Journal für Hirnforschung: Band 19, Heft 2 1978 [Reprint 2021 ed.] 9783112519707, 9783112519691

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Journal für Hirnforschung

Heft 2 -1978 Band 19

Internationales Journal für Neurobiologie Begründet von Cécile und Oskar Vogt

Redaktion: J. Anthony, Paris • A. Hopf, Düsseldorf W Kirsche, Berlin • J.Szentägothai, Budapest Wiss.Sekretär:J. Wenzel, Berlin

Akademie-Verlag • Berlin EVP 25,-M- 32105

Begründet von Cécile und Oskar Vogt Unter Mitwirkung des Cécile und Oskar Vogt Instituts f ü r Hirnforschung in Düsseldorf und der Arbeitsgemeinschaft f ü r vergleichende Neuroanatomie der Fédération mondiale de Neurologie (World Fédération of Neurology)

Mitherausgeber: H . ADAM, S a l z b u r g — W i e n O . S. ADRIANOW, M o s k a u J . ARIÉNS KAPPERS,

Amsterdam

E . CROSBY, A n n A r b o r A . DEWULF, C o r b e c k - L o J . ESCOLAR, Z a r a g o z a R . HASSLER, F r a n k f u r t a. M. E . HERZOG, S a n t i a g o J . JANSEN, O s l o J . KONORSKI, W a r s c h a u ST. KÖRNYEY, P é c s

Le J O U R N A L F Ü R H I R N F O R S C H U N G publiera des études sur la morphologie normale (anatomie, histologie, cytologie, microscopie électronique, histochimie), sur le développement du système nerveux ainsi que des études anatomiques expérimentales. On acceptera aussi des t r a v a u x du caractère de la coopération entre des domaines différents à condition qu'ils contiennent des résultats morphologiques obtenus p a r les méthodes de la neuromorphologie et de la neurophysiologie ou de la neuropharmacologie et de la neurochimie. Les t r a v a u x doivent contenir des acquisitions nouvelles sur l'action réciproque entre la structure et la fonction. Des études neuropathologiques seront seulement acceptées quand elles contribuent à la conaissance des structures normales, des changements structurels ou de leur signification fonctionelle. Des études sur la localisation cérebrale de phénomènes expérimentaux ou cliniques d'excitation ou de déficit (doctrine des localisations) seront également publiées par le J O U R N A L F Ü R H I R N FORSCHUNG. Une partie spéciale sera réservée à la neurobiologie comparée.

M. MARIN-PADILLA, Hanover-New Hampshire J . MARSALA, K o s i c e H . A . MATZKE, L a w r e n c e

D. MISKOLCZY, Tirgu Mures

G . PILLERI, W a l d a u - B e r n T . OGAWA, T o k y o B . REXED, U p s a l a H . STEPHAN, F r a n k f u r t a. M. W . J . C. VERHAART, L e i d e n M. VOGT, C a m b r i d g e F . WALBERG, O s l o K . G. WINGSTRAND, K o p e n h a g e n E . WINKELMANN, L e i p z i g W . WÜNSCHER, B e r l i n A . D . ZURABASHVILI, Tbilissi

I m J O U R N A L F Ü R H I R N F O R S C H U N G werden Arbeiten aus dem Gesamtgebiet der normalen Morphologie (Anatomie, Histologie, Cytologie, Elektronenmikroskopie, Histochemie) und der Entwicklungsgeschichte des Nervensystems unter Einschluß experimentell-anatomischer Arbeiten veröffentlicht. E s werden auch Arbeiten multidisziplinären Charakters aufgenommen, sofern sie morphologische Ergebnisse beinhalten, die mit neuromorphologischen und neurophysiologischen oder neuropharmakologischen bzw. neurochemischen Methoden gewonnen wurden und einen Erkenntnisgewinn hinsichtlich der Wechselwirkung zwischen S t r u k t u r und Funktion beinhalten. Neuropathologische Arbeiten werden nur angenommen, wenn sie Beiträge zur normalen Struktur, den Strukturwandlungen oder deren funktionellen Bedeutungen enthalten. Zum Publikationsgebiet des J O U R N A L F Ü R H I R N F O R S C H U N G gehören auch Arbeiten, die sich mit der Zuordnung experimenteller Reiz- und Ausfallerscheinungen bzw. klinischen Symptomen zu bestimmten Strukturen des Gehirns („Lokalisationslehre") befassen. Als spezielles Publikationsgebiet ist die vergleichende Neurobiologie vorgesehen.

The J O U R N A L F Ü R H I R N F O R S C H U N G will publish studies on normal morphology (anatomy, histology, cytology, electron microscopy, histochemistry), on t h e development of t h e nervous system, as well experimental anatomical studies. Papers of multidisciplinary character will also be included so far as t h e y contain morphological results which were obtained using neuromorphological and neurophysiological or neuropharmacological and neurochemical methods and provide further information on t h e interaction between structure and function. Neuropathological studies will only be published if they contribute to t h e knowledge of normal structures structurals changes or their functional significance. Papers dealing with the cerebral localization of experimental excitation and deficit phenomena or clinical symptoms (localization theory) will also be published b y the J O U R N A L F Ü R H I R N F O R S C H U N G . A special p a r t of t h e publication is reserved for comparative neurobiology.

Bezugsmöglichkeiten des „ J o u r n a l für Hirnforschung". Bestellungen sind zu richten — in der D D R an eine Buchhandlung oder an den AkademieVerlag, D D R - 108 Berlin, Leipziger Straße 3 - 4 — im sozialistischen Ausland an eine Buchhandlung für fremdsprachige Literatur oder an den zuständigen Postzeitungsvertrieb — in der B R D und Westberlin an eine Buchhandlung oder an die Auslieferungsstelle K U N S T U N D WISSEN, Erich Bieber, 7 S t u t t g a r t 1, Wilhelmstraße 4 - 6 — in Österreich an den Globus-Buchvertrieb, 1201 Wien, Höchstädtplatz 3 — im übrigen Ausland an den Internationalen Buch- und Zeitschriftenhandel; den Buchexport, Volkseigener Außenhandelsbetrieb der Deutschen Demokratischen Republik, D D R - 701 Leipzig, Postfach 160, oder an den Akademie-Verlag, D D R - 108 Berlin, Leipziger Straße 3 bis 4

Journal f ü r Hirnforschung Herausgeber: I m Auftrag des Akademie-Verlages von einem internationalen Wissenschaftlerkollektiv herausgegeben. Verlag: Akademie-Verlag, D D R - 1 0 8 Berlin, Leipziger Straße 3 - 4 ; F e r n r u f : 2236 229 und 2236 221; Telex-Nr.: 114420; B a n k : Staatsbank der D D R , Berlin, Kto.-Nr.: 6836-26-20712. Schriftleitung: Prof. Dr. W. Kirsche, Berlin. Wissenschaftlicher Sekretär: Dr. Jürgen Wenzel, Berlin. Veröffentlicht unter der Lizenznummer 1326 des Presseamtes beim Vorsitzenden des Ministerrates der Deutschen Demokratischen Republik. Gesamtherstellung: V E B Druckhaus „Maxim Gorki", D D R - 74 Altenburg. Erscheinungsweise: Das Journal f ü r Hirnforschung erscheint jährlich in einem Band mit 6 Heften. Bezugspreis eines Bandes 180,— M zuzüglich Versandspesen (Preis f ü r die D D R 1 5 0 , - M); Preis je H e f t 3 0 , - M (Preis f ü r die D D R 2 5 , - M). Bestellnummer dieses Heftes 1018/19/2. © 1978 by Akademie-Verlag Berlin. Printed in t h e German Democratic Republic. AN (EDV) 60315

ISSN 0 0 2 1 - 8 3 5 9

Journal

J . Hirnforsch. 19 (1978) 1 0 1 - 1 0 8

•

m

Internationales Journal für Neurobiologie H e f t 2 • 1978 • B a n d 19

^jj p

Hirnforschung

Aus dem Anatomischen I n s t i t u t des Bereiches Medizin (Charité) der Humboldt-Universität zu Berlin (Direktor: Prof. Dr. sc. med. W . KIRSCHE)

F l u o r e s z e n z h i s t o c h e m i s c h e und neurohistologische U n t e r s u c h u n g e n z u r a d r e n e r g e n I n n e r v a t i o n des Cortex pyriformis der Ratte 1 ' 2

H e r b e r t DANNER u n d C l a u s PFISTER

M i t 5 Abbildungen (Eingegangen am 11. J u n i 1977)

Summary: T h e pyriform region of normal adult rats was investigated b y fluorescence histochemistry and Golgirapid-impregnation technique. Fluorescence microscopically noradrenaline and dopamine fibers could be described in typical arrangement and distribution. I t is suggested t h a t various Golgi-"beaded"-axons in the cortex pyriformis ma} 7 be equivalent to the fluorescence histochemically demonstrated varicose catecholamine axons. This would raise t h e possibility of interpreting, in t h e Golgi impregnation technique, kinds of axons in a functional manner. Zusammenfassung: Die Regio pyriformis der normalen juvenilen R a t t e wurde fluoreszenz-histochemisch und mittels der Golgi-Rapid-Imprägnations-Technik untersucht. Fluoreszenzoptisch konnten Noradrenalin- und Dopamin-Fasern in charakteristischer Anordnung und Verteilung dargestellt werden. E s wird vermutet, daß es sich bei den nach Golgi-Rapid-Imprägnation dargestellten unterschiedlichen varicösen Axonen im Cortex pyriformis um das Äquivalent zu den fluoreszenzhistochemisch nachweisbaren CatecholaminAxonen handelt. D a m i t bestünde die Möglichkeit, bestimmte Axontypen in der Golgi-Rapid-Darstellung funktionell zu interpretieren.

Einleitung

handlung nach

Mit Hilfe der hochsensitiven Glyoxylsäure-Technik nach LiNDVALLet al. (1974 a) ist es möglich, Catecholaminfasern in ihrem gesamten Verlauf bis zu ihrem Terminationsgebiet zu verfolgen und morphologisch dopaminerge Axone deutlich von noradrenergen Axonen zu unterscheiden (LINDVALL and B J Ö R K L U N D 1974 a). Die nach Anwendung der Paraformaldehyd-Be-

1 9 6 2 ; F U X E et al. 1 9 7 0 ; BJÖRKLUND et al. 1972) er-

1 2

FALCK

und

HILLARP

(FALCK

et al.

hobenen Befunde hinsichtlich der catecholaminergen Innervation des Cortex cerebri konnten präzisiert und erweitert werden (LINDVALL and B J Ö R K L U N D 1974b; B E R G E R et al. 1974). Da es die Glyoxylsäure-Technik nach L I N D V A L L et al. (1974 a) ermöglicht, Noradrenalin- und Dopaminfasern mit Sicherheit zu erkennen und zu ver-

Mit dankenswerter Unterstützung durch einen Forschungsauftrag des Ministeriums für Wissenschaft und Technik der DDR. Herrn Prof. Dr. J . Szentagothai zur Vollendung des 65. Lebensjahres m i t herzlichen Glückwünschen gewidmet.

Hirnforschung, Bd. 19, Heft 2

8

102

D a n n e r , HL, u n d C. P f i s t e r

Abb. 1. Catecholamin-Innervation des Cortex pyriformis (Fissura rhinalis) der R a t t e . Äußere corticale Schichten; vorwiegend Noradrenalin-Fasern. A b b . - M a ß s t a b : 380:1

folgen (LINDVALL and B J Ö R K L U N D 1974b; B E R G E R et al. 1974, 1976), war es naheliegend, derartig fluoreszenzhistochemisch definierbare Axontypen mit Axonen ähnlicher Morphologie nach Golgi-Imprägnations-Darstellung zu vergleichen.

Autofluoreszierende G r a n u l a u n d Gefäßendothelien w a r e n leicht a n ihrer organge-gelben Fluoreszenzfarbe e r k e n n b a r . 2. Golgi-Rapid-Imprägnations-Technik n a c h V A L V E R D E (1970) Zur U n t e r s u c h u n g gelangten 6 u n b e h a n d e l t e R a t t e n beiderlei Geschlechts im Alter v o n 6 — 8 Wochen. S c h n i t t d i c k e : 200 (xm. Die Mikrophotos w u r d e n m i t der Kleinbildkamera E x a k t a R T L 1000 angefertigt. F i l m m a t e r i a l : O R W O N P 27 bzw. N P 15

Befunde Material und Methode 1. Glyoxylsäure-Technik n a c h L I N D V A L L et al. (1974a): Zur U n t e r s u c h u n g gelangten 12 männliche, u n b e h a n d e l t e R a t t e n im Alter von 10 — 12 Wochen. Herstellung v o n H i r n serienschnitten a m Vibratom® (Oxford I n s t r u m e n t s , California). 1 S c h n i t t d i c k e : 25 — 35 (im. Die Kontrolle auf Spezifität der R e a k t i o n erfolgte a) d u r c h A p p l i k a t i o n 3 Stunden);

von

Reserpin

(5 mg/lcg,

i. p „

b) d u r c h Weglassen der Glyoxylsäure-Gasbehandlung. U n t e r s u c h u n g der Schnitte n a c h E i n d e c k e n in Entellan® (MERCK) m i t d e m Fluoreszenzmikroskop F L U O V A L ® ( V E B Carl Zeiss, Jena) bei Auflichtfluoreszenz u n t e r Verw e n d u n g eines Kardioid-Kondensors. Erregerfilter: B G 12/4 g; Sperrfilter OG 1. Die Catecholaminfasern zeigten u n t e r den angegebenen Bedingungen eine charakteristische blau-grüne Fluoreszenz. 1 Diese Arbeiten w u r d e n a m Biological Research Center der Ungarischen A k a d e m i e der Wissenschaften in Szeged a u s g e f ü h r t . F ü r ständige freundliche U n t e r s t ü t z u n g sei besonders Dr. F . J o o , K . M O H A C Z I u n d Dr. I . T O T H gedankt.

1. Corticale Strukturen (Regio pyriformis, Übergangsgebiet an der Fissura rhinalis) nach Glyoxylsäure-Behandlung Der pyriforme Cortex zeigt im Fluoreszenzbild ein ausgeprägtes adrenerges Innervationsmuster (Abb. 1, 2). Die einzelnen Rindenschichten sind nicht gleichmäßig innerviert, vielmehr sind deutliche Unterschiede sowohl hinsichtlich der allgemeinen Innervationsdichte wie auch der speziellen Axonmorphologie sichtbar. Zumindest zwei differente Axontypen sind zu unterscheiden: a) Axone mit relativ dicken, knopfförmigen, gleichmäßig angeordneten Varicositäten Axone dieses Typs sind in allen Schichten des Cortex darstellbar, sie steigen von den tiefen Anteilen in leicht kurvenförmigen Verläufen zur pialen Oberfläche auf, nehmen in den medialen Laminae eine häufig vertikale Richtung (Abb. 3a, c; 4a), um in den oberflächlichen Schichten (LI) mehr oder minder parallel zur Oberfläche zu verlaufen. Die Fasern

Adrenerge Innervation des Cortex p i r i f o r m i s 103

Abb. 2. Catecholamin-Innervation des Cortex pyriformis (Fissura rhinalis) der R a t t e . Tiefere corticale Schichten; überwiegend Dopamin-Fasern. Abb.-Maßstab: 380:1

dieses Typs haben ihre größte Dichte im Stratum moleculare. Zwischen den Varicositäten sind deutlich fluoreszierende intervaricöse Abschnitte erkennbar.

b) Axone mit relativ kleinen, rundlich bis oval erscheinenden, unregelmäßig angeordneten Varicositäten Für die Fasern dieses Typs ist charakteristisch, daß sie als Einzelfaser weniger deutlich darstellbar sind und in ihren Afferenzgebieten plexusartige Endverzweigungsmuster bilden. Sie durchsetzen die Laminae nicht gleichmäßig, sondern sind in der weitaus größten Dichte in den tieferen corticalen Schichten lokalisiert, während sie im Stratum moleculare nur in geringer Zahl vertreten sind (Abb. 2). Die Cortices der reserpinierten Tiere zeigten, von

104

Danner, H., und C. Pfister

Abb. 3. Axondarstellung nach Glyoxylsäure-Behandlung (a und c) und Golgi-Rapid-Imprägnation (b und d). a und c) aufsteigende Noradrenalin-Axone (Pfeil) (Typ-aAxone) und feiner Dopamin-Faser-Plexus (c) Abb.-Maßstab 150:1 b) Aufsteigende varicöse Axone (Pfeil) (Typ-a-Axone) ; vgl. glatte, nicht-varicöse Axone (Kopfpfeil) Abb.-Maßstab: 320:1 d) Geflecht aus feinsten heterogen erscheinenden Axonen mit unregelmäßigen Varicositäten (Typ-b-Axone). Abb.-Maßstab : 450:1

autofluoreszierenden Endot'helien abgesehen, • keine Fluoreszenz. Nach Weglassen der Schnitt-Begasung war eine geringere Fluoreszenzintensität zu verzeichnen; außerdem machten die Schnitte übersichtsmäßig einen „fleckig-schmutzigen" Eindruck, so daß die fluoreszierenden Strukturen sich weniger deutlich gegen einen dunklen Hintergrund abhoben. 2. Corticale Strukturen (Regio pyriformis, Übergangsgebiet an der Fissura rhinalis) nach GolgiRapid-Imprägnation Nach Golgi-Rapid-Imprägnation sind neuronale Elemente sehr gut darstellbar. Auffallend ist der reiche Spine-Besatz u. a. der Dendriten der inneren und äu-

ßeren Pyramiden-Neurone (Abb. 5a, b). Anhand dieser Spines sind die Dendriten auch in ihren periphersten Aufzweigungen zu erkennen und deutlich gegen Axone abgrenzbar. Auf Grund ihrer Morphologie konnten wir drei Axontypen unterscheiden: a) Axone mit regelmäßig angeordneten, runden, gleichmäßig geformten großen Anschwellungen, die ihnen ein perlschnurartiges Aussehen verleihen: varicöse Axone (Abb. 3b; 4a, b; 5). b) Axone mit sehr unregelmäßig angeordneten Varicositäten unterschiedlicher Größe, jedoch von geringem Durchmesser als die des Typs a. Diese Axone bilden in ihren Terminationsgebieten ein

Abb. 4. Corticale Strukturen nach Golgi-ImprägnationsTechnik und Glyoxylsäure-Behandlung a—c) obere Laminae; d—e) tiefere Laminae a) Übersicht, Abb.-Maßstab: 320:1 b) Vertikal verlaufendes dick-varicöses Axon (Pfeil) (Typa-Axon). Abb.-Maßstab: 680:1 c) Vertikal verlaufende grob-varicöse Noradrenalin-Axone Abb.-Maßstab: 390:1 d) Plexus von feinvaricösen, heterogenen Axonen b-Axone) . Abb.-Maßstab: 320:1 e) Fasern verschiedenster Art (varicös, nicht-varicös). Abb.-Maßstab: 320:1

(Typ-

Adrenerge Innervation des Cortex pyriformis

105

106

Danner, H., u n d C. P f i s t e r

Abb. 5. Cortex pyriformis. a) Obere corticale Schichten m i t apikalen D e n d r i t e n u n d A x o n e n verschiedenen Typs. Acodendritischer Längsk o n t a k t zwischen einem grob-varicösen Axon u n d dendritischen Spines in H ö h e L I I — I I I (Pfeil) Abb.-Maßstab: 320:1 b) Axodendritische L ä n g s k o n t a k t e (Pfeile) Abb.-Maßstab: 1360:1

dichtes geflechtartiges Muster. Im Vergleich zu den sehr gleichmäßig erscheinenden Axonen des Typs a zeigen sie ein sehr heterogenes Erscheinungsbild: heterogene Axone (Abb. 3d). c) glatte Axone ohne Anschwellungen: nicht-varicöse Axone (Abb. 3b; 4a, b).

Für die varicösen Axone konnten axo-dendritische Längskontakte an Dendriten von Lamina-V-Neuronen wahrscheinlich gemacht werden (Abb. 5b). In den tieferen Rindenschichten war eine gezielte Verfolgung von Axonen auf Grund der großen Zahl von imprägnierten Zellen und des dichten Fasergeflechtes schwieriger.

Diskussion Mit der Einführung der Glyoxylsäure-Methode durch et al. (1974 a) konnte gezeigt werden, daß auch die corticalen Strukturen durchweg eine außerordentliche hohe und differenzierte adrenerge Innervation aufweisen. Neben der noradrenergen InnerLINDVALL

Adrenerge I n n e r v a t i o n des C o r t e x pyriformis

vation, die mittels der klassischen Technik nach u n d HILLARP (FALCK e t al. 1 9 6 2 ; F U X E e t al. BJÖRKLUND ( F U X E et al.

FALCK 1970,

et al. 1972) dargestellt werden konnte 1965,

1968,

1970;

BLACKSTADT et

al.

1967; U N G E R S T E D T 1971, konnte nunmehr auch eine dopaminerge Innervation belegt werden (LINDVALL and B J Ö R K L U N D 1974b; B E R G E R et al. 1974, 1976), was bereits durch T H I E R R Y et al. (1973 a) auf Grund biochemischer Daten vermutet worden war. Die Ursprungsneurone der dopaminergen Axone liegen in mesencephalen Kerngebieten (LINDVALL et al. 1974b); die noradrenerge Termination stammt aus dem Locus coeruleus ( U N G E R S T E D T 1971). Die Glyoxylsäure-Methode erlaubt wegen ihrer hohen Sensitivität eine Axon-Differenzierung (LINDVALL and B J Ö R K L U N D 1974a). So sind auf Grund der Morphologie zwei Haupttypen von Axonen darstellbar : 1. relativ dicke Axone mit eng beieinanderliegenden, regelmäßig angeordneten, runden Varicositäten. Bei diesen Axonen handelt es sich um noradrenerge Fasern aus dem Locus coeruleus; sie kommen innerhalb der Rinde besonders reichlich im Stratum moleculare vor. 2. dünne Axone mit unregelmäßig angeordneten, spindelförmigen Varicositäten. Es handelt sich hierbei um dopaminerge Fasern. Sie liegen u. a. in tieferen corticalen Schichten ( L I N D V A L L and B J Ö R K LUND 1974b; L I N D V A L L et al. 1974b; B E R G E R et al. 1974). Die Durchmesser der Dopamin-Varicositäten betragen ca. 0,3 —0,6 (xm; die der NoradrenalinAxon-Varicositäten in der Regel über 1 ¡j.m (HILLARP et al. 1966; F U X E et al. 1970). Die Varicositäten enthalten in besonders hoher Konzentration den spezifischen Transmitter (FUXE et al. 1970), widerspiegeln also funktionelle Zustände. Sie werden als praesynaptische Bildungen angesehen (HILLARP et al. 1966). Varicöse Axone ("beaded axons", V A L V E R D E 1 9 7 0 ) sind auch aus der Golgi-Imprägnations-Darstellung bekannt. R A M O N - M O L I N E R ( 1 9 7 0 a) faßt die Varicositäten als zufällige Erscheinungen auf, die im Ergebnis der schlechten Steuerbarkeit der Golgi-Imprägnations-Technik entstehen sollen. F o x ( 1 9 7 0 ) hält sie nicht für Artefakte; er verweist auf Axone mit Konstriktionen und Dilatationen, die in der gleichen Weise sowohl im Golgi-Bild wie auch elektronenmikroskopisch darstellbar sind. Axone mit Varicositäten sind in unserem Untersuchungsmaterial mit absoluter Regelmäßigkeit vorhanden; sie prägen wesentlich das Erscheinungsbild nach Anwendung der Golgi-Imprägnations-Technik (Abb. 3 b ; 4a, b; 5a). Infolge ihrer morphologischen Charakteristika sind sie stets von glatten Axonen ohne Varicositäten (Abb. 3 b ; 4a) wie auch von zarten dendritischen Endästen mit vereinzelten Spines abgrenz-

107

bar. Ein Vergleich der varicösen Axone im GolgiBild mit den fluoreszenzhistochemisch definierbaren adrenergen Axonen ( L I N D V A L L and B J Ö R K L U N D 1974a; B E R G E R et al. 1974, 1976) (s. a. Abb. 1, 2, 3a, 3 c, 4 c) ist somit aus morphologischer Sicht naheliegend. Für eine mögliche Identität sprechen auch Verlauf und Verteilung dieser Fasern in der Regio pyriformis : 1. Typ-a-Fasern (varicöse Axone): Diese Axone sind in der Regel über längere Strecken verfolgbar. Sie steigen vertikal oder in Bögen zum Stratum moleculare auf, verlaufen dort überwiegend in tangentialer Richtung und weisen dort auch ihre größte Dichte auf. Fasern dieses Typs könnten den Noradrenalin-Axonen nach Glyoxylsäure-Behandlung entsprechen (Abb. 1, 3a, c, 4c). 2. Typ-b-Fasern (heterogene Axone): Axone dieses Typs sind auf Grund ihrer Zartheit und ihres unregelmäßigen Verlaufs im Terminationsgebiet nur über kurze Strecken darstellbar. Sie weisen ihre größte Dichte in den tieferen corticalen Schichten auf. Fasern dieses Typs (Abb. 2) könnten den DopaminAxonen nach Glyoxylsäure-Behandlung entsprechen. 3. Für die glatten Axone ohne Varicositäten (Typc-Fasern) (Abb. 3b) konnte keine Entsprechung im Fluoreszenzbild gefunden werden. Eine Gegenüberstellung von einander entsprechenden Laminae (L V—VI) im Bereich der Regio pyriformis an der Fissura rhinalis und des Cortex pyriformis im engeren Sinne zeigt bereits im Übersichtsbild deutlich voneinander verschiedene Faserplexus (Abb. 4d, e). Den Typ-b-Fasern in der Gegend der Übergangszone (Abb. 4e) entspricht eine reiche dopaminerge Innervation (Abb. 2). Die varicösen Noradrenalin-Axone gehen im Bereich des Cortex cerebri wahrscheinlich axodendritische Längskontakte ein ( H I L L A R P et al. 1966; F U X E et al. 1970). Elektronenoptisch-autoradiographische Befunde ( D E S C A R R I E S and L A P I E R R E 1973) unterstützen derartige Vorstellungen. Auch bei den von uns dargestellten engen räumlichen Beziehungen zwischen varicösen Axonen und Pyramiden-Neuronen (Abb. 5) könnte es sich um axodendritische Längskontakte handeln, wobei diese Frage letztlich nur die elektronenmikroskopische Untersuchung entscheiden kann (s. a. R A M O N - M O L I N E R 1970b). Auf Grund des Vergleichs von fluoreszenzhistochemisch erhaltenen Befunden mit denen nach GolgiRapid-Imprägnation möchten wir die Vermutung aussprechen, daß es sich bei den gleichmäßig-varicösen Noradrenalin-Axonen und den varicösen Axonen im Golgi-Bild (Typ-a-Axone) einerseits sowie den heterogen-varicösen Dopamin-Axonen und den heterogenen Axonen im Golgi-Bild (Typ-b-Axone) an-

108

Danner, H „ und C. Pfister

dererseits um funktionell identische Fasern handelt. Diese Vermutung ist durch experimentelle Untersuchungen weiter zu objektivieren.

Literatur BERGER, B.,

J . P . TASSIN,

G. BLANC,

M. A. MOYNE

and

: Histochemical confirmation for dopaminergic innervation of the rat cerebral cortex after destruction of the noradrenergic ascending pathways. Brain Res. 81, 3 3 2 - 3 3 7 (1974). A. M. THIERRY

B E R G E R , B . , A. M . THIERRY, J . P . TASSIN a n d M. A. MOYNE:

Dopaminergic innervation of the rat prefrontal cortex: A fluorescence histochemical study. Brain Res. 106, 1 3 3 - 1 4 5 (1976). B J Ö R K L U N D , A . , B . F A L C K and C H . O W M A N : Fluorescence microscopic and microspectrofluorometric techniques for the cellular localization and characterization of biogenic amines. In: Methods in investigative and diagnostic endocrinology ( J . E . R A L L and J . J . K O P I N , eds.) NorthHolland-Publ. Comp., pp. 3 1 8 - 3 6 8 (1972). B L A C K S T A D T , T . , K . F U X E and T . H Ö K F E L T : Noradrenaline nerve terminals in the hippocampal region of the rat and guinea-pig. Z. Zellforsch. 78, 463 — 473 (1967). DECARRIES, L . and Y . L A P I E R R E : Noradrenergic axon terminals in the cerbral cortex of rat. I. Radioautographic visualization after topical application of DL- 3 H-norepinephrine. Brain Res. 51, 1 4 1 - 1 6 0 (1973). F A L C K , B., N.-A. H I L L A R P , G. T H I E M E and A. T O R P : Fluorescence of catecholamines and related compounds condensed with formaldehyde. J . Histochem. Cytochem. 10, 3 4 8 - 3 5 4 (1962). F o x , C. A., Diskussion zu R A M O N - M O L I N E R in: Contemporary research methods in neuroanatomy. (W. J . H. N A U T A and S. O. E . E B B E S O N , eds.) p. 53, Berlin —Heidelberg — New York, Springer "Verlag 1970. F U X E , K., B . H A M B E R G E R and T. H Ö K F E L T : Distribution of noradrenaline nerve terminals in cortical areas of the rat. Brain Res. 8, 1 2 5 - 1 3 1 (1968). FUXE, K.,

T . HÖKFELT,

G . JONSSON

and

U.

UNGERSTEDT:

Fluorescence microscopy in neuroanatomy. In: Contemporary research methods in neuroanatomy. (W. J . H . N A U T A and S. O. E . E B B E S O N , eds.), pp. 275 — 314 Berlin —Heidelberg—New York, Springer Verlag 1970. FUXE, K.,

T . HÖKFELT,

O . JOHANSSON,

G . JOHANSSON,

P.

and A . L J U N G D A H L : The origin of dopamine nerve terminals in limbic frontal cortex. Evidence for meso-cortico dopamine neurons. Brain Res. 82, 349 — 355 (1974). LIDBRINK

N.-A., K. F U X E and A. D A H L S T R Ö M : Demonstration and Mapping of central Neurons containing Dopamine, Noradrenaline and 5-Hydroxytryptamine and their reactions to Psychopharmaca. Pharmacol. Rev. 18, 727 — 741 (1966). L I N D V A L L , O., and A. B J Ö R K L U N D : The Glyoxylic Acid Fluorescence Histochemical Method: a Detailed Account of the Methodology for the Visualization of Central Catecholamine Neurons. Histochemistry 39, 97 — 127 (1974a). L I N D V A L L , O . , and A. B J Ö R K L U N D : The organization of the ascending catecholamine neuron systems in the rat brain as revealed by the glyoxylic acid fluorescence method. Acta physiol. scand, Suppl. 412, 1 - 4 8 (1974b). L I N D V A L L , O . , A. B J Ö R K L U N D and L.-A. S V E N S S O N : Fluophore Formation from Catecholamines and Releated Compounds in the Glyoxylic Acid Fluorescence Histochemical Method. Histochemistry 39, 197 — 227 (1974a).

HILLARP,

LINDVALL, O.,

A. BJÖRKLUND,

R . Y . MOORE

and

U.

STE-

NEVI: Mesencephalic dopamine neurons projecting to neocortex. Brain Res. 81, 325 — 331 (1974b). R A M O N - M O L I N E R , E . : The Golgi-Cox-Technique. In : Contemporary research methods in neuroanatomy. (W. J . H. N A U T A and S. O. E . E B B E S O N , eds.), pp. 32 — 50 B e r l i n Heidelberg—New York, Springer Verlag 1970. R A M O N - M O L I N E R , E . : Diskussion zu V A L V E R D E . I n : Contemporary research methods in neuroanatomy. (W. J . H. N A U T A andS. O. E . E B B E S O N , eds.), p. 30 Berlin —Heidelberg—New York, Springer Verlag 1970b. THIERRY, A. M.,

L . STINUS, G . BLANC

and

J.

GLOWINSKI:

Some evidence for the existence of dopaminergic neurons in the rat cortex. Brain Res. 50, 230 — 234 (1973a). THIERRY, A. M.,

G . BLANC,

A. SOBEL,

L . STINUS

and

J.

: Dopamine terminals in the rat cortex. Science 182, 4 9 9 - 5 0 1 (1973b). U N G E R S T E D T , U . : Stereotaxic mapping of the monoamine pathways in the rat brain. Acta physiol. scand., Suppl. 367, 1 - 4 8 (1971). V A L V E R D E , F . : The Golgi method. A tool for comparative structural analyses. I n : Contemporary research methods in neuroanatomy. (W. J . H. N A U T A and S. O. E . E B B E S O N , eds.), pp. 12 —28 Berlin —Heidelberg —New York, Springer Verlag 1970. GLOWINSKI

Anschrift

der

Verfasser:

D r . m e d . H . DANNER D r . rer. n a t . C. PFISTER

Anatomisches Institut des Bereiches Medizin (Charité) der Humboldt-Universität zu Berlin D D R - 104 Berlin, Philippstraße 12

J . Hirnforsch. 19 (1978) 109—131

División d e Paleontología de Vertebrados, F a c u l t a d de Ciencias N a t u r a l e s y Museo, Universidad Nacional de L a P l a t a (Director: Prof. Dr. Rosendo P A S C U A L )

The Tectum opticum of Pantodactylus (Teiidae, Lacertilia, Reptilia)

schreiberii

Wiegmann

By J u a n C. QUIROGA 1

W i t h 12 figuies (Received J u n e 20, 1977)

Summary: W e s t u d y t h e histology of t h e optic lobe of Pantodactylus schreiberii W i e g m a n n . I t has 14 layers w i t h characteristic cells in each of t h e m . W e s t u d y with emphasis t h e elements of t h e superficial layers, discussing some physiologic implications. W e described four s t r a t a of optic terminals. To t h e s t r a t a 1 and 3 reach fibres f r o m layer 14. To t h e s t r a t u m 4, f r o m t h e layer 12 a n d t o t h e s t r a t u m 2, fibres f r o m b o t h layers 14 and 12. Some optic fibres f r o m layer 12 send collaterals which reach t h e s t r a t u m 4 of optic terminals. Zusammenfassung: Die mikroskopische A n a t o m i e des T e c t u m o p t i c u m v o m Pantodactylus schreiberii Wiegm a n n w u r d e beschrieben. Das T e c t u m o p t i c u m b e s t e h t aus 4 Schichten m i t charakteristischen Zellen in jeder Schicht. Die oberflächlichen Schichten u n d deren B a u e l e m e n t e w u r d e n eingehend u n t e r s u c h t u n d einige physiologische W i d e r s p r ü c h e erörtert. E s w u r d e n 4 Schichten m i t optischen E n d i g u n g e n nachgewiesen. Z u m S t r a t u m 1 u n d 3 verlaufen F a s e r n aus der Schicht 14. Z u m S t r a t u m 4 verlaufen F a s e r n aus Schicht 12 u n d z u m S t r a t u m 2 ziehen F a s e r n aus den beiden Schichten 14 u n d 12. Einige optische F a s e r n v o n der Schicht 12 senden Kollateralen aus, die das S t r a t u m 4 der optischen E n d i g u n g e n erreichen.

Introduction

CUTT

1971,

BUTLER

1976), experimental embryology

(EICHLER 1 9 7 1 , RAFFIN 1 9 7 2 , KRANZ u n d

The study of the tectum opticum has captivated the attention of many workers in the past, who have realized excellent descriptions of its histology, that reached its zenith with the works of the Spanish School in the age that we could call "The Gold Era of the Golgi's Method". Has been also the study of this nervous centre that permited to Don Santiago Ramón y Cajal to make the first and fundamental steps in the theory of the neuron, and based in the study of the cellular types and the optic terminations in this formation, he published the first ideas on the dynamic polarization and he attacked the reticularists. Only ultimately, after an oversight of several years, have the workers gone back to lend attention to this centre, that in fishes, amphibians, reptiles and aves, performs a function very superior to that its homologous does in mammals. The studies of the tectum opticum have been reundertaken from several viewpoints. In this manner is studied the fibre connections (BUTLER and N O R T H 1

To L u c i a n o HERRERO, teacher a n d friend.

RICHTER

1971), matrix cell dynamics (RICHTER und K R A N Z 1970), histology (BUTLER and E B B E S S O N 1975) and physiology (MATURANA et al. 1960). Also is focused the study of this centre for phylogenetic studies (SENN 1966). On the other hand, Senn derived his concepts on the stratification of the reptilian nervous system (SENN 1968a and b, 1970) from the embryology and anatomy of the tectum opticum. In respect to the study of this formation in reptiles, the descriptions of P . R A M Ó N ( R A M Ó N 1891, 1894, 1896) in Lacerta agilis, Emys europaea and Chamaleon, are the more complete, and in them we shall base the descriptions of our materials. B U T L E R and E B B E S S O N (1975) have studied this centre in Tupinambis nigropunctatus with the Golgi's method, basically in respect to the dendrite distribution and the disposition in layers of the cellular types, together with the afferent and efferent bundles that the degeneration methods have discovered. Huber and Crosby dedicated special attention

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to the differencies between different orders and suborders in Reptilia, summarized in the great work of A R I E N S K A P P E R S , H U B E R a n d CROSBY ( 1 9 6 7 ) .

In our description we shall employ the division in laj'ers, using numbers, like P. R A M Ó N does it in the chamaleon (RAMÓN 1896). This methodology can or can not be correct, and de facto it is confirmed by the existence of differencies in different regions of the same lobe, because if in the central region of the formation we can appreciate 14 well differentiated layers, it does not occur in the anterior or posterior portions, and also in the medial and lateral zones of the lobe. Even knowing this, we shall describe the more differentiated region, in which exists the 14 layers, leaving annotated their inconstancies along the formation. Due to these inconstancies in the conformation of the layers in the optic lobe in different species, Huber and Crosby (ARIENS K A P P E R S , H U B E R and C R O S B Y 1 9 6 7 ) and S E N N ( 1 9 6 8 a and b) have proposed new clasifications, which, in general manner, can include the different types. We only quote here, summarily, the correspondencies of the three clasifications. 1. The periventricular layer of Senn corresponds to the strata 1 and 2 of Huber y Crosby, namely, stratum fibrosum periventriculare and stratum griseum periventriculare; and to the layers 1 to 5 of Ramón. 2. The central layer of Senn includes the strata 3 and 4 of Huber and Crosby, namely, stratum album centrale and stratum griseum centrale; and the layers 6 and 7 of Ramón. 3. The superficial layer of Senn comprises the strata 5 and 6 of Huber and Crosby, respectively stratum fibrosum et griseum superficiale and stratum opticum; and the layers 8 to 14 of Ramón. After the description of the cellular types and the fibrillar components, we shall do some physiologic inductions that the morphology can arrive for the knowledge of the internal functioning of this cortex.

Material and Methods W e h a v e employed e x e m p l a r s of Pantodactylus schreiberii W i e g m a n n , obtained f r o m t h e Localidad de Berazategui, Provincia d e B u e n o s Aires. As histologic m e t h o d s we used t h e v a r i a n t s of t h e silver c r o m a t e m e t h o d proposed b y R i o HORTEGA

(1956)

and

VAISAMRUAT

(VAISAMRUAT a n d

HESS

which p e r m i t t h e utilization of formalin fixed m a t e rial, t h e first in a direct m i x t u r e in 10% formalin a n d t h e second in destilled water. A f t e r c u t t h e material, we applied t h e procedure of L A V I N A 1 ( L A V I N A 1 9 4 3 ) , for stabilizing 1953),

1

Lavilla is t h e real n a m e of t h e author, graphic mistake.

LAVISA

is a t y p o -

t h e reaction, which offers t o t h e p r e p a r a t i o n s a n e x c e / / e n c o n t r a s t and clearness, staining t h e nuclei of n o n - i m p r e g n a t e d cells, permiting in this w a y a v e r y good general image. B o t h procedures h a v e given good results, a l t h o u g h t h a t of Rio H o r t e g a was more specific for t h e fibres of t h e w h i t e substance, in special t h e myelinized ones, which takes w i t h difficulty t h e reaction. Moreover, we used t h e reduced silver n i t r a t e m e t h o d of Cajal in its v a r i a n t s for formalin fixed m a t e r i a l ( C A J A L y C A S T R O 1972), w i t h which we obtained good images of t h e fibrillar t r a c t s and t h e nuclear distribution.

Description The layers that superpose from the ventricular ependym to the surface, suffer along the width of the lobe great variations, and it is only in determinate regions where it is possible to appreciate it very well differentiated and presenting clearly the 14 layers. From the median mesencephalic line to the lateral part, we observe, at the level of the commissura tecti optici that layer 1 is represented by the subcommissural organ, the layers 2 to 5 by a fibrillar stratum, corresponding to layer 2, followed by a cellular one that comprises those that laterally shall form the layers 3, 4 and 5. Following to the surface, we find now the layer 6, that at this level forms the body of the commissure tecti optici. Above this layer, exist counted or none nervous cells and one only sees the processes of the cells of the subcommissural organ and those of the ependymal cells of the vecinity. From here, these latter cells ever populate the first layer, and do not suffer variations. Penetrating in the mass of the lobe, one observes that the cellular strata 3 to 5, differentiate little by little until acquire evident characteristics. Appear also cellular groups of elements of great size, that do not spread in a great extent laterally, and never pass the middled third of the lobe, and represent the mesencephalic root of the V nerve. Layer 6, without varies in with, does not show distinctive characteristics. The superficial layers begin to insinuate, without much clarity. Appears, above layer 6, an homogeneous stratum that laterally subdivides in the layers 7 and 8. Above this stratum, one sees a zone with scanty cells that represents the layer 9, followed by a cellular fringe that will form the layers 10, 11 and 12. Beneath the surface one observes the layer 13 and the stratum of the optic fibres or layer 14. Toward the middle third of the lobe, we can clearly appreciate the 14 layers, that from the ebendj^m to the external surface conform the optic lobe. In this zone one observes the greater histologic differentiation 2 . 2 As histologic differentiation we u n d e r s t a n d t h e complexity, as well cytologic as structural, of a given f o r m a t i o n .

The tectum opticum of the Pantodactylus

Toward the external third, which is also in an inferior position, the periventricular layers lose gradually their stratification in order to transform in the periventricular gray and fibrillar substances of the tegmentum. Layer 6 expands in a fanlike manner in order to form the projection bundles of the tectum; layers 7 and 8 lose uniformity to continue forward, in a more or less abrupt manner, with the internal celluar stratum of the pretectal geniculate nucleus, and behind and below, now losing insensibly their mass, with the reticular substance of the tegmentum. The superficial layers 9 to 11 contimue forward with the molecular zone of the pretectal geniculate nucleus. Layer 12 continues with the optic tract, in the same form as does it the 14, while layer 13 remains enclosed by the unification of the two latter. If we study the modifications of the layers from the anterior to the posterior regions of the lobe, we obtain relatively the same picture. According as we advance toward the more central region of the lobe, we observe the major histological differentiation, which gradually disappear when more posterior is the observed zone. Therefore we can conclude that the middle third of the lobe, as much antero-posterior as medio-laterally, we find the major histologic differentiation, represented in this lizard by 14 layers rather good differentiated. The basic characteristics of each of these layers are as follow: Layer 1. Ventricular Ependym. Formed it by one or two cellular rows, it does not offer variations in all its length, save in the medial region where it is represented by the subcommissural organ.

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root of the trigeminal nerve. These cells can see also in layer 6, although generally near the layer 5. It is separated clearly from the following layer, Layer 6. Fibrillar. It is the deeper white substance. Zone of central afferent and efferent bundles of the lobe. Layer 7. This layer permits to be subdivided in two regions, an upper and a lower one, without a precise boundary. The upper region is eminently cellular, while the lower is as much cellular as fibrillar. In this latter one sees the fibres that, sometimes in form of bundles, gain the superior region, or through which travel those fibres which from the superficial layers descend to the deeper white substance. Layer 8. Cellular. It is formed by one or two rows of cells that because its regularity detaches very well above the layer 7. Layer 9. Molecular. With scanty cells in it, it differentiates clearly from the former. Layer 10. Sparce row of small cellular elements, which does not show neat limits with the molecular layers that surround it. Layer 11. Molecular. More feeble that layer 9. Populates it few cells. Layer 12. Through it elapses a great amount of the optic fibres that reach the lobe. This layer is called by SENN stratum opticum internum (SENN 1966). One finds also cellular ele-

ments in relative abundance. Therefore, it is a cellulo-fibrillar layer.

Layer 2. Molecular. Without evident variations. It represents the periventricular fibrillar layer of the mesencephalon at the level of the optic lobes. It enlarges notably toward the postero-inferior part of the lobe, where it forms part of the toris semicircularis.

Layer 13. With scanty cells, is this layer fundamentally molecular.

Layer 3. Cellular. It is conformed by one or two cellular rows that unifies lateral and medially with layer 5. It will form together with the latter layer the periventricular cellular layer of the mesencephalon.

As one can see, we have divided the afferent and efferent fibres of the optic tectum in superficial and profound. This is justified by the efferent fibres of the cells with ascendent axons to the optic tract. After this brief description of the basic characteristics of each layer, we pass now to the cytologic details.

Layer 4. Molecular. More feeble than layer 2, it has not the extension of the latter, and exists only in determinate zones. Layer 5. Cellular. In all similar to layer 3. Toward the medial regions one observes in its mass the cellular groups, formed by elements of great size, that form the mesencephalic

Layer 14. Layer of the optic fibres. Through it pass the bulk of optical afferents and the superficial efferent fibres of the optic lobe.

Layer 1. The epithelial neuroglial cells that form the layer (Figure 1, L), with its body radicated in this layer, run through its peripheral process all the height of

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Fig. 1. Semischematic drawing of the cells of the lower layers and some of layer 7. 1 to 14. Layers of t h e tectum. A. Elements of ascendent axon with an apical and a basal dendritic trunks which cover t h e whole width of t h e lobe. B. Element similar to A b u t with dendritic trunks more ramifyed. C. Crook axon cell with a long basal dendrite. D. Cell with slender basal dendrites and a long apical dendritic t r u n k which reaches the layer 13. E. Crook axon cell of layer 5. F. A variant of t h e cells of ascendent axon. G. Cell of great size with an apical dendritic t r u n k amplely ramified, and an axon t h a t forms a large net in layer 7. H . Medium-size element with short apical dendrites and an axon t h a t disappears in layer 6. I. Cell similar to G b u t in layer 3. J . Short apical dendrite cell with an axon remified in t h e lower layers. K. Long crook axon cell of layer 3. L. Epithelial neuroglial cell, a) axons.

the lobe, for terminate in the external surface in little widenings. In layer 13 and even within the 14, we have seen that some processes bifurcate one or two times before reach the surface. Toward the ventricular lumen, emit these cells small fringes that may be the representation of the flagella, common in these cells. The body presents around it little buds that offer to the cell an irregular aspect. Along the length of the peripheral process, one can appreciate filaments of different sizes, generally finished in a small terminal button, that can vary in quantity as we take a cellular or a molecular zone, increasing its complications in the latter. Layer 6 is the poorest in them, but possessing the longest. Layer 2. The dendritic plexus existing in this layer is formed basically by the basal dendrites of the cells of layer 3, although in several oportunities we have seen processess that reach this layer from layer 5.

The tectum opticum of the Pantodactylus

Elements of the superior layers send dendrites to this plexus, and such a behavior we have observed in elements of layer 7 (Figure 1, A and B), and in layer 6 (Figure 1, D; Figure 2, E). Together with the dendritic plexus one observe slender fibres that travel this layer as much along as in wide. These fine axons can present little enlargements in their trajectory, and form with the dendrites the delicate plexus that offers to this layer its molecular structure. On the origin of these fibres, in many cases we know little, in others we can affirm that they belong to cells of layer 3, as is the case of the element J of the figure 1, or they can be the terminal resolution of fibres that reach this layer from the deeper white matter. Layer 3. Within the morphologic diversity of the elements of this layer we shall describe the more common types we found, just as some less frecuents. They basically correspond with those described by RAMON (1891, 1894, 1896).

a. Element with a long apical dendritic trunk, that reaches the layer 13 (Figure 1, K), and whose axon, of the type of crook axon (axon en cayado) of CAJAL (1972), starts in layer 7 and after a curve, descends to the deeper white matter where disappears. Toward the base of the cellular body originate one or two small protoplasmic trunks that dividing progressively, already in layer 2, form an ample dendritic net, formed by delicate prolongations garnished with lateral spines. The thick apical dendritic trunk travels through the superior layers without give any collateral, presenting only littles enlargements in its trajectory. At the level of layer 9, and specially at the superior limit of layer 8, we have observed that this trunk sistematically emits small collaterals that ramify broadly in that region, often more than the figure shows. These collaterals form a plexus of distorted filaments, which finish in a little widening. In the layer 9 suffers the trunk its first bifurcation, for produce, after it, others until, reach the layer 13, where it displaies more manifestly. In layer 12 and 13, and specially in the latter, forms the trunk a plexus of prolongations very similar to that described for layer 9, but much ample. The more superficial fillets contact with layer 14 and, while some of them appear to penetrate it, the majority double in a right angle to pursue a space patched under it. The axon commonly emits some collaterals after produce the descendent curve. b. This second cellular type is entirely similar to

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those described by RAMÓN in the quoted works as cells with ascendent axons to the optic tract. This element possesses basal dendrites which resenble those described for the anterior type. A strong apical dendritic trunk ascends undivided until pass the layer 6, after which it bifurcates giving two or three dendritic branches that can climb as far as layer 13. At the level of this division origins the axon, which in ascendent march, and giving sometimes collaterals, reaches the optic tract. Variants of this cellular type can be the elements, whose apical trunk remains undivided until the strata 8 or 9, where it divides, and in which it was difficult to us to find the axon. Some of the ascendent axons of these cellular types, disappear when they reach the layer 12, and may be it the case that they leave the tectum together with the optic fibres that travel through this layer. c. Show these elements a more voluminous body than the former, just as a stronger apical dendritic trunk. The basal dendrites resembles those of the preceding types (Figures 1, I). The powerful apical dendritic trunk reaching the inferior limit of layer 6, expands in an ample fan producing two, three or four secondary trunks, some vertical and others oblique or horizontal, that after a more or less long stretch, bend in an ascendent direction. In order to ascend, each of these prolongations divide little by little until reach the upper strata, leaving, specially in the molecular layers, small collaterals which resolve in a plexus of complicate sprouts. The axon originates at the level of the first division, and ascends until reach the layer 7, where it starts to ramify producing an ample plexus. We have pursued its more superficial twigs until the superior limit of layer 7, or the inferior regions of layer 8, but we think that it is probable they reach superior levels that the histologic method has not wanted to show us. d. Present these elements a medium-size cellular body (Figure I, H,) with basal dendrites and an apical trunk that ascends undivided until reach the layer 6, where it resolves in several secondary trunks that rarely exceed the layer 6. Some dendritic branches are horizontal and travel parallelly to the surface that limits the layers 5 and 6. They offer little lateral spines, specially in the slenderer sprigs. The axon, originated sometimes from the dendritic trunk in its ascendent portion, as is the case of the figure, and other times at the level of the division, ascends a stretch until reach the layer 6, in which it disappears after offer, occasionally, an almost right angle. e. This type differs in small degree of the former in what concerns its dendrites, save in that one frecuently observes the division of the apical trunk in

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Fig. 2. Semischematic picture of cells of some superficial and profound layers. 1 to 14. Layers of the tectum. A. Polygonal cell of layer 12. From this the point of view it appears as an horizontal triangular element with long opposite dendrites. B , C and E . Plumed cells which inhabit the layers 7 and 6. D. Element of the mesencephalic root of the trigeminus nerve. a) axons. b) ascendent collaterals of the axon of the plumed cell E . c) collateral of the axon of the same cell that runs in opposite direction, towards the commissura tecti.

the layer 4 (Figure 1, J). The differential characteristic is its axon. It originates in the basal portion of the cellular body and bifurcates almost inmediately in T; one of the branches gains the second layer where disappears, while the other branch ascends a space until reach the layer 5, through which travels a more or less long stretch for to go back to layer 2.

Between all types, the a, b, and c were the more frecuents, in special the first two. One must be care for not confound the types c and d, with other types wrong impregnated3. Layer 4. Its appearance is entirely similar to layer 2, although the major dendritic affluents pertain to the cells of layer 5. Fine axonal fillets travel this layer forming a very delicate plexus. Layer 5. This layer repeats the cellular types described for layer 3, with very little variations. As we see in the figure 1, the cell E of layer 5 resembles to the K of layer 3, just as the G the I respectively. The elements of ascendent axons exist also here as we described 3 I t is evident that spearnk of frecuency of a cellular type in Golgi material is touching very much the error, due to the "personality" of this technique, and any reference to frecuencies we quote, is in consequence, highly relative.

The tectum opticum of the Pantodactylus

Fig. 3. Semischematic drawing carried out with a camera clara. One observes the location, dendritic field and the course of the axon of a plumed and a ganglion cells. The collaterals of the plumed cell's axon reach the nucleus lateralis profundus mesencephali. 6. deeper white matter of the tectum. A. Ganglion cell, whose axon, after divides in T, reaches through its peripheral process the level of the nucleus lateralis profundus mesencephali, where it bends antero-posteriorly. B. Plumed cell, a) axons. VM. Ventriculus mesencephalicus. NLPM. Nucleus lateralis profundus mesencephali.

them precedingly. A variant of this type can be the element F of the figure 1, in which the arrangement of the apical dendritic branches maintains the described distribution, but its axon, instead of depart at

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the level of the bifurcation of the dendritic trunk, it originates from the cellular body, and ascends, then, almost rightly to gain the optic tract. The cellular elements of the mesencephalic root of the trigeminus nerve are conspicuous in this layer through its great size, its scanty basal dendrites which reach the layer 4, and its apical dendritic trunks, that in a number of two or three, ascend ramifying little by little until reach the layer 12. During its passage through the different layers, these trunks emit small collateral branchlets (Figure 2, D). The axon, generally departing from the cellular body inmediately gains the layer 6 where it takes an anteroposterior direction after a short transverse course. Layer

6.

Through this layer travel the profound afferent and efferent fibres of the optic lobe.

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The fibres that reach the tectum through this way, distribute in different layers by terminals and collaterals. We divide them in two types, those that ascend to the upper layers and those that descend to the lower ones. Ascendent fibres From layer 6. these fibres reach the layer 7 where resolve in a delicate fan of branches that sometimes can reach the layer 8. Some of them, without sending an}' collateral, terminate in the upper region of the layer 7 (Figure 11, J). Others give during their course through layer 6, some collaterals before ending. These collaterals spread out at different levels of the layer 7, some in the upper and others in the lower regions (Figure 11, b). The fibres that send these collaterals, after doing it, end freely in the layer 7 (Figure 11, y). Other fibres, as is the case of the fibre L of the figure 11, give some collaterals that ramify within the layer 6 (Figure 11, c). Deascendent fibres These fibres give collaterals that reach, from the layer 6, the lower layers 5, 4 and 3, and sometimes the layer 2 (Figure 11, d), and then, they terminate in a delicate penache in the same manner as their collaterals (Figure 11, z). The efferent axons that gain this layer pertain to the preceding mentioned cellular tipes of the lower layers, and fibres that from elements located in superior strata, come down to this layer for transforming in projection fibres. Amóng these cells we count those of crook-axons of the layers 7 and 8, the plumed cells of Ramón and the great ganglion elements that populate the layer 7, which frecuently one can find submerged between the fibres of this layer (Figure 2, E ; figure 3, A and B). One finds also, elements of the mesencephalic root of the V nerve displaced from the layer 5 and droped between the fibres of this layer (Figure 2, D). Moreover, we have found elements of medium-sieze body, which lie almost in contact with layer 5, and that one can consider as displaced in this stratum (Figure 1, D). The apical dendritic trunk of this cell type, suffers few divisions until arrive the superficial layers, ramifaying in them, giving off little collaterals in its passage through these layers, which, specially in the layer 9, form a plexus of complicate branchlets that finish always in a terminal enlargement. The basal dendrites, originated in one or two trunks, ramify in layers 4 and 2 after run through the strata 5 and 3. They are garnished with lateral spines in all its trajectory. The axon, issued from the cellular body, disappears among the fibres of layer 6.

The plumed and ganglion cells shall be described in the next layer, in which one finds the great majority. Layer 7. As we said above, this layer can be subdivided in an upper region, eminently cellular, in which inhabit elements of medium-size body, pyriform or ovoid, and in a lower region, fibro-cellular, in which we find the great majority of the ganglion and plumed cells. a. The ganglion cells are elements of giant size, with a triangular or polygonal body, from which start three or four strong protoplasmic trunks, among which, some acquire a vertical direction, and others an horizontal or oblique one. The verticals ascend dividing as travel the layer 7. The width of the dendrites decreases little by little from the point of departure of each bifurcation until be very slender, almost filamentous, in the upper strata, where they end, already in the layers 9, 11 or 13, in a brush of distorted branchlets, finished in small enlargements. The horizontal trunks, after travel a more or less long distance, bend for climb to the upper layers, with the same characteristics as the verticals. In this manner, only one ganglion cell dominates a amplest receptive field that anyone other element of the lobe, and summarizes the information into an only one efferent fibre (Figure 3, A). The axon originates in the basal part of the cellular body or near the point of departure of one of the protoplasmic trunks, and descends until reach the layer 6, in which penetrates in order to tranform in a projection fibre. Occasionally, as shows the figure 3, we have seen it dividing in T, taking one of the branches a direction towards the commissure of the lobe, and the other a peripheral direction. This last will form part of the efferent fibres of the tectum. b. The plumed cells (Figure 2, B, C and E ; figure 3, B), with an ovoidal body and smaller size than the ganglion elements, emit from their poles two horizontal protoplasmic trunks, which after a strecth, start to ramify giving ascending dendrites that climb to the upper layers where resolve in penaches of small fibrills which originate as collaterals or as the resolution of a very slender dendritic terminal. R A M O N (1896) affirms that the terminal penaches of these dendrites end all in the same layer, and he draws a cell of this type in his figure 14. We have not had the opportunity to observe similar elements. The terminals end in the upper strata, but in different layers. This discordance can be take in mind as interspecific differencies. Some dendritic trunks emit, before ascend, collate-

The tectum opticum of the Pantodactylus

Fig. 4. Semischamatic drawing that shows cell types of layer 8 and the upper regions of layer 7. 7 to 13. Layers of the tectum. A. Crook axon cell. In this element the axon descends until reach layer 8 where resolves, reaching some processes the upper regions of layer 7. B. Crook axon cell of the upper region of layer 7. This element is also frecuent in layer 8. Through a strong apical dendritic trunk, which sends powerful collaterals during its ascendent course, reaches the cell the layer 13. The axon gives a multitude of collaterals during its passage through layer 8 in its descendent course. C. Cell of descendent basal axon. D. Cell of ascendent axon which ramifies in layer 13. During the passage through the layer 9, the apical dendritic trunk of this cell gives some collaterals which unifies to the dense plexus of this layer. E. Cell with an horizontal dendrite and a descendent basal axon. a) axons. b) Strong dendritic collaterals of the apical trunk of the cell B.

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rals that runing through the layer 6, penetrate in the periventricular layer of Senn, reaching some of them the layer 2 (Figure 2, E and B). The axon descends to the deeper white matter, through which runs, giving collaterals along its trajectory (Figure 2, b) that ascend to layer 7, just as fibres that starting from the main process and then bending on the opposite direction, travel toward the commissura tecti optici (Figure 2, c). We can pursue the axon of this cellular type long way towards the tegmental regions. The figure 3, carried out with camera clara, shows a case in which the axon of the plumed cell B sumes up with the fibrillar contingent that will give origin to the medial tectobulbar tract. We see that at the level of the nucleus lateralis profundus mesencephali (Figure 3, NPLM), it emits a series of collaterals, which in distorted trajectory, leave terminals in the region, and then, giving a curve, some of them seek the periventricular gray substance. In the same figure, one observes the axon of the ganglion cell A, that takes place, after its bifurcation in T, externally respect the 9

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former. We can pursue this last axon until reach the level of the nucleus lateralis profundus mesencephali, where it takes an antero-posterior direction, We can also abserve these plumed cells in more superficial zones of layer 7, but in these cases, although without varying their morphology, the elements are smaller than the profounds. The receptor field sweeping by the dendrites of these cells can be as great as those of the ganglion elements, specially that of the profound cells, although frecuently this field is smaller. In the upper region of layer 7 one finds a diverse series of elements that we can summarize, based in the dendritic and axonic distribution, in two basic types. c. Crook axon cell (célula de axon en cayado). With a medium-size and ovoid form body, this element takes place fundamentally in the median and upper regions of the layer 7 (Figure 1, C; figure 4, B). Through its basal part it emits one or two fine dendritic trunks, which ramifying in slender fillets, distribute within this layer. As generally occurs with the short basal dendrites, they have spines along its extension. In other cases, one of the basal trunks is more powerful (Figure 1, C), which after runs through the lower layers, reaches the plexus of layer 4, in which intermingles. The apical dendritic trunk behaves in a different form depending on it pertains to superficial or profound elements. In the latter, the trunk ascends undivided, presenting only some enlargements, until reach the layer 9 where bifurcates, giving off two trunks that, without ramifying, arrive to layers 12 and 13 where they resolve in a plexus of little fillets. During the passage through layer 9, 11 and 12, leave, these trunks, collateral branchlets, which sume up to the plexus of these strata. A characteristic case is represented by the cell C of the figure 1. The more superficial elements, on the other hand, have the apical trunk with an aspect more similar to that of the cells that populate the layer 8. These trunks, in its ascendent trajectory, emit great collaterals and reaching the layer 11, show a bifurcation, whose resulting dendrites reach the layers 12 and 13, where end in little distorted branchlets, whose terminal sprouts show, as in the majority of the cases, small enlargements in their extremities. The cell B of the figure 4 represents an element of this type. The great collateral branches originate from the apical trunk at the level of the layer 8 or in the limit with the 9. They offer a flexuous trajectory, in some cases very long; commonly they give secondary colla-

terals and show in all their length small fillaments, finished in terminal widenings, that offer them their characteristic aspect (Figure 4, b). Above the level of the collaterals, and now in the middle of the layer 9, originate from the apical trunk some smaller collaterals that ramify, sometimes more, sometimes less, in the plexus of this stratum. The axons of these cells offer variations, as well as the apical trunk, according to the cells inhabit profound or superficial regions. In the profound regions, the axon originates from the apical trunk before its bifurcation within the layer 7. After a curve descends until gain the layer 6 where disappears. In some cases we have seen it to emit some collaterals that do not escape out the layer 7; in other opportunities it descends without collaterals (Figure 1, C). In the superficial elements, the axon departs from the apical trunk during its passage through the layer 9, showing inmediately a curve that gives it a descendent direction. In few cases, the axon was directly descendent. With a flexuous trajectory, it runs through the lower layers to end in the deeper white substance where disappears (Figure 4, B). In its passage through layer 8, emits the axon a series of ascendent collaterals, that with a sinuous trajectory, we can pursue until tha strata 10 and 11, ending some of them in the layers 8 and 9. They show few secondary branches and show a rosary-like aspect produced by small enlargements along its course. They can cover rather spacious fields. The cell B of the figure 4 shows an axon with the described features. d. Cells of ascendent axon. This elements can show different dendritic distribution (Figure 1, A and B). Some cells show two dendritic trunks, one basal and other apical, both similar, undivided in the most part of their trajectory, ramifying towards the terminal points. The apical trunk reaches the more superficial layers, resolving in little branchlets in the strata 12 and 13. The basal one, arrives to the layers 4 and 2 where it ends in slender thorny dendrites (Figure 1, A). The element a that inhabits the layer 7 of the figure 5B, pertains to this cellular type. Other cells, on the other hand, offer a basal trunk that descends, giving some fine collaterals, until reach the level of layer 6, where bifurcates, producing two branches that descend to the layer 2, in which they spread out in an ample plexus of thorny dendrites. Apically show these cells one or two dendritic trunks that inmediatelly ramify and reach the superficial strata where they end. The axon of these elements originates from the apical trunk before its ramification in the first type, and from one of the collateral apical branches in the

The tectum opticum of the Pantodactylus

second type. It ascends vertically until reach the stratum opticum, without we can observe any collateral. Layer 8. The cellular elements that populate this layer can be summarized in four fundamental types. a. Crook axon cells. Basically similar in what concerns the dendritic distribution, the axonal behavior permits us to separate these cells in two varieties. The first type is all in all similar to that described in the former layer which inhabits the upper regions, represented by the cell B of the figure 4, and in consecuence we do not describe it. In what concerns the second type, the axon originates in more superficial layers than the first type, and we have observed it as high as the stratum 11. Along its flexuous descent, emits the axon collaterals, and then it resolves in a plexus at the level of layer 8. Some branchlets reach the upper extreme of layer 7. This axon shows enlargements in its trajectory and the collaterals and terminals do not travel through a great extension (Figure 4, A). The figure 5A shows an element of this type. b. Cells of basal descendent axon. With a body similar to the former, show these elements an apical trunk that ascends towards the level of the zone 11 where it bifurcates, arriving its terminals to the layers 12 and 13. Provided of little collateral branchlets, resembles this trunk, in general, those described for the former types. From the basal pole of the cellular body departs an axon that penetrates in the layer 7, showing short collaterals and spines in its trajectory. Within the layer 7 shows the axon a curve that confers it an oblique direction in respect to the surface of the layer. In this condition it follows forward until reach the deeper white substance in a distant region from that in which inhabits the cellular body that originates it. c. Cells of ascendent axon. With a medium-size body and short thorny basa dendrites, show these elements a flexuous apical trunk, garnised with collaterals that form an ample plexus in the layer 9 and ascend to the more superficial layers (Figure 4, D). Through the behavior of the axon one can describe two types. In the first case, the axon departs from the apical trunk during its passage through the layer 9 and making a curve, ascends until disappears in the second case, the course of the axon resembles the former until reach the layer 13 where, instead of continue and penetrate in the 14, divides in several terminal branches, which in a number of two or three, and giving secondary and tertiary branches

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which are garnished with small enlargements and show a rosary-like aspect, run through long distances in the layer 13 (Figure 4, D). Sometimes, we have observed a delicate prolongation that originated very height in a dendrite, within the layer 11, and that progressing a space until reach the layer 12, it resolves in a complicate plexus within this last layer, reaching some fillets the layer 13. The figure 6 shows one of these cases. We incline to think that this can be the axon of the cell. d. Cells of horizontal type. With a size resenbling that of the former, offers this element a dendritic trunk that originated from its apical pole, inmediatelly shows a curve, and then runs a more or less long distance through the boundary between the layers 8 and 9 (Figure 4, E). From the cellular body depart also small dendrites which ramify loosely in the neighbourhood. From the lower pole or from one of the sides of the cellular body originates the axon, which has a descendent direction, and giving only some collaterals, it follows forward until reach the layer 7 where disappears. Layer 9. Basically plexiform, this layer shows few cells. a. Near the limit between this layer and the 8, there are elements of ovoid form, from their body originates several dendritic trunks, some horizontal, some vertical. The horizontal ones, after a stretch, during which emit ascendent collaterals, show a curve until gain the upper layers. All these dendrites reach, tranformed in very slender fibres, the layers 12 and 13 where they resolve in several branches with small terminal fillets ending in enlargements. The axon originates from the cellular body and runs through certain distance along the limit between the layers 8 and 9, giving collaterals in both layers, although more ample in the 9 (Figure 7, c). The figure 7 shows in E an elements of this type. b. With a pyriform body, lie these cells in the middle of the layer 9; they possess an apical dendritic trunk that ascends until reach the layer 11, in which resolves in several terminal fillets. From the basal pole originates other trunk, shorter than the former, which finishes in a plexus that arrives to layer 7. From this trunk departs the axon, which after runs through layer 8, disappears in the 7. During its course through the layer 8, emits the axon a strong collateral (Figure 7, d), that running in the limit with layer 9, divides after certain distance in two or three branches which cover the width of the layer 8. After a space, some of the fillets pass to more superficial regions of layer 7, where they finish. The other 9*

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fillets end within layer 8. Along their trajectories, show these collaterals a rosary-like aspect. In the figure 7, the element F is an exponent of this cellular type. In this picture we have not drawn the terminal dendrites for not overstock the figure. Layer 10. The elements which inhabit this layer, are generally small and their dendritic irradiations, except some cases, is relatively sparing. We divide them for their description in two basic cellular types, according to the axon ends within the superficial layers or reach the stratum 6. a. Short axon cells. Of small body, possess these cells several dendritic trunks, ascendent ones and descendent others. The former ascend directly or after a short horizontal course. They branching several times and reach the layer 12 and 13 where resolve in little branches after giving some collaterals for the layers 10 and 11 (Figure 8, E ; figure 9). The descendent trunks branching profusely in layer 9, giving some processess which reach the layer 10 and also the 11. The fillets originated in its ramification are tortuous, supplyed with enlargements and garnished with lateral bits which commonly finish in terminal knobs (Figure 9; figure 6B, b). The cell b of the figure 5B, shows clearly its basal dendrites, but not the origin of the apical ones because they are out of the plane of the photograph, but they appear in an upper place, reaching some of them the layer 12 where desappear. At this level the impregnation of the layer 12 has been as complete that appears black in the photograph. Two are the ways that can take the axons of these cells, and we describe them separately. I. As one can observe in the cell E of the figure 8, the axon originates from one of the dendritic trunks and acquires an horizontal course at the limit bet.ween the layers 10 and 11 (Figure 8, d), emiting some short ascendent collaterals to the zone 11, and reaching some of them the layer 12. Along the trajectory show as well the axon as its collaterals, little widenings which confer them a rosary-like aspect. The extension traveled by the axon can be relatively long. II. An example of this axonal type shows the figure 9. Departing from one of the basal poles of the cellular body, the axon takes an opposite direction to that that acquire the dendrites. It descends a stretch to reach the layer 8, where resolves in several terminal branches, giving before few collaterals. The terminal branches, some ascendent, others descendent, travel through trajectories more or less long. The longer of the ascendent ones climb until reach the layers 10

and 11, while the shorter ones resolve in the layers 8 and 9, emiting some collaterals that spread out forming part of the plexus of the zone 9 or, on the other hand, they run horizontally through layer 8. The descendent terminals penetrate into layer 7, ending in several free fibres at different levels in the upper region of this layer. Likely as the axons already described, show these a rosary-like aspect due to small enlargements existing in their courses, specially in the terminal branches.

b. Long axon cells. Through their dendritic distribution one can describe three types. The axonal distribution, save little exceptions, is similar in all the three types. I. Represented by the element F of the figure 8, these cells possess an apical dendritic trunk which bifurcate at the level of layer 11 and end in the layer 12 through little branchlets. In the sides of the cellular body originate dendrites that resolve within the layer 10, reaching several fillets the layer 11. The whole surface of the cellular body is garnished with small short knobs. The axon descends with a flexuous course until reach the layer 6. At the level of the stratum 8 emits the axon one or two collaterals, which in an ascendent direction, reach the layer 10 where they end (Figure 8, e). In the layer 7 appear collaterals, which run tortuously between the cells of this stratum (Figure 8, c). This axon also shows the rosary-like aspect of those of the short axon cells. II. Shows this type an ovoid cellular body, with an apical and a basal dendritic trunks. The apical one ascends vertically crossing the layer 11, and in the boundary between this layer and the 12, resolve into two or three trunks that ascend until end, some in layer 12 and others in the 13, in a few fillets or in small distorted branches (Figure 8, G). The basal dendritic trunk descends to the layer 9 in which ramifies diffusely in branches which, little by little, each time more slender, end in small fillets supplyed with terminal enlargements. The plexus formed by the ramification of the basal dendritic trunk is ample and profuse. The axon of this element, originated from the basal trunk near its depart from the cellular body, descends vertically, running through the lower layers, until desappear in the deeper white substance. During its course, the axon emits collaterals which take different ways. Near its origin, this axon gives an ascendent prolongation which reaches the layer 12 (Figure 8, f). In the lower regions we find fillets which adopt an horizontal course, penetrating some of them in the layer 8 (Figure 8, g). These collaterals can

The tectum opticum of the Pantodactylus

show secondary branches of ascendent course (Figure 8, h). We have not observe in this axon the rosary-like aspect that characterizes the other axons, save in its more distant collaterals. III. The prevailing feature of this element is the distribution of the basal dendrites. Of triangular body, possesses this cell an apical dendritic trunk that we have seen reaching the layer 12. From its basal region, originate from the cellular body two thick dendritic trunks which adopting opposite directions, spread out amplely in the layer 9 (Figure 8, z). Through all its course emits each trunk collaterals that resolve in distorted branchlets supplyed with fine terminal fillets. We have also seen ascendent collaterals which, originated from one of the basal trunks, can reach the layer 12. The terminal prolongations of these trunks can distribute in the layer 9 or in the 8, as in the case of the trunk z of the figure 8. The behavior of the axon of this element is similar to that of the former type. After its origin from one of the basal dendritic trunks, emits the axon, in the lower region of the layer 9, an ascendent collateral which ascends to the layer 12 where disappears (Figure 8, i). Inmediately originates the axon other collateral that adopts an horizontal direction and penetrates in the layer 8, through which runs it in an extense course, and finishes in few terminal branches. The rosary-like aspect is less evident in these collaterals. After follow it through the layer 7, where it does not emit collaterals, we have lost this axon in the deeper white matter. Layer 11. The cells that inhabit this layer, although scanty, show very diverse forms. All of they are near the limit with the layer 12. a. Showed by the element C of the figure 8, has this cell an important basal dendritic trunk that resolves into a plexus in layer 11, reaching its terminal branches the layers 9 and 12. We have not drawn them in the figure for not overload the picture. Few short apical dendrites resolve in the layer 12. In the cellular body, one appreciates little filamentous branchlets. The axon, originated from the basal trunk, descends without showing any collateral until reach the layer 7 where resolves in few terminal branches. These branches run through more or less tortuous courses within this layer ending freely between its cells. They can descend until reach the lower regions of layer 7.

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b. With the body in the limit with layer 12, shows this element a basal and an apical dendritic trunks (Figure 8, D). The apical trunk crosses the layer 12, giving in it some short collaterals, and ends in the layer 13 through a fan of terminal branches. The basal trunk descends until reach the layer 9, in which resolves in a delicate plexus of distorted branchlets. The axon, originated in the layer 9 from the basal trunk, resolves also in a plexus in this layer, mixed with the basal dendrites. Few has been the occasions in which we can observe this axon with clarity. In general, masked by the plexus resulting from the basal trunk, we can only observe it at its origin and pursue it during a very short course. c. With small and stellate body, possesses this element several dendrites, most of them of ascendent courses, that crossing the layer 12 and giving collateral branches during the passage through it, end in the layer 13 in delicate tortuous branchlets. Commonly, originates from one of the basal poles of the cellular body a dendritic trunk which internates in the layer 11, and, giving then a curve, ascends showing the same behavior as the other dendrites. From this last dendritic trunk generally originates the axon, which, horizontal through a short trajectory, terminates in a T bifurcation (Figure 7, B). The ascendent branch reaches the layer 12 in whose fibrillar jungle disappears. The descendent branch, in a vertical course, reaches the upper regions of the layer 7 in which appears to finish. In its descent emits the axon a multitude of collaterals, which with ascendent and descendent courses, form an ample plexus in the layer 9, reaching sometimes the 10 (Figure 7, b). Show these collaterals, as others we have seen above, the characteristic rosary-like aspect. d. With a size smaller than the former, has this cell a multipolar form (Figure 7, C). with dendrites that take predominantly both apical and basal directions. The apical ones, in a number of two or three, give some fine tortuous collaterals in the layer 12, finishing in the 13 through a delicate plexus of fine branchlets. The basal dendrites, in a number of one or two trunks, divide near their origin and resolve within the layers 11 and 10 in a dense plexus formed by fillets of tortuous courses and garnished with lateral knobs. This cells has in this manner, an ample fan of reception. The axon, originated at the base of the cellular body, or from one of the lower protoplasmic trunks, descends, masked in the basal dendritic net, until reach the layer 10, where resolves in several terminals. During its course through the layer 11, gives

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the axon collaterals of predominantly horizontal trajectories which end freely near its origin. e. Representated by the element D of the figure 7, has this cell a small body with very long dendritic branches of horizontal course. These dendrites, which can be one or two at earch side, show a rugose aspect with some small collaterals, and they finish freely in delicate fillaments in a distant region from the cellular body. The axon, originated from one of the dendritic trunks, resolves inmediately in an ample plesus of fillets with rosary-like aspects and tortuous courses, limited to layer 11. Layer 12. Of cellulo-fibrillar contitution, shows this layer little morphologic variety in its cellular elements which lie mixed among the optic fibres. We shall describe these optic fibres together with those of the layer 14. The predonimant cellular type is an element of medium-size body, polygonal form and horizontal position, garnished of great and extensive dendritic trunks, which diverging in all directions cover an ample horizontal surface of the layer. Observing it in profile, offers this element a triangular form, from whose basal poles originate the dendritic trunks (Figure 2, A). On the other hand, if we observe it from the top, it shows clearly its polygonal morphology (Figure 10, A). The dendritic trunks, after suffer several bifurcations, resolve in a delicate terminal penache (Figure 10, b). The axon originated commonly from one of the dendritic trunks, distributes horizontally along the layer, covering a more or less wide surface. A variant of this cellular type is the element B of the figure 10, of small body, bipolar, and that possesses two dendritic trunks, which covers with its ramifications an ample horizontal surface. The behavior of the axon resembles that of the former type. The cell A of the figure 5C, represents one of the elements described in first place, and due to it is in a different plane of that of the photograph, we do not appreciate its details. Layer 13. From a strict morphologic viewpoint, three are the cellular types we have seen in this layer. a. Possesses this element a small body, bipolar, from whose poles originate two strong dendritic trunks, which run through extensive trajectories along the layer. Presenting few divisions, have these

trunks a rugose aspect produced by small protoplasmic knobs, some of them longer, likely fringes. They end commonly through little distorted branchlets which unify to the dense plexus of this layer (Figure 7, A). Its axon, departing from one of the protoplasmic trunks, offers collaterals for this layer and resolves in the zone 12 into fillets which, due to the existing net of this layer, we can not pursue it in a long stretch. b. Through its dendritic distribution, is this element similar to the former, but its body, of greater size, located in contact with the layer 14, is pyriform, presenting a lower pole from which originate two dendritic trunks which taking opposite directions, ramify along the layer. The axon, differing from the former type, resolves horizontally only in this layer (Figure 8, B). c. Of small body, shows this element one or two short dendrites that resolve near the cellular body into a multitude of distorted branchlets which form a dense dendritic net, limited to this layer. Unfortunately, we can not observe its axon. Layer 14. In this place we describe the optic fibres that reach the tectum opticum through this and 12 layers, and we shall do it according to their points of arrival. a. F i b r e s of t h e l a y e r 14. These fibres descend vertically until reach the layers 13, 12 and 11. Those fibres that arrive to layer 13 (Figure 11, A and B) resolve in this layer through a penache of delicate and distorted branchlets, with small knobs in their course. This terminal penache can, sometimes, be very long, as is the case of the fibre B, and comprises large extensions. The fibres that descend to the layer 12 (Figure 11, C), are less frecuent and they run through a short longitudinal distance, and their terminal penache is less dense than the former. The fibres that reach the layer 11 (Figure 11, D and E) are similar to those that arrive to layer 13, and possess an ample terminal penache, which forms large longitudinal nets intermingled with the dendritic mesh of the cells of this and the lower layers. Sometimes, we have observed that after offers the fibre a terminal penache of delicate fibrills, a slender prologation that departs from it, travels through a more or less long space for end forming a secondary penache at certain distance from the first (Figure 11, a). Some fibres, as is the case of the fibre F of the figure 11, descend until reach the layer 11, and witout

The tectum opticum of the Pantodactylus

Fig. 5 . A : Crook axon cell. The apical dendritic trunk ascends in a sinuous course until bifurcate at the level of layer 10. The resulting branches reach layer 12 where they disappear. One observes the slender basal dendrites, but not the axon because it is too fine. 8 to 12. Layers of the tectum. B : This photograph shows two elements located in different layers. The element a lies in the upper region of layer 7 and gives a basal and an apical dendritic trunks. The apical trunk, gives some collaterals during its course, and then it reaches the layer 13. The element b lies in the layer 10 and shows a conspicuous basal dendrite, that ramifies in layer 9. This cell possesses also an apical dendrite that we only see as a blurred shadow, which ascends ramifying in layer 11 and reaching its upper branches, of which we observe some of them, the layer 12. 7 to 13: Layers of the tectum. Layer 12 because its great impregnation, appears black in the figure. C. This photograph shows optical fibres and the strata they form. a) stratum 1 (layer 13). b) stratum 3 (layer 11). c) stratum 4 (layer 9). In the stratum 2 one only can observe few fibres. A. Polygonal cell of layer 12. 11 to 14 : Layers of the tectum.

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take a longitudinal direction, spread out in a terminal plexus of delicate fibrills, which, in part, reach the layer 10. b. F i b r e s of t h e l a y e r 12. Two are the fibre types we observed in this layer, namely, those that end in it, and those that descend to layer 9. The former end in the layer 12 through a short penache of delicate fibrills and distorted branchlets (Figure 11, I). This was the less frecuent type. Those fibres that finish in the layer 9 can be collaterals (Figure 11, G) or terminals (Figure 11, H). In the layer 9, they ramify in 2 or 3 principal trunks, which little by little resolve in each time more slender branches, and end forming an ample and dense plexus of distorted fillets, which show small knobs ih their courses and possess lateral spines garnished with terminal enlargements. These fibres descend at different levels of layer 9, and the plexus that they form covers perfectly the whole width of the layer, unifying to the dense dendritic plexus that exists there. As we can see, four are the strata formed by the optic fibres, namely S t r a t u m 1: It covers the layer 13, and is formed by the fibres arrived from the layer 14 (Figure 5C, a).

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10

Fig. 6. This figure shows the ramification in layer 12 of an axon, which originates in a very high level from a dendritic branch of a cell of layer 8.

Fig. 7. Semischematic drawing of the cells of the superficial layers. 7 to 13: Layers of the tectum. A. Horizontal cell with long and rugous dendritic branches. I t is located in the layer 13 and its axon ramifies in this and the 12 layers. B . Stellate cell of layer 11 with descendent axon. C. Cell with apical and basal tortuous dendrites which ramify in the adjacent layers. The axon, originated from the basal part of the cellular body, ramifies in the layers 11 and 10. D. Horizontal element of layer 11 with long opposite dendrites and an axon that form a very complicate net within the layer. E . Element located in the lower region of layer 9 with long ascendent and horizontal dendrites. One of the ascendent dendrites reaches the layer 13. The axon with a main horizontal course, travels through layer 8 sending some collateral to layer 9. F. Pyramidal element of layer 9. Its apical dendritic trunk, which reaches the layer 13, is cut for not overload the figure. Its axon, of descendent course, gives collaterals during its passage throughlayer 8, which travel in an horizontal direction. a) axons. b) collaterals to layer 8 of the axon of the cell B . c) collateral to layer 9 of the axon of the cell E . d) axonic collateral of cell F. X . cellular ghosts.

The tectum, opticum of the Pantodactylus

Fig. 8. Semischematic picture of the cells of the superficial layers. 6 to 13: Layers of the tectum. A. Small element of layer 13 with very ramified and tortuous dendrites. B. Pyriform cell with two basal dendrites and an horizontal axon. The dendrites take opposite directions and travel through long distances in layer 13. C. Medium-size element of the layer 11 with small apical dendrites, which resolve in layer 12, and a basal dendritic trunk, which divides in T giving ascendent and descendent branches. The axon, which originates from the basal trunk, with a descendent course, resolves in layer 7. D. Cell with a rugous cellular body, that possesses a basal and an apical dedritic trunks which ramify in layers 9 and 13 respectively. The axon, originated from the basal trunk, ramifies in layer 9. E. Small cell whose dendrites resolve in layer 12. The axon, in an horizontal course, travels through the boundary of layers 11 and 10, giving some collaterals to layers 11 and 12. F. Small element with descendent axon. This cell shows an apical dendrite resolved in layer 12 and slender ones finished near the cellular body. The axon gives some collaterals in the upper region of layer 7 and in layer 8, which take ascendent or horizontal tortuous directions.

125

G. Pyramidal cell of layer 10. The dendritic trunks, one apical and the other basal, ramify in layers 13 and 9 respectively. The apical gives collaterals during its course through layer 12. The axon, of the descendent course, sends collateral during its passage through layer 9. H. Triangular element of layer 10. The apical dendrites reach the layer 12. The strong basal dendrites travel through layer 9 long distances, where give collaterals which ramify profusely forming an ample and dense net that sumes up to the plexus of this layer. The descendent axon gives during its passage through layer 9 an ascendent collateral for layer 12, and then, in layer 8, send other collaterals which with an horizontal course, travel through long distances in layer 8. a) axons. b) horizontal axonic collateral for layer 8 of cell H. c) axonic collateral for layer 7 of cell F. d) horizontal course of the axon of cell E. e) ascendent axonic collateral of cell F. f) ascendent axonic collateral of cell G. g) horizontal axonic collateral of cell G. h) secondary branch of g. i) ascendent axonic collateral of cell H. z) strong descendent dendrite to layer 9 of cell H. X . cellular ghosts.

126

Quiroga, J . C.

Fig. 9. Stellate cell of layer 10 whose dendritic branches cover the layers 9, 10, 11, 12 and 13. The axon, taking an almost opposite direction, ramifies profusely in the layers 7, 8 and 9. 7 to 13: Layers of the tectum, a) axon.

The tectum opticum of the Pantodactylus

Fig. 10. Cells of layer 12 that we see in their plane bution. The cell A, of polygonal form, has dendritic which after a long courses resolve in a penache of fillets. Cell B, bipolar, shows similar features in its branches. The axons of both cells ramify in the layer 12. a) axons. b) terminal dendritic penaches of cell A. c) terminal dendritic penaches of cell B.

of distribranches tortuous dendritic plane of

S t r a t u m 2: It fills the layer 12 and is formed by fibres that arrive from this and the 14 layers. S t r a t u m 3: It occupies the layer 11 and part of the 10, and is formed by fibres of the layer 14 (Figure 5C, b). S t r a t u m 4: It covers the layer 9 and is formed by fibres arrived from layer 12 (Figure 5C, c).

127

In the figure 5C one does not observe with clarity the stratum 2, although some fibres run through it. Only in few cases, we have observe the terminals of this stratum, which were the less frecuent.

Discussion

Our findings coincide in general to those of R A M Ó N ( R A M O N 1891, 1894 and 1896). In what import to the cells of the layers 3 and 5, save the greater ones (Figure 1, G and I), those described by Ramón are similar to those of Pantodactylus. The same is for the ganglion and plumed cells, except in the fact that the terminal penaches of the dendrites of the latter, do not finish at the same level in the superficial layers as is the case in Chamaleon. The cells of layer 8, as well the crook axon cells as those of ascendent axon, are similar to those described by Ramón. However, there are elements with some differencies, such as those with an ascendent

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Quiroga, J . C.

axon which resolves in the layer 13 (Figure 4, D) and those whose axon ramifies in the layer 12 (Figure 6). The superficial layers, specially the 9, 10 and 11, are the layers with the major cellular diversity, as much that in many cases is it a very difficult task to say if one element is assimilable to this or to that cellular type. Ramón has done mention to this great diversity of cellular types, and described some of them. We think that, as well in the Chamaleon, Lacerta agilis ( R A M O N 1 8 9 1 , 1 8 9 4 and 1 8 9 6 ) , as, through the figures of B U T L E R and E B B E S S O N ( 1 9 7 5 ) , also in Tupinambis nigropunctatus, exists this polimorphismus in the superficial layers. We can not say the same words for Emys europaea. It pertains to other order and are necessary further studies. B U T L E R and E B B E S S O N ( 1 9 7 5 ) refer that there are not, in Tupinambis nigropunctatus, cells that located in the layers 3 or 5 reach the superficial layers through an apical dendritic trunk. Tupinambis pertains, as Pantodactylus, to the Family Teiidae, and therefore, we think, in first instance, that here there is a technical problem. The G O L G I ' S method needs many essays to produce good results, and these authors have employed very few animals for their work. It is evident, on the other hand, the relationship between this cellular polimorphism and the optic terminals. The strata formed by the optic fibres fill the layers 9 to 13, and is in these layers where we find this great variablity. This cellular polimorphismus can be observed in the superficial layers of the regions, save structures as the nucleus opticus basalis, are under the influence of the tractus opticus (QUIROGA 1977). In this manner, we have observed cells of similar morphology to those of the layers 13 and 11 in the nucleus geniculatus lateralis (QUIROGA 1977) and in the nucleus geniculatus pretectalis (QUIROGA, unpublished observations) . On the other hand, there are some features which deserve to be commented in the cells of each superficial layer. Layer 13 possesses elements in which the axon covers a suface that form an horizontal plane (Figure 8, B) ; similar is the case in layer 12 (Figure 10, A and B). Layer 11, on the other hand, shows elements in which the distribution of the axon covers, forming a net, a volume in the layer (Figure 7, C and D). In layer 10 we find elements, as is the case of the cell E of the figure 8, whose axon, with a linear course, connects in series the cells of this layer or the dendrites that run through it. Other example is that represented in the figure 9. In this case, the

axon, descending towards lower layers, adopts a distribution almost opposite to that of the dendrites. In this manner, while the cell receives impulses from a determinate vertical column, affects through its axon, an adjacent column. This can have certain physiologic value. In the layer 9, the element E of the figure 7, of horizontal type, connects in series the cells of the layer 8. In the layers 9, 10 and 11 exist cells whose axon is descendent. Some of these axons emit a multitude of collaterals that form a tridimensional net (Figure 7, B), while others show collaterals which travel, in a more or less linear courses, within the layer 8 (Figure 7, F; figure 8, F, G and H). The cells of short axon which ramifies within the superficial layers, such as the elements E and C of the figure 8, the E of the figure 7 and the represented in the 9, can have excitatory or inhibitory functions and they can act as the stellate cells of the cerebral cortex or the basket cells of the cerebellum (ECCLES 1969).

Fig. 11. Semischematic p i c t u r e of t h e superficial, t h a t is t o say optical, a n d p r o f o u n d a f f e r e n t s of t h e t e c t u m o p t i c u m . 1 to 14: Layers of t h e t e c t u m . A a n d B. Optic fibres which end in layer 13, f o r m i n g t h e first s t r a t u m of optic terminals. C. Optic fibre for t h e second s t r a t u m (layer 12). D a n d E . Optic fibres which resolve in t h e layer 11 a n d f o r m t h e third s t r a t u m . F. F i b r e t h a t ends in t h e s t r a t u m 3, within t h e layers 11 a n d 10.

G. Optic collateral t h a t descends until reach t h e layer 9, where is t h e f o u r t h s t r a t u m of optic terminals. H . T e r m i n a l optic fibre for t h e s t r a t u m 4. I. T e r m i n a l p e n a c h e of a fibre of layer 12 in t h e second stratum. J a n d K. Deep fibres for layer 7. T h e fibre J a p p e a r s to end w i t h o u t collaterals. I t s t e r m i n a l p e n a c h e reaches t h e layer 8 w i t h slender branches. On t h e o t h e r h a n d , fibre K gives some collaterals d u r i n g its course t h r o u g h layer 6 which r a m i f y a t different level of layer 7. L. F i b r e t h a t sends collaterals t o layers 7 a n d 6. M. D e e p fibre t h a t ends in t h e lower layers a f t e r give one collateral. a) Long slender branchlets t h a t d e p a r t i n g f r o m t h e t e r m i n a l p e n a c h e of t h e optic fibre D in layer 11, f o r m s a secondary a n d smaller p e n a c h e a t certain distance f r o m t h e former. b) c) d) y) z)

ascending collaterals of t h e deep fibres. collateral of fibre L t h a t ends within layer 6. descending collateral of fibre M. t e r m i n a l resolution of fibre K. t e r m i n a l penache of fibre M.

The tectum opticum of the Pantodactylus

129

130

Quiroga, J . C.

Fig. 12. Schematic representation of the interconnections of the elements of the superficial layers. 6 to 14: Layers of the tectum. A. Opposite axon cell of layer 10. B. Descendent axon cell of layer 10 with an horizontal collateral to layer 8. C. Horizontal axon cell of layer 10. D. Horizontal axon cell of layer 9. E. Cells of ascendent axons to the optic tract. F. Crook axon cells. a) axons. b) horizontal axonic collateral of cell B. o) optic fibres.

In the figure 12 we have represented some of these cellular types and their interconnections. The physiologists shall be who put the symbols + or — in the synapses of the cells of the layers 9 and 10. For the cellular connections in other layers, may be consulted C A J A L ( 1 9 7 2 ) . As the same manner that with the stellate cells of the superficial layers, we do not yet know the function of the cells with ascendent axons to the tractus opticus. In what concerns to the distribution of the optic fibres, it is important to note that, while the strata 1 and 3 (layers 13 and 11) are formed only by fibres of the layer 14, the stratum 4, on the other hand, only receives optic afferent from layer 12. To the stratum 2, which is the more slender and contains

The tectum opticum of the Pantodactylus

the lesser quantity of optic terminals, arrive fibres of both the layers 14 and 12. MATURANA et al. ( 1 9 6 0 ) r e l a t e o n t h e e x i s t e n c e of

5 types of ganglion cells in the retina of the frog, whose axons finish in four strata in the superficial layers of the optic lobe. Perhaps, a physiologic study of this type in reptiles can correlate the here described optic strata with different types of retinal ganglion cells. In what concerns to the optic collaterals (Figure 11, G), only further studies can show the magnitude of its importance.

Bibliography ARIENS

KAPPERS, C. U.,

G . C. H U B E R

and

E.C.CROSBY:

The comparative anatomy of the nervous system of vertebrates, including man. Vol. 2. Hafner Publishing Company, New York (1967). Originally published in 1936. B U T L E R , A. B . , and R . G. N O R T H C U T T : Ascending tectal efferent projections in the lizard Iguana iguana. Brain Res. 35, 5 9 7 - 6 0 1 (1971). B U T L E R , A. B., and O. E . E B B E S S O N : A Golgi study of the optic tectum of the tegu lizard, Tupinambis nigropunctatus. J . Morph. 146, 215 228 (1975). B U T L E R , A. B . : Ascending efferent tectal projections in the lizard Gekko gecko. Neurosci. Abstr. 2 (1976). C A J A L , S. R . y : Histologie du système nerveux de l'homme et des vertébrés. Consejo Superior de Investigaciones Científicas, Madrid (1972). Originally published in 1911. C A J A L , S. R. y, y F. de C A S T R O : Elementos de técnica micrográfica del sistema nervioso. Salvat, Madrid (1972). Originally published in 1933. E C C L E S , J . C. : The inhibitory pathways in the central nervous system. Liverpool University Press (1969). E I C H L E R , V. B . : Neurogenesis in the optic tectum of larval Rana pipiens following unilateral enucleation. J . Comp. Neurol.'141, 3 7 5 - 3 9 6 (1971). K R A N Z , D., und W. R I C H T E R : Autoradiographische Untersuchungen zur Regeneration des Tectum opticum von Lebistes reticulatus (Teleostei). Z. mikrosk.-anat. Forsch. 84 4 2 0 - 4 2 8 (1971). LAVIÑA, J . C . : Estabilización de las coloraciones cromoargénticas. Arch. Hist. norm. pat. 1, 441 — 442 (1943). MATURANA, H . R . ,

W. H.

PITTS:

J . Y . LETTVIN,

W . S . MCCULLOGH,

and

Anatomy and Physiology of vision in the

131

frog [Rana pipiens). J . gen. Physiol. 4 i, Suppl. 2, 129 bis 175 (1960). Q U I R O G A , J . C.: E l cuerpo geniculado lateral y el principio de las capas en el tálamo de Pantodactylus schreibereii Wiegmann (Lacertilia, Reptilia). Obra del Centenario del Museo de La Plata (in press, 1977). R A F F I N , J . - P . : Quelques effets de l'ablation uni- ou bilatérale de l'ébauche optique sur le tectum superficiel du poulet (Gallus domesticus). Acta Embryol. Exp. 45 -63 (1972). R A M O N , P. : E l encéfalo de los reptiles. Tipografía de la Casa Provincial de Caridad, Barcelona (1891). R A M O N , P. : Investigaciones micrográficas en el encéfalo de batracios y reptiles. Cuerpos geniculados y tubérculos cuadrigéminos de los mamíferos. Establecimiento tipográfico " L a Derecha", Zaragoza (1894). R A M O N , P. : Estructura del encéfalo del camaleón. Rev. trimest. micrográf., 1, 46 — 82 (1896). R I C H T E R , W., und D. K R A N Z : Die Abhängigkeit der DNSSynthese in den Matrixzonen des Mesencephalons vom Lebensalter der Versuchstiere (Lebistes reticulatus — Teleostei). Autoradiographische Untersuchungen. Z. mikrosk.anat. Forsch. 82, 7 6 - 9 2 (1970). R I O H O R T E G A , P. del: Tercera aportación al conocimiento morfològico e interpretación funcional de la Oligodendroglia. Arch. Hist. norm. pat. 6, 1 3 2 - 1 8 3 (1956). SENN, D. G.: Über das optische System im Gehirn squamater Reptilien. Acta anat. Suppl. 52 = 1 ad Vol. 65 (1966). SENN, D. G. : Bau und Ontogenese von Zwischen - und Mittelhirn bei Lacerta sicula (Rafinesque) Acta anat. Suppl. 55 = 1 ad Vol. 71 (1968a). SENN, D. G. : Der Bau des Reptiliengehirns im Licht neuer Ergebnisse. Verhandl. Naturi. Ges. Basel 79, 25 - 4 3 (1968b) SENN, D. G. : The stratification in the reptilian nervous system. Acta anat. 75, 5 2 1 - 5 5 2 (1970). V A I S A M R U A T , V., and A . H E S S : Golgi imprégnation after formalin fixation. Stain tech. 28, 303 — 304 (1953).

A ddress : Dr. Juan C . Q U I R O G A Division de Paleontología de Vertebrados Facultad de Ciencias Naturales y Museo Universidad Nacional de L a Plata Paseo del Bosque 1900 L a Plata Buenos Aires Argentina

j . Hirnforsch. 19 (1978) 133 — 14i

Department of Anatomy and Laboratory of Electron Microscopy, Institute of Medical Biology, School of Medicine, Gdansk, Poland

The Distribution of Axon Terminals with Flattened Vesicles in the Nuclei of the Amygdaloid Body of the Cat By O. NARKIEWICZ,

Jolanta

JURANIEC

and Teresa

WEZOIKOWA

With 11 figures and 1 table (Received January 30, 1977) Summary: The morphology of synapses in the amygdaloid nuclei was studied in 10 cats. On the basis of the percentage of axon terminals with flattened vesicles (F-type) nuclei were distinguished, in which these terminals are as sparsely distributed as in most areas of the central nervous system, from other nuclei in which they are abundant (about one-third to one-half of all synaptic boutons). The lateral, basal dorsal and basal ventral nuclei belong to the first, the medial and central nucleus and the anterior amygdaloid area — to the second group. The cortical nucleus, which generally has a small number of boutons of F-type has some parts seemingly belonging to the first, and others to the second group. In all amygdaloid nuclei axon terminals of F-type form symmetrical synaptic contacts. In nuclei with a low percentage of F-type terminals these boutons are predominantly small and synapse either with perikarya or with large dendrites. The amygdaloid nuclei having numerous F-type terminals contain not only small but also larger terminals with flattened vesicles. Both, the larger and smaller axon terminals form in these nuclei synaptic contacts with various parts of dendrites even with very small ones and with dendritic spines. The subdivision of amygdala into two parts, one with a low and another with a high number of F-type boutons would seem to support the hypothesis that amygdala may be subdivided physiologically into a dorsomedial — "excitatory" and basolateral — "inhibitory" portion.

Introduction

Publications of H A L L (1972) and W A K E F I E L D and (1974 a) and our own studies ( J U R A N I E C et al., 1974b, 1975; N A R K I E W I C Z et al., 1975) have shown that various parts of the amygdaloid body differ in respect to amount of F-type synapses. We tried to establish, whether these differences are consistent in various amygdaloid nuclei and if so, how they correlate with the cytoarchitectonics of the amygdala. On the other hand it would be interesting to see whether the frequency of occurrence of F-type synapses had any relation to the putative functional significance of individual nuclei of the amygdaloid body in relation to the behaviour of animals. Behavioral studies on changes in drives resulting from stimulation or damage of the amygdaloid body suggest that there are striking differences between its main parts. According to some authors (FONBERG, 1965, 1966, 1967, 1969, 1971, 1974; RICHARDSON, 1973) it is even possible to differentiate two antagonistic portions of the amygdaloid body; one predominantly excitatory and the other inhibitory. Attempts to establish the relation between functions and synaptic structure of these portions seem to be of essential significance for explaining some functional mechanisms of the limbic system. HALL

Synaptic boutons with predominantly flattened (Ftype) and spherical (S-type) vesicles have been distinguished since the description of UCHIZONO ( 1 9 6 5 ) . Despite some observations, showing that the shape of vesicles depends on fixation and subsequent treatment of the material ( B O D I A N , 1 9 7 0 ; L U N D andWESTRUM, 1 9 6 6 ; W A L B E R G , 1 9 6 5 ) the differences between both types of axon terminals seem to be important not only from a morphological but also from a physiological point of view. As was suggested by Uchizono axon terminals containing spherical vesicles are often found to be excitatory, while those with flattened vesicles inhibitory. This suggestion, although not having been proved unequivocally, has much evidence in support. In most areas of the central nervous system F-type boutons are far less numerous than those of S-type; generally they do not exceed 1 0 — 1 5 % of all axon terminals. However, there are some areas of brain and spinal cord in which boutons with flattened vesicles are more numerous and in some even predominant ( B O D I A N , 1 9 6 6 ; R I N V I K and G R O F O V A , 1 9 7 0 ; NAKAJIMA, 1 9 7 1 ; WESTRUM a n d BLACK, Hirnforschung, Bd. 19, Heft 2

1971).

10

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Narkiewicz, O., J . Juraniec and T. Wrzolkowa

Fig. 1. On the left side (A) frontal sections of the amygdala from rostral to caudal showing topography of the amygdaloid nuclei. A — anterior amygdaloid area, B D — basal dorsal nucleus, B V — basal ventral nucleus, CL — lateral part of central nucleus, CM — medial part of central nucleus, CO — cortical nucleus, L — lateral nucleus, M — medial nucleus, TO — nucleus of the lateral olfactory tract. On the right side (B) on respective cross —sections places from which specimes were taken showing percentage of the boutons with flattened vesicles, x 2, x 3 , x 4 show number of specimens taken from the same place. Note striking difference in percentage between the basolateral and dorsomedial part of the amygdala.

OD/ â < 20% 20% - 3 5 %

IP 35% - 50% Ä - > 50%

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Narkiewicz, O., J . J u r a n i e c and T . Wrzolkowa

Material and Methods 10 healthy adult cats both sexes were used in the study. Their weights ranged 2,5 — 3,5 kg. Under Nembutal anaesthesia (45 mg/kg) cats were perfused through the left ventricle of the heart with 0 , 9 % NaCl containing 0 , 5 % solution of xylocaine at 37 °C. This was followed mmediately b y about 1 500 ml of 4 % paraformaldehyde in 0,05 M phosphate buffer according to KAWANA et al. (1971). T h e perfusate was adjusted to pH 7,4 at room temperature. T h e procedure performed under controlled pressure lasted about 30 minutes. As soon as perfusion fixation was achieved the brain was removed from the skull and cut into 1 — 2 mm thick coronal slices b y means of razor blades. In chosen slices a needle of 1 mm diameter was driven bilaterally through the various amygdaloid nuclei: lateral, basal dorsal, basal ventral, medial, cortical, both parts of the central nucleus and anterior amygdaloid area. E a c h core of amygdaloid tissue removed with the needle (45 cores) was transferred directly into a glutaraldehyde solution. T h e rest of each section was stored in 1 0 % formaline for control under the light microscope. Control sections 15 to 20 (im thick were stained with Cresyl — violet. T h e localization of cores was indicated in the appropriate diagrams and shown in Fig. 1. Specimens encompassing parts of more than one cytoarchitectonic subdivision were discarded. Pieces of amygdaloid tissue taken for electron microscopic investigation were kept in 6 , 5 % glutaraldehyde a t 0 °C in 0,1 M phosphate buffer at p H 7,4 with 8 % sucrose for 1 hour. N e x t they were washed for 15 minutes in a phosphate buffer of the same parameters and postfixed in 1 % osmium tetroxide in the same buffer for 1 hour. After dehydration through a series of graded alcohols and propylene oxide the blocks were embedded in Epon 812. Thin sections were cut with glass knives with an L K B microtome and stained with uranyl acetate (WATSON, 1958) and lead citrate (REYNOLDS, 1963). T h e specimens were examined and photographed with a J E M 7A electron microscope. F o r quantitative analysis the axon terminals were counted on electron micrographs in about 42,000 final magnification. A total of 17020 axon terminals of F - t y p e and S-type were

counted on micrographs at random from all sections. T h e mean values and standard deviations were determined separately for each nucleus (Table 1).

Results The subdivision of the amygdaloid body of the cat into various nuclei was first described by F o x (1.940). We adopted this nomenclature with some modifications (NITECKA et al., 1973). On a mainly phylogenetic basis it is customary to subdivide the amygdala into a basolateral and a corticomedial part. The basolateral part consists of the (a) lateral, (b) basal dorsal and (c) basal ventral nucleus. The corticomedial part contains (a) the central nucleus, which may again be subdivided into two or even three parts — subnuclei, (b) the medial nucleus and (c) the cortical nucleus. The latter has distinct morphological features making it partly similar to the prepyriform cortex and partly to the basal ventral nucleus. For this reason the structure of the cortical nucleus will be discussed separately; the remaining nuclei — the medial and central will be decribed as the dorsomedial part of the amygdala. The latter merge in the anterior direction with the anterior amygdaloid area, that can be considered either as a separate structure or as a part of the corticomedial complex. The topographic relations of the amygdaloid nuclei and their positions on the frontal sections are shown in Fig. 1. 1. Basolateral part of the amygdala The percentage of synaptic boutons with flattened vesicles in all the nuclei of this group (lateral, basal

T a b l e 1. Frequency of occurrence of terminals with flattened vesicles in the nuclei of the amygdaloid body

Basolateral part of amygdala

Dorsomedial part of amygdala

Nucleus

Number of cases

Number of terminals

% of terminals with flattened vesicles

Basal dorsal nucleus B a s a l ventral nucleus Lateral nucleus

4

2091

11,9

±1,24

5

1739

13,3

±3,30

6

3381

9,9

±3,20

5

1083

39,2

±8,08

5

1102

30,8

±5,56

6

1736

39,4

±6,13

Cortical nucleus

5

2 797

17,7

±9,74

Anterior amygdaloid area

9

3 091

51,5

±12,75

Medial part of the central nucleus Lateral part of the central nucleus Medial nucleus

standard deviation of the mean

The distribution of axon terminals in t h e amygdaloid body

Fig. 2. Synaptic bouton containing flattened vesicles (F) form symmetrical synaptic contact with a dendritic shaft. Lateral nucleus. Fig. 3. Asymmetrical synapse with several contact areas. Large bouton filled with spherical vesicles (S) contacts with dendrite of medium size (D). Lateral nucleus. Fig. 4. An axosomatic synapse; axon terminal contains mostly flattened vesicles (F). P — perikaryon. Medial p a r t of t h e central nucleus.

137

dorsal and basal ventral) is small, and differences between particular nuclei are in this respect statistically insignificant. Also within each of the above nuclei the number of boutons with flattened vesicles is rather constant regardless from where the specimen is taken from. Individual specimens taken from the basolateral complex differ, within the relatively narrow limits of 7,1.% to 17,4%.

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Narkiewicz, O., J. Juraniec a n d T. Wrzolkowa

Boutons containing flattened vesicles are generally small — 0,6 ^m to 1,5 (im in diameter. The vesicles are rather uniformly distributed only partly clustering near presynaptic membrane. In general they are flat, of similar size and shape, although vesicles only slightly oval or spherical were also observed in places. Synaptic contacts of boutons with postsynaptic structures are relatively short, usually without visible density on the postsynaptic side. These contacts correspond generally to the usual description of symmetrical synapses (Fig. 2). Boutons with flattened vesicles generally form synapses with perikarya of neurons or with large proximal parts of dendrites. Sometimes they contact thinner dendritic shafts, b u t were never found synapsing with dendritic spines. Since the terminals of F-type are so few in all nuclei of the basolateral part of amygdala, the vast majority of boutons are those containing spherical vesicles (boutons of S-type). Their appearance is, on the whole, typical and they form asymmetrical synapses with pronounced postsynaptic densities. Spherical vesicles are present not only in small boutons contacting secondary dentritic shafts and their spines b u t also in larger ones of over 2 ¡xm diameter. In the lateral nucleus the large axon terminals surround dendritic shafts, forming several synaptic contact areas (Fig. 3). 2. Dorsomedial part of the amygdala In contrast to the basolateral nuclei, we observed m a n y boutons with flattened vesicles in the dorsomedial part of amygdala (medial and central nucleus; the latter subdivided into medial and lateral subnucleus). They contribute from 27,1% to 50% of the total. The differences between the means for each nucleus or its parts are not very considerable, although in the medial nucleus and the medial part of the central nucleus they are more numerous than in the lateral. In the latter there are many more t h a n in the neighbouring structures: the putamen and the nuclei of the basolateral complex of the amygdala. Unlike in the basolateral complex, these boutons are often large and occasionally very elongated. The vesicles are distributed comparatively densely, some are only slightly oval others even spherical. Boutons with flattened vesicles establish symmetrical contacts usually. Some of them with the perikarya (Fig. 4), b u t there are m a n y more on the dendritic shafts and on thinner branches of the dendrites. Boutons with flattened vesicles m a y also establish contacts with dendritic spines, generally by means of symmetrical synapses. As a rule, one synaptic bouton was in contact with one spine (Fig. 5), rarely one spine was connected with several boutons (Fig. 6).

At the base of the dendritic spine contacted b y an asymmetrical synapse with a bouton containing spherical vesicles a bouton containing flattened vesicles was encountered forming an asymmetrical synapse with a dendritic shaft. However, in the dorsomedial part of the amygdala dendritic spines are not numerous; the neuropil consist mainly of smooth dendrites of varying calibers surrounded by boutons containing flattened or spherical vesicles. In the central and medial nucleus, similarly to the basolateral part of the amygdala, axon terminals synapsed with one postsynaptic structure only. In some places F-type terminals were observed to form synapses with two postsynaptic elements (Fig. 7, 8). Hower, since these observations were made on random single sections and not on series, not much can be deduced from this. Axon terminals of S-type are similar to those in the basolateral part of amygdala. 3. Cortical nucleus I n the cortical nucleus the number of boutons with flattened vesicles varied in different specimens. It depended probably on the place from which the samples were t a k e n : most of these boutons were found in samples from the anterior part of the cortical nucleus near the anterior amygdaloid area (36,1%), fewer in specimens taken from its posterior part (9,8%—19%). Apparently the superficial and deep parts of the cortical nucleus also differ in this respect. Because of the heterogeneity of the cortical nucleus and varied synaptic structure of its parts, further detailed studies based on larger material are necessary. Nevertheless, on the basis of our investigations it would seem t h a t in the cortical nucleus we m a y distinguish large areas in which the synaptic structure resembles t h a t in the dorsomedial part of the amygdala. Boutons with flattened vesicles in the cortical nucleus are generally small, particularly in samples

Fig. 5. A dendritic spine (DS) is contacted by axon terminal containing predominantly flattened vesicles; a symmetrical synapse showing, however, slight postsynaptic density (pd). Medial p a r t of t h e central nucleus. Fig. 6. Three synaptic boutons containing flattened vesicles (F) form symmetrical synaptic contacts with a dendritic spine (DS). Medial nucleus. Fig. 7. Bouton containing flattened vesicles (F) synapses with two dendrites of medium size (D). Medial nucleus. Fig. 8. Bouton containing flattened vesicles makes synaptic contacts both with perikaryon (P) and dendritic s h a f t (D). Medial nucleus.

lug. 5 - 8

140 Narkiewicz, O., J. Juraniec and T. Wrzolkowa

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The distribution of axon terminals in the amygdaloid body

taken from its posterior part. In the anterior part, in which boutons with flattened vesicles are more numerous, some of them are of larger size. Axon terminals with flattened vesicles form symmetrical synapses on the surfaces of the perikarya and on the proximal and distal parts of dendrites (Fig. 9). Boutons with spherical vesicles were found mainly on dendritic spines, although at times we observed them even on the surface of the perikarya, their synapses having always a distinct postsynaptic density. 4. Anterior amygdaloid

area

A characteristic feature of this area is the presence of many synaptic boutons with flattened vesicles. They are even more numerous here than in the dorsomedial part of the amygdala, although in this respect we found major differences between samples. In the most superficial part the percentage of boutons with flattened vesicles was the lowest (26,8%), the remaining samples taken from the depth of anterior amygdaloid area contained 42,8% to 72,9% of such boutons. In the anterior amygdaloid area small synaptic boutons with flattened or spherical vesicles predominate, like in other nuclei of amygdala. As a rule large boutons of over 1,5 ¡¿m in diameter and up to 3,5 ^m, contained flattened vesicles. This type of very large boutons was not found in the basolateral part of amygdala and was less numerous in its dorsomedial part. As in other amygdaloid nuclei, boutons containing flattened vesicles usually form typical symmetrical synapses, most often with dendrites of varying thickness. Similar as in the nuclei of the dorsomedial complex of amygdala we often found in the anterior amygdaloid area dendritic shafts of medium size cut transversally or longitudinally (Fig. 10), that were surrounded by numerous boutons containing flattened vesicles. Boutons with flattened vesicles are also found on the perikarya and the proximal parts of dendrites mainly singly, more rarely as groups of axon terminals. Among them one could see single ones with spherical vesicles. In the anterior amygdaloid area, as it was the case in the central and medial nuclei, boutons of F-type were observed to synapse

Fig. 9. Axodendritic synapse in contact with a dendritic shaft (D); two symmetrical contact areas (arrows). Axon terminal containing fattened vesicles (F). Cortical nucleus. Fig. 10. Longitudinal section of a dendrite (D) surrounded by boutons containing mostly flattened vesicles (F). Anterior amygdaloid area. Fig. 11. Axon terminal with a large number of granular vesicles. Anterior amygdaloid area.

141

with dendritic spines. In boutons with predominantly flattened or spherical vesicles there are a relatively small number of nucleated vesicles, but some axon terminals are filled with them (Fig. 11).

Discussion It is apparent from these observations that the dorsomedial part of the amygdala differs significantly from the basolateral part with regard to frequency of boutons with flattened vesicles. In the basolateral nuclei of amygdala as in the majority of other telencephalic structures i.e. claustrum J U R A N I E C et al., (1971, 1974a), striatum (Fox et al., 1971; K A W A N A et al., 1971), visual cortex (COLONNIER, 1968), prepyriform cortex (WESTRUM, 1970) boutons with flattened synaptic vesicles contribute only to a small percentage of the total number of boutons in the neuropil, while in the dorsomedial part of amygdala they comprise over 30%. This agrees, on the whole, with the results of H A L L (1972), and W A K E F I E L D and H A L L (1974a) who found a large number of Ftype boutons in both parts (subnuclei) of the central nucleus but few only in the lateral nucleus. Our results concerning the medial nucleus of the amygdala differ somewhat, but this may be explained by the use of a different fixation technique. Such large numbers of boutons with flattened vesicles are found only in certain parts of the central nervous system. B O D I A N ( 1 9 6 6 ) stated that in ventral horns of spinal cord they comprise 46% of all the boutons. In the spinal trigeminal nucleus according to W E S T R U M and B L A C K ( 1 9 7 1 ) they contribute to about 58%, in the pars reticulata of the substantia nigra even to 9 0 % ( R I N V I K and GROFOVA, 1970).

It is generally thought that the shape of synaptic vesicles depends mainly on the method of fixation. According to L U N D and W E S T R U M ( 1 9 6 6 ) and W A L B E R G ( 1 9 6 5 ) the vesicles flatten during perfusion after the use of aldehyde fixatives. N A K A J I M A ( 1 9 7 1 ) found that paraformaldehyde causes flattening of vesicles to a greater degree than glutaraldehyde. B O D I A N ( 1 9 7 0 ) on the other hand, supposes that it depends on the rinsing in cacodylate buffer and not on the method of fixation. According to V A L D I V I A ( 1 9 7 1 ) the osmolarity of fixative would be the decisive factor. Irrespective of these factors/rigid maintenance of the same conditions of fixation and futher procedure, permits comparing the frequency of occurrence of boutons with flattened vesicles in different structures. This is confirmed by the constancy of our results in various samples taken from the same area or nucleus. It has not yot been explained, which is (or are) the

142

Narkiewicz, O., J . Juraniec and T. Wrzolkowa

inhibitory transmitters in the amygdala that might be localized in the F-type boutons. No publications, so far have given any evidence for larger amount of one or another of the putative inhibitory transmitters in the dorsomedial part of the amygdala, where F-type boutons are numerous. According to B E N - A R I et al., ( 1 . 9 7 5 ) the concentration of norepinephrine in the anterior amygdaloid area and corticomedial part of the amygdala is lower than in the basolateral part. The level of dopamine is high only in the central nucleus and low in the medial nucleus. This would speak against the relation of the boutons with flattened vesicles to catecholamines in the amygdaloid body. The investigations of B R O W N S T E I N et al., ( 1 9 7 4 ) on putative transmitters in amygdala also do not permit correlating our electron microscopic results with their biochemical studies. Histochemical methods showre different intensities of acetylcholinesterase activity in various nuclei of amygdala, the strongest reaction being observed in the basal dorsal nucleus. In other nuclei the intensity of reaction differs greatly, and does certainly not show any correlation with the amount of boutons with flattened vesicles. B E N - A R I et al., ( 1 9 7 5 ) claime that the highest activity of choline acetyltransferase was found in the basolateral amygdala which contained about three times the activity found in the corticomedial part. Taking into consideration the hypothesis that flattened vesicles would contain an inhibitory substance, it might also be assumed that the transmitter is gamma-aminobutyric acid ( K I M et al., 1 9 7 1 ; P F E N NINGER,

1973;

STORM-MATHISEN,

1974;

UCHIZONO,

1 9 7 5 ) , however, nothing about its distribution in the nuclei of amygdala is known. Differentiation of amygdaloid nuclei into those containing few or many boutons with flattened vesicles is in accord with the results suggesting that in the amygdala two parts might be distinguished having antagonistic functions ( F O N B E R G , 1 9 6 5 , 1 9 6 6 , 1 9 6 7 , 1969,

1971, 1974;

RICHARDSON,

1973,

1974).

Damage to the dorsomedial part including the central nucleus is supposed to decrease both, the emotional reactions and the aggressiveness of the animal while damage to the basolateral part causes reverse reaction, expressed in excitement and hyperphagia. Stimulation of these areas evokes generally opposite results. The effects are most apparent in the dog, but occur in other animals as well. These findings support the hypothesis that the basolateral portion of the amygdala plays an inhibitory role and the dorsomedial — an excitatory one. Although this hypothesis is not universally accepted, many data speak in its favour.

It is still unknown where the parent cells of axon terminals with flattened vesicles, found in the amygdala, are situated. The investigations fo W A K E F I E L D and H A L L ( 1 9 7 4 ) and our unpublished data seem to support the view that some of them lie in the preoptic lateral area. But even extensive damage to the preoptic area and hypothalamus causes degeneration of relatively few synaptic boutons. The remainder are probably terminals of amygdaloid intrinsic neurons. In this respect it is interresting that according to TOMBOL a n d SZAFRANSKA-KOSMAL (1972) a n d

KAMAL

and T O M B O L ( 1 9 7 5 ) , the dorsomedial part of the amygdala shows less interneurons than its basolateral portion. Therefore it seems that some terminals of F-type, which are so numerous in the dorsomedial part of the amygdala, have parent cells in the basolateral nuclei. Since interneurons have inhibitory functions in many parts of the brain, it may be assumed that in this way the neurons of the basolateral "inhibitory" nuclei may inhibit the activity of the dorsomedial "excitatory" part of the amygdala.

Literature R. E . ZIGMOND and K . E . M O O R E : Regional distribution of tyrosine hydroxylase, norepinephrine and dopamine within the amygdaloid complex of the rat. Brain Research 87, 96—101 (1975). BODIAN, D.: Synaptic types on spinal motoneurons: an electron microscopic study. Bull. J . Hopkins Hosp. 119, 10 bis 45 (1966). BODIAN, D . : An electron microscopic characterization of classes of synaptic vesicles by means of controlled aldehyde fixation. J . Cell Biol. 44, 1 1 5 - 1 2 4 (1970).

BEN-ARI, Y . ,

BROWNSTEIN, M . , J . M . SAAVEDRA a n d M . PALKOVITS:

Nor-

epinephrine and dopamine in the limbic system of the rat. Brain Research 79, 4 3 1 - 4 3 6 (1974). COLONNIER, M . : Synaptic patterns on different cell types in the different laminae of the-cat visual cortex. An electron microscope study. Brain Research 9, 2 6 8 - 2 8 7 (1968). FONBERG, E . : Effect of partial destruction of the amygdaloid complex on the emotional-defensive behaviour of dogs. Bull, de l'Acad. Pol. Scien. CI. II, 13, 4 2 9 - 4 3 2 (1965). F O N B E R G , E . : Aphagia, produced by destruction of the dorsomedial amygdala in dogs. Bull, de l'Acad. Pol. Scien. CI. II, 14, 7 1 9 - 7 2 2 (1966). F O N B E R G , E . : The role of the amygdaloid nucleus in animal behaviour. Progr. Brain Research 22, 2 7 3 - 2 8 1 (1967). E . : The role of the hypothalamus and amygdala in food intake, alimentary motivation and emotional reactions. Acta Biol. Exp. 29, 3 3 5 - 3 5 8 (1969). F O N B E R G , E . : Hyperphagia produced by lateral amygdalar lesions in dogs. Acta Neurobiol. Exp. 31, 1 9 - 3 2 (1971). FONBERG, E . : Amygdala functions within the alimentary system. Acta Neurobiol. Exp. 34, 4 3 5 - 4 6 6 (1974). Fox, C. A.: Certain basal telencephalic centers in the cat. J . Comp. Neurol. 79, 2 7 7 - 2 9 5 (1940). FONBERG,

F o x , C. A . ,

A . N . ANDRADE,

D. E.

HILLMAN

and

R . C.

SCHWYN: The spiny neurons in the primate striatum: A. Golgi and electron microscopic study. J . Hirnforsch. 3. 1 8 1 - 2 0 1 (1971).

The distribution of axon terminals in t h e amygdaloid body HALL, E . : Some aspects of the structural organization of the amygdala. I n : T h e Neurobiology of the Amygdala. B. E. Eleftheriou, ed. Plenum Press, New York, 95 — 121 (1972). J U R A N I E C , J . , O . N A R K I E W I C Z and T . W R Z O L K O W A : Axon terminals in t h e claustrum of the c a t : an electron microscope study. Brain Research 35, 277 — 282 (1971). J U R A N I E C , J . , T . W R Z O L K O W A and O . N A R K I E W I C Z : Types of synapses in t h e claustrum of t h e cat. Acta Neurobiol. E x p . 34, 2 3 3 - 2 5 2 (1974 1 ). J U R A N I E C , J . , T . W R Z O L K O W A , and O. N A R K I E W I C Z : Synaptic organization of some nuclei in the amygdaloid complex. Annals Med. Sect. Pol. Acad. Sci. 19, 1 1 5 - 1 1 6 (1974"). J U R A N I E C , J . , T . W R Z O L K O W A and O . N A R K I E W I C Z : Ultrastructure of the " e x c i t a t o r y " area of the amygdaloid body. Annals Med. Sect. Pol. Sci. 20, 85 — 86 (1975). K A M A L , A. M . , and T . T O M B O L : Golgi studies on the amygdaloid nuclei of t h e cat. J. Hirnforsch. 16, 1 7 5 - 2 0 1 (1975). K A W A N A , E., K . A K E R T and H . B R U P P A C H E R : Enlargement of synaptic vesicles as an early sign of terminal degeneration in the r a t caudate nucleus. J . Comp. Neurol. 142, 297 bis 307 (1971). K I M , J . S., I . J . B A K , R . H A S S L E R aind Y . O K A D A : Role of y-aminobutyric acid (GABA) in the extrapyramidal motor system. 2. Some evidence for the existence of a t y p e of GABA — rich strio-nigral neurons. Exp. Brain Res. 14, 9 5 - 1 0 4 (1971). L U N D , R. D., and L. E. W E S T R U M : Synaptic vesicle differences after primary formalin fixation. J. Physiol. (Lond.), 185, 7 P - 9 P (1966). N A K A J I M A , Y . : Fine structure of the medial nucleus of the trapezoid body of t h e b a t with special reference to two types synaptic endings. J . Cell Biol. 50, 121 — 134 (1971). NARKIEWICZ, O.,

J. JURANIEC

and

T. WRZOLKOWA:

The

ultrastructure of synapses and functional differentiation of the amygdaloid body, (in Polish) Ann. Acad. Med. Gedan. 5, 4 3 - 6 0 (1975). N I T E C K A , L., H . Z A W I S T O W S K A and J . B I A L O W A S : Nuclei of t h e amygdaloid body in cats — structure and a c e t y l cholinesterase activity. Folia Morph. (Warszawa), 32, 39 bis 50 (1973). P F E N N I N G E R , K . H . : Synaptic morphology and cytochemistry. Progr. Histochem. Cytochem. 5, 1 — 86 (1973). R E Y N O L D S , E . S . : The use of lead citrate a t high p H as an electronopaque stain in electron microscopy. J. Cell Biol. 17, 2 0 8 - 2 1 3 (1963). R I C H A R D S O N , J. S.: The amygdala: historical and functional analysis. Acta Neurobiol. Exp. 33, 623 — 648 (1973). R I C H A R D S O N , J. S.: Basic concepts of psychopharmacological research as applied to the psychopharmacological

Note added in proof: Since this manuscript was finished, several papers on putative transmitters in amygdala and activity of their enzymes have ben published. B E N - A R I et al. (1976) found t h a t t h e central and medial nuclei have significantly higher activities of the glutam a t e decarboxyase t h a n other amygdaloid areas. Gamma-aminobutyric acid was also more concentrated

143

analysis of t h e amygdala. Acta Neurobiol. Exp. 34, 543 bis 562 (1974). R I N V I K , E., and I. G R O F O V A : Observations on fine structure of the substantia nigra in the cat. Exp. Brain Res. 11, 2 2 9 - 2 4 8 (1970). S T O R M - M A T H I S E N , J . : GABA as a transmitter in the central nervous system of vertebrates. J. Neural Transmission, Suppl. XI, 2 2 7 - 2 5 3 (1974). T O M B O L , T., and A. S Z A F R A N S K A - K O S M A L : A Golgi study of t h e amygdaloid complex in the cat. Acta Neurobiol. Exp. 32, 8 3 5 - 8 4 8 (1972). U C H I Z O N O , K . : Characteristics of excitatory and inhibitory synapses in the central nervous system of t h e cat. N a t u r e (Lond.), 207, 6 4 2 - 6 4 3 (1965). U C H I Z O N O , K . : Excitation and inhibition. Synaptic morphology. Tokyo — Elsevier Scientific Publishing Company. Amsterdam. Oxford. New York 1975. V A L D I V I A , O.: Methods of fixation and the morphology of synaptic vesicles. J . Comp. Neurol. 142, 257 — 274 (1971). W A K E F I E L D , C., and E. H A L L : Some observations on t h e ultrastructure of the central amygdaloid nucleus in t h e cat. Cell Tiss. Res. 151, 4 8 9 - 4 9 8 (1974 1 ). W A K E F I E L D , C., and E. H A L L : Hypothalamic projections to the amygdala in t h e cat. A light and electron microscopic study. Cell Tiss. Res. 151, 4 9 9 - 5 0 8 (1974 b ). W A L B E R G , F . : Elongated vesicles in terminal boutons of the central nervous system, a result of aldehyde fixation. Acta Anat. 65, 2 2 4 - 2 3 5 (1965). W A T S O N , M. L.: Staining of tissue sections for electron microscopy with heavy metals. J . Biophysic. Biochem. Cytol. 4, 4 7 5 - 4 7 8 (1958). W E S T R U M , L. E. Observatious on initial segments of axons in t h e prepyriform cortex of t h e rat. J. Comp. Neurol. 139, 3 3 7 - 3 5 6 (1970). W E S T R U M , L. E., and R. G. B L A C K : Fine structural aspects of t h e synaptic organization of the spinal trigeminal nucleus (pars interpolaris) of the cat. Brain Research 25, 2 6 5 - 2 8 7 (1971).

Authors

adress:

O. NARKIEWICZ J.

JURANIEC

T.

WRZOLKOWA

Department of Anatomy and Laboratory of Electron Microscopy, Institute of Medical Biology, School of Medicine ul. Dgbinki 1. 80 — 211 Gdansk Poland

in these nuclei w h a t supports hypothesis t h a t it m a y be the t r a n s m i t t e r of F - t y p e boutons in the amygdaloid body. B E N - A R I , Y., J . K A N A Z A W A and R. E. Z I G M O N D : Regional distribution of glutamate decarboxylase and G A B A within the amygdaloid complex and stria terminalis system of t h e rat. J. Neurochem. 26, 1279-1283 (1976).

J Hirnforsch 19 (1978) 145 - 1 58

1st Department of Anatomy, Semmehveis University Medical School, Budapest

Quantitative histological studies on the lateral geniculate nucleus in the cat. I. Measurements on Golgi material By Terez TOMBOI., Magda

MADARASZ,

F

HAJDU,

Gy.

SOMOGYI,

J.

GERLE

With 12 figures (Received 7 t h July 1977)

Summary: Numerical data of neuron and fiber elements were observed and measured on Golgi preparations The aim of the quantitative Golgi analysis was to obtain data on the spatial arrangement of arely neurons, their relations to the interneurons, on the spatial architecture of the arborizations of optic fibers and their relations to the mam neuron types The quantitative data on arely neurons include numbers of dendrites, of their chief points of ramification as the main targets of the optic afferents, geometric data on the "specific active sphere" of the dendritic tree, on the arborization of initial collaterals. The data on interneurons refer both to numeric and geometric parameters of their axonal arborizations, and to those of various characteristics of their dendritic processes The data on optic fibers include the size of their arborization space, the numbers of bouton clusters and preterminal bouquets per arborization.

Introduction

1 9 7 3 , MOREST 1 9 7 1 , MOREST 1 9 7 5 , PASIK et al 1 9 7 3 ,

Numerous studies have been dealing with the Golgi architecture of the lateral geniculate nucleus (LGN) since the classical description by RAMÓN y C A J A L (1911). Two main types of neurons have been generally distinguished ( F A M I G L I E T T I and P E T E R S 1 9 7 2 ) , P A S I K , P A S I K , HÁMORI, SZENTÁGOTHAI 1 9 7 3 , T Ó M B Ó L

1968,

one being the thalamocortical relay (TCR) neurons and local interneurons (IN) the other. However, further subdivisions had to be made in both types ( G U I L L E R Y 1 9 6 6 , 1 9 6 9 , H A Y H O W 1 9 5 8 , SZENTÁGOTHAI 1 9 7 3 , T O M B O L 1 9 6 8 ) . The two types of the TCR neurons were: The large tufted TCR neurons (Class I.), found predominantly close to the laminar borders, and the medium sized TCR (principal) neurons (Class II.). The interneurons were also divided into two subgroups (TOM BOL 1969): (a) interneurons with intralaminar connexions only and; (b) those with interlaminar connexions by means of their longer axons. Golgi observations were gaining progressively in importance in view of corresponding findings in the EM level. The dendrites of Class I and Class I I neurons are obviously the main postsynaptic elements in the synaptic glomeruli ( F A M I G L I E T T I , P E T E R S 1 9 7 2 , WONG-RILEY

1972),

G U I L L E R Y 1 9 6 9 , SZENTÁGOTHAI 1 9 7 3 , SZENTÁGOTHAI

et al. 1 9 6 6 ) . The peculiar dendritic appendages of the interneurons correspond to the presynaptic dendritic profiles that contain synaptic vesicles (FAMIGLIETTI, P E T E R S 1 9 7 2 , G U I L L E R Y 1 9 6 9 , LIEBERMANN

RALSTON,

HERMANN

STERLING

1970,

1969,

R I N V I K , GROFOVA

SZENTAGOTHAI

1973,

1974,

WONG-RILEY

1972).

There is a considerable body of new evidence on the presynaptic nature of interneuron dendrites (FAMIG L I E T T I , P E T E R S 1 9 7 2 , HAMORI e t a l . 1 9 7 4 ,

LIEBER-

MANN 1 9 7 3 , L U N D 1 9 6 9 , M O R E S T 1 9 7 1 , M O R E S T 1 9 7 5 , P A S I K e t a l . 1 9 7 3 , RALSTON 1 9 7 1 , R I N V I K , GROFOVA 1 9 7 4 , W O N G 1 9 7 0 , W O N G - R I L E Y 1 9 7 2 ) and on earlier unknown but rather strictly determined combinations of synaptic contacts between the specific afferents, the relay neurons and interneurons (SZENTAGOTHAI

1963,

SZENTAGOTHAI

et

al.

1966,

TOMBOL

at thalamic level. These speculations about the possible functional significance (for a comprehensive review see S H E P H E R D 1 9 7 2 ) calls for a deeper insight into the quantitative relations, at several levels of organization, between the various neuronal elements of LGN. In the present paper the results of representative measurements, made on Golgi specimens, are presented that are a prerequisite for a functional interpretation of the quantitative cytological analysis to be presented in the following papers of this series. 1 9 6 8 , TOMBOL 1 9 6 9 )

Materials and Methods The study was carried out on serial sections of about three hundred cat brains stained by Golgi-Kopsch perfusion method Adult (approximately 3 month old) cats were used.

Fig. 1. Thalamocortical relay neurons; Golgi-Kopsch preparates. a) large relay neuron with axon (ax). On the dendrites and soma dendritic appendages of Golgi I I type interneuron are seen (arrows). b) medium sized (principal) relay neuron whith axon (ax); the axon (ax) emits initial collateral (ic). At the ramification of the dendrites protrusions are seen (arrows), c) the both types of relay neurons are seen together (600 X ) .

Q u a n t i t a t i v e histological studies on L G X

Fig. 2. Drawing (a) represents the principal relay neuron, (b) R a p i d Golgi picture shows a relay neuron. T h e axon emits initial collateral (ic), which ramifies (arrows).

Most of the serial sections ware oriented in t h e coronal and some in the sagittal plane. T h e sections were of 120 —130(i.m thickness. T h e unpredictable selectivity of t h e impregnation makes Golgi material unsuited for anything b u t representative measurements. Only neurons were used for measurements t h a t appeared to be stained with satisfactory completeness. T h e measurements include the following characteristics: cell body size of various kinds of neurons, two dimensions in the plane of sectioning of dendritic and axonal arborizations, t h e number of branching points of the dendritic trees and the distance of these points from the cell body, t h e number of specific dendritic appendages, particularly those of the l X - s . T h e sizes of t h e arborizations of t h e optic fibers were measured and t h e number of optic terminals per arborization were counted. T h e third dimension of dendritic and axonal fields could be reconstructed partly from t h e thickness of the section and b y comparing t h e d a t a observed in coronal and sagittally oriented sections.

Results (1)

TCR neurons

The T C R neurons have been classified by G U I L L E R Y (19(H)) into Class I and Class I I (• = principal) neu-

147

rons. Both types send their axons into the primary visual cortex ( G A R E Y and P O W E L L 1967, G A R E Y and P O W E L L 1.971, G L I C K S T E I X et al. 19(57), while the Class I neurons are thought to have additional terminals in area 18 ( G A R E Y and P O W E L L 1.9(57, G A R E Y and P O W E L L 1971, P O W E L L 1973). The Class I neurons (GUILLERY 1966) are large multiangular cells (TOMBOL 19(58) (Fig. :1a). The diameter of the cell somata is of 35 um (4-2.5 fxm) in average. The cell body is generally multiangular, occasionally rounded in consequence of the considerable number (8 — 10) of principal dendrites that take origin directly from the cell somata. The principal or main dendrites are thick (3—5 ¡xm) and are branching at various distances (15—30 ¡¿m) either dichotomically or in a fashion somewhat resembling the pattern called " t u f t e d " . Secondary and tertiary ramifications occur generally at acute angles (GUILL E R Y 1.966, 1969, SZEXTAGOTIIAI 1.9(53, TOMBOL 1.968). The length of the dendrites varies from 1.50 to 300 ¡xm. Their course is rather straight and radially oriented. The diameter of the entire dendritic tree is of 300 to (500 fxm and of fairly symmetric spheroid shape. The surface of the dendrites is rough, covered by irregular processes of different shape. The Class I neurons can be found in all the main layers of dorsal LGX, scatte-

148

Tombol, T., M. Madarasz, F. Hajdu, G. Somogyi, J . Gerle

Fig. 3. Golgi Il/a type interneuron (a, b, c, d). The axon (ax) ramification and further arborization (e) are well seen. On the dendrites the dendritic processes (dp) are well impregnated (600x). Higher magnification represents the dendrites of a Golgi Il/a type interneuron on which there are some dendritic appendages (dp)-complex spine-apparates (800x).

Quantitative histological studies on LGN

red irregularly with some preference to the rostral and dorso-lateral parts of the nucleus and the laminar ' borders. The dendrites of the large neurons do not respect the borders of the layers, which they cross freely whenever their position is sufficiently close. Although the primary divisions of the principal dendrites occur often in tufts, it is the repeated dichotomic branching of the dendrites, which is most characteristic for these neurons. The Class I I or according to our nomenclature principal neurons (Fig. lb) are medium size cells with an elongated or ovoid body (26 versus 18 ¡Am, ± 6 (im, of the two diameters). The few (4—6) principal dendrites originate mostly in groups at the two poles of the cell body, giving characteristic all over appearance and also an orientation to these cells. The principal dendrites are of 10—30 [xm length and ramify several times usually in tufted manner. At these points of ramification — the number of which varies between 14—25 per cell — 2—8 dendritic eccrescences emerge normally from the initial part of the secondary and tertiary dendrites. These are large (1 — 2 [i.m) spheric or ovoid bulges and are often arranged in grape-like clusters. Single spines or bulbs may occur also on the distal parts of the dendrites. The spaces occupied by the dendritic trees are ovoid and predominantly perpendicular, with their long axis to the surface of the cell layers. This orientation characterizes the principal neurons in the layers A and A-L with the exception of the neurons near the laminar surfaces. The distal portions of the dendrites may cross the laminar borders but the special branching-points with bulges are restricted to the cells own layer. The lengthes of the dendrites (140 to 180 ¡xm) as well as the diameters of dendritic trees (280—360 ¡Am) are smaller than those of the Class I neurons. The ramification points on the dendrites are at different distances from the cell body (Fig. 2a).. The closest branchings are at 10—30 (Am, the most distant ramification points are at 80—100 [Am distances. The grape-like clusters proved to be the main target of the optic terminals (TOMBOL 1968, 1969). Taking this into consideration one may surmise a slightly ovoid space around the cell body of a diameter of 80 and 100 [xm respectively as the "active" part of the dendritic tree (called "specific active dendritic space" of 2,143,576 with 80 ¡Am diameter and 4,186,666 ¡Am3 with 100 [Am diameter respectively). The axons of TCR neurons give rise from the cell body with well defined cones and enter the optic radiation leaving the nucleus dorsalward. The initial axon collaterals of the principal neurons have to be mentioned as functionally important elements, Hirnforschung, Bd. 19, Heft 2

149

however, their poor impregnation does not allow us to get exact numerical data about this component of the principal neurons. The only features known about them are their ramification near the parent cell and their number of contacts with IN-s observed in some rapid GOLGI specimens (Fig. 2 b). (2) The interneurons

(IN)

The interneurons of LGN are diyided into two subgroups: GOLGI I I a and GOLGI l i b types of I N - s (TOMBOL 1968, 1969). This classification is based primarly on the size of their axonal arborization fields, although there are also other less conspicuous differences between the two cell types. The axon of the GOLGI I I a I N (Figs 3 and 4) arborizes locally within one layer. The axonal ramification is usually arranged in a parallel disc with the dendritic tree but shifted either in transversal or in

axon (ax). 11

150

Tombol, T „ M. Madarasz, F . Hajdu, G. Somogyi, J . Gerle

Fig. 5. Golgi I l / b type interneuron is demonstrated in different depths. On the dendrites single spines and the axon (ax) — arborization in different position can be seen. GolgiKopsch preparate (600 x ) .

because the dendrites of cells compressed to a smaller volume are more tortuous. A characteristic feature of GOLGI II a IN dendrites is their surface properties by being endowed with a rich population of appendages (FAMIGLIETTI and

antero-posterior direction. There is only an insignificant overlap between the axonal and the dendritic arborization. The axon divides successively five to six times, usually in a dichotomic manner. The number of the claw-shape axon terminals is about 40—50 per cell, establishing contacts predominantly with relay neurons; usually with the characteristic bulges of the dendrites. The dendritic tree of the GOLGI II a IN is oriented in perpendicular direction to the laminae. Four or five dendrites originate from the rounded cell body, some of them ramify. All the dendrites are rather thick, their course is irregular and tortuous or wavy. Neither the axon nor the dendrites of the Golgi II a cells cross the laminar borders. The longer dendrites are bent back at the border of the layer. The dendritic tree is cylindric if the cell happens to be located in the middle of the layer and hence not subjected to restriction by the laminar border. — The vertical diameter of the dendritic tree ranges between wide limits: from 300—800 |xm. The transversal diameter is of 150—200 [xm. Differences in size of the dendritic trees do not entail corresponding differences in the total length of the dendrites (about 1 7 0 0 - 2 0 0 0 (xm),

processess or appendages can be observed regularly: some of them are bunches of 6—8 drumstick spines originating from a thickening of the dendrite, others are large (3 — 5 (im) irregular terminal swellings, still others originate as sidebranches with long delicate axon-like stalks bearing flower-like appendages. These various appendages and processes of the dendrites make the GOLGI II type IN-s easily recognizable even if only fragments are stained. The number of dendritic appendages is 40—60 per cell, their allocation does not show any obvious regularity along the dendrites. The GOLGI l i b IN-s (Figs. 5 and 6) are larger than the Golgi II a, their cell body is ovoid with diameters 15 x 18 and 12 x 15 [I.m respectively. The dendrites originate with a thick initial part and ramify dichotomically. They are covered with regular drumstick spines. The dendritic tree does not seem to show any preferential orientation. The dendrites are 200 to 250 ¡j.m long. The axonal arbozation of this neuron type — as already previously described (TOMBOL 1968, 1969) — may establish (axo-axonic) connections in the layer of origin with the proximal portions of relay cell axons (see Fig. 4 in the III. paper of this series)

PETERS 1 9 7 2 , TOMBOL 1 9 6 8 , 1 9 6 9 ) . V a r i o u s t y p e s of

Quantitative histological studies on LGN

151

380/»

Fig. 6. Drawing of Golgi I l / b type interneuron. Both the characteristics of dendrites and the axonarborization (ax) are demonstrated.

and with the principial dendrites of the TCR neurons. The main branch, however, crosses the laminar border, obviously for establishing contacts in the other layer (SANDERSON 1 9 7 1 , SANDERSON et al. 1971, SUSUKI and KATO 1966). In the sagittal plane — observed recently — relatively long (600—800 (xm) sidebranches run parallel with the direction of the lamina giving off several short side-branches at regular intervals. These terminal branches of beaded appearance are somewhat like the basket cell axons in the cerebellar cortex. The main cell types hitherto described can be found also in lamina B (corresponding to lamina C, CJ^ and C 2 according to GUILLERY, however, neither in appearance nor in orientation of their branches are the differences so well characterized than in lamina A and Ax.

(3) Fusiform

neurons

At the laminar borders, especially between layers A and A 1 and also between Ax and B there are medium size fusiform neurons oriented in parallel with the layers, preferentially in the sagittal direction. These neurons resemble those of the perigeniculate nucleus albeit of somewhat smaller size (20x9(xm). The pattern of their dendritic arborizations suggests that these cells might belong to the reticular system, perhaps to the reticular nucleus of the thalamus. Their axons have not been stained successfully in this material. (4) Axons of extrageniculate source and their terminal ramifications The study of fiber arborizations raises important questions concerning useful criteria for the identification of various types of afferents: viz. determination of the shape and size of their arborization spaces and of the number, size, and approximate density of synaptic terminal portions. Determination of such parameters depends heavily on the quality of the

152

Tómból, T., M. Madarász, F. Hajdu, G. Somogyi, J . Gerle

Fig. 7. Microphotographs and drawing from optic fibers arborization. B o t h the preterminal bouquets (terminal units) and bouton clusters (terminal portions) with knobs can be seen (arrows).

G O L G I stain. I t is widely known that the classical rapid procedure does not always stain the terminal portions but is a very convenient method to demonstrate the total ramification, including the preterminal portion of the fiber. Conversely, the perfusion

Golgi-Kopsch procedure is more advantageous for staining the synaptic terminals but one can seldom find a completely stained arborization. Some of the afferents of the dense neuropil could be identified unequivocally, however, there are still types of arborizations that have not been identified and classified according to their source. 4.1.

Retinal

afferents

The ramification of the optic fibers in LGN has been described first by R A M Ó N y C A J A L (191 J) and was

Quantitative histological studies on LGN

Fig. 8a—b. Details from terminations of retinal afferents (OF) and their relation with characteristic synaptic sites of principal relay neurons. (OT = optic terminals, ax = axon of principal relay neuron). Golgi-Kopsch preparates (800 x ) .

completed in some essential details by O ' L E A R Y (1940). Both descriptions dealt with the large calibred fibers. The arborizations were described as dense brushes of conic shape confined to a single lamina and somewhat compressed into the antero-posterior direction. The dense "terminal nests" according to RAMÓN y C A J A L encompass about 10 cell bodies. In the G O L G I - K O P S C H material of the mature L G N the terminal portions of the optic fibers can be seen rather well in many of their details. The "dense brushes" of C A J A L (1911) correspond probably to the entire arborization consisting of different size of units according to the cascade-like arborization. The optic fiber arborizes into 3 — 4 preterminal branches each of which develop 2—3 preterminal bouquet (previously called terminal units). The preterminal bouquet (terminal unit) consists of several (5 — 15) so called bouton clusters (terminal portions). They are 1 0 - 2 0 ¡xm long (TÓMBOL 1969) (Figs. 7 a n d 8).

In the bouton clusters the boutons are arranged like grains of a corn-ear. The bouton clusters contact the dendritic tufts of Class I I relay cells (Fig. 8). This was first recognized by SZENTÁGOTHAI (1963), confirmed

153

by G U I L L E R Y (1966) analyzed in detail and assembled in a synthetic picture by T O M B O L (1968, 1969) in G O L G I - K O P S C H specimens. The "dense brushes of conic shape" are rarely stained in the G O L G I - K O P S C H material. The largest size retinal axon arborizations observed and measured in this study are cylindric in shape and about 300 X 200 x 150 (xm. Most of the impregnated optic fibers are only fragments of the "total" arborization. They appear to have different sizes: consisting of from one preterminal bouquet (terminal unit) to several first-order preterminal branches. The largest retinal axonarborization observed consist of 3—4 first-order preterminal branches, of about 10 preterminal bouquets (terminal units) and of about 80 bouton clusters (terminal portions). The bouton clusters (terminal portions) may sometimes consist of a single knob only, more frequently it is a set of 8—15 large bulges of 2 —3 ¡xm diameter. The arborization disk of optic fiber is mostly oriented in the sagittal plane with its longer diameter. Its smallest extension is in the perpendicular direction. But one can also find cases in which the arborization space of a single fiber spans the entire layer in perpendicular direction as was observed already by O ' L E A R Y (1940). Optic terminals were found also in layer B. The arborization pattern was similar, its size was smaller than thos in layers A and Aj.

i54

T o m b o i , T., M. Madarasz, F. H a j d u , G. Somogyi, J . Gerle

Fig. 9a—b. Arrows p o i n t on cortico-geniculate fibers. GolgiKopsch p r e p a r a t e s ( 6 0 0 x ) . 4.2.

Descending corticogeniculate

fibers

The corticogeniculate fibers are known to originate from the striate cortex (BERESFORD 1962, FLECHSIG 1 8 9 6 , GUILLERY 1 9 6 7 , MONTERO a n d GUILLERY 1 9 6 8 , NAUTA e t a l . 1 9 5 4 , PROBST 1 9 0 2 , SZENTAGOTHAI 1 9 6 3 ,

1975). Their major source is the 6th layer of the primary visual cortex (GILBERT and K E L L Y 1974, TOMBOL et al. 1975). These fibers enter the L G N through its dorsal surface. The entering fibers ramify at a narrow angle, their branches traverse the nucleus in perpendicular direction. Along their course both in the main and in the side branches small drumstick shape spine-like terminal processes emerge. They contact all cell types (Fig. 9). Numerical data about these fibers are not yet available.

Together with the cortigo-geniculate fibers another medium sized axon enters the LGN (Fig. 10). Their course and arborization pattern is rather different from that of the corticogeniculate fibers. Their arborizations seem to be restricted to a single layer. They have long varicose terminal branches, which can often be seen to run parallel with the terminal branches of an optic fiber. They might have their origin in the perigeniculate nucleus. — Neither the size of their arborization, nor the number of their terminal branches are known.

Discussion The neuronal and fiber architecture of LGN is demonstrated in drawings. Figs 11 and 12 show the neuronal and fiber arrangement on sagittal and

Quantitative histological studies on LGN

155

Fig. 10a—b. Medium sized fibers with varicous side-branches (arrows). Golgi-Kopsch preparates (600 x ) .

and K A T O 1 9 6 6 ) , the binocular interaction correlated with the Golgi l i b interneurons (SANDERSON 1971, SANDERSON et al. 1 9 7 1 ) . Recent studies have described the presynaptic dendrites which belong to the

coronal sections, respectively. Both pictures are composed from few closely neighbouring sections. It is important to emphasize that on both pictures the different elements are represented according to their relative proportional sizes and original orientations. The perpendicular orientation of the neurons and fibers is the most prominent organization principle of the LGN. The numerical ratio of different cell types in the diagrams does not, of course, reflect the real situation, owing to the inherent random selectivity of the Golgi impregnation technique. In spite of the large quantity of the material studied, the parameters shown in this paper are considered open to further studies. New materials may disclose other relations more fortunate impregnations may complete the numerical data of this Golgi material. The intrinsic connections of LGN have been discussed in earlier studies: the complex inhibitory interaction explained by the presence of local Golgi I I type neurons ( B U R K E and S H E F T O N 1 9 6 6 , SANDERSON

I N - s ( F A M I G L I E T T I a n d P E T E R S 1 9 7 2 , HÂMORI e t a l .

e t al. 1 9 7 1 , SINGER a n d CREUTHFELDT 1 9 7 0 ,

SUSUKI

1 9 7 4 , LIEBERMANN 1 9 7 3 , MOREST 1 9 7 1 , et

al.

1973,

RALSTON

1971,

RINVIK

1975,

and

PASIK

GROFOVA

1 9 7 4 , W O N G 1 9 7 0 , W O N G - R I L E Y 1 9 7 2 ) emphasizing their complex action in the optic transmission. The numerical data of Golgi impregnated neuronal and fiber elements are significant in themselves, however, an appraisal of their functional significance have to be reserved until the data of general quantitative cytological analysis of LGN, to be given in the subsequent papers of this series, can be considered together with the GOLGI data.

References BERESFORD, W. A.: A Nauta and gallocyanin study of the

corticogeniculate projection in the cat and monkey. J . Hirnforsch. 5 2 1 0 - 2 2 8 (1962). .

BURKE, W . , and A. J . SHEFTON: I n h i b i t o r y m e c h a n i s m s in

lateral geniculate nucleus of rat. J . Physiol. (Lond.) 187 2 3 1 - 2 4 6 (1966).

CAJAL, S., y RAMON: Histologie du S y s t è m e N e r v e u x de

l'Homme et des Vertébrés, Vol. 2. Maloine, Paris/1911.

FAMIGLIETTI, E . V., and A. PETERS: The synaptic glomeru-

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Tömböl, T., M. Madarâsz, F. H a j d u , G. Somogyi, J. Gerle

A

B

^

Fig. 11. Coronal section of LGN. The neuronal elements are drawn in original places and direction.

T

,

t

1 = large neuron, p = principal relay neuron, a = Golgi I I / a interneuron, b = Golgi I I / b interneuron, c = optic fiber arborization, c = corticogeniculate fiber, r = medium sized varicous fiber.

lus and the instrinsic neuron in the dorsal lateral geniculate nucleus of t h e cat. J. comp. Neurol. 144, 285 — 334 (1972). F L E C H S I G , P . v o n : Weitere Mitteilungen über den Stabkranz des menschlichen Gehirns. Neurol. Centralbl. 15, 2 - 4 (1896). G A R E Y , L . }., and T . P. S. P O W E L L : The projection of the lateral geniculate nucleus upon cortex in the cat. Proc. Roy Soc. B. 169, 1 0 7 - 1 2 6 (1967). G A R E Y , L . J . , and T . P . S. P O W E L L : An experimental study of the termination of the lateral geniculo-cortical p a t h w a y in the cat and monkey. Proc. Roy. Soc. B. 179, 41 — 63 (1971). G I L B E R T , C., and J . P . K E L L Y : The use of peroxidase transport to study the connections of the cat's visual system. Soc. Neurosci. Abstr., 227 (1974). GLICKSTEIN, M., R . A . KING,

J. MILLER a n d M.

BERKELEY:

Cortical projections from the dorsal lateral geniculate nucleus of cats. J. comp. Neurol. 130, 55 — 76 (1967). G U I L L E R Y , R. W . : A study of Golgi preparations from the dorsal lateral geniculate nucleus of the adult cat. J. comp. Neurol. 128, 2 1 - 5 0 (1966). G U I L L E R Y , R. W . : Patterns of fiber degeneration in the dorsal lateral geniculate nucleus of the cat following lesions in t h e visual cortex. J . comp. Neurol. 130, 197 — 222 (1967).. GUILLERY, R . W.: The organization of synaptic interconnections in the laminae of the dorsal lateral geniculate nucleus of the cat. Z. Zellforsch. 96, 1 — 38 (1969).

HAMORI, J.,

T . PASIK,

P . PASIK

and

J . SZENTÂGOTHAI :

Triadic synaptic arrangements and their possible significance in the lateral geniculate nucleus of t h e monkey. Brain Research, 80, 3 7 9 - 3 9 3 (1974). H A Y H O W , W . R . : The cytoarchitecture of t h e lateral geniculate body in the cat in relation to t h e distribution of crossed and uncrossed optic fibers. J. comp. Neurol. 110, 1 - 6 3 (1958). L I E B E R M A N , A. R . : Neurons with presynaptic perikarya and presynaptic dendrites in the r a t lateral geniculate nucleus. Brain Research 59, 35 — 59 (1973). L U N D , R . D . : Synaptic patterns of t h e superficial layers of the superior colliculus of the rat. J. comp. Neurol. 135, 1 7 9 - 2 0 8 (1969). M O N T E R O , V . M . , R . W . G U I L L E R Y : Degeneration in the dorsal lateral geniculate nucleus of the r a t following interruption of the retinal or cortical connections. J. comp. Neurol. 134, 2 1 1 - 2 4 2 (1968). M O R E S T , D. K. : Dendrodendritic synapses of cells t h a t have axons: The fine structure of the Golgi type I I cell in t h e medial geniculate body of the cat. Z. Anat. Entwickl.Gesch. 133, 2 1 6 - 2 4 6 (1971). M O R E S T , D. K. : Synaptic relationships of Golgi t y p e II cells in the medial geniculate body of the cat. J. comp. Neurol. 162, 1 5 7 - 1 9 4 (1975). N A U T A , W . J . H . , and V . M . B U C H E R : Efferent connections . of the striate cortex in the albino rat. J. comp. Neurol. 100, 2 5 7 - 2 8 1 (1954).

Quantitative histological studies on LGN

Fig. 12. Sagittal section of LGN. The neuronal elements are drawn in original places and direction. 1 = large relay neuron, p = principal relay neuron, a — Golgi I l / a interneuron, b = Golgi I l / b interneuron, c = optic fiber arborization, c = corticogeniculate fiber, r = medium sized varicous fiber.

157

E., and J . G R O F O V A : Light and electron microscopical studies of the normal nuclei ventralis lateralis and ventralis anterior thalami in the cat. Anat. Embryol. 146, 5 7 - 9 3 (1974). S A N D E R S O N , K. J . : Visual field projection columns and magnification factors in the lateral geniculate nucleus of the cat. Exp. Brain Res. 13, 1 5 9 - 1 7 7 (1971). RINVIK,

SANDERSON, K . J . , P . O. BISHOP a n d I . DARIAN-SMITH : T h e

A structural analysis of the lateral geniculate nucleus of the cat. J . comp. Neurol. 73, 405 — 430 (1940).

O'LEAEY, J. L.:

PASIK, P.,

T . PASIK,

J . HÄMORI

and

J.

SZENTAGOTHAI:

Golgi type I I interneurons in the neuronal circuit of the monkey lateral geniculate nucleus. Exp. Brain Res. 17, 1 8 - 3 4 (1973). P O W E L L , T. P . S.: The organization of the major functional areas of the cerebral cortex. Symp. Zool. Soc. h. No. 33, 2 3 5 - 2 5 2 (1973). P R O B S T , M.: Über den Verlauf des zentralen Sehfasern (Rindensehhügelfaser) und deren Endigungen im Zwischen- und Mittelhirn und Sehsphäre. Arch. Psychiat. 35, 2 2 - 4 3 (1902). R A L S T O N , H. J . I I I . : Evidence for presynaptic dendrites and a proposal for their mechanism of action. Nature (Lond.), 230 5 8 5 - 5 8 7 (1971). R A L S T O N , H . J . , and M . M . H E R M A N : The fine structure of neurons and synapses in the ventrobasal thalamus of the cat. Brain Research 14, 77 — 97 (1969).

properties of the binocular fields of lateral geniculate nucleus. Exp. Brain Res. 13, 1 7 8 - 2 0 7 (1971). S H E P H E R D , G . M.: The neuron Doctrine: A revision of functional concepts. Yale Journal of Biology and Medicine 45, 5 8 4 - 5 9 9 (1972). S I N G E R , C . , and O . D . C R E U T Z F E L D T : Reciprocal lateral inhibition of on and off centre neurones in the lateral geniculate body of the cat. Exp. Brain Res. 10, 311 — 330 (1970). S T E R L I N G , P . : A light and electron microskopic study of the superficial gray of the cat superior colliculus. Anat. Rec. 166, 383 (1970). S T O N E , J., and B . D R E H E R : Projection of X and Y-cells of the cat's lateral geniculate nucleus to areas 17 and 18 of visual cortex. J . Neurophysiol. 36, 551 — 567 (1973). S U S U K I , H . , and E . K A T O : Binucular interaction at cat's lateral geniculate body. J . Neurophysiol. 29, 909 — 920 (1966). S Z E N T A G O T H A I , J . : The structure of the synapse in the lateral geniculate body. Acta Anat. 55, 166 — 185 (1963). S Z E N T A G O T H A I , J . : Neuronal and synaptic architecture of the lateral geniculate body. In R. Jung (Ed.) Handbook of Sensory Physiology, Vol. VII/3. B. Springer Verlag, Berlin, 1973. pp. 1 4 1 - 1 7 6 .

158

Tömböl, T., M. Madaräsz, F. Hajdu, G. Somogyi, J . Gerle

The "module-concept" in cerebral cortex architecture. Brain Research 95, 475 — 496 (1975). S Z E N T Ä G O T H A I , J., J . H Ä M O R I and T . T Ö M B Ö L : Degeneration and electron microscope analysis of the synaptic glomeruli in the lateral geniculate body. Exp. Brain Res. 2, 2 8 3 - 3 0 1 (1966). TELLO, J . F . : Disposition macroscopica y estructura de cuerpo geniculado externo. Trab. Lab. Invest. Biol. Univ. Madrid 3, 3 9 - 6 2 (1904). T Ö M B Ö L , T . : A thalamus specificus magjainak synapticus architekturäja (Synaptic architecture of specific thalamic nuclei). Thesis, Budapest (1968). In Hungarian. T Ö M B Ö L , T . : Two types of short axon (Golgi 2nd) interneuSZENTÄGOTHAI, J . :

rons in the specific thalamic nuclei. Acta morph. Acad. Sci. hung. 17, 2 8 5 - 2 9 7 (1969). T O M B O L , T . , F . H A J D U and Gy. S O M O G Y I : Identification of the Golgi picture of the layer VI. cortico-geniculate projection neurons. Exp. Brain Res. 24, 107 — 110 (1975). W O N G , M . T . T . : Somato-dendritic synapses in the squirrel monkey lateral geniculate nucleus. Brain Research 20, 1 3 5 - 1 3 9 (1970). W O N G - R I L E Y , M . T . T . : Neuronal and synaptic organization of the normal dorsal lateral geniculate nucleus of the squirrel monkey, Saimiri sciureus. J. comp. Neurol. 144, 2 5 - 6 0 (1972).

J . Hirnforsch. 19 (1978) 1 5 9 - 1 6 4

1st Department of Anatomy, Semmelweis University Medical School, Budapest

Quantitative histological studies on the lateral geniculate nucleus in the cat II. Cell numbers and densities in the several layers By M a g d a MADARASZ, J . GERLE, F . HAJDU, G y . SOMOGYI and T e r e z TOMBOL

With 3 figures and 3 tables (Received 7 t h July 1977) Summary: In the cat lateral geniculate nucleus all cells of all types were counted: their absolute numbers, their densities and their relative distribution. The nuclear volume and the volume of each layer was measured. The data of volume was applied on living material. The average densities of cells were calculated from the volume of LGN. The density of the cells was combined with the "specific active dendritic sphere" of relay neurons and it could be estimated that there may be a large number of spatially overlaping of ramification points around the cell body of relay neurons.

Introduction Previous quantitative cytoarchitectonic studies in the lateral geniculate nucleus (LGN) were directed mainly at establishing the total number of cells and of the volume of the nucleus. In the cat LGN the cell number obtained by B I S H O P ( 1 9 5 3 ) was about 4 5 0 , 0 0 0 cells, other studies of similar nature were made in the rat, the monkey and in man (CHOW et al. 1 9 5 0 , L A S H L E Y 1 9 3 9 , L E R A N T H et al. 1 9 7 5 ) . Generally, quantitative studies in the CNS are aimed at gaining information on absolute numerical data and on the volume proportion in the tissue of cellular and neuropil constituents. In most cases the absolute and relative ( = densities) numerical data and also the ratios of nerve cells and afferent and/or efferent fibers are important. The LGN is relatively favourable material for such studies, mainly due to its clear boundaries and well defined cellular layers. Although quantitative cytoarchitectonic data are only very elementary steps on the long and circuitous route towards the elucidation of the fundamental questions of the neuron coupling — notably convergence and divergence — they are absolutely essential for any forther penetration into the problem. This paper presents as a first approximation the absolute numbers of nerve cells, regardless of their types and some data on their distribution ( = differential densities) in various parts of the nucleus. Finally the average densities of nerve cells were calculated from the volume of LGN. Various corrections have, of course, to be made in order to arrive at realistic results; these corrections have been elaborated at much detail in the literature on quantitative histo-

logy ( F L O D E R U S 1944, P A L K O V I T S et al. 1971a, b, c, 1972). It is also advisable to calculate all values for the non fixed state, i.e. the "living animal", particularly in order to make structural data directly applicable in the physiological experiments. This applies, of course, only to the metric data, since neither the absolute, nor the relative numbers of neurons and fibers change with the shrinkage of the tissue during the histotechnique procedures. Materials and Methods General methods Adult (non-albino and non-Siamese) cats' brains were fixed by perfusion, under anesthesia, with 10% and subsequently immersed in 4% formol. The brains were then embedded into paraffin and coronal serial sections of 10 (xm thickness were prepared. Every tenth section of an uniterrupted series was collected for further processing. The thickness of the sections was controlled continuously. For staining procedure Haematoxylineosin, N I S S L and K L U V E R - B A R R E R A (1953) stains were used. Calculation of shrinkaeg For the determination of shrinkage linear measurements were used. The distance between two arbitrarily chosen but clearly defined points of fresh brains was marked before fixation and also measured in the final preparation. The results were similar to the data of earlier studies ( P A L K O V I T S et al. 1971, 1972); the linear shrinkage was found to be about 20%.

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Madarasz, M., J . Gerle, F. Hajdu, G. Somogyi and T. Tömböl

The number of the neurons in the LGN The absolute number of neurons was determined by counting all cell nuclei with visible nucleolus in the whole series. To aid systematic counting and in order to avoid repeated counting of the same nuclei, a rectangular grid was placed into the eye-piece of the microscope. Since only every tenth section of the series was processed, the final raw value is the sum of all nuclei with nucleolus x 10 X the correction factor according to F L O D E R U S (1944). This correction factor was 0.8322 for three cats and 0.8249 for one cat. (The Floderus correction is necessary to compensate for the effect of "double-counting" due to sectioned nucleoli that would appear in two neighbouring sections. The smallest nucleolar cap recognisable in the sections had a diameter of 0.88 ¡xm, the average radius of the nucleoli was 1.1 ¡xm. From these data of counted nucleoli in a section corresponded to a section thickness of 11.70 ¡xm). The densities of different

neurons

An overall density was calculated from the volume of LGN and from the total number of all kinds of neurons. The relative density was established by counting the number of cell nuclei with visible nucleoli in the fields of a rectangular grid of 250 x 250 ¡xm field size (-•- (52,500 ¡xm2) placed into the eye-piece of the microscope and the results were marked on a map of each section. The densities were determined separately for the several layers and corrected for living state. Fig. 1. Coronal sections of TGN in different levels;

Volume of LGN The volume of LGN was determined on the same section series. On each processed section of the total LGN the confines of the main layers were traced under low magnification of the screen of the Visopan (Reichert) microscope and the surfaces were measured by using a planimeter. The real surface area is then obtained by dividing the data of the measurements with the square of magnification. Calculation of the volume is based on the formule

a) in the rostral, b) in the middle and c) in the caudal part of the nucleus. The five groups of density of cells are signed with five slightly different shade. The dense (dark) region is in layer A and Aj the less density of cells was found in layer B.

Results

The quantitative examination was carried out on four cats. In two cats the neurons were counted without regard for the laminar borders. In these a, an cases only the total number of neurons was deter1 0 0 — -f a, . . . a n - - 1 f — 2 2 2 mined (Table I). Taking into consideration the distribution of optic fibers in the several laminae and its where (a) is the surface area, 1 n the number of functional significance, it was deemed necessary to counted sections, 100 is the distance between two obtain separate data for the main layers. The direct measured surfaces in um (•— the thickness of ten sec- counting of cells from section to section in the different tions). The volumes were corrected for the fresh layers shows the varying densities of neurons (Figs, l a, ( living) state by multiplication with the cube of l b , c). the correction factor. The numbers obtained from the several layers of two cats (No 1 and No 2) in layer A: 282,507, in

Q u a n t i t a t i v e histological studies on LGN

161

)4-% B

A

10—

25 °/e

Ai

U %

10 % Ai

'///. ,1%

I. 1-6,3 cells

16.7-24,9

Fig. 2. Histogram of the different groups of density. The percentage of the five density groups is illustrated in t h e three m a i n layers of LGN.

layer A ^ 204,346 and in layer B: 72,285 in cat-No 1 and in te second cat (No 2) A: 278,204, A x : 212,840 and in layer B : 69,490. The total cell number- of the LGN in these two animals was 559,138 and 560,534 respectively (Table I). The total number of neurons in the two cases in which the count was not taken separately according to the layers — were 563,080 and 545,320 respectively. The standard deviation of all numbers in the four cats is ±7388.2,- the relative deviation is 1.3% altogether. The difference between the several counts can be considered insignificant and can be due both to individual variations and to flaws of the counting procedure. The interlaminar nuclei were not counted separately but were lumped together with the main laminae. The numerical differences between layers A and A1 might be partly due to this fact but more probably related to the higher number of crossed fibers. In the arrangement of the neurons no obvious regularity was observed. The distances from cell body to cell body vary between 15 [xm and 65 ¡xm. Considering that the linear shrinkage is 20% the distance in

V. GROUP OF DENSITY 33,3 — i n 625,000 ¿ima

living animal between different cells can be assumed to be realistically 18 [xm to 78 [xm. Most of the measured distances are ranging between the two above values, while there are cell bodies next to each other in the dense areas, and at larger distances in the less dense regions. The measurement was made with an ocular micrometer. The density was found to vary extensively not only in the several layers but also in different parts of the same layer. The fields of the 250 X 250 [im rectangular grid used for measuring the densities correspond to unit fractions of tissue space of 625,000 fxm3 (1,080,000 [xm3 in the living material) for which the cell number, i.e. local densities could be determined separately. Five densitygroups (I—V) were arbitrarily chosen according to the number of cells found in one unit: groups I—IV contain 1 - 1 0 , 1 1 - 2 0 , 2 1 - 3 0 , 3 1 - 4 0 neurons, while the fifth density class corresponds to more than 40 cells per unit. Applying the Floderus correction, the number of cells in one unit, in the five density groups.is more realistically 1—8.3, 8.4—16.6, 16.7 to 24.9, 2 5 - 3 3 , 2 and above 33.3 cells. On the basis of this classification the local density of cells could be demonstrated pictorially (Fig. 1) for representative sections and the percentual distribution of density groups could be displayed for the several laminae. It appears from Fig. 2 that in layer " A "

162

Madârâsz, M., J . Gerle, F. Hajdu, G. Somogyi and T. Tömböl

about 3 5 % of the space units contain more than 20 cells. The distribution of the neurons does not seem to follow any obvious pattern, although density appears to increase in the caudal part of the LGN. The overall cell density of layer " A j " is somewhat smaller than of layer " A " , an increase in density in the caudate part is less pronounced than in layer " A " . In the layer B the overall density is the smallest, virtually the whole 8 5 % of this layer is belonging to the first and second group of density (Fig. 2). It also seemed necessary to calculate a "theoretical independent space" available to one cell in the different groups of density (of course, if there were no interpénétrations of each other's spaces by the dendrites) . In the first group the autonomous space of one cell would be 148,809 ¡¿m3, in the second group 50,000 ¡i.m3, in the third group 30,048 [¿m3, in the fourth group 21,477 fxm3, in the fifth group 15,625(xm3. (The specific active dendritic space of one neuron is

about 2,143,573 |xm3, as minimum and 4,186,666|xm3 as maximum respectively (Fig. 3). In the previous paper the space around the cell body, containing the part of dendritic arborization, that is receiving the majority of optic afferents, was labeled as "specific active dendritic space" which does not change with the density of cells.) Considering the "specific active dendritic space" of one relay neuron and combining this with the five groups of density, it is possible to calculate how many cells, relay and interneurons are sharing in the "specific active dendritic space" of one relay neuron in the various groups of density ("specific active dendritic space" per the theoretical autonomous space in the given group of density). The results of this calculation shows that in the first group of density 14, in the second 42.8, in the third 71.3, in the fourth 99.8 and in the fifth group of density 132.2 cell bodies have to be located in (i.e. sharing the space with) the "specific active dendritic space" of one relay neuron. The total volume of LGN and the volume of layers was also measured. The volume of layer " A " is 9.471 mm 3 , that of layer "A/' is 6.883 mm 3 and of the layer " B " is 3.485 mm 3 . The total volume of LGN is 19.839 mm 3 . This value concerns the shrunken material, and, corrected for living state the volume of " A " would be 18.468 mm 3 of " A ^ 13.421 mm 3 , and of layer " B " is 6.795 mm 3 (Table II). The overall average density was calculated from the total cell number and the volume of LGN both in shrunken and living material (Table III).

Discussion

Fig. 3. The "specific active dendritic sphere" of relay neuron is shown diagrammatically. The radius of this sphere is 80 to 100 ¡xm. The ramification of the dendrites, both the primary and secondary branchings are found in this sphere. At the ramification points the dendritic protrusions (bulges) are found. The 15 rim and 65 ¡xm distances point on the about smallest and largest distances among the relay neurons.

The total number of neurons counted in four cats with the appropriate corrections are quite uniform and are at variance with previous data (Bishop 1953). The rather small differences between the total numbers of cells in the four animals (1.3%) might be attributed to flaws in the counting procedure and/or individual differences, perhaps with more likelyhood of the first. These numerical data might be compared with those in the anterior thalamic nuclei obtained by POWELL e t al. ( 1 9 5 7 ) . T h e i n d i v i d u a l differences were

T a b l e 1. The number of cells in the different layers of LGN Layer A Cat, Cat 2 Cat 3 Cat,

282,507 278,204

Layer A l 50.54% 49.65%

204,346 212,840

Layer B 36.54% 37.97%

72.285 69.490

Total 12.92% 12.39%

559,138 560,534 563,080 545,320

100% 100%

Quantitative histological studies on LGN

163

T a b l e I I . Volume of LGN (mm3) Layer A Shrunken material Living material

9.471 18.468

Layer Aj 47.73% 47.74%

6.883 13.421

Layer B 34.69% 34.69%

3.485 6.795

Total 17.57% 17.56%

19.839 38.684

100% 100%

T a b l e I I I . Cell density (cells/mm 3 ) of LGN Layer A Shrunken material Living material

29,828,634 15,295,452

Layer B

Layer Aj 106% 105%

29,688,507 15,225.84

much larger in the anterior thalamic nuclei. The largest deviation from the average cell number was observed in the anterior ventral nucleus, and was about 14%. In the supraoptic nucleus (LERANTH et al. 1975) the individual variations in cell numbers were in the range of 23%. The average densities in the two main layers are rather similar, the difference being only 140 cells per mm 3 in the processed material. This difference is less than '1%. From the average density it is easy to calculate the distances among the cells under the assumption of an even spatial distribution. The results of this calculation are similar to the data obtained by direct measurement. The measured values are more accurate but also the calculated numbers are in good agreement with the measured data. On the basis of average density the distance between two neurons is about 28 |i.m (34 iim in living animal). The density of neurons is of impartance in judging the spatial overlap of ramification points of the dendritic trees of the relay neurons. In the preceding paper the dendritic ramifications of the relay neurons were described in quantitative terms on the basis of Golgi stained material. Since the two different histological procedures upon which the two sets of data are based have some difference in shrinkage and the cats used for the studies were not quite of the same age groups — young, although near adult size in the preceding, and adults in this study — the two sets of data can be compared only with certain reservations. However, since the shrinkage in the GOLGI procedure, practical in this Department, is smaller than in paraffin embedding (PALKOVITS et al. 1971), it is reasonable to assume that the small original size difference of the brains is compensated. The following reasonings are based also on representative measurements of some apparently crucial parameters of the dendritic trees, hence allowance has to be given for a considerable margin of error. In spite of this it is felt that such calculations offer important insights into the structure of the neuron network.

105% 105%

20,741.75 10,637.969

Total 74% 73%

28,183.77 14,453.986

100% 100%

The ramification points of dendrites of relay neurons (in both types) are arranged in a sphere around the cell body at about 80—100 ¡xm. Taking into consideration the ratio (approximately 2:1, see the next paper) between the relay and interneurons and counting with the now determined density of the neurons it can be estimated that there has to be a major spatial overlap in the ramification points of relay neurons. The sphere of ramification points belonging to one relay neuron ("specific active dendritic space") is roughly about 2 —4 x 106 ¡xm3. According to the average cell density there are about 64 and 125 neurons respectively, located in such spheres. Reckoning with the five density groups these data change, as was described above: 14, 42.9, 71.3, 99.8, 132.2 (28, 85.6, 142.6, 199.6, 264.4 respectively) other cell bodies are located within the "specific active dendritic space" of any given relay cell. There is a spatial overlap of dendrites which depends on the number of dendritic branches too. The quantitative analysis of Golgi pictures lead to an estimate of the number of principal dendrites of relay neurons (8—10 of large, 4—6 of medium size relay neurons). In one dendritic arborization of both types of relay neurons about 14 — 25 (18 on average) ramification points were found. In the specific active dendritic space, according to the average density may be a coincidence of these ramification points can be occur. The intersections of ramification points of dendrites depend (1) on the density of relay neurons, (2) of the number of ramification points belonging to one relay cell and (3) on spatial arrangement-possibility of the ramification points related to the specific and other afferents. On the basis of this speculation all the dendritic ramification points of the relay neurons, which are located within the "specific active dendritic space" of any given neuron, might intersect eachother. The coincidence of the specific parts of relay dendrites is a basic factor from the point of view of divergence.

164

Madarasz, M., J. Gerle, F. Hajdu, G. Somogyi, T. Tömböl

References P. O . : Synaptic transmission. An analysis of the electrical activity of the lateral geniculate nucleus in the cat following optic nerve stimulation. Proc. Roy. Soc, B. 141, 362 (1953). C H O W , K A O L I A N G , L . S . B L U M and R . A . B L U M : Cell ratios, in the thalamocortical visual system of macaca mulatta. J. comp. Neurol. 92, 2 2 7 - 2 3 9 (1950). L E G R O S C L A R K , W . E . : The lateral geniculate body in the platyrrhine monkey. J. Anat. 76, 131 — 140 (1941). F L O D E R U S , S.: Untersuchungen über den Bau der menschlichen Hypophyse mit besonderer Berücksichtigung der quantitativen mikromorphologischen Verhältnisse. Acta path, microbiol. scand. 53, 276 (1944). K L Ü V E R , H . , and E. B A R R E R A : A method for the combined staining of cells and fibers in the nervous system. J. Neuropath. Exp. Neur. 12, 4 0 0 - 4 0 3 (1953). L A S H L E Y , K. S.: The mechanism of vision V I . The functioning of small remnants of the visual cortex. J. comp. Neurol., 70 4 5 - 6 7 (1939). L E R Ä N T H , C S . , L . Z Ä B O R S Z K Y , J . MARTONandM. P A L K O V I T S : BISHOP,

Quantitative studies on the supraoptic nucleus in the rat. I. Synaptic organization. Exp. Brain Res., 22, 509 — 523 (1975). P A L K O V I T S , M . , P . M A G Y A R and J . S Z E N T A G O T H A I : Quantitative histological analysis of the cerebellar cortex in the cat. I. Number and arrangement in space of the Purkinje cells. Brain Research 32, 1 - 1 3 (1971). P A L K O V I T S , M . , P . M A G Y A R and J . S Z E N T A G O T H A I : Quantitative histological analysis of the cerebellar cortex in the cat. II. Cell numbers and densities in the granular layer. Brain Research 32, 1 5 - 3 0 (1971). P A L K O V I T S , M., P . M A G Y A R and J . S Z E N T A G O T H A I : Quantitative histological analysis of the cerebellar cortex in the cat. I I I . Structural organization of the mulecular layer. Brain Research 34, 1 — 18 (1971). P A L K O V I T S , M . , P . M A G Y A R and J . S Z E N T A G O T H A I : Quantitative histological analysis of the cerebellar cortex in the cat. IV. Mossy fiber-Purkinje cell numerical transfer. Brain Research 45, 15 — 29 (9172). POWELL, T . P . S „

R . W . GUTLLERY

and

W . M . COWAN:

A

quantitative study of the fornix mamillothalamic system. J. Anat. 91, 4 1 9 - 4 3 7 (1957).

J. Hirnforsch. 19 (1978) 165-176

D e p a r t m e n t of N e u r o a n t o m y , I n s t i t u t e of H i g h e r Nervous Activity, Osaka University Medical School, O s a k a / J a p a i i

Organization and projections of the neurons in the dorsal tegmental area of the rat By M a s a y a T O H Y A M A , Keiji and Toru ITAKURA

SATOH,

Tetsuro

SAKUMOTO,

Yasuhiko

KIMOTO,

Yasuyuki

TAKAHASHI,

Kazumi

YAMAMOTO

W i t h 8 figures (Received 2 n d A u g u s t 1977)

S u m m a r y : T h e organization a n d projection of t h e neurons in t h e dorsal t e g m e n t a l area a t t h e locus coeruleus level of t h e r a t h a v e been investigated b y m e a n s of t h e horseradish peroxidase ( H R P ) m e t h o d , combined w i t h m o n o a m i n e oxidase staining t o i d e n t i f y t h e noradrenaline (NA) neurons. J u d g i n g f r o m t h e a n a t o m i c a l aspects elucidated in this study, this area is composed of various different cell groups. Although t h e f u n c t i o n a l role of these neuronal groups remains t o be determined, t h e present s t u d y strongly suggests t h a t t h e dorsal t e g m e n t a l area p a r t i c i p a t e s in i m p o r t a n t regulation of several brain functions. R é s u m é : L'organization et les projections des neurones d a n s le t e g m e n t u m dorsal o n t été étudiées p a r le marq u a g e r é t r o g r a d e à la p e r o x y d a s e ( H R P ) , combiné à la réaction de la m o n o a m i n e oxidase, ceci, afin d'identifier les neurones c o n t e n a n t de la noradrenaline. Le t e g m e n t u m dorsal chez le r a t se compose de g r o u p de neurones n o m b r e u x e t a u x caractéristiques a n a t o m i ques différentes. Le rôle d e ces différentes groupes du t e g m e n t u m dorsal, bien que l ' i m p o r t a n c e de cette s t r u c t u r e d a n s le f u n c t i o n n e m e n t cérébral ne fasse pas de doute, reste à préciser.

Abberiviations AH

cell g r o u p a of M E E S S E N and anterior h y p o t h a l a m i c area

B

B A R R I N G T O N nucleus

a

OLSZEWSKI

cerebral cortex cerebellum decussation of superior cerebellar peduncle H hypothalamus 1 cell g r o u p 1 of M E E S S E N and O L S Z W S K I LC locus coeruleus LCD dorsal p a r t of locus coeruleus LCV v e n t r a l p a r t of locus coeruleus LH lateral h y p o t h a l a m i c area MH medial h y p o t h a l a m i c area n o n noradrenergic subcoeruleus n e u r o n nsc nts nucleus t r a c t u s solitarii PB p e r i - B A R R I N G T O N area pbl nucleus parabrachialis lateralis pbm nucleus parabrachialis medialis PT nucleus lateralis posterior t h a l a m i a n d lateral geniculate b o d y P V M nucleus p e r i a q u e d u c t u s ventralis mesencephali R some neurons in nucleus reticularis p o n t i s oralis RD nucleus r a p h e dorsalis sc noradrenergic subcoeruleus neuron SEE subcoeruleus complex SCP superior cerebellar peduncle SP spinal cord TD nucleus t e g m e n t i dorsalis (Gudden) TLD nucleus laterodorsalis t e g m e n t i T L D G magnocellular p a r t of nucleus laterodorsalis t e g m e n t i CC Ce DSC

Hirnforschung, Bd. 19, Heft 2

T L D P parvocellular p a r t of nucleus laterodorsalis t e g m e n t 1 TV nucleus tegmentalis ventralis (Gudden) V ventricle VM

nucleus t r a c t u s mesencephali n. trigemini

Introduction Recent anatomical and electrophysiological evidence has established that the dorsal tegmental area contains very important cell group, such as tegmental nuclei, locus coeruleus (LC) and nucleus raphe dorsalis (nRD). The degeneration silver method has demonstrated that the tegmental nuclei innervate the mammilar body (BAN and ZYO, 1963; B U C H E R a n d BURGI, 1 9 5 5 ; BURGI a n d BUCHER, 1 9 6 0 ; GUILLERY, 1 9 5 6 ; MORIN, 1 9 5 0 ; ROUSSY a n d MONSINGER,

1946). Furthermore histofluorescence method has clearly revealed that the LC and nRD contain noradrenaline (NA) and serotonin (5HT) respectively, and give rise to widespread ascending and descending projections (ANDEN et al., 1966; B J Ò R K L U N D et al., 1 9 7 3 ; CHU a n d BLOOM, 1 9 7 4 ; DÀHLSTROM a n d F U X E , 1964;

LINDVALL

et

al.,

1974;

MAEDA

et

SHIMIZU,

1 9 7 2 ; MAEDA e t a l . , 1 9 7 4 ; OLSON a n d F U X E , SATOH

et

al.,

1977;

TOHYAMA

et

1971;

al., 1 9 7 4 ; TOHY-

AMA et al., 1974; TOHYAMA, 1976; UNGERSTEDT, 1971). Additional confirmations were recently made by the degeneration silver method (SHIMIZU et al., 1974; S H I -

12

166

T o h y a m a , M., K. Satoh, T. Sakumoto, Y. Kimoto, Y. T a k a h a s h i , K . Y a m a m o t o a n d T. I t a k u r a

Fig. 1 (1—8). Stereotaxic atlas showing t h e organization of t h e dorsal tegmental area f r o m caudal mesencephalon t o t h e rostral pons of t h e rat.

MIZU et al., 1975). T h e r e t r o g r a d e tracer technique

(KIMOTO e t a l . , 1 9 7 7 ; SAKUMOTO e t a l . , 1 9 7 7 ; SATOH

using horseradish peroxidase (HRP) has given several indications that this area at the LC level might be composed of various nuclear groups with complicated fiber

e t al., 1977).

connections

ZEWSKI ( 1 9 4 9 ) , TABER (1961) a n d VALVERDE (1962).

(KIMOTO e t a l . , 1 9 7 7 ; SAKUMOTO e t a l . ,

The terminology used follows, unless otherwise s t a t e d , t h a t of BERMAN ( 1 9 6 8 ) , MEESSEN a n d OLS-

1 9 7 7 ; SATOH e t al., 1 9 7 7 ; SHIMIZU, 1 9 7 7 ; TOHYAMA

et al., 1977). However, our knowledge of the organization and fiber connections of the cells situated in this area is still incomplete. Accordingly in the present study, using the H R P method, we have attempted to differentiate the various cell groups within the dorsal tegmental area at the LC level from the point of view of fiber connections and neurotransmitters. The results have been partly reported previously

Material and Methods The H R P method was recently introduced into t h e investigations of t h e fiber connections of t h e aminergic system in t h e central nervous system (FREEDAN et al., 1975; KIVIEX a n d KUYPERS,

1975;

LEGER e t a l . ,

1975;

LYUNGDAHL e t

al.,

1975; SAKAI et al., 1976). However using H R P m e t h o d alone, t h e n e u r o t r a n s m i t t e r of t h e labeled neurons such as NA could n o t be identified. I n t h e present study, monoamine oxidase (MAO) staining (GLENNER et al., 1957) and t h e H R P method

Dorsal tegmental area of the rat

were combined in order to introduce neurotransmitter specificity into the H R P method (SATOH et al., 1977). I t has been shown that the cells stained by MAO corespond to NA neurons as seen by the histofluorescence technique (MAEDA, 1 9 7 0 ; SHIMIZU e t al., 1 9 5 9 ) . T h e r e f o r e ,

MAO-posi-

tive neurons can probably be regarded as NA neurons. More than 200 male rats weighing 150—200 g were used. H R P (Sigma type VI, 30 — 50% solution in saline) was injected using a micropippete system into various regions, such as the cerebral cortex, nucleus lateralis posterior thalami and lateral geniculate body (PT), cerebellum, hypothalamus, ' nucleus tractus solitarii(nts) and spinal cord. In the case of H R P injections into the hypothalamus and PT, we used a double syringe micropippete system in order to prevent H R P leaking to other brain regions such as the cerebral cortex, hippocampus and thalamus.

1.67

The animals were perfused transcardially with 0,4% paraformaldehyde — 1.25% glutaraldehyde in 0.05 M phosphate buffer (pH 7.2, 4°C). The brain and spinal cord were removed and postfixed with the same fixative for 4 — 6 hours, followed by washing overnight in 0.1 M phosphate buffer containing 30% sucrose. The brains were forzen and the serial frontal sections, 20—30 (xm thick, were cut. The sections were incubated alternatively in the medium of Tris-HCLbuffer containing 3,3'-diaminobenzidine and H 2 0 2 for revealing H R P in nerve cells and in the medium of 0.1 M phosphate buffer containing 0.3% tryptamin hydrochloride and 0.025% nitro-blue tetrazolium for 45 — 60 min (37 °C). In some cases both methods were performed on the same sections.

168

Tohyama, M., K . Satoh, T. Sakumoto, Y . Kimoto, Y . Takahashi, K . Yamamoto and T. Itakura

m

. t' P V M -

r *"• V

K

r ^ . z *

R D

*

- 2

Fig. 2. Dark-field photomicrograph of H R P positive neurons in the n R D and PVM following injection of H R P into the PT. x 100

rous neurons in the nPVM were storongly labeled by HRP (Fig. 2). These neurons were situated The dorsal tegmental area can be divided into two in the PGM between the level of the rostral pole regions: (1) the part located in the periaqueductal of the nucleus tegmentalis ventralis (nTV) and gray matter and (2) the part located ventral and the caudal level of the decussation of the superior lateral to the periaqueductal gray matter. cerebellar peduncle (SCP) (Fig. 1-1). Some of the nPVM neurons were also labeled by HRP after injection into the cerebral cortex, spinal cord or (I) Dorsal tegmental area located in the periaqueductal into the nts (Fig. 8-1). From the Nissle staining it is slightly difficult to distinguish the nPVM from the gray matter neighbouring territories. This nucleus, as will be Nucleus raphe dorsalis (nRD). The nRD can be described below, is composed of medium sized cells easily recognized in the ventral region of the peri- which were mainly fusiform in shape, while surroundaqueductal gray matter (PGM) by Nissle staining ing areas were composed of exclusively small sized (Fig. 1 — 1 ~ 7 ) . This nucleus extends from the caudal cells (Fig. 1-1 ~ 2 ) . pole of the oculomotor nucleus to the caudal pole Following administration of HRP into the medial of the nucleus tegmentalis dorsalis (nTD). The cells part of the hypothalamus (AH and medial preoptic are medium sized and fusiform. Numerous nRD area (POM)), a large number of labeled neurons were neurons were labeled by HRP injections into: 1) cere- found in the PGM from the level of rostral part of the bral cortex; 2) preoptic area; 3) anterior hypothalamus "BARRINTON nucleus" (BARRINGTON, 1925; SATOH, (AH); 4) lateral hypothalamus (LH) (Figs. 2, 3, 4 1976) to the caudal part of the decussation of the SCP. and 8—3). Only a few labeled cells were found in the These labeled cells were almost exclusively small and nRD when the HRP was injected into the cerebellum. oval in shape, and thus could be distinguished from It should be worthwhile to note that following injec- those in the nPVM (which were labeled by HRP tion of HRP into the hypothalamus, neurons in the injections into the PT) (Figs. 1-1 ~ 2 and 8-1 ~ 2 ) . nRD were labeled mainly ipsilaterally (Figs. 3 and 4). Just medial to the VM, HRP labeled cells constiThe cell group occupying the area lateral to the nRD tute a closely grouped cluster. At the rostral pole and medial to the nucleus nervi trigemini mesen- of the nTV, these neurons merged laterally with cephalicus (VM) in the PGM and its adjacent neuronal the small sized HRP labeled cells situated in the nucgroup. Following the HRP injections into various leus parabrachialis lateralis (pbl) (Fig. 3 arrows). At regions of the brain (see below), we have noticed the rostral pole of the "BARRINGTON nucleus", an important group of neurons, which occupies the these neurons fused caudally with the small sized area lateral to the nRD and medial to VM in the HRP containing neurons in the central gray matter PGM (Fig. 1-1). We call this hitherto unrecognized (TLDP). neuronal group in the PGM 'the nucleus periaqueNucleus later odor salis tegmenti (TLD). Following ductus ventralis mesencephali (nPVM)' in this study. injection of HRP into the cerebral cortex, PT Following the injection of HRP into the PT, nume- or hypothalamus, medium sized cells labeled Results

Dorsal tegmental area of the rat

I

R D

W4 'W y,:

iJ

TLDp'

/

P b l

y,

S C P

.

3T

«

.

,

-

Fig. 3. Dark-field photomicrograph of H R P positive neurons in the nRD, T L D P and pbl following injection of H R P into the right POM. Note that small sized H R P labeled neurons which lie in an area just medial to the VM merged laterally (arrows) with H R P labeled pbl neurons. Note also that the nRD was labeled with marked ipsilateral predominance, x 70

.J v

• #

9 f '-

^

169

* V

T

.

*

"

.

T L D G »

a • Í Sgi Fig. 4. Dark-field photomicrograph of H R P positive neurons in the nRD, TLDG and T L D P (indicated by arrows) following injection of H R P into the AH. XlOO

flm

by H R P were found in the central gray matter from the caudal pole of the nTV to the rostral pole of the LC (TLDG) (Figs. 1-3 ~ 8 and 4). It should be noted that H R P injection into the hypothalamus resulted in labeling both small sized cells (TLDP) and medium sized cells (TLDG) (Fig. 4), while injections into the cerebral cortex or P T failed to label these small neurons. Furthermore, it should be emphasized that only injection of H R P into the LH succeeded in labeling the small cells arround the "BARRINGTON nucleus" (PB) (Fig. 5 a). Accordingly the area which lies lateral to the nRD and medial to the VM (called 'nucleus laterodorsalis tegmenti (TLD)' by CASTALDI in the cat) can be subdivided in the rat into three regions: 1) parvo-

C e

B

tPB 5 o

Fig. 5 a, b and c. a) Dark-field photomicrograph of H R P positive neurons in the P B area following injection into the LH. x 100; b) Dark-field photomicrograph showing faintly labeled cells in the " B " following injection into the spinal cord, x 100; c) Dark-field photomicrograph showing strongly labeled neurons in the " B " following injection of H R P into the spinal cord of the 6-hydroxydopa pretreated rat. x 100

170

Tohyama, M., K. Satoh, T. Sakumoto, Y . Kimoto, Y . Takahashi, K. Yamamoto and T. Itakura

Fig. 5 b and c

cellular part of the TLD (TLDP) which innervates mainly the AH. 2) magnocellular part of the TLD (TLDG) which innervates the hypothalamus, cerebral cortex and PT. 3) peri-BARRINGTON area (PB) which innervates mainly the LH. It should be mentioned that although CASTALDI included the LC and "BARRINGTON nucleus" in the TLD, we excluded these regions from the TLD. "Barrington nucleus". The "BARRINGTON nuccleus" as defined here (Fig. 1-7 ~ 8 ) extends from the level of the rostral disappearance of the LC to the level of the caudal pole of the TLD. These neurons have been included in the TLD, but recent study (SATOH, 1976) indicated that this nucleus can be separated from the surrounding TLD. This nucleus is composed of medium sized cells of oval shape occupying a zone medial to the VM and also a laterocaudal part of the TLD. When the HRP was injected into the spinal cord, this nucleus was faintly labeled (Fig. 5 b). However, this nucleus was strongly labeled after pretreatment with 6-hydroxydopa (Fig. 5 c) which produces selective degeneration of NA nerve terminals (CORRODI e t al. 1 9 7 1 ; ONG et al., 1 9 6 9 ; SACHS a n d JOHNSSON, 1 9 7 2 ; SACHS et al., 1 9 7 3 ; TOHAYMA et al., 1 9 7 4 ; THOYAMA e t al., 1 9 7 4 ; TOHYAMA, 1 9 7 6 ) .

Cell group 0 and tx of Meessen and Olszewski. Following injection of HRP into the lateral part of the hypothalamus (lateral preoptic area (POL) and LH) several HRP positive neurons could be found in the central gray matter at the mid-point

of the LC. These cell groups are composed of small sized neurons and might correspond to the cell group 0 a n d oc of MEESSEN a n d OLSZEWSKI (1948) ( F i g s . 1 - 1 0

and 8-10). It should be noted that none of the neuronal groups described above showed MAO activity. Locus coeruleus. (LC) The LC extends from the mid-point of the VM to the caudal level of the nTD (Fig. 1-8 ~ 1 0 ) and the neurons are reported to be almost exclusively noradrenergic (ANDEN et al., 1966; DAHLSTROM a n d F U X E , 1 9 6 4 ; LINDVALL e t al., 1 9 7 4 ; MAEDA et SHIMIZU, 1 9 7 2 ; OLSON a n d F U X E ,

1972;

TOHYAMA et al., 1974) and stain for MAO activity (HASHIMOTO e t

al.,

1962;

MAEDA,

1970;

SHIMIZU

et al., 1959). The dorsal part of this nucleus is composed of small cells. Most of the neurons labeled by the HRP with injections into the cerebral cortex, cerebellum or PT, while only scarrered labeled cells were found in this area with injections of HRP into the spinal cord, nts, LH or POL. The ventral part of the LC, on the other hand, is composed of relatively large sized and multipolar neurons. Injections of HRP into the spinal cord, cerebellum, nts or lateral part of the hypothalamus (LH and POL), always caused several HRP containing neurons in this area. It is worthwhile to note that labeled neurons in the LC were concentrated in its ventral part, when the HRP was injected into the spinal cord (Figs. 6a and b).

Dorsal tegmental area of the rat

171

Fig. 6 a and b. Photomicrographs showing a double reaction of MAO and DAB on the same section. MAO staining (b) and DAB reaction observed in the dark-field (a). Arrows indicate that the same neurons show MAO and DAB reactions. Note that H R P labeled neurons in the LC are concentrated in its ventral part, x 100

Fig. 7. Dark-field photomicrograph of H R P positive neurons in the pbl and pbm following injection into the LH. x 100

(II)

Dorsal tegmental area located ventral and lateral to the periaqueductal gray matter

Subcoeruleus complex (SCC). The SCC (Fig. 8-9 ~10) extends from the level of LC to the "BARRINGTON nucleus". The SCC is composed of neurons which show strong MAO activity and of other neurons which do not show MAO activity. HRP injections into the spinal cord or cerebellum labeled, in the SCC, medium

to large sized and multipolar shaped neurons showing strong MAO activity (Fig. 6 a and b). In the case of cerebellum injections, several neurons which did not show MAO activity were also labeled in this area. Although HRP injection into the hypothalamus in this study failed to label MAO-positive neurons in the SCC and other pontine regions, these results should be "false-negative", since histofluorescence analysis clearly showed that these neurons also send their

172

T o h y a m a , M., K . Satoh, X. S a k u m o t o , Y. K i m o t o , Y. T a k a h a s h i , K. Y a m a m o t o a n d T. I t a k u r a

Fig. 8 (1—4). Schematic represent a t i o n of t h e organization a n d projections of t h e neurons in t h e dorsal t e g m e n t a l area.

axons to innervate the hypothalamus (OLSON and FUXE,

1971;

OLSON

and

FUXE,

1972;

MAEDA

et

SHIMIZU, 1 9 7 2 ; SAKUMOTO e t a l . , 1 9 7 7 ) .

Reticular formation rostromedial to SCC (R). Injection of H R P into the spinal cord also gave evidence that neurons in the reticular formation (R) situated slightly rostromedial to the SCC also send their axons to the spinal cord. These neurons do not stain for MAO activity. Cell group I of Meessen and Olszewski. Cell group 1 of MEESSEN and OLSZEWSKI lies just ventrolateral to the fasciculus longitudinalis medialis at the level of the caudal pole of the nTD (Fig. 1-9). This nucleus is characterized by its densely arranged medium sized

cells. Following injection of H R P into the cerebellum, a large number of these cells were labeled. Nuclei parabrachialis lateralis (pbl) and medialis (pbm). Following injection of H R P into the medial part of the hypothalamus (POM and AH), a large number of labeled neurons were found in the pbl. This neuronal group is composed of small cells and extends from the rostral pole of the "BARRINGTON nucleus" to the caudal part of the decussation of the SCP (Fig. 7). These H R P labeled neurons fused rostrally with small sized H R P containing neurons situated just medial to the VM at the mid-point of the nVT. When H R P was injected into the POL or LH, on the other hand, a moderate number of H R P posi-

Dorsal tegmental area of the rat

tive neurons were found in the pbm at the level of the rostral pole of the " B A R R I N G T O N nucleus". Furthermore injection of H R P into various hypothalamic areas resulted in labeling several mantle cells of the SCP at the caudal level of its decussation, while after injection of H R P into nts a cluster of labeled neurons occured in the nucleus reticularis pontis oralis just ventral to the SCP at the level of the caudal end of the nTV. This H R P labeled cell group contains mainly small cells. No H R P labeled neurons, however, were found in the pbl and pbm ( T A K A H A S H I et al., in preparation).

173

Organization and projection of the neurons in the dorsal tegmental area at the LC level are schematically drawn in Fig. 8. Discussion In the cytoarchitectonic study, c ell bodies which show a characteristic arrangement are readily identified as a nucleus such as the LC, n R D , nTV, nTD, and VM in the dorsal tegmental area. In other parts of the dorsal tegmental area, however, cells are more loosely arranged so that the identification of any nucleus

174

Tohyama, M., K. Satoh, T. Sakumoto, Y . Kimoto, Y . Takahashi, K. Yamamoto and T. Itakura

group is difficult when based only upon the cytoarchitectonic characteristic of the nucleus. On this point, the present study indicates that the combination of retrograde tracer techniques, such as the HRP and histochemical techniques, such as MAO staining is a useful tool for this kind of investigation. The present study gives evidence that the dorsal tegmental area can be subdivided into various neuronal groups. We will mainly discuss some of these neuronal groups. The present findings indicate that a cluster of medium sized cells (which lies in the PGM lateral to the nRD and medial to the VM) (nucleus periaqueductus ventralis mesencephali (nPVM)) give rise to widespread ascending projections like the neurons of the LC and nRD. But it should be emphasized that this group of cells is composed exclusively of nonnoradrenergic neurons. The present study also provides evidence that the TLD can be subdivided into three components: TLDG, TLDP, and PB. Medium sized cells in the TLD (TLDG) (Figs. 1 and 8), which lies just rostromedial to the LC, innervates monosynaptically the cerebral cortex, hypothalamus and the PT (Fig. 8 - 4 ^ 6 ) . On the other hand, small sized cells in the TLD (TLDP) innervate mainly the AH, while neurons in the peri-BARRINGTON area (PB) innervate mainly the LH (Fig. 8-4 ~ 8 ) . It should be noted that the LC and SC (which contain NA), and the PVM and "BARRINGTON nucleus" (which do not contain NA) both innervate the spinal cord. It should be also noted that the neurons in the SC are composed of NA and non-NA neurons, and both types of neurons have a similar projection pattern. In the present study, neurons of the "BARRINGTON nucleus" were intensively labeled by the H R P when the animals were pretreated by 6-hydroxydopa. The "BARRINGTON nucleus" was first described b y BARRINGTON (1.925) in t h e c a t as t h e ' p o n t i n e m i c -

turition reflex center', since the bilateral destruction at the dorsal tegmental area of the cat resulted in an urinary retention. Many subsequent electrophysiological and morphological studies concerning the central control of micturition have been reported (BORS and

KOMMAR,

1971;

KURU,

1965;

LANGWORTHY

•et al., 1 9 4 0 ; ROUSSEL e t al., 1 9 7 6 ) , b u t precise locali-

zation of this nucleus remained to be determined. Recently SATOH identified the "BARRINGTON nucleus" in the TLD by lesion experiments in the rat (1976). In this study we have demonstrated the monosynaptic fiber connections to the spinal cord from this nucleus as well as from the LC. It is of great interest to note that the amount of H R P transported through the axons to the perikaryon of this nucleus remarkably increased by pretreatment of 6-hydroxy-

dopa. This observation might indicate that central NA systems also share in the control of the micturition reflex, but that the main control of this function is played by "BARRINGTON nucleus" which does not contain NA. Possible interactions between NA and non-N A neuronal system have been suggested previously (JOUVET, 1972). The present findings together with some previous e l e c t r o m i c r o s c o p i c studies (KODA a n d BLOOM, 1 9 7 7 ; ITAKURA et al., 1 9 7 7 ; MAEDA et al., in p r e s s ;

SAKUMOTO et al., 1977) are also in favor of possible interactions among different neurotransmitter systems. This paper is dedicated to Prof. Nobuo S H I M I Z U , who retired from the Osaka University Medical School on May 31, 1976.

References ANDEN, N . - E . ,

A . DÄHLSTROM,

K. FUXE,

K . LARSSON,

L.

and U . U N G E R S T E D T : Ascending monoamine neurons to telencephalon and diencephalon. Acta physiol. Scan., 67, 3 1 3 - 3 2 6 (1966). B A R R I N G T O N , F. J . T . : The effect of lesion of the hind- and mid-brain on micturition in the cat. Quart. J . Exptl. Physiol. 15 8 1 - 1 0 2 (1925). BAN, T., and K. ZYO: Experimental studies of the mamillary peduncle and mamillotegmental tracts in the rabbit. Med. J . Osaka Univ. 13, 2 4 1 - 2 7 0 (1963). B E R G M A N , A. L . : The brain stem of the cat. Cytoarchitectonic atlas with stereotaxic coordinates. University of Wisconsin Press, (1968). B J Ö R K L U N D , A. O., R. Y. M O O R E , A. NoBiNandU. S T E N E V I : The organization of tubero-hypophyseal and reticuloinfundibular catecholamine neuron system in the rat brain. Brain Research 51, 1 7 1 - 1 9 1 (1973). B O R S , E . , and A. E. C O M A R R : Neurological urology. Univ. Park Press, (1971). B U C H E R , N. M., und S. M. B U R G I : Über einige rhinencephale Verbindungen des Zwischen- und Mittelhirn. Dtsch. Zschr. Nervenheilk. 174, 8 9 - 1 0 6 (1955). B U R G I , S. M., und V. M. B U C H E R : Markhaltige Faserverbindungen im Hirnstamm der Katze. Monogr. Neurol. Psychiat. 87 (1960). C A S T A L D I , L . : Studi sulla struttura e sullo sviluppo del mesencefalo. Ricerche in Cavia cobaya. I I I . Arch. ital. Anat. Embriol. 23, 4 8 / - 6 0 9 (1926). CHU, N. S., and F. E. BLOOM: The catecholamine containing neurons in the cat dorsolateral pontine tegmentum; distribution of cell bodies and some axonal projections. Brain Research 66, 1 - 2 1 (1974). OLSON

CORRODI, H . ,

W.G.CLARK

and

D . I. MASUOKA:

The

syn-

thesis and effects of DL-6-hydroxydopa. 6 hydroxydopamine and catecholamine neurons. Eds. by M A L F O R M S , J . , and E. T H O E N E N . North-Holland Amsterdam —London 1 8 7 - 1 9 2 (1971). D Ä H L S T R O M , A., and K . F U X E : Evidence for existence of monoamine containing neurons in the central nervous system. Acta physiol. Scan. 62, Suppl. 232, 1 — 55 (1964). F R E E D M A N , R., S. L. F O O T E and F. E. B L O O M : Histochemical characterization of neocortical projection of the nucleus

Dorsal tegmental area of the rat

175

ITAKURA, T . ,

Central dual innervation of arterioles and capillaries in the brain. Stroke, 8, 360- 365 (1977) The role of monoamines and acetylchoJOUVET, M . : line containing neurons in the regulation of the slepe waking cycle. Ergebniss der Physiologie. 64, 1 6 6 - 3 0 7 (1972).

K . YAMAMOTO, M . TOHYAMA a n d N . SHIMIZU:

coeruleus noradrenaline neuron: The cerebellar innervation. Brain Research 28, 1 6 5 - 1 8 1 (1971). OLSON, L . , and K . F U X E : Further mapping out of central noradrenaline neuron system: Projections of the "subcoeruleus" area. Brain Research 43, 289 - 2 9 5 (1972). O N G , H., C. R. C R E V E L I N G and J . D A R Y : The synthesis of 2,4,5-trihydroxy-phenylalamin(6 hydroxydopa). A centrally active norepinephrine deeping agent. J . med. Chem. 12, 4 5 8 - 4 6 1 (1969). R O U S S E L , B., J . F. P U J O R , et M. J O U V E T : Effects des lesion du tegmentum ponti pontique sur les etats de sommeil chez le rat. Arch. ital. Biol. 114, 188- 209 (1976). R O U S S Y , G . , et M. M O S I N G E R : Traite de neuroendocrinologie. Manson (1946). S A C H S , CH., and J . J O N S S O N : Selective 6-hydroxydopa induced degeneration of central and peripheral noradrenaline neurons. Brain Research 40, 563 — 568 (1972). S A C H S , C H . , G . J O N S S O N and K . F U X E : Mapping of central noradrenaline pathways with 6-hydroxydopa. Brain Research 63, 2 4 9 - 2 6 1 (1973).

KIMOTO, Y . ,

K . SATOH,

SAKAI, K . , M . TOURET,

locus coeruleus in the squirrel monkey. J . Comp. Neurol. 164, 2 0 9 - 2 3 2 (1975). GLENNER, G. G.,

H. J . BURTNER

and

G. W . J R . BROWN :

The histochemical demonstration of monoamine oxidase activity by tetrazolium salts. J . Histochem. Cytochem 5, 5 9 1 - 6 0 0 (1957). GUILLERY, R . W . : Degeneration in the postcommisural fornix and the mamillary peduncle of the rat. J . Anat (Lond.). 90, 350--370 (1956). HASHIMOTO, P . I L ,

T . MAEDA,

K . TORII

and

N . SHIMIZU:

Histochemical demonstration of autonomic regions in the central nervous system of the rabbit by means of a monoamine oxidase staining. Med. J . Osaka Univ. 12, 425 — 465 (1962).

T . SAKUMOTO,

M . TOHYAMA

and

Afferent fiber connections from lower brain stem with special reference to noradrenergic iieuron system. J . Hirnforsch. 1 9 , 8 5 - 1 0 0 (1978). K I V I E T , J . , and H . G . J . M . K U Y P F . R S : Subcortical afferents to frontal lobe in the rhesus monkey studied by means of retrograde horseradish peroxidase transport. Brain Research 85, 2 6 1 - 2 6 6 (1975). K O D A , L. Y., and F. E. B L O O M : A light and electron microscopic study of noradrenergic terminals in the rat dentate gyrus. Brain Research 120, 327 — 335 (1977). L A N G W O R T H Y , O . R . , L . C . K O L B and L . G . L E W I S : Physiology of micturition. Baltimore, (1940). N . SHIMIZU:

LEGER, L.,

K . SAKAI, D . SALVERT, M . TOURET a n d M .

JOU-

VET: Delineation of dorsolateral geneculate afferents from the cat brain stem as visualized by horseradish peroxidase technique. Brain Research 93, 490 — 496 (1975). L I N D V A L L , O . , and A. B J Ö R K L U N D : The organization of the ascending catecholamine neuron system in the rat brain: As revealed by glyoxilic acid fluorescence method. Acta physiol. Scan. Suppl. 412, 1 - 4 8 (1974). M A E D A , T . : Histochemical consideration on the relationship between monoamine oxidase and biogenic monomaines in the brain. Advance in Neurological Sciences 13, 812 bis 820 (1970). MAEDA, T . ,

A. KASHIBA,

M . TOHYAMA,

M. HORI,

T . ITA-

and N . S H I M I Z U : Aminergic terminals and their contacts demonstrated by in situ fixation of endogeneous monoamine with potassium permanganate. Brain Research in press. KURA

MAEDA, T . , C. PIN, D . SALVERT, M . L I G I E R a n d M .

JOUVET:

Les neurones contenant des catecholamine du tegmentum pontique et leur voies de projection chez le chat. Brain Research 57, 1 1 9 - 1 5 2 (1973). M A E D A , T., et N . S H I M I Z U : Projection ascendante du locus coeruleus et d'autres neurones aminergiques au niveau du prosencephale du rat. Brain Research 36, 19 — 35 (1972). M E E S S E N , H . , and J . O L S Z E W K I : A cytoarchitectonic atlas of the rhombencephalon of the rabbit. Karger, Basel. (1948). M O R I N , F . : An experimental studies of the hypothalamic connections in the guinea pig. J . Domp. Neurol. 92, 91 bis 115 (1950). O L S O N , L . , and K . F U X E : On the projection from the locus

D . SALVERT, L . L E G E R a n d M .

JOU-

VET: Afferent projections to the cat locus coeruleus as visualized by the horseradish peroxidase technique. Brain Research 120, 1 — 21 (1976).

SAKUMOTO, T . ,

M . TOHYAMA,

K . SATOH,

T . ITAKURA,

T.

KINUGASA, T . TANIZAWA, K . YAMAMOTO, K . K U R A C H I a n d

N. S H I M I Z U : Fine structure of noradrenaline terminals in the inner layer of the median eminence of the rat. J . Hirnforsch., in press. SAKUMOTO, T . ,

M . TOHYAMA,

K . SATOH

and

N . SHIMIZU:

Afferent fiber connections from lower brain stem to hypothalamus with special reference to noradrenaline neuron system. Exp. Brain Res. 30 (1977) in press. S A T O H , K . : Experimental morphological studies on the micturition reflex center in the pontine tegmentum in the rat. Brain and nerve 29, 63 — 71 (1976). (in japanese). SATOH, K . ,

M . TOHYAMA,

K . YAMAMOTO, T . SAKUMOTO

and

N. S H I M I Z U : Noradrenaline innervation of the spinal cord by the horseradish peroxidase method combined with monoamine oxidase staining. Exp. Brain Res. 30, 1 7 5 - 1 8 6 (1977). S H I M I Z U , N., N. M O R I K A W A and K. O K A D A : Histochemical studies of monoamine oxidase of the brain of rodents. Z. Zellforsch. 49, 3 8 9 - 4 0 0 (1959). SHIMIZU, N . ,

S . OHNISHI,

M . TOHYAMA

and

T.

MAEDA:

SHIMIZU, N . ,

S . OHNISHI,

M . TOHYAMA

and

T . MAEDA:

Demonstration of ascending projection from locus coeruleus by degeneration silver method. Exp. Brain Res. 20, 1 8 1 - 1 9 2 (1974).

Demonstration of ascending projection of amine neurons in the brain stem by degeneration silver method. 10th Int. Cond. Anat. (Abstract) 135 (1975). SHIMIZU, N.: Morphology of locus coeruleus. From the memorial book of the retirement of Prof. SHIMIZTJ from Osaka Univ. Med. School 1977, in japanese. T A B E R , T . : The cytoarchitecture of the brain stem of the cat. I Brain stem nuclei of the cat. J . Comp. Neurol. 116, 161 - -187 (1961). T O H Y A M A , M.: Comparative anatomy of the cerebellar catecholamine innervation from teleosts to mammals. J . Hirnforsch. 17, 4 3 - 6 0 (1976). TOHYAMA, M.,

T . MAEDA,

A. KASHIBA

and

N . SHIMIZU:

Fluorescence and electron microscopic analysis of axonal change of coerulo-cortical noradrenaline neuron system following destruction of locus coeruleus and administra-

176

Tohyama, M., K. Satoh, T. Sakumoto, Y. Kimoto, Y. Takahashi, K. Yamamoto and T. I t a k u r a

tion og 6-hydroxydopa. Med. J. Osaka Univ. 24, 205 bis 221 (1974). T O H Y A M A , M . , T . M A E D A and N . S H I M I Z U : Detailed noradrenaline pathways of locus coeruleus neuron to the cerebral cortex with use of 6-hydroxydopa. Brain Research 79, 1 3 9 - 1 4 4 (1974). TOHYAMA, M.,

K . SATOH,

T . SAKUMOTO

and

N.

SHIMIZU:

Possible interaction between noradrenergic and nonnoradrenergic neuron system. Acta Anat. nippon., (Abstract), in japanese, in press. U N G E R S T E D T , U . : Stereotaxic mappings of the monoamine pathways in the r a t brain. Acta Physiol. Scan., Suppl. 367, 1 - 4 8 (1971).

VALVERDE, F. : Reticular formation of the pons and medulla oblongata. A golgi study. J. Comp. Neurol. 116, 71 — 99 (1961).

Address: D r . M . TOHYAMA

Department of Neuroanatomy Institute of Higher Nervous Activity Osaka University Medical School 3 - 5 7 Nakanoshima 4 chome Kitaku Osaka 530 J a p a n

J . Hirnforsch. 19 (1978) 177 - 187

Paul-Flechsig-Institute of Brain Research, Department of Neuroanatomy, Karl-Marx-University Leipzig, German Democratic Republic (Head Prof Dr sc med. E W I N K E L M A N N )

Phylogenetical changes and functional specializations in the dorsal lateral geniculate nucleus (dLGN) of mammals1-2 3 By Kurt

BRAUER,

Wilfried

SCHOBER

and Ernst

WINKELMANN

With 16 figurs (Received 2 nd August 1977) Summary Depending on their visual specialization and their place in the zoological system the mammalian species show some peculiarities in the morphology of the dorsal lateral geniculate nucleus (dLGN). The lamination of dLGN can be find in mammals with a highly developed visual system and a proportionally big part of ipsilaterally projecting retmogemculate fibres Number and sequence of ipsi- and contralaterally innervated laminae varies between species of different taxonomical categories Hence these features depend on phylogenetical trends which could be traced back to the beginning of the "mammalian radiation" in the cretaceous period and the earliest tertiary Ancestral conditions may be found m mammals with an unlaminated dLGN and a medially located ipsilaterally innervated part It may be registrated that a cytoarchvtectomcal separation of laminae (concerning the ipsi- and contralateral innervation) is the secondary step after fibre anatomical separation. The most primitive status may be realized in mammals having a zone of mixed ipsi- and contralateral fibre input. Some conclusions are made with regard to the suitability of lab mammals for generalization of results in the dLGN morphology. Zusammenfassung In Abhängigkeit von ihrer visuellen Speziahsation und ihrem Platz im zoologischen System weisen die Säugetierarten einige Besonderheiten im Bau des dorsalen Kerns des Corpus geniculatum laterale (dLGN) auf Eine Lammierung des dLGN wird bei Säugern mit einem hoch entwickelten visuellen System und einem vergleichsweise großen Anteil ipsilateral projizierender retinogemculärer Fasern angetroffen. Anzahl und Reihenfolge ipsi- und contralateral innervierter Laminae variieren zwischen Species, die unterschiedlichen taxonomischen Kategorien angehören Daraus folgt, daß diese Merkmale von phylogenetischen Trends abhängig sind, welche bis zum Beginn der "Säuger-Radiation" m der Kreidezeit und dem frühesten Tertiär zuruckverfolgt werden könnten Ancestrale Bedingungen zeigen möglicherweise Säugetiere mit einem unlammierten dLGN und einem medial lokalisierten ipsilateral innervierten Anteil. Es kann festgestellt werden, daß die cytoarchitektonische Abgrenzung der Laminae als sekundärer Schritt der faseranatomischen — in bezug auf die ipsi- und contralaterale Innervierung — folgt Das primitivste Stadium wird bei Säugetieren anzutreffen sein, bei denen eine Zone gemischten ipsi- und contralateralen Inputs existiert Es wurden einige Schlußfolgerungen über die Eignung von Labortieren für eine Verallgemeinerung von morphologischen Befunden am dLGN gezogen. Introduction

a n d fibre-preparations. Investigators utilize as most i m p o r t a n t characteristics size a n d shape of nerve cell

Numerous c o m p a r a t i v e investigations concerning t h e m a m m a l i a n dorsal l a t e r a l geniculate nucleus (dLGN) indicate so m u c h differences in morphological characteristic properties rarely t o observe in a n y other s u b c o r t i c a l s t r u c t u r e . T h e r e are not only considerable size differences in t h e d L G N of various species, but also specific peculiarities in its histological structure a b o u t which a lot of literature has been published. T h e s e findings are based in the first line on Nissl-

s o m a t a , t h e presence or absence of their

laminar

arrangement as well as especially during t h e last years — the demonstration of an ipsi — or c o n t r a l a t e r a l afferentiation for characterizing the laminae or for t h e proof of lamination if the l a t t e r cannot be vizualized b y the Nissl-method. Comparing the d L G N structure in various orders or species it m a y be of i m p o r t a n c e t o ask for the causes of these striking differences. W e will t r y to find some references or answers in this paper a n d b y t h e w a y t o the usefulness of some lab m a m m a l s for

1 Sponsored by a grant of the Ministry of Science and Technology of the GDR 2 Prof. Dr J Szentagothai dedicated with the best wishes to the 65 birthday 3 The authors wish to thank Mrs M A T T H E S and Dr B I G L for critical reading the english text.

experiments concerning the visual system, too. dLGN structure o f various m a m m a l s T h e following review of t h e most i m p o r t a n t characteristics will include only placental m a m m a l s since —

178

Brauer, K,, W. Schober and E. Winkelniann

on the one hand — the homologies of the "LGNa" and "LGNb" of Monotremata are not completly settled ( C A M P B E L L and H A Y H O W 1 9 7 1 ) and — on the other — the Marsupialia investigated until now show relations which are mainly in accordance with those of basal and speciallized placental mammals (HAYHOW, 1 9 6 7 , CAMPBELL 1 9 7 2 ) .

A detailed investigation of the homologous structures of the dLGN in other vertebrates is available by EBBESSON

(1972).

Insectívora (fig. 1, 2) This order includes numerous olfactory orientating species with highly reduced visual system but also species, where the eye is developed rather good. But there are also a few tree-living forms with a good organized visual system. While we do not have any results on moles and shrews representing insectivores with poor developed visual apparatus, the dLGN of hedgehog (Erinaceus europaeus) a species having a better developed visual system has been investigated (fig. 1). No cytoarchitectonic lamination could be observed: After enucleation the medial and dorsal sections of the ipsilateral dLGN show a sharply outlined islelike area with degenerating terminals. These areas are degeneration-free on the contralateral side (CAMPBELL 1 9 7 2 ) . This fact can be considered as primitive stage of a beginning lamination which was already concluded from fibre-anatomical studies but could not yet be shown cytoarchitectonically (CAMPBELL 1972).

One of the insectivores with highly developed visual system is the tree-shrew (Tupaia) (fig. 2). In the dLGN six laminae may be detected cytoarchitectonically as well as after degeneration of retinal fibres ( G L I C K S T E I N 1 9 6 2 , C A M P B E L L 1 9 7 2 , H U B E L 1 9 7 5 ) . According to the widely accepted enumeration of laminae from lateral to medial, used for the laminated dLGN, the laminae 1, 3, 4 and 5 receive a contralateral input and only laminae 2 and 6 an ipsilateral one. According to H U B E L ( 1 9 7 5 ) there are differences in the projection of these laminae onto different cortical layers. Chiroptera (fig. 3, 4) Our material as well as the investigations of P E N T N E Y and C O T T E R ( 1 9 7 6 ) show that the dLGN of bats who orientate primarily acoustically is well developed. Nevertheless it does not show any lamination or a subdivision in different cell regions (fig. 3). Regarding the termination zone of ipsilateral fibres no studies are available. Unfortunately, no results for the Macrochiroptera who orientate mainly optically are published in lite-

rature. After analyzing own preparations we can now say there may exist cytoarchitectonically a lamination in the dLGN (fig. 4). Primates (fig. 5, 6, 7, 8) Members of this order dispose of laminated dLGN. The lamination is already visible cytoarchitectonically. Regarding the number of laminae of various species opinions are partly contradictory. H A S S L E R ( 1 9 6 7 ) described four laminae at Callithrix, Aotes, Saimiri and Tarsius, but at prosimians he found six (fig. 5 ) . J O N E S ( 1 9 6 6 ) who investigated Aotes, agrees with H A S S L E R (fig. 6 ) . Contrary to H A S S L E R , CAMPOSO R T E G A and H A Y H O W ( 1 9 7 0 ) as well as T I G G E S and T I G G E S ( 1 9 7 0 ) indicate a seventh lamina at Galago, a prosimian. At Cebus and Macaca also seven, at Papio even- nine laminae are distinguished (CAMPOSO R T E G A a n d H A Y H O W 1 9 7 0 ) . CAMPBELL ( 1 9 7 2 )

des-

cribes in Saimiri a seventh lamina too (0), but he found six at the prosimian Nycticebus. At Galago and Nycticebus it is unanimously stated that laminae 2, 3 and 4 get input from ipsilateral, laminae 1, 5 and 6 from contralateral retina (CAMPOSO R T E G A a n d HAYI-IOW 1 9 7 0 , C A M P B E L L 1 9 7 2 , T I G G E S a n d TIGGES 1 9 7 0 ) .

According to J O N E S ( 1 9 6 6 ) the two inner laminae contrary to the dorsal and ventral adjacent outer laminae of Aotes are innervated ipsilaterally. CAMP O S - O R T E G A and G L E E S ( 1 9 6 7 ) as well as C A M P B E L L ( 1 9 7 2 ) described in Saimiri 6 laminae after fibredegeneration whereas cytoarchitectonically only 4 laminae can be distinguished. W O N G - R I L E Y ( 1 9 7 2 ) and T I G G E S and O ' S T E E N ( 1 9 7 4 ) also indicate six laminae for Saimiri and mention that the distribution of optic fibres is the same as in rhesus monkeys and men, i.e. the laminae 1, 4 and 6 are contralaterally innervated, laminae 2, 3 and 5 ipsilaterally. Compared with that Hylobates, a pongide-like species, has only 4 laminae (KANAGASUNTI-IERAM and K R I S H N A M U R T I

Fig. 1. Erinaceus europaeus (hedgehog); left LGN; coronal section, Cresylviolet and Luxol-fast-blue stained, x 50; dLGN = dorsal lateral geniculate nucleus; vLGN = ventral lateral geniculate nucleus. Fig. 2. Tupaia glis (tree shrew); left LGN; coronal section, Cresylviolet and Luxol-fast-blue stained, x 50; 1 — 6 = laminae 1 — 6 of dLGN. Fig. 3. Artibeus jamaicensis (fruit-vampire); left dLGN, coronal section, Cresylviolet stained, x 30. Fig. 4. Pteropus giganteus (flying fox); left LGN, coronal section, Cresylviolet stained, x 50; 1 —3 = laminae 1 — 3 of dLGN.

The dorsal lateral geniculate nucleus of mammals

Fig. 1 - 4

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Brauer, K., W . Schober and E . Winkelmann

Fig. 5. Galago crassicaudatus (giant galago); left dLGN, coronal scction, Cresylviolet and Luxol-fast-blue, x25; 1 — G — laminae 1 - 6 of the dLGN. Fig. G. Aotes trivirgatus (owl monkey), left dLGN, coronal section. Cresylviolet and Luxol-fast-blue; X 25; 1 — 4 = laminae 1 — 1 of the dLGN. Fig. 7. Macaca mulatta (rhesus monkey), left dLGN, coronal section, Cresylviolet and Luxol-fast-blue; x 25; 1 — 6 = laminae 1 — 6 of the dLGN Fig. 8. Pan troglodytes (chimpanzee), left dLGN coronal section; x 25; 1 —G = laminae 1 — G of the dLGN.

1970). Detailed investigations in the projection of the dLGN to the visual cortex of the rhesus monkey showed that the individual laminae project on different sublayers of layer I V ( H U B E L and W I E S E L 1 9 7 2 ) .

KAAS et al. (1972) suggest that all primates except the prosimians dispose of 4 laminae only, of which especially the parvocellular but in some cases also the magnocellular laminae form so-called "leaflets". The latter, however, do not correspond to true laminae. These authors take into consideration that also the prosimians — as well as other primates — ascend from ancestors with 4 laminae. The supply of the "central" laminae with ipsilateral retinal input points to a possible relationship to basal insectivores (Erinaceus). Lagomorpha

(fig. 9, 10)

The hare-like animals are mammals with well developed visual apparatus. For that reason the dLGN occupies a relatively greater volume as, for instance,

The dorsal lateral geniculate nucleus of mammals

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in most of the myomorphic rodents. ROSE (1935) did not indicate any lamination in the rabbits dLGN but discriminated four "subnuclei" (