX, methotrexate; PACNS, primary angiitis of the central nervous system; PAN, polyarteritis nodosa; PE, plasma exchange; RTX, rituximab; SHARE, Single Hub and Access Point for Paediatric Rheumatology in Europe; SV, small-vessel; TAK, Takayasu arteritis; TNF, tumor necrosis factor.

anti-inflammatory properties includes the ability to decrease vascular permeability, inhibit the migration of inflammatory cells to sites of injury or untoward inflammation, inhibition of PMN) and mononuclear cell function, and a variety of mediators important in the inflammatory response such as kinins, histamine, and prostaglandins. They are potent inhibitors of the immune response with broad effects on antigen processing and immune activation including lymphocyte proliferation mediator release, as well as lymphocyte trafficking. There is general agreement on the beneficial and deleterious effects of differing dosing schedules CS. First, therapeutic efficacy and toxicity are related to the administered dose, duration of therapy, and frequency of administration with more serious side effects during sustained daily long-term therapy without tapering. Second, divided doses of daily prednisone are probably more potent than a single high morning dose of greater than 80 mg of prednisone. Third, daily low-doses of 15 mg or less of prednisone incur many of the same problems as high-dose therapy in particular hypothalamic pituitary

681

adrenal (HPA) axis suppression with incipient adrenal failure. Fourth, pulse therapy of 1000 mg of methylprednisolone daily for several days is generally well tolerated and is associated with fewer effects than long-term therapy with an equivalent degree of immunosuppression and short-term anti-inflammatory benefit. Fifth, alternate day therapy, typically reserved for patients whose disease activity is under good control reduces long-term toxicity, in particular HPA suppression, CS myopathy, and osteoporosis. Sixth, while the different therapeutic options for CS administration generally relates to the best ratio of benefits to risk, when the disease is life-threatening or fulminant as in systemic vasculitides, it is most reasonable to institute pulse therapy followed by daily highdoses if monotherapy is employed, while the inverse may be more applicable if CS are combined with an alkylating cytotoxic agent such as CYC. Early intensive therapy has been suggested in patients with LVV including TAK and GCA (also known as large-vessel GCA [LV-GCA]), to induce remission. Corticosteroids are the mainstay of therapy in GCA, however, their use is associated with predictable and occasionally serious side effects even with an initial dose of 0.5 to 0.7 mg/kg/day of prednisone in the absence of eye involvement, or 1 mg/kg/day in the presence thereof, both of which are continued for 1 month before gradual tapering (Salvarani et al., 2008c; Mukhtyar et al., 2009). There are no clinical trials of the efficacy of CS in patients with MVV and SVV (Mukhtyar et al., 2009). Corticosteroids employed in almost all clinical trial and cohort studies to obtain remission or induce cure alone or in association with other agents for AAV, are given at the dose of 1 mg/kg/day of prednisone for 3 to 4 weeks according to the EUVAS study group (Jayne et al., 2003). The FSVG (Guillevin et al., 1990) recommends pulse methylprednisolone for life-threatening organ involvement because of its rapid onset of action and favorable safety index. The use of CS in the treatment of many types of vasculitides other than LVV and systemic necrotizing vasculitides is controversial. Pulse methylprednisolone was alternative therapy given over 1 to 3 days in children with KD is probably comparable to one or more IVIg infusions to alleviate fever and acute inflammation (Newburger et al., 2004). A prospective randomized open-label Japanese study of KD (Kobayashi et al., 2012) that employed IVIg therapy plus CS showed significantly less coronary artery abnormalities than those treated with IVIg and aspirin. Side effects of CS in PAN and associated HBV infection included enhancement of viral replication and progressive cirrhosis. Such patients treated with PE and antiviral therapy in addition to CS to avert severe life-threatening manifestations

682

D.S. YOUNGER

allows discontinuation of CS. They are safely administered with IFN-a to treat HCV-related cryoglobulinemia (Dammacco et al., 1994).

Cyclophosphamide Cyclophosphamide is a member of the alkylating class of cytotoxic drugs that covalently binds across linked deoxyribonucleic acid (DNA) strands and interferes with mitosis and cell replication. The immunosuppressive effects of CYC include absolute suppression of B- and T-cells, as well as suppression of both cell-mediated and humoral immunity. Interest in the use of CYC emerged in early studies by Fauci and Wolff of GPA (1973) so treated with combination prednisone and oral CYC at doses of 2 mg/kg/day evidencing that improved long-term benefit despite severe kidney disease and treatment-related morbidity. The latter included increased propensity for infection, hemorrhagic cystitis, bladder fibrosis, bone-marrow suppression, ovarian failure, bladder cancer and hematologic malignancies (Hoffman et al., 1992). A prospective, multicenter, randomized trial comparing steroids and pulse CYC versus steroids and oral CYC in GPA (Guillevin et al., 1997) demonstrating equal efficacy of pulse CYC in achieving initial remission of GPA, with fewer side effects and lower mortality. However, treatment with pulse CYC did not maintain remission or prevent relapses as well as oral CYC. A meta-analysis by the EULAR (Mukhtyar et al., 2009) concluded that pulsed CYC was more likely to result in remission status than continuous oral therapy, with a lower risk of side effects. A meta-analysis of three studies of intravenously pulsed CYC (de Groot et al., 2001) demonstrated that pulsed regimens reduced cumulative CYC exposure by 50%, and were at least as effective at inducing remission, with fewer infective and myelosuppressive side effects however there was possible increased risk of relapse. The recommended initial dose of CYC varies from 0.5 to 0.7 g/m2 at 2-week intervals given initially on days 1, 15 and 30, and continued every 3 weeks until remission is obtained, followed later by maintenance therapy. The CYCLOPS study (de Groot et al., 2009) randomized 149 patients with generalized AAV to receive either pulse CYC 15 mg/kg at 2-week intervals for the first three doses and every 3 weeks thereafter, or daily oral CYC 2 mg/kg/day noting that pulse therapy was equally effective as daily oral CYC with a lower cumulative dose and fewer instances of leukopenia. Patients aged 65 years and older with newly diagnosed PAN, GPA, MPA, and EGPA who receive low-dose intravenous pulse CYC with faster CS dose-tapering, have reduced rates of severe adverse events and similar remission and relapse rates.

Apart from AAV, CYC has been used in other systemic vasculitides including severe IgAVand CValthough these indications are controversial. A multicenter, prospective, randomized open-label trial (Pillebout et al., 2010) found that the addition of CYC to CS provided no further benefit compared to steroids alone in treating adult patients with severe IgAV. In patients with HBV-related PAN and HCV-related CV, treatment with PEG-IFNa plus ribavirin was associated with a good prognosis whereas immunosuppressive agents including CS, were associated with a poor outcome and increased mortality (Terrier et al., 2011).

Azathioprine Azathioprine is an antimetabolite and purine analogue that interferes with DNA synthesis. Long-term AZA immunosuppression leads to decreased numbers of B- and T-cells, as well as decreased B-cell proliferative responses and antibody synthesis. It also inhibits natural killer (NK) cell activity (Maltzman and Koretzky, 2003). The Cyclophosphamide versus Azathioprine for Early Remission Phase of Vasculitis (CYCAZAREM) trial (Jayne et al., 2003) studied patients with a new diagnosis of GPA and MPA and a serum creatinine concentration of 5.7 mg per deciliter (500 mm/L) or less. All patients received at least 3 months of therapy with oral CYC and prednisolone. After remission, patients were randomly assigned to continued CYC therapy of 1.5 mg per kilogram of body weight per day or a substitute regimen of AZA of 2 mg per kilogram per day. Both groups continued to receive prednisolone and were followed for 18 months from study entry. Relapse was the primary end point. It concluded that patients with at least 3 months of oral CYC and prednisolone for AAV, and randomly assigned to continued CYC therapy at the dose of 1.5 mg per kilogram of body weight per day or a substitute regimen of AZA of 2 mg per kilogram per day, in addition to prednisolone, and followed 18 months from study entry, with an endpoint of relapse, showed no increase in the rate of relapse. The relapse rate was lower among the patients with MPA than among those with GPA (P ¼ 0.03) and the withdrawal of CYC and the substitution of AZA after remission did not increase the rate of relapse. Thus, the duration of exposure to CYC may be safely reduced with the addition of AZA after initial remission in AAV.

Methotrexate Methotrexate is an antimetabolite that inhibits folic acid and has been used in LVV and AAV. A meta-analysis by Mahr et al. (2007) of 3 randomized placebo-controlled trials in patients with newly diagnosed GCA (Spiera et al., 2001; Jover et al., 2001; Hoffman et al., 2002)

ADULT AND CHILDHOOD VASCULITIS in whom treatment consisted of initial high-dose CS and randomly assigned oral MTX therapy of 7.5–15 mg/ week versus placebo, and a comparison of time-to-event, and continuous outcomes. The authors (Mahr et al., 2007) found that adjunctive treatment of GCA with MTX lowered the risk of relapse and reduced exposure to CS, thus MTX could be considered as a therapeutic option in addition to standard-of-care treatment with CS. A randomized, multicenter trial that enrolled 98 patients from 16 with newly diagnosed GCA to determine if MTX reduced relapses and cumulative CS requirements and diminished disease- and treatment-related morbidity did not support the adjunctive use of MTX to control disease activity or to decrease the cumulative dose and toxicity of CS in patients with GCA. Findings from the Wegener’s granulomatosis-Entretien Trial (WEGENT) in 2008 (Pagnoux et al., 2008) suggested that AZA or MTX could effectively maintain remission of GPA or MPA. A subsequent long-term study with 10 years of followup for 112 of the 126 original trial participants found no between-treatment differences with regard to rates of relapse, adverse events, damage, survival without severe side effects, and survival without relapse and severe side effects. Thus, AZA and MTX are comparable treatment options for maintaining remission of GPA or MPA.

Mycophenolate mofetil Mycophenolate mofetil is a prodrug of mycophenolic acid that inhibits inosine monophosphate dehydrogenase required for the synthesis of DNA thereby impacting proliferating T- and B-cells (Fulton and Markham, 1996). The open-label RCT International Mycophenolate Mofetil Protocol to Reduce Outbreaks of Vasculitides (IMPROVE) Trial (Hiemstra et al., 2010) randomly assigned 156 patients to AZA starting at 2 mg/kg/day or MMF starting at 2000 mg/day after induction of remission with CYC and CS. The patients were followed for a median of 39 months. Relapses were more common in the MMF group compared with the AZA group however severe adverse events did not differ significantly between groups. Thus, among patients with AAV, MMF was less effective than azathioprine for maintaining disease remission, however both had similar adverse event rates. Between 2007 and 2013, the Clinical Trial of Mycophenolate Versus Cyclophosphamide in ANCA Vasculitis (MYCYC) (Jones et al., 2019) evaluated MMF in comparison to intravenous CYC in the induction of remission in new cases of AAV, noting that MMF was not inferior to CYC. However, the inferiority of MMF to AZA in maintenance therapy will probably reduce the use of MMF only in patients with AAV refractory to intravenous and oral CYC, and RTX. A recent comparison of guidelines and recommendations on managing AAV (Geetha

683

et al., 2018) found that patients with nonsevere and non–organ-threatening disease should be recommended a milder regimen than CYC or RTX with the British Society for Rheumatology (BSR) and British Health Professionals for Rheumatology (BHPR) (Ntatsaki et al., 2014) and EULAR (Yates et al., 2016) including CS with either MTX or MMF (grade B recommendation for MTX and grade C recommendation for MMF), where grade A is highest and D is lowest (Dougados et al., 2004).

Leflunomide The pyrimidine synthesis inhibitor LEF was evaluated in an open-label study to demonstrate improvement in disease activity and acute phase reactants with 20 mg/day of leflunomide in patients TAK who were refractory or intolerant to conventional therapy with CS and immunosuppressive agents (de Souza et al., 2012) noting that LEF was safe with steroid-sparing effect. A multicenter, prospective randomized controlled clinical trial (Metzler et al., 2007) of patients with GPA were treated either with oral LEF 30 mg/day or oral MTX (starting with 7.5 mg/week reaching 20 mg/week after 8 weeks) for 2 years following induction of remission with CYC. The primary endpoint was the incidence of relapses. The authors concluded that LEF was effective in the prevention of major relapses in GPA, however, this was associated with an increased frequency of rapidly progressive glomerulonephritis, pulmonary hemorrhage, and cerebral granuloma adverse events.

Plasma exchange Plasma exchange was initially used in systemic vasculitides considering the contributory pathogenic role of ANCA and antiglomerular basement membrane (GBM) antibodies, cryoglobulins, cytokines, and immune complexes. It is presently employed as a second-line agent in the treatment of PAN refractory to conventional regimens. Plasma exchange improves renal survival in patients with severe renal disease as defined by a serum creatinine >500 mmol/L when used as an adjunct to daily oral CYC and CS in the prospective randomized MEPEX trial (Jayne et al., 2007). A total of 137 patients with a new diagnosis of AAV confirmed by renal biopsy and serum creatinine >500 mmol/L (5.8 mg/dL) were randomly assigned to receive seven PEs or 3000 mg of intravenous methylprednisolone; both groups received oral CYC and oral prednisolone. The primary endpoint was dialysis independence at 3 months. Plasma exchange was associated with a reduction in risk for progression to end stage renal disease (ESRD) of 24% at 12 months compared to methylprednisolone (95% CI 6.1%–41%). Patient survival and severe adverse event rates at 1 year were 76% and 48% in the intravenous methylprednisolone

684

D.S. YOUNGER

group compared to 73% and 50% in the PE group. The risk reduction of 24% for ESRD with PE was clinically important in view of the cost, morbidity, and mortality of end-stage renal failure, such that the additional costs of PE were outweighed by these savings. The improvement in renal recovery rates with PE was consistent with the hypothesis that PE was most likely to be of benefit in those with the most severe disease. This study excluded patients who had been dialysis-dependent for >2 weeks because they were considered to have little chance of renal recovery. The protocol of the Plasma Exchange and Glucocorticoid Dosing in the Treatment of Anti-Neutrophil Cytoplasm Antibody Associated Vasculitis (PEXIVAS) Trial (Walsh et al., 2013) is a two-by-two factorial randomized trial begun in 2013 that has been evaluating adjunctive PE and two oral CS regimens in severe AAV. Patients receive PE or not and a standard or reduced oral CS dosing regimen. All patients receive immunosuppression with either CYC or RTX. The primary outcome is the time to the composite of all-cause mortality and endstage renal disease. The PEXIVAS study was due to report its findings in 2018. The primary composite outcome, death from any cause or end-stage renal disease occurred in 28% patients receiving PE compared to 31% in the no-PE group (hazard ratio 0.86, 95% CI 0.65–1.13; P ¼ 0.27). The primary outcome occurred in 28% of patients in the reduced CS group and 26% in the standard CS group (absolute risk difference 2.3%, 90% CI 3.4% to 8.0%), meeting the noninferiority hypothesis. Serious infections in the first year occurred less often in the reduced CS group compared to the standard group (incidence rate ratio 0.70, 95% CI 0.52–0.94; P ¼ 0.02). The investigators concluded that in the largest ever trial in AAV, a reduced dose of CS was noninferior to a “standard dose” and resulted in fewer serious infections. Levy et al. (2001) examined the long-term outcome of severe anti-GBM antibody disease who received PE, prednisolone and CYC. Those with a serum creatinine less than 5.7 mg/dL had patient and renal survival respectively of 100% and 95%, and patient and renal survival of 84% and 74% at last follow-up. Those with a creatinine 5.7 mg/dL but did not require immediate dialysis, and were treated with PE, prednisolone and CYC had comparative patient and renal survival of 83% and 82% at 1 year, and 62% and 69% at last follow-up. Patients with the anti-GBM disease and severe renal failure should be considered for urgent immunosuppression therapy, including plasma exchange, to maximize the chance of renal recovery.

Intravenous immune globulin High-dose IVIg is a safe and well-tolerated therapy compared with standard CS and immunosuppressive therapy.

Its use in systemic vasculitides was first established in the prevention of coronary artery aneurysms and in the reduction of systemic inflammation in children with KD (Furusho et al., 1984; Newburger et al., 1986). A later study Newburger et al. (1991) showed the superiority of a single infusion of IVIg as compared with four infusion in the treatment of acute KD. Improvement of patients with AAV was associated with a mean reduction in pretreatment ANCA levels by 51% in patients so treated (Jayne et al., 1991). It was alternative treatment in patients with GPA and MPA without threatened systemic organ involvement (Jayne and Lockwood, 1996), leading to full clinical remission lasting at 1 year before commencing conventional immunosuppressive therapy. A placebo-controlled trial of a single course of 2 g/kg per kg of IVIg in AAV with persistent disease activity (Jayne et al., 2000) noted reduced disease activity as judged by a reduction in the Birmingham Vasculitis Activity Score (BVAS), along with CRP and ANCA levels in subjects randomized to IVIg compared to placebo. The effect of a single course of IVIg was not maintained beyond 3 months however, and mild reversible side effects following therapy were frequent. A multicenter, prospective, open-label study (Martinez et al., 2008) for relapses of GPA and MPA during treatment or in the year following discontinuation of CS or immunosuppressive therapy led to complete remissions some lasting up to 2 years after treatment with high-dose IVIg for up to 6 months.

Rituximab Rituximab is a genetically engineered chimeric murinehuman monoclonal IgG1k that is directed against the CD20 antigen expressed on the surface of B-cells. In 2010, two randomized clinical trials, RTX in ANCAassociated vasculitides (RAVE) (Stone et al., 2010), and RTX versus CYC for ANCA-associated vasculitides (RITUXVAS) (Jones et al., 2010) provided initial RCT evidence that at 6–12 months of follow-up respectively, RTX was as safe and effective as conventional immunosuppressive therapy to control active MPA and GPA. A subgroup of 63 of 197 enrolled patients in RAVE with either GPA or MPA treated with 375 mg/m2 of body surface area (BSA)/week for four weeks, as compared with 2 mg/kg/day of CYC in those who reached the primary endpoint of remission of disease without use of prednisone at 6 months, showed that RTX was not inferior to daily CYC for induction of remission in severe AAV; and might even be superior to CYC in relapsing disease. A single course of RTX was as effective as continuous conventional immunosuppressive therapy with CYC followed by AZA for the induction and maintenance of remission of patients with severe organ-threatening

ADULT AND CHILDHOOD VASCULITIS AAV over the course of 18 months in the RAVE-ITN Trial (Specks et al., 2013). The primary outcome was complete remission of disease by 6 months, with remission maintained through 18 months. Guillevin and colleagues (2014) studied 115 patients with newly diagnosed or relapsing GPA, MPA and renal-limited AAV in complete remission after a CYC-CS regimen who were randomly assigned to receive either 500 mg of RTX on days 0 and 14 and at months 6, 12, and 18 after study entry or daily AZA until month 22. The primary end point at month 28 was the rate of major relapse (the reappearance of disease activity or worsening, with a Birmingham Vasculitis Activity Score > 0, and involvement of one or more major organs, diseaserelated life-threatening events, or both). More patients with AAV had sustained remission at month 28 with RTX than with AZA. At month 28, major relapses had occurred in 29% of the AZA group compared to 5% in the RTX group, with similar frequencies of severe adverse events. A 24-month phase III randomised-controlled trial of 115 patients over time who received RTX or AZA for AAV maintenance therapy and completed a Health Assessment Questionnaire (HAQ), scores (Pugnet et al., 2016) showed mean improvements from baseline to month 24 that were significantly better for the RTX than the azathioprine group. Zaja et al. (2003) found that RTX was a safe and effective alternative to standard immunosuppressive therapy of type II mixed cryoglobulinemia (MC). Sansonno et al. (2003) studied patients with MC and HCV-positive chronic active liver disease resistant to INFa therapy noting that 80% of patients treated with an intravenous infusion of 375 mg/m2 of RTX once a week for four consecutive weeks showed a complete response characterized by rapid improvement of clinical signs and decline of the cryocrit and anti-HCV antibody titers. Roccatello and colleagues (2004) found that RTX was a safe and effective option in symptomatic patients with HCV-associated MC and glomerulonephritis and signs of systemic vasculitis. Saadoun et al. (2008) found that 94% of patients with severe refractory HCV-related MC vasculitis showed clinical improvement, 63% of who were complete responders with undetectable HCV RNA and serum cryoglobulins. Terrier et al. (2009) found that RTX combined with PEG-INFa-2b plus ribavirin (RBV) induced a complete and partial clinical response respectively in 80% and 15%; a complete and partial immunologic response was respectively noted in 67% and 33% of patients, and a sustained virologic response in 55% of patients so treated, making RTX combined with PEG-INFa-2b plus RBV safe and effective treatment in severe refractory HCV-associated mixed CV. Ignatova et al. (2017) studied the efficiency of traditional CS and CYC and selective RTX therapy of HCV-associated CV over an average follow-up period of 2.8 years noting

685

that combined therapy of RTX and antiviral therapy was most effective in patients with severe forms of vasculitis.

Anti-TNFa The anti-TNFa monoclonal antibody infliximab or analogue of its receptor, etanercept, has been proposed to treat primary systemic vasculitides. Infliximab, a chimeric anti-TNFa monoclonal antibody in combination with conventional therapy led to clinical remission in 88% of the patients with acute or persistently active AAV enrolled in an open, prospective trial (Booth et al., 2004). In 2002, Bartolucci et al. (2002) reported their findings of einfliximab treatment in 7 patients with severe refractory GPA all of whom obtained complete or partial remissions, with cutaneous eruption being the only adverse effect. Etanercept, another TNFa blocker, comprised of a soluble protein derived from the p75 TNF receptor fused to the Fc portion of IgG, has been tested in AAV and in conjunction with conventional therapy to reduce relapse rate. The WGET trial (Wegener’s Granulomatosis Etanercept Trial Research, 2005) compared etanercept to placebo in addition to receiving standard therapy of CS plus CYC or MTX. After sustained remission the primary outcome was defined as a BVAS of 0 for at least 6 months and tapering of standard medications according to an established protocol. Of 174 patients, 72% had a sustained remission however there were no significant differences between the entanercept and control groups in the rates of sustained remission (69% versus 75%, P ¼ 0.39) or in the relative risk of disease flares per 100 person-years of follow-up. The RATTRAP trial (de Menthon et al., 2011), which compared efficacy and tolerance of infliximab versus RTX to treat refractory GPA, demonstrated the usefulness of infliximab to obtain remission of refractory GPA with a trend at 12 months favoring RTX. During long-term follow-up, RTX was better able at obtaining and maintaining remission. Seror et al. (2014) studied the effect of adding a ten week course of 40 mg on alternate weeks of subcutaneous adalimumab for ten weeks to a standard treatment of 0.7 mg/kg/day of prednisone, with a primary end point of the percentage of patients in remission on less than 0.1 mg/kg of prednisone at week 26. Among near equal numbers of study and control subjects, there was no difference between the adalimumab compared to prednisone in increasing the number of patients in remission on less than 0.1 mg/kg of CS at 6 months. Mekinian and others (2012), who reported the findings of a multicenter retrospective study tolerance study of infliximab in refractory TAK, noted a significant decrease in clinical biological activities within 3 months, with a fall in the median CS dose at 12 months.

686

D.S. YOUNGER

Tocilizumab Tocilizumab, a humanized monoclonal antibody against the interleukin (IL)-6 receptor, was evaluated in retrospective series in patients with refractory large vessel vasculitides demonstrating evidence clinical and serologic improvement in patients with refractory and relapsing disease. The multicenter, randomized double-blind, placebo-controlled study, GiACTA (Stone et al., 2017) randomly assigned 251 patients, in a 2:1:1:1 ratio, to receive subcutaneous tocilizumab (at a dose of 162 mg) weekly or every other week, combined with a 26-week prednisone taper, or placebo combined with a prednisone taper over a period of either 26 weeks or 52 weeks. The primary outcome was the rate of sustained glucocorticoid-free remission at week 52 in each tocilizumab group as compared with the rate in the placebo group that underwent the 26-week prednisone taper. The key secondary outcome was the rate of remission in each tocilizumab group as compared with the placebo group that underwent the 52-week prednisone taper. Tocilizumab, received weekly or every other week, combined with a 26-week prednisone taper was superior to either 26-week or 52-week prednisone tapering plus placebo with regard to sustained glucocorticoid-free remission in patients with giant-cell arteritis. Longer follow-up is necessary to determine the durability of remission and safety of tocilizumab.

Insights into treatment RANDOMIZED CLINICAL TRIALS The past 20 years has witnessed tremendous progress in the conduct of clinical trials in vasculitis. This progress has occurred due to several factors. First, the widespread adoption of standardized definitions (Jennette et al., 2013) and classification criteria (Hunder et al., 1990; Watts and Scott, 2009) for the vasculitides, leading to greater standardization of eligibility criteria. Second, the use of validated outcome measures for vasculitis (Direskeneli et al., 2011) beginning with the Birmingham Vasculitis Activity Score (BVAS) (Luqmani et al., 1994) for a weighted score, based on symptoms and signs of disease activity due to vasculitis in nine separate organ systems, and other validated tools such as the five factor score (FFS) (Guillevin et al., 2011) to evaluate prognosis at diagnosis and direct the therapeutic choices in necrotizing vasculitis (Cohen et al., 2007; Ribi et al., 2008). However, many forms of vasculitis still lack well-validated measures of disease activity or state for use in clinical trials including primary CNS and PNS vasculitides. The Vasculitis Working

Group of the Outcome Measures in Rheumatology (OMERACT) initiative has been pursuing a variety of projects to advance development of valid measures in multiple forms of vasculitis (Merkel et al., 2014). Third, the establishment of national and international research networks to facilitate collaborations between academic institutions with expertise in clinical care and research in vasculitis within which many of the most significant multicenter trials in vasculitis have been conducted by such groups or the core personnel within them.

ANCA-ASSOCIATED VASCULITIDES The treatment AAV notably in GPA and MPA has evolved over the last four decades from CS alone to extended use of CYC (Fauci and Wolff, 1973) to several different treatment protocols based on disease severity, disease stage, and including substantially reduced cumulative doses of CYC or the avoidance of this medication altogether (Mukhtyar et al., 2009). For purposes of treatment, severe disease flare in AAV has been defined as the presence of active disease manifestations that are lifethreatening and organ-threatening. Current and future treatment strategies in AAV are addressing optimization of existing therapies and the introduction of novel agents. The approach to EGPA differs from other AAVs due to several factors including recognition that many patients have a monocyclic course, identification of clinical prognostic factors influencing course of treatment, and the impact of asthma on the clinical course will not be considered. However, the main principles can be summarized as follows. Corticosteroids are the mainstay of therapy for EGPA and are included in all treatment regimens and are often the sole treatment for cases without poor-prognosis factors (FFS ¼ 0) (Ribi et al., 2008), or in combination with intravenous CYC when the FFS 1. In a trial of superiority of six versus twelve pulses of CYC (600 mg/m2 per pulse) in patients with a poor prognosis, defined as a FFS 1, the longer duration of treatment exceeded shorter treatment for control of disease (Cohen et al., 2007). Patients treated with CS alone had more frequent relapses suggesting the need for a revision in this treatment approach. A RCT of AZA and CS as first-line treatment in EGPA without factors mitigating poor-prognosis demonstrated no additional benefits for remission rates, relapse risk, sparing of CS, or diminishing the EGPA asthma/rhinosinusitis exacerbation rate over CS alone. Although the trials of EGPA mainly involve use of intravenous CYC, many centers will apply the treatment regimens used for GPA and MPA for patients with severe EGPA and prescribe oral CYC for several months, transitioning afterward to AZA or MTX for remission maintenance;

ADULT AND CHILDHOOD VASCULITIS in fact, MTX combined with CS is a regularly used remission-induction regimens to treat nonsevere EGPA (Metzler et al., 2004). With a greater understanding in the disease pathogenesis, and the advent of highly specific biological molecules, a targeted approach is now possible, the ultimate goal of which is to control the underlying disease process while eliminating or minimizing disease side effects associated with broad spectrum immunosuppression. Although the desire to minimize CYC exposure has dominated clinical studies for the last 20 years, CYC remains a relatively safe and effective induction agent in AAV, and is a component of the standard of care in consensus guidelines for the treatment of generalized disease (Mukhtyar et al., 2009). Between 2003 and 2009, three notable adjustments in the administration of CYC in clinical trials demonstrated that it could be given more safely. Frist, the sequential replacement of CYC by AZA once remission was achieved as in the CYCAZAREM study (Jayne et al., 2003). Second, the replacement of CYC by MTX for early systemic disease without critical organ manifestations in the NORAM study (De Groot et al., 2005). Third, the use of pulsed intravenous CYC with dose reductions for those aged over 60 years and renal impairment rather than daily oral CYC in the CYCLOPS study (de Groot et al., 2009), enabling a cumulative dose reduction of approximately one-half.). In long-term follow-up of patients in the CYCLOPS study (Harper et al., 2012) reduced CYC was associated with a higher risk of relapse, while MTX was associated with less effective disease control than CYC induction therapy in the NORAM study (Faurschou et al., 2012). In neither however were long-term morbidity and mortality increased. Induction therapy for severe initial presentations or flares and relapses of disease in AAV currently involves the use of CS in combination with a choice of CYC or RTX. Two clinical trials compared CYC to RTX for remission-induction and found these two treatments to have similar efficacy in inducing remission when used in combination with CS (Stone et al., 2010; Jones et al., 2010). The standard dose of oral CYC for remissioninduction of AAV is 2 mg/kg/day for 3–6 months. Pulse intravenous CYC therapy is a reasonable alternative to oral administration. A randomized trial (the “CYCLOPS” study) demonstrated equivalence of the two routes of administration of CYC for remission-induction (de Groot et al., 2009) and the use of intravenous CYC has been supported by other studies (de Groot et al., 2001). However, the dosing of CYC in the “CYCLOPS” regimen was 15 mg/kg every 2 weeks for 3 infusions then once every 3 weeks. The duration of CYC in the major trials was 4–6 months prior to transition to another agent. The dose

687

of oral or intravenous CYC should always be adjusted down among patients with renal insufficiency, in response to leukopenia, and in older patients. The cumulative exposure to CS and immunosuppressive drugs that contributes to organ damage, raised concerns of CYC-related toxicity involving chronic myelosuppression, infection, urothelial malignancy, and infertility (Hoffman et al., 1992; Talar-Williams et al., 1996). Remission-maintenance treatment in AAV following achievement of remission with CYC involves the use of intravenous RTX 500 mg every 6 months. Azathioprine at the dose of 2 mg/kg/day or MTX at doses of 20 to 25 mg/week can also be used for at least 1–2 years after CYC and often much longer or indefinitely for patients at high risk of relapse (Jayne et al., 2003; De Groot et al., 2005). One RCT found no difference in the efficacy of AZA and MTX for maintaining remission (Pagnoux et al., 2008). The choice between these two agents depends on individual patient factors and preferences; methotrexate should not be used by patients with renal insufficiency. The IMPROVE trial in AAV (Hiemstra et al., 2010) showed that following induction of remission with CYC, AZA was superior to MMF for remission-maintenance making the latter a third choice for this indication, after azathioprine and methotrexate. The equivalence of RTX to CYC in the induction of remission of AAV (Stone et al., 2010; Jones et al., 2010), and the licensing of RTX for treatment of AAV have been significant achievements. The induction regimen used in the trials was 375 mg/m2 given intravenously weekly for 4 consecutive weeks. Rituximab is a reasonable alternative to CYC for treatment of severe AAV. However, the high cost of this drug has resulted in wide variation internationally in the use of rituximab. In some countries it is a first-line treatment option, especially to treat younger patients seeking to preserve their fertility. In many countries use of rituximab is restricted to patients with relapsing disease, especially those who have had prior courses of cyclophosphamide who comprise a subset of patients for whom rituximab was found superior to CYC in the RAVE trial (Specks et al., 2013). The optimal treatment strategy following RTX induction still needs to be addressed. In the RAVE trial (Specks et al., 2013) one course of RTX was equally efficacious in maintaining remission as 18 months of CYC and AZA. Single-center retrospective cohort studies provided evidence that repeated doses of RTX helped maintain remission in many patients (Cartin-Ceba et al., 2012; Smith et al., 2012). However, due to the expense of RTX and concerns about the impact of repeated dosing and hypogammaglobulinemia, consideration has since been given to using more standard immunosuppressive agents such as AZA or MTX following RTX.

688

D.S. YOUNGER

Different treatment strategies for maintenance of remission in AAV after induction with rituximab are currently being studied in a multinational clinical trial (RITAZAREM) (ClinicalTrials.gov Identifiers: NCT01697267). Rates of cardiovascular disease in AAV that increase such as accelerated atherosclerosis (de Leeuw et al., 2005) raised concerns that there may be nontraditional risk factors such as endothelial activation and excessive vascular remodeling to take into account as predominant causes of death, rather than uncontrolled vasculitis (Little et al., 2010). It is uncertain whether increasing use of RTX can remedy these unforeseen problems. The role of PE in the treatment of AAV has been highly controversial. In one RCT of patients with AAV and severe rapidly-progressive renal impairment at diagnosis, the addition of PE reduced the frequency of progression to end-stage renal disease but not overall mortality (Jayne et al., 2007). Other studies have not clearly demonstrated the benefit of PE in AAV (Walsh et al., 2011), including for the treatment of alveolar hemorrhage. A recent large multinational randomized clinical trial involving 704 patients with GPA and MPA (Walsh et al., 2013) found that PE did not reduce the rate of end-stage renal disease or death. Adjunctive therapy with a single course of high-dose IVIg therapy reduced disease activity in persistent AAV but this affect was not maintained beyond 3 months making it an alternative treatment for persistent disease only after standard therapy (Jayne et al., 2000). The optimal duration of therapy for remission maintenance in GPA and MPA remains unknown. However, the trend has been toward longer courses of therapy with the remission-maintenance agent often continued for at least 12–18 months for treatment of new-onset disease and even longer or indefinitely for relapsing disease. Affected patients that demonstrate ANCA with specificity to proteinase 3 have substantially higher rates of relapse than those patients positive for antibodies to myeloperoxidase; relapse in AAV is itself predictive of future relapse (Specks et al., 2013). Insights into disease pathogenesis have also influenced the approaches to treatment. Genetic susceptibility and environmental exposures are now known to contribute to the autoimmune etiopathogenesis of AAV. Animal models of AAV show that the transfer of murine MPOANCA IgG without functioning B- or T-cells results in a pauci immune, necrotizing crescentic glomerulonephritis similar to that seen in AAV in humans (Xiao et al., 2002). There are also several lines of investigation linking infections to ANCA formation through molecular mimicry (Little et al., 2012). Fimbriated bacteria induce novel ANCA antibodies to human lysosome membrane protein-2 (LAMP-2), which in turn leads to

crescentic glomerulonephritis in animals (Kain et al., 2008) and infection with Staphylococcus aureus is associated with relapse of GPA (Popa et al., 2002b). Proteinase 3 ANCA binding levels are predictive of outcome with a rise in antibody titers before clinical relapse. However, patients who are consistently ANCA-seronegative fit the clinical phenotype of AAV and the efficacy of B-cell depletion with rituximab is not associated with ANCA status. Despite the “pauci immune” nature of the histology in ANCA vasculitis, there is increasing evidence for the role of both cell- and humoral immunity with immune complex deposition and complement activation in renal involvement. Components of the alternative complement pathway are detected in glomeruli and small blood vessels in kidney biopsy tissue specimens from patients with AAV which colocalize with C3d and the membrane attack complex (MAC) (Xing et al., 2009). The contribution of cell-mediated immunity is exemplified by activation of circulating T- and B-cells, with infiltration of plasmoblasts into affected tissues (Popa et al., 2002a). Autoreactive B-cells, necessary for the development of autoantibody producing cells, appear to play the important roles of supporting autoreactive T-cell activity through antigen presentation, costimulation, and direct production of proinflammatory cytokines including IL-6 and TNFa. In view of their role as precursors of ANCA secreting plasma cells, B-cells are a therapeutic target in AAVand T-cells play an important role in the eventual pathogenesis of AAV (Weyand and Goronzy, 2003). Class-switched, IgG autoreactive antibodies receive cognate T-cell help, and T-cells cause damage via direct cytotoxicity and recruitment and activation of macrophages (Lamprecht, 2005). However, early outcome results for the treatment of childhood AAV, in particular GPA, reported by Morishita and colleagues on behalf of ARCHiVe Investigators Network and the Pediatric Vasculitis (PedVas) Initiative (Morishita et al., 2017) was less encouraging. Among 105 children with AAV, mainly GPA, who received CS, CYC, MTX, or RTX for remission-induction, and PE in conjunction with CYC and/or RTX, 42% achieved remission at 12 months (Pediatric Vasculitis Activity Score [PVAS] of 0, CS dose 7 days of mechanical ventilation in medical and surgical ICUs (Sharshar et al., 2009) employing the Medical Research Council (MRC) score (from 0 to 60) (Kleyweg et al., 1991) adjusted for patients with only 3 or 2 accessible limbs (Garnacho-Montero et al., 2001; De Jonghe et al., 2002) to score the severity of weakness at the time of awakening. Their results showed that nonsurvivors had significantly lower scores and higher rates of ICU paresis, and higher hospital and ICU mortality after multivariate risk adjustment. This affirmed the importance of the ICUAW and the urgent necessity of uncovering new interventions and strategies to reduce the short- and long-term mortality. The most urgent call for understanding of ICUAW accompanied the severe acute respiratory syndrome-coronavirus-2 (SARS-nCoV-2) 2019 disease (COVID-19) pandemic coronavirus-2 infection that was associated with prolonged invasive ventilation and consequential morbidity and mortality due to direct infection and a pronounced systemic inflammatory immune response syndrome (SIRS) referred to as a cytokine storm (Younger, 2021a, 2021b). It was unsurprising that 69% of COVID-19 ICU survivors were found to have significant limb and respiratory weakness in a cross-sectional study of 2 tertiary hospitals, of whom 44% were unable to walk 100 m after ventilator weaning (Zhou et al., 2020).

NOSOLOGY Of the many descriptive terms used to identify motor syndromes in critically ill patients (entering, residing in, or emerging from the ICU), critical illness–associated weakness (CI-AW or CIAW) seems to be the most useful (Younger and Bolton, 2005) reflecting a single starting

710

D.S. YOUNGER

point for clinicians in assessing a given patient’s weakness from among the spectrum of causative disorders from all levels in the peripheral nervous system (PNS) and across the motor unit (MU) (Vianello et al., 2022). Moreover, the authors preference for this nosology (Younger and Bolton, 2005) reflected the difficulty in separating the influence of the ICU due to the overlap with competing etiopathophysiological processes (Latronico, 2003) and the potential flux that came with redefining causation due to CIP and CIM after more extensive electrophysiological evaluation (Trojaborg et al., 2001). In addition, ICUAW appears to be a dynamic process affecting cohorts and individuals differentially as shown in a recent systematic review and meta-analysis of 12 prospective cohort studies published from 2005 to 2021 (Yang et al., 2022). The authors noted a multifactorial etiology of ICUAW associated with female sex, days of mechanical ventilation, older age, longer length of ICU stay, an infectious illness, use of renal replacement therapy or aminoglycoside drugs, the sepsis related organ failure assessment (SOFA) score, and hyperglycemia, but inconsistent evidence of corticosteroids or neuromuscular blockers, or sepsis. Employing ICUAW or ICU-associated paresis (AP) as an end-point for prospective randomized studies may have the appeal of identifying patients with CIP and CIM and related diseases states (Kress and Hall, 2014) especially after the exclusion of subjects with deconditioning and dysfunction that appears to be diminishing with acceptance and implementation of evidencebased strategies of early mobilization in the ICU (Bailey et al., 2007). However, prospective randomized clinical trials (RCTs) scarcely show clear cause-and-effect relationships for a given disease trigger (Schweickert et al., 2009; Farhan et al., 2016), suggesting at least for the present, that ICUAW may be a marker for the severity of illness or an independent covariable that contributes to eventual adverse clinical outcomes, although the truth most probably lies somewhere in between (Van der Jagt and Kompanje, 2017).

NEUROMUSCULAR ASSESSMENT History and physical examination If an affected child or adult patient is alert and able to write, an adequate history can be obtained by having the patient respond by handwriting. It is important to consider preexisting causes of weakness when considering a patient with ICUAW (Stevens et al., 2009). Patients or relatives should be asked about a previous history of neuromuscular disease. Spinal and radicular pain, bulbar and limb weakness, breathing difficulty, impaired sensation, bladder dysfunction, muscle cramping, fasciculation, and fatigability are all useful clues to an underlying

neuromuscular disorder that may follow a major surgical procedure, for example, surgery on the thoracic aorta that may accompany spinal cord ischemia and resulting paraplegia. Neuromuscular blocking agents, even shortacting ones such as vecuronium, can have an action of several hours in the setting of renal failure. It is important to recognize the reasons for intubation and mechanical ventilation, whether for airway protection or weakness of the respiratory muscles, and the type of mechanical ventilation, frequency of intermittent mandatory ventilation, degree of pressure support, blood gas results, and the ability to breathe while off the ventilator. General inspection of the skin may reveal localized edema and redness at sites of compression that are likely to occur in patients with drug-induced coma lasting hours. Pressure at the site of an underlying peripheral nerve can cause mononeuropathy. Signs of sympathetic insufficiency such as increased temperature, redness, and dry skin in a paretic lower limb compared with the opposite limb suggest a lesion of the lumbar plexus because sympathetic fibers bypass the caudal nerve roots. Ptosis, asymmetric ocular palsies, and facial weakness require exclusion of a neuromuscular transmission defect. The response to a tracheal tug on the endotracheal tube assesses the ability to swallow and cough. Abnormalities of this type are seen in motor neuron disease (MND), GBS, and neuromuscular junction (NMJ) disorders. Generalized or focal muscle wasting may be difficult to determine in the presence of limb edema. An affected patient can have voluntary or partial voluntary movements in bulbar and limb muscles in a pattern that suggests hemiplegia, quadriplegia, or paraplegia. If there are no such movements, compression of the nail beds with a pencil may induce one of several involuntary motor responses. Simple flexion movement of the stimulated ipsilateral or contralateral limbs occurs on a reflex basis at the level of the spinal cord level and even in brain death. More complex movements of a voluntary nature require intact cerebral function. If the opposite hand accurately moves in a coordinated way to attempt to remove the painful stimulus, reasonable function of that limb must be present. In addition, the patterns of limb movement may offer further clues as to the presence or absence of coexisting neuromuscular disease. Painful stimulation that induces vigorous facial grimacing without limb movement implies normal afferent and efferent cranial conduction of the painful stimulus along the peripheral nervous system (PNS) and CNS pathways. Similar findings are often seen in high cervical cord lesions, but because severe pain impulses are not transmitted through the spinal cord, facial grimacing is not present. Vigorous upward movement of the great toe on plantar stimulation may be absent on the side of a peroneal nerve palsy; similarly, there may be focal absence of deep tendon reflexes

CRITICAL ILLNESS–ASSOCIATED WEAKNESS AND RELATED MOTOR DISORDERS (DTRs) suggesting lumbosacral plexopathy with particular involvement of the femoral nerve.

Laboratory evaluation The laboratory evaluation of a patient with critical illness weakness should be guided by the presenting clinical features and likeliest diagnoses. In addition to routine ICU studies, the consulting neurologist or neurointensivist may elect to perform brain, spinal cord, regional or body neuroimaging, as well as a routine serum CK, electrophysiological studies, pulmonary function tests (PFTs), dynamic evaluation of the diaphragm, and nerve and muscle biopsy, depending upon the presumptive diagnosis and available technical expertise.

NEUROIMAGING Brain, spinal cord, regional body and whole-body magnetic resonance imaging (MRI) are widely used modalities in patients with critical illness motor disorders to facilitate a diagnosis and formulate a treatment plan. The intravenous contrast agent gadolinium crosses the blood–brain barrier (BBB) and is associated with few side effects. It shortens T1 and T2 relaxation times of spin-echo images and accumulates in lesions as areas of increased signal intensity compared with precontrast images. Spinal MRI has supplanted myelography in the evaluation of patients with spinal cord disorders. Muscle imaging (myo-MRI) has been used to image the cross-sectional planes of individual limbs both at sites of focal muscle wasting due to active myopathy, as has MRI neurography (MRN) to detect abnormalities along selected nerves of the lumbosacral plexus. All patients with critical illness weakness should undergo MRI neuroimaging of the neuroaxis including those with encephalopathy. Whole-body or regional muscle MRI employing short tau inversion recovery (STIR) images has emerged as a logical extension of this diagnostic modality in patients with critical illness weakness to examine the upper legs, which are otherwise difficult to assess by conventional EMG and nerve biopsy. Maramattom (2022) compared whole body MRI in 7 cases each of ICU-acquired weakness with mixed myopathic and axonal sensorimotor features and noted diffuse and extensive muscle edema in all 7 (100%) patients compared to 1 of 7 (14%) patients with GBS and demyelinating features on electrodiagnostic studies in the third week of illness. Chhabra et al. (2011) suggested MRN employing 3 T, high-resolution multiplanar structural sequences to image both myopathic and neuropathic changes in pelvifemoral regions and along particular nerve distributions in patients with CIAW. Standardized approaches to ultrasonography using special algorithms have been developed for the bedside

711

evaluation of ICUAW (Gruber et al., 2022) obtaining images of single-nerve cross-sectional areas and summing score values to differentiate pure CIM from CIP. Neuromuscular ultrasound incorporating indices of muscle thickness, echo intensity and homogeneity, as well as nerve cross-sectional area, thickness and vascularization of the area under the curve of the receiver operating characteristic curve (ROC-AUC) were employed in a cohort of ICUAW on the 9th day of ICU hospitalization and mechanically ventilated for at least 48 h (Witteveen et al., 2017). Their findings showed a relatively low diagnostic accuracy of 51%–68% for muscle parameters, and 51%–66% for nerve parameters, making neuromuscular ultrasound an unreliable modality relatively early in the course of the illness and certainly not at the time an affected patient first awakened.

ELECTROPHYSIOLOGICAL STUDIES Electrophysiological studies are important in the evaluation of patients with CIAW and ICUAW (Latronico et al., 2009) to investigate suspected myopathy, NMJ disorders, peripheral neuropathy, entrapment neuropathies, plexus and root disorders. Nerve conduction studies Serial nerve conduction studies (NCS) and conventional EMG should be obtained in all patients using standard techniques (Ferrante, 2022) referenced to published normative (Rosenfalck and Rosenfalck, 1975) or reference values for the individual laboratory. Although electrophysiological studies should ideally be timed to allow for maturation of the underlying lesions, neither should it be postponed so as to delay timely diagnosis. Some experts advocate them as soon as 72 h after the onset of the ICU hospitalization to ascertain very early features of inexcitability (Khan et al., 2006), with plans to repeat them if there are normal findings (Argov and Latronico, 2014). Only precipitous weakness will be promptly recognized by physicians and staff in the ICU, whereas most other affected patients will certainly have a less well documented onset and duration of their neuromuscular condition as a result of prolonged unconsciousness and limb restraints. A simplified electrophysiologic algorithm that evaluated the peroneal (fibular) nerve showed initial promise as a rapid, highly sensitive diagnostic test for CIP (Latronico et al., 2007), however it did not distinguish between neuropathy and myopathy particularly at the outset since both disorders may be characterized by low amplitude CMAPs and sparse spontaneous activity in the form of fibrillation potentials and positive sharp waves (PSWs). Moreover, the differentiating feature may rest on the presence of normal SNAPs, while are unreliable in the presence of leg edema.

712

D.S. YOUNGER

Technical challenges to electrophysiological studies arise in the ICU due to electrical interference from adjacent machines, poorly grounded plug-ins, inadequate shielding of cables, and other electrical devices. The skin should be adequately prepared to reduce resistance. The 60-cycle notch filter on the EMG machine may have to be used. It may not be possible to electrically stimulate certain nerves due to the presence of intravascular lines, surgical wounds and dressings, casts, and splints. Cardiac arrhythmias may be induced if an electrical stimulus is applied along a limb with an intravascular line that resides distally in the heart, in which case it may be wise to test the opposite side. The ground electrode should be on the same side of the body that is being stimulated to avoid transmission of the electrical impulse through the heart. Virtually all EMG studies including those of the diaphragm are quite safe. In our experience, these various technical challenges can be easily overcome, and a complete electrophysiologic assessment can almost always be achieved. Repetitive nerve stimulation Repetitive nerve stimulation (RNS) at distal and proximal sites is potentially useful in the evaluation of severely weak ICU patients to disclose a NMJ disorder (Huijbers et al., 2022). Such disorders present in the ICU with variable fluctuating or fixed weakness involving limb and bulbar weakness, often with impending or frank respiratory failure (Birch, 2021). Abnormal findings with varying frequencies (Hz) of RNS result from the loss of the safety factor for NMJ transmission. Diagnostic testing may be flawed in weak patients due to failure to actively participate with even brief exercise. A decremental response of 12%–15% or more of successive CMAP after 3-Hz stimulation with aggravation of the block for several minutes after brief exercise, is highly indicative of a postsynaptic defect typical of MG. Maximal exercise after 15 s causes transient improvement of the decrement termed postactivation facilitation as does an injection of edrophonium chloride. Prolonged exercise for 1 min if possible, followed by repetitive trains of nerve stimulation at 1-min intervals, worsens the block, termed postactivation exhaustion. Several diseases affect the NMJ leading to severe weakness. Ingested toxin of Clostridium difficile in contaminated foods results in binding of receptors along the terminal axon with incipient clinical and electrophysiological manifestations of botulism. Lambert–Eaton myasthenic syndrome (LEMS) a rare autoimmune presynaptic disorder commonly associated with small cell lung cancer (SCLC) that results from the blocking action of antibodies directed against presynaptic P/Q-type voltage-gated calcium channels (VGCCs) (Schoser et al., 2017). Both

botulism and LEMS show low-amplitude CMAP on conventional NCS, which increases by 100%–200% after 20 Hz or more RNS, indicative of the underlying presynaptic defect. Neuromuscular junction blockade by nondepolarizing neuromuscular blocking drugs (NDNMBDs) that cause slowly reversible paralysis due to antagonisms of acetylcholine (ACh) binding at ACh receptors (AChRs) can be diagnosed and monitored by RNS at 2–3 Hz (Fig. 29.2).

Needle electromyography After completion of NCS, a needle examination is performed using a concentric needle electrode referenced to the cannula, or a monopolar needle insulated with Teflon and referenced to a surface electrode at a frequency setting of 2–10k Hz leads. The primary aim of the EMG examination is to determine whether weakness is due to myopathy or to neurogenic lesion. Appropriately chosen proximal and distal weak muscles are examined at rest for spontaneous muscle fiber activity at rest. In control and affected muscle, the end-plate zone shows miniature EPP and spontaneous muscle fiber discharges with a sharp negative onset. Outside the end-plate zone, propagated action potentials are rarely recorded

Fig. 29.2. Top: Example of tracings from serial neuromuscular transmission studies in the ulnar motor territory (2 Hz stimulation) of a patient with a prolonged neuromuscular transmission deficit following vecuronium use. Bottom: Note that the CMAP (compound muscle action potential) is also reduced in amplitude in the recovery trace. The patient subsequently was noted to have evidence of CIM when the neuromuscular transmission deficit cleared. Reproduced from Koshy K, Zochodne DW (2013). Neuromuscular complications of critical illness. Handb Clin Neurol 115: 759–780, with permission.

CRITICAL ILLNESS–ASSOCIATED WEAKNESS AND RELATED MOTOR DISORDERS in normal muscle. In myopathy, fibrillation activity is found when there is muscle fiber necrosis associated with myositis and some types of muscular dystrophy, rarely in mitochondrial myopathy and thyrotoxicosis, whereas denervation activity indicative of failure of reinnervation is seen in neurogenic disorders, which may endure for some time after the injury has occurred as in poliomyelitis. Fibrillation potentials, PSW activity, and complex repetitive discharges (CRD) arise from groups of myofibers in both myopathic and neurogenic lesions. Fasciculation potentials, recognized as irregular discharges of groups of muscle fibers that can occur in normal muscle and in those with myositis and neurogenic lesions, have longer discharge intervals. Specialized electrophysiologic studies A solution to increasing the yield of conventional electrophysiology in separating myopathy and neuropathy and therefore CIM from CIP has been to add other more specialized electrophysiological tests including: CMAP waveform analysis, QMUP, measurement of muscle fiber excitability by DMS, and MUNE. Not only does each technique require a higher level of performance and interpretative expertise, but as suggested by Trojaborg (2006), they have further value when incorporated as a diagnostic strategy for the electrophysiological evaluation of ICUAW. Analyze compound muscle action potential waveforms. This important overlooked electrophysiological measure to separate myopathy from neuropathy was suggested in a 1997 Case Study of the Massachusetts General Hospital “Weekly Clinicopathological Exercise” (1997b). The patient in question suffered from chronic obstructive pulmonary disease and was hospitalized for respiratory failure and longstanding history of alcoholic abuse. After a 1-month stay in the hospital, he became tetraplegic with absent deep tendon reflexes. Nerve conduction studies showed low amplitude CMAPs and marked prolongation of the muscle responses without signs of desynchronization. Motor nerve conduction velocities (NCV) and F-wave responses were normal as were all sensory-evoked responses. EMG revealed no spontaneous activity. Despite profound weakness, a full recruitment pattern was readily obtained. The distal CMAP duration on the 34th hospital day was 9.4 and 10.0 ms for the abductor pollicis brevis and adductor digiti minimi, respectively, and increased to 12.7 and 14.0 ms a month or so later. When muscle strength improved, amplitudes returned to normal followed by gradual correction of the dispersion of the CMAPs. The clinical, laboratory and electrophysiological features of the case were compatible with CIM-associated myosin deficiency. Biopsy of the right deltoid muscle showed marked diffuse fiber

713

atrophy with numerous small, angulated fibers, and electron microscopy (EM) revealed a striking loss of thick myosin filaments compatible with critical illness myopathy. The dispersion of the CMAP could not be explained by the histological findings but was likely related to mechanisms affecting excitable membranes either in the distal part of the motor axons or the sarcolemma as no evidence of demyelination of intramuscular motor axons was noted. It was suggested that dispersion could be caused by slowing of MFCV as a result of impairment or blockade of certain voltagegated ion channels. Park et al. (2004) later reviewed 9 cases examined in the ICU between 1999 and 2003 with possible CIM diagnosed by electrophysiological evaluations. The CMAPs evoked by stimulation of the median, ulnar, peroneal, and tibial nerves showed markedly reduced amplitude and increased CMAP negative phase duration, the highest value being more than 300% of the upper limit of normal; however, the waveforms remained synchronous and smooth. Moreover, CMAP duration was not significantly different between proximal and distal stimulation sites. The authors concluded that determination of the CMAP duration was a quick and simple diagnostic bedside test preferable to other electrophysiological tests to distinguish CIM from CIP. Quantitative motor unit potential analysis. To avoid errors in the diagnosis of myopathy, it is important to analyze the mean duration of simple MUPs using a standardized sampling technique (Stalberg et al., 1994; Simonetti et al., 2022). Twenty different MUPs are collected by random insertion of a concentric needle electrode into several regions of the muscle under examination. The mean duration and amplitude of the collected MUP from each muscle are compared to normal age matched values, and the duration expressed as a percentage deviation from normal age-matched values for all collected MUP, with even greater accuracy obtained by selection of nonpolyphasic MUP for analysis. Mild disease and heterogeneous affection of the muscle by myositis and monoradicular disease can lead to outlier abnormal mean duration and amplitude values (Stalberg et al., 1994). It is important to obtain signals uncontaminated by activity from adjacent MUPs and surrounding artifacts especially when in the ICU and to control for temperature. Analysis of MUPs at weak effort has a central position in the EMG examination because the duration, amplitude, and shape of the MUAP recorded with a concentric or a monopolar needle electrode reflects the architecture of the motor unit. Trojaborg et al. (2001) used previously described methods (Trojaborg, 1990) to sample 20 MUPs in a cohort of patients with CIAW, from which they calculated mean

714

D.S. YOUNGER

duration and amplitude of the collected MUPs from each muscle and compared to normal age-matched values (Rosenfalck and Rosenfalck, 1975) expressed as a percentage deviation from normal age-matched values for all collected MUPs. The number of motor units was estimated using a standard technique (Daube, 1995), and modification of the original technique described by McComas et al. (1971) in the biceps brachii, tibialis anterior, abductor pollicis brevis (APB), or abductor digiti minimi (ADM). In their cohort of CIAW, in all individual muscle tested, the mean MUP duration was 0.5 (Rich et al., 1997). Evoked response amplitude and muscle fiber conduction velocity are also measured such that CMAP amplitudes evoked by DMS 0.5 in all, indicative of myopathy. In two other cases, the muscle was inexcitable following DMS using subdermal electrodes, although a response was obtained with a concentric needle electrode. The authors advocated DMS especially in the absence of adequate voluntary MUPs for analysis. Among 30 consecutive patients with CIAW, Lefaucheur et al. (2006) studied responses of the right deltoid and tibialis anterior muscles to DMS and to motor

nerve stimulation (MNS) with comparison to conventional NCS and concentric needle EMG using an original algorithm for differential diagnosis. They first considered the amplitude of the responses to DMS, then the MNS to DMS amplitude ratio, and finally the amplitude of the SNAPs recorded at the lower limbs. Evidence of neuropathy and myopathy was found respectively in 57% and 83% of patients. Pure or predominant myopathy was found in 19 patients. Other results were consistent with neuromyopathy (n ¼ 5) and pure or predominant neuropathy (n ¼ 2). Four patients had normal results with stimulation techniques, but spontaneous EMG activity and raised plasma creatine kinase suggesting necrotic myopathy. Accordingly, a myopathic process was found

CRITICAL ILLNESS–ASSOCIATED WEAKNESS AND RELATED MOTOR DISORDERS in a majority of cases due to reduced muscle fiber excitability, which may be suspected by a combination of DMS, needle EMG and serum CK levels.

Motor unit number estimation. This technique described 30 years ago by McComas et al. (1971) is use to determine the number of functioning MUs in a given muscle. The method is based on the concept that the number of MU can be determined by dividing the CMAP amplitude of the entire muscle by the estimated amplitude of a single motor unit action potentials (S-MUAP). Most modern MUNE methods are useful for identifying and tracking progressive lower motor neuron (LMN) disease (McComas, 1991), and because they require no volitional activation, they can easily be performed in comatose patients. This technique has a direct appeal given that several levels of the peripheral nervous system may cause reductions in the CMAP amplitude, and since EMG gathers only limited information in a poorly cooperative subject, routine electrodiagnostic methods may not reliably distinguish between nerve and muscle injury. Measurements are typically performed in a representative weak proximal and distal limb muscles such as the biceps brachii and tibialis anterior and abductor pollicis brevis (APB) or ADM using the statistical technique described by Daube (1995) and Shefner et al. (1999) Trojaborg et al. (2002) studied a cohort of 22 patients with CIAW and DMS in the ICU in the tibialis anterior muscle and found significant reduction in the maximal CMAP and S-MUAP amplitudes, and a small but insignificant decrease in MUNE with increasing age. DMS and QEMG showed reduced muscle fiber excitability in 17 patients and QEMG that were both compatible with myopathy, that was correlative with muscle biopsy features of CIM in 4 cases, and significant muscle fiber atrophy affecting type II fibers in 3, without signs of neurogenic affection. Because reduced motor unit size is a hallmark of myopathic injury, the findings of normal MUNE and reduced CMAP and S-MUAP amplitudes supports a predominantly myopathic process. Conversely, when MUNEs decline with decreasing CMAP amplitude and there is a stable to increasing S-MUAP size as a result of collateral innervation, a neuropathic process is suggested. Other clinical studies have demonstrated reduced motor unit numbers in neuropathy (Brown, 1973; Feasby and Brown, 1974). The finding of severe fiber type atrophy in 3 patients reported by Trojaborg et al. (2002) was especially significant given the frequent occurrence of this histological finding in both myopathy and neuropathy, yet it should not alter MU parameters or muscle excitability in the absence of underlying myopathy as in this cohort of CIAW.

717

Another technique to discriminate myopathy from neuropathy is the calculation of the average value of the mean step area of individual MUPs when using MUNE. Baslo et al. (2003) found an average value of mean step area of 2.46  0.88 mVms in normal control subjects compared to 36  1.94 mV/ms in patients with motor neuronopathy and 1.06  0.61 in patients with myopathy. In five patients with histologically verified myopathy with loss of thick myosin filaments, the mean step area was 0.34  0.07 mVms. Although the number of cases examined in this way is few, it is a promising but not commonly used test in the ICU, likely because it is difficult and rather time consuming to perform and analyze; DMS has instead been considered a better electrophysiologic test to separate CIM from CIP in the ICU (Bird and Rich, 2002). Cerebrospinal fluid analysis Among the many potential reasons for contemplating examination of CSF and a lumbar puncture in patients with critical illness associated weakness is the presumptive diagnosis of GBS. The diagnosis typically rests on the presentation of symmetrical weakness of more than one limb, that progresses over 4 weeks and absent or reduced DTRs, with typical CSF findings of albuminocytologic dissociation (with elevated protein content and 10 or less white blood cells (WBCs), and suggestive findings of primary demyelination on NCS and EMG) (Wiederholt et al., 1964; Ropper, 1992).

MUSCLE AND NERVE BIOPSY The indications for muscle and nerve biopsy broaden as the evaluative process has improved. Patients planning to undergo the procedures should have an assessment of the serum CK assay and detailed electrodiagnostic studies to guide the choice of tissues that are clinically and electrophysiologically affected but not overly wasted muscle to avoid end-stage samples. The surgical procedures should ideally be performed by a neuromuscular neurologist or an experienced surgeon at centers with a dedicated neuromuscular pathologist. The vastus lateralis and biceps muscles are ideal for disorders accompanied by proximal weakness, and the gastrocnemius and peroneal longus muscles can be sampled if distal myopathy and neurogenic disease are suspected. Patients with CIAW should ideally undergo combined muscle and cutaneous nerve biopsy especially those with presumptive CIP, because of the possibility of unsuspected or incidental CIM, further guided by the pattern of weakness and electrodiagnostic studies. There are available distal and proximal sources of tissue that are guided by the presumptive diagnosis and distribution of clinical signs including, the sural, superficial fibular sensory nerve or femoral

718

D.S. YOUNGER

intermedius nerves and underlying soleus, peroneus longus or rectus femoris muscle, that can be accomplished with a single incision (Fig. 29.5). There is a technique for biopsy of the gracilis motor nerve and adductor muscle tissue in the medial thigh that does not lead to a noticeable deficit (Corbo et al., 1997); however, it entails a deeper dissection. At least three specimens of muscle and 2 centimeter (cm) of nerve should be removed taking care to incise the tissue samples without imposing tissue handling artifact, and in the case of the nerve specimen, leaving the often weakly adherent epineurial vascular elements intact. Separate pieces of muscle and nerve are placed in moist saline gauze, 10% formalin, and glutaraldehyde and transported directly to the laboratory for processing. Full-thickness nerve biopsy is preferable to fascicular biopsy; however, some still prefer the latter technique to minimize dermatomal sensory loss and afford regeneration across the missing gap of tissue. Recent developments in the analysis of nerve biopsies include preparation of plastic-embedded semithin tissue sections and teased nerve fiber analysis to demonstrate the presence or absence of segmental demyelination and remyelination. A panel of standard and investigational

immunohistochemical, immunoperoxidase, immunofluorescent, and immunocytochemical analyses available at most neuromuscular centers can identify abnormalities in macrophages, T-cell and B-cell subsets, major histocompatibility complex (MHC) type 1 and 2 antigens, immunoglobulins, complement proteins, cytokines, myofibrillar and cytoskeletal proteins (ubiquitin, desmin, calpain, cathepsin B, titin, nebulin, actin), selected proteases, myosin isoforms, and ultrastructural morphology to diagnose and differentiate CIM (Ramsay et al., 1993) CIP (Zochodne, 2005), AQM (Showalter and Engel, 1997), and IMNM (Merlonghi et al., 2022). A 3-mm punch biopsy of glabrous skin at standard sites in the distal lateral calf and proximal thigh immunostained with protein gene product (PGP) 9.5 reveals intraepidermal nerve fibers (IENFs) that can analyze for density and qualitative branching patterns with comparison to age matched controls in the thigh (McArthur et al., 1998) and calf (Joint Task Force of the E and the PNS, 2010) in patients with neuropathic pain and autonomic dysfunction (Latronico et al., 2013; Skorna et al., 2015); due to small fiber neuropathy. Dermal myelinated nerve fibers can be identified with deeper dermal tissue biopsy to retrieve small myelinated fiber bundles (Fig. 29.6A–D) (Li et al., 2005).

Fig. 29.5. Superficial fibular sensory nerve and underlying peroneus longus muscle biopsy procedure. (A) The nerve is palpated laterally along the distal third of the legs along a line between the fibular head and lateral malleolus, providing the markings for the incision. (B) Under monitored anesthesia care, an incision is made and the area is dissected, revealing the nerve (n) obliquely traversing the field (arrow). (C) Incising the aponeurosis reveals the underlying muscle tissue (m) in addition to nerve (n, and arrows) available for biopsy. (D) After the specimens are removed, and the distal end of the nerve is buried to avoid a neuroma, the entire site is irrigated, and interrupted sutures close the aponeurosis, and subcuticular absorbable sutures close the skin margins.

CRITICAL ILLNESS–ASSOCIATED WEAKNESS AND RELATED MOTOR DISORDERS

719

Fig. 29.6. Dermal myelinated nerves are identifiable in the glabrous skin. Glabrous skin from a control subject stained with toluidine blue. The dermal myelinated nerve fibers within a small nerve bundle are shown in (A) running parallel to the skin surface. In (B), a longitudinal section of myelinated nerve fiber is shown (arrowheads). These nerve fibers can also be identified with immunohistochemistry by using polyclonal antibodies to neuron-specific ubiquitin hydrolase (PGP9.5), a pan-axonal marker (black arrowheads in (C) and (D)). These nerves that innervate dermal mechanical receptors are also myelinated and run in the vertical direction in a one-to-one relationship with the individually innervated receptors behaving as naturally occurring teased nerve fibers making them particularly suitable for studying the molecular architecture of myelinated and unmyelinated nerve fiber disease. Reproduced from Li J, Bai Y, Ghandour K, et al. (2005). Skin biopsies in myelin-related neuropathies: bringing molecular pathology to the bedside. Brain 128: 1168–1177., with permission.

RESPIRATORY ASSESSMENT Bedside evaluation The ability to breathe independently is assessed at the bedside by discontinuing or altering mechanical ventilation by stopping intermittent mandatory ventilation while keeping the patient on pressure support or continuous positive airway pressure to overcome airway/ ventilator resistance; as a maximum of 15 min maintains reasonable oxygenation. Mechanical ventilation is routinely continued until the patient meets standard criteria for extubation (Feeley and Hedley-Whyte, 1975) baring episodes of respiratory distress, arterial oxygen saturation of 4.87 30% increase in the duration of the proximal compared to the distal CMAP to distinguish TD due to demyelination from axonal loss (Van Asseldonk et al., 2005) (Table 29.3). Uncini et al. (2017) also provided modified criteria, in

reconsidering the single study approach, responding to calls that began in 2012 (Uncini and Kuwabara, 2012). The need for serial studies reflects less the need for its practical impact on treatment, but in understanding the underlying pathophysiology and prognostication of its varied clinical and electrophysiological presentations, based primarily on early changes that may mimic

Table 29.3 Electrodiagnostic criteria for GBS Hadden’s criteria (Hadden et al., 1998) Rajabally’s criteria (Rajabally et al., 2015)

Uncini’s criteria (Uncini et al., 2017)

(1) AIDP (1) AIDP (1) Primary demyelinating At least one of the following in at least two At first or second study at least one of the nerves: At least one of the following in at least two nerves: ● MCV 130% ULN nerves, or at least two ● F-response latency >120% ULN, or ● dCMAP duration >120% ULN of the following in 1 ● pCMAP/dCMAP duration >130% nerve if all others >150% ULN (if dCMAP 120% ULN inexcitable and of LLN) dCMAP OR OR one of the above in one nerve PLUS: ● F-wave absence in two nerves with ● Absent F waves in two nerves with 10% LLN ● MCV