Exp. Brain Res. 29, 233-248 (1977)
Experimental Brain Research
@ Springer-Verlag1977
The Pontine Projection to the Cerebellar Anterior Lobe. An Experimental Study in the Cat with Retrograde Transport of Horseradish Peroxidase P. Brodal and F. Walberg Anatomical Institute, University of Oslo, Karl Johansgt. 47, Oslo 1, Norway
Summary. By use of the retrograde axonal transport of horseradish peroxidase (HRP) the projection to the anterior lobe from the pontine nuclei was mapped in detail, In 18 cats 0.1-0.5 ~tl of a,50% suspension of Sigma VI or Serva H R P was injected in the anterior lobe under visual guidance or stereotaetically. The main findings are as follows: I. The projection to the vermis of the anterior lobe is bilateral with a contralateral preponderance (about 3/4). 2. The vermis of the anterior lobe receives afferents from a restricted, laterally located region in the caudal part of the pons. Dorsal parts of this region project anteriorly (lobules I-III), ventral parts posteriorly (lobulus
V). 3. The projection to the intermediate-lateral part is almost exclusively contralateral and is considerably heavier than the projection to the vermis. 4. The intermediate-lateral part receives afferents from two pontine regions. One is located laterally and coincides caudally with the region projecting to the vermis. The other is located medially and projects only sparsely to the vermis. 5. Within the projection from the lateral pontine region to the intermediate part there is a somatotopical pattern corresponding to that within the projection to the vermis. Within the medial pontine region the somatotopical pattern is less clear, but there is a tendency for cells projecting to lobules III-IV to be located more ventrolaterally than those sending their fibres to lobulus V. 6. The pontine regions projecting to the anterior lobe seem to coincide closely with those receiving fibres from the primary sensorimotor cortex, particularly the lateral part of the anterior sigmoid gyrus and medial part of the posterior sigmoid gyrus, as determined previously (P. Brodal, 1968a). Key words: P o n s - Cerebellum - Pontocerebellar projection - HRP method - Cat
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Introduction
Among the various cerebellar connections, the cortico-ponto-cerebeUar pathway is remarkable on account of its much higher number of fibres than the others (Tomasch, 1968). Its functional significance presumably increases during evolution in parallel with its quantitative increase. However, little is understood of the information brought to the cerebellum via the pontine nuclei (see Evarts and Thach, 1969; Allen and Tsukahara, 1974), and our knowledge of the anatomical organization of this large pathway is insufficient. The first link in the cerebro-ponto-cerebellar pathway, the corticopontine projection, has recently been studied in some detail using small cortical lesions and silver impregnation methods (P. Brodal, 1968a, b, 1971a, b, 1972a, b, c). It was shown that the pattern of organization is very complex, and that there is a higher degree of topographical order in this projection than previously found. In general, each small part of the cerebral cortex projects onto several discrete pontine regions, often arranged as longitudinal columns, while each of these receives fibres from several cortical spots. Some of the corticopontine projections show a somatotopical localization. Due to the utilization of axonal transport of macromolecules for mapping purposes, it has now been possible to study the pontocerebellar projection in corresponding detail (Hoddevik, 1975; Hoddevik et al., 1977). Using the retrograde axonal transport of horseradish peroxidase (HRP), Hoddevik (1975) found that the paramedian lobule receives afferents from several discrete parts of the pons, mainly arranged as longitudinal columns. The classical visual receptive area of the cerebellum (lobules VI-VIII) receives fibres from similarly arranged, but mainly differently located cell columns (Hoddevik et al., 1977). In the present study we have used the HRP method in order to map the pontocerebellar projection to the anterior lobe. Since there is no agreement in the literature as to which parts of the pons do project to the anterior lobe (see Brodal and Jansen, 1946; Voogd, 1964), it is hardly surprising that details concerning projections to subdivisions of the anterior lobe are lacking. Physiologically, however, the anterior lobe is one of the most completely studied parts of the cerebellum. Thus, the somatotopically organized relationship between the sensorimotor cortex and the anterior lobe is well documented (Adrian, 1943; Hampson, 1949; Snider and Eldred, 1951; Provini et al., 1968; Allen et al., 1974; Oka et al., 1976). Since the corticopontine projection from the sensorimotor cortex is somatotopically organized (P. Brodal, 1968a, b) there is strong reason to believe that the somatotopically organized mossy fibre potentials evoked in the anterior lobe after cortical stimulation are relayed, at least mainly, in the pontine nuclei (Allen and Tsukahara, 1974). The main aims of the present study are, therefore, to decide which parts of the pontine nuclei project to the anterior lobe, and to see whether there is a sumatotopical localization within this projection. If such a localization is present, it will be of interest to compare its pattern with that of the corticopontine projection determined previously. Since the anterior lobe exhibits a longitudinal organization in addition to the transverse (Voogd, 1964; Korneliussen, 1968; Oscarsson, 1973; Armstrong et al., 1974; Brodal and
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W a l b e r g , 1977), we tried to decide w h e t h e r this principle is e v i d e n t also in the pontocerebellar projection.
Material and Methods Altogether 18 young and adult cats have used in the present study. Most of them belong to a larger material prepared for the study of olivocerebellar connections (Brodal and Walberg, 1977). Under Nembutal anesthesia the HRP solution was injected slowly (about 30 min) with a Hamilton syringe. Usually from 0.1-0.5 ~tl of a 50% buffered solution of Sigma VI or Serva HRP was injected. The animals survived from one to 3 days, and were then perfused through the heart under deep Nembutal anesthesia with a buffered mixture of 0.4% paraformaldehyde and 1.25% glutaraldehyde. After the perfusion the brain and brain stem were dissected out, divided into appropriate blocks and stored overnight in the same fixative at 4 ~ C. Then the blocks were stored in a phosphate buffer solution containing 30% sucrose for another 24 hrs. Sagittal sections of the cerebellum and transverse sections of the pons were cut at 50 ~tm (in a few cases the pons was cut at 20 ~m) on a freezing microtome. The sections were collected into groups of five consecutive ones, and two from each group were treated according to Graham and Karnovsky (1966). One series was left unstained, the other weakly stained with cresyl violet. From one or both of the series the sections of the pons were drawn using an electronic pantograph, and the labeled cells (as identified with bright field and/or dark field microscopy) were plotted as dots. In some cases the plotted cells were counted, in order to determine the proportion of cells projecting contralaterally and ipsilaterally. The diagram of the pontine nuclei in Figure 2 shows the topography of the pons in a particular cat (B.St.L. 700) as seen in equidistant sections throughout its rostrocaudal extent. The findings made in the other cases were transferred to corresponding levels of this diagram (Figs. 3-6). The subdivision of the pontine grey into nuclei is that used by Brodal and Jansen (1946). This division is used here only to facilitate the description.
Results T h e e x t e n t of the s t a i n i n g following i n j e c t i o n s of H R P in the a n t e r i o r lobe varies c o n s i d e r a b l y from case to case. I n the p r e s e n t m a t e r i a l t h e r e was always s t a i n i n g of the m o l e c u l a r a n d g r a n u l a r layers a n d o f t e n also of the a d j a c e n t white m a t t e r . I n the diagrams the staining of the c e r e b e l l a r cortex as it a p p e a r s o n the surface is i n d i c a t e d w i t h o u t d i f f e r e n t i a t i n g b e t w e e n i n v o l v e m e n t of the m o l e c u l a r a n d g r a n u l a r layers. T h e i n t r a c e r e b e l l a r n u c l e i were u n s t a i n e d in all cases except o n e (B.St.L. 665). I n this case the fastigial n u c l e u s was slightly involved, b u t according to H o d d e v i k et al. (1977) this does n o t receive a p o n t i n e p r o j e c t i o n . O n e to t h r e e days after i n j e c t i o n of H R P in the a n t e r i o r lobe l a b e l e d cells c a n be f o u n d in the p o n t i n e n u c l e i (Fig. 1). T h e i r d i s t r i b u t i o n did n o t d e p e n d u p o n w h e t h e r Sigma V I or Serva H R P was used. N e i t h e r did the d i s t r i b u t i o n of l a b e l e d cells a p p e a r to d e p e n d u p o n w h e t h e r superficial or deep parts of the l o b u l i were stained, a l t h o u g h this q u e s t i o n was n o t studied systematically. T h e size a n d d e n s i t y of the H R P g r a n u l e s in the p o n t i n e cells m a y vary c o n s i d e r a b l y , u s u a l l y heavily l a b e l e d cells are f o u n d i n t e r m i n g l e d with m o r e lightly l a b e l e d ones. T h e n u m b e r of l a b e l e d cells seems to vary r o u g h l y i n p r o p o r t i o n to the e x t e n s i o n of the H R P s t a i n i n g in the c e r e b e l l a r cortex. A l t h o u g h m o s t l a b e l e d cells are f o u n d w i t h i n well-restricted areas (Fig. 1A), their d i s t r i b u t i o n does n o t respect the b o r d e r s b e t w e e n different p o n t i n e s u b n u c l e i o u t l i n e d o n a c y t o - a r c h i t e c t o n i c a l basis ( B r o d a l a n d J a n s e n , 1946).
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Fig. 1. Horseradish peroxidase labeled cells in the pontine nuclei. A A group of labeled cells in the lateral pontine nucleus 2 days following injection of HRP into lobules I and II of the vermis in cat B.St.L. 700 (cp Fig. 2). Dark field. X 112. B Labeled cells from the same group as in A as seen in the interference microscope. X 1200
In the following some representative cases showing the main features of the projection will be described in detail, while others with corresponding findings will be presented briefly. Due to the spread of staining along the cerebellar folia, the present material is not suited for a detailed analysis of possible differences in the projection between sagittal zones of cortex, while the more precise localization within the olivocerebellar projection permits an analysis of this kind for t~le projection from the oliva to the anterior lobe (Brodal and Watberg, 1977). Therefore, the cases will be separated into two main groups only: 1. those with injections mainly or exclusively restricted to the vermis, and 2. those with injections of the intermediate-lateral part of the anterior lobe.
Injections of the Vermis Lobules I and II. In cat B.St.L. 700 (injected stereotactically 0.5 ~1, survival time 2 days, Fig. 2) the staining involves the medial parts of these lobules on both sides. In the pontine nuclei labeled cells are found almost exclusively in a somewhat twisted band in the caudal fourth of the pons (sections 2 - 4 ) on both sides. It is located dorsally in the peduncular and lateral nuclei and extends from
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Fig. 2. The pontine projection to the lobules I and II of the anterior lobe vermis. The extent of staining in the cerebellar cortex is indicated in black in a schematized drawing of the unfolded cerebellum. Labeled cells are indicated as dots in drawings of transverse sections through the ports. All labeled cells observed in the represented sections are indicated in this and the following figures. Note that labeled cells are mainly confined to a transversally oriented band in the caudal part of the pons lateral to the peduncle. In order to facilitate comparisons, this series of drawings of transverse sections is used in the following figures also
Abbreviations Br.p.: Brachium pontis L.m.: Medial lemniscus N.d.: Nucleus dorsolateralis N.I.: Nucleus lateralis N.p.: Nucleus peduncularis N.pm~: Nucleus paramedianus N.v.: Nucleus ventralis Ped.: The continuation through the ports of the cerebral peduncle
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Fig. 3. Organization of the pontine projection to the anterior lobe vermis. Four representative cases with injections in lobules I-V. The extent of the staining is shown in black in the cerebellar diagrams at the top of the figure. Below the distribution of labeled cells in the pontine nuclei is shown as dots in drawings of transverse sections through the pons. Since after vermal injections the distribution is symmetrical on the two sides of the pons, one side only is represented. Only the caudal third of the pons is represented, since very few labeled cells were found in more rostral parts (cp. Fig. 2). Rostral lobules (hindlimb representation) of the vermis are supplied from more dorsal parts of the pons than caudal lobules (forelimb representation)
the lateral aspect of the peduncle to the c a u d a l m o s t part of the dorsolateral nucleus (Fig. 2). O n l y a few labeled cells are f o u n d medial to the peduncle. Closely c o r r e s p o n d i n g findings are m a d e in two o t h e r animals (cats B.St.L. 665, Fig. 3, and 715, not illustrated, injected 0.25 and 0.15 ~tl, survival time 2 days). I n cat B.St.L. 715 the staining is restricted to lobulus II, mainly on the right side. T h e r e are s o m e w h a t m o r e labeled cells in the left pontine nuclei than in the right. Lobule III. W e have no case with staining restricted to lobule III. H o w e v e r , the only difference in distribution of labeled cells w h e n injections involve lobule I I I in addition to lobulus II is that the transverse b a n d tends to extend s o m e w h a t m o r e ventrally than following injections of lobules I and II (cats B.St.L. 701, not illustrated, and 725, Fig. 3, injected stereotactically 0.25 and 0.20 ~1, survival time 1 and 2 days, respectively). Lobule IV. I n cat B.St.L. 709 (injected stereotactically 0.2 ~tl, killed after 2 days, Fig. 3) the staining is mainly restricted to the right vermal part of lobulus IV, but extends s o m e w h a t over the midline. L a b e l e d cells are distributed symmetrically in the left and rightpontine nuclei, but the m a j o r i t y (approx. two thirds) is f o u n d
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on the left side. The labeled cells are restricted to the caudal fourth of the pons, and are distributed lateral to the peduncle in a transversally oriented band in the dorsal part (Fig. 3). Corresponding observations are made in two other cases with injections of lobulus IV (cats B.St.L. 720 and 723, not illustrated, 0.15 and 0.2 ~tl, survival time 2 and 3 days, respectively). In cat B.St.L. 723 with practically unilateral staining, three fourths of the labeled cells are found contralateral to the injection. The band of labeled cells extends somewhat more ventrally in these cases with injections of lobule IV than in those affecting lobules I - I I I (Fig. 3). Lobule V. In five cases the staining is restricted to the vermal part of lobulus V. Even if different folia are involved in different cases, the distribution of labeled cells in the pontine nuclei is not significantly different. In cat C.Co. L. 192 (injected 0.15 ~!, survival time 1 day, Fig. 3) the staining extends more laterally on the left than on the right side. It is restricted to the vermis. In the rightpontine nuclei labeled cells are found in an area lateral to the peduncle in the caudal third of pons. A few labeled cells are also found in the caudalmost part of the dorsolateral nucleus (section 4) and medial to the peduncle. In the left pontine nuclei a smaller number of cells show a closely similar distribution. In three other cases, (cats B.St.L. 648,654 and 689, injected 0.25, 0.075 and 0.4 ~1, survival times 2, 1 and 2 days, respectively, not illustrated), mainly the medial zone of the vermis of lobule V is stained, but the distribution of labeled cells is the same as in C.Co.L. 192 (Fig. 3). In the fifth case (cat C.Co.L. 193, injected 0.2 ~tl, survival time 1 day, not illustrated), the staining involves in addition to the vermis the medialmost strip of the intermediate region, but the findings in the pons do not differ from those made when the staining is restricted to the vermis. In this case the staining extends only slightly across the midline, and three fourths of the labeled cells are found in the contralateral pontine nuclei.
Comments Following injections restricted to the vermis of the anterior lobe, labeled cells are concentrated in a well restricted area lateral to the peduncle in the caudal third to fourth of the pons. The main difference between cases with injections of the rostral (lobules I, II and III) and the caudal (lobule V) parts of the anterior lobe is that the band of labeled cells is found more ventrally in the latter (Fig. 3, see also Fig. 6). Judging from cases with almost unilateral staining of the cortex, the pontocerebellar projection to the anterior lobe vermis appears to be mainly contralateral (about 3/4), however with a significant (1/4) ipsilateral contribution.
The Intermediate-Lateral Part of the Anterior Lobe Due to the small size of the intermediate-lateral part in the rostral part of the anterior lobe, injections without involvement of the vermis or the deep cerebellar nuclei have only been obtained in lobules V and III-IV.
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Fig. 4. The pontine projection to the intermediate part of lobules V. The extent of staining in the lobulus V is indicated with black in the diagram of the cerebellum (the staining involves to a lesser degree also lobulus VI). Labeled cells are represented with black dots in drawings of transverse sections through the pontine nuclei. One group of cells is found medial to the peduncle, another lateral to it. Both have the shape of longitudinal columns. The labeled cells within the circle are due to the involvement of lobulus VI by the injection (cfr. text)
Lobule V. I n cat C. Co. L. 190 (0.2 ~1, survival t i m e 1 day, Fig. 4) t h e c a u d a l folia o f t h e right i n t e r m e d i a t e p a r t o f l o b u l e V a r e stained, b u t in a d d i t i o n t h e r e is s o m e slight staining o f t h e a d j o i n i n g folia o f l o b u l e VI. In t h e contralateral pontine nuclei n u m e r o u s l a b e l e d cells (up to 80 cells in o n e 40 ~m section) a r e d i s t r i b u t e d in two c o l u m n - s h a p e d a r e a s in t h e c a u d a l 2/3 o f the p o n s , o n e m e d i a l , t h e o t h e r l a t e r a l to t h e p e d u n c l e . T h e m e d i a l c o l u m n is f o u n d d o r s a l l y close to t h e p e d u n c l e . In a d d i t i o n to t h e s e two c o l u m n s , a m i n o r g r o u p of l a b e l e d cells is l o c a t e d close to t h e v e n t r a l a s p e c t o f t h e p e d u n c l e (circles w i t h b r o k e n line in Fig. 4). This g r o u p is n e v e r f o u n d in cases with i n j e c t i o n s r e s t r i c t e d to t h e a n t e r i o r lobe, a n d c o i n c i d e s with a r e g i o n f o u n d b y H o d d e v i k et al. ( 1 9 7 7 ) to p r o j e c t to l o b u l u s VI. I n t h e ipsilateral pontine nuclei o n l y a few
Pontocerebellar Projeetion to the Anterior Lobe
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Fig. 5. Organization of the pontine projection to the intermediate part of the vermis. No convincing differences are apparent between the distribution of pontine cells projecting to lobulus IV (hindlimb) and lobulus V (forelimb) judged from cases with unilateral injections (B.St.L. 623 and C.Co.L. 195). However, in C.Co.L. 198, with bilateral injections, a somatotopical pattern is evident. Dorsal parts of the lateral region project to lobules III-IV, while ventral parts supply lobulus V. Within the medial region there is a slight tendency for cells projecting to lubules III-IV to be located more ventrolaterally
labeled cells are f o u n d (about 1 % of the contralateral n u m b e r ) . In two o t h e r cases with injections of the intermediate-lateral part of the lobulus V (cats C.Col.L. 189, not illustrated, and 195, Fig. 5, 0.3 and 0.1 ~1, respectively, survival time 2 days) the distribution of labeled cells c o n f o r m s to the description given above, except that the v e n t r o m e d i a l c o l u m n is lacking. Lobule IV. I n cat B . S t . L . 623 (injected 0.5 ~1, survival time 3 days, Fig. 5) the staining is restricted to the left intermediate-lateral part o f lobulus IV. I n the contralateral pontine nuclei a considerable n u m b e r of labeled cells (up to 110 cells in one 50 [~m section) are distributed mainly into two regions in the caudal
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2/3 of the ports, extending caudo-rostrally, medial and lateral to the peduncle, respectively. In the ipsiIateral pontine nuclei only a few labeled cells are found. Corresponding observations are made in another case with a closely similar injection (cat B.St.L. 710, injected stereotactically 0.2 ~tl, survival time 3 days, not illustrated). However, the labeled cells are on the whole located somewhat more dorsally than in cat B.St.L. 623.
Bilateral Injection of Lobules I l l - I V and V In order to compare in more detail the projection to the intermediate part of the different lobules, bilateral injections were made in one case. In cat C.Co.L. 198, 0.1 ~tl H R P was injected in lobule V and 0.1 ~1 stereotactically in lobules I I I - I V (survival time 2 days, Fig. 5). On the left side the staining is found in the intermediate part of lobules III and IV. The intracerebellar nuclei are not involved. In addition, there is a minute area of staining in the intermediate part of lobule V where the cannula has entered. On the right side the staining is confined to the intermediate part of lobule V. In thepontine nuclei labeled cells are distributed on both sides in a pattern in agreement with the findings in cases with corresponding unilateral injections (Figs. 4 and 5). However, certain differences between the two sides are obvious. Thus, in most sections the group of labeled cells lateral to the peduncle is located more ventrally on the left side than on the right. The group of labeled cells medial to the peduncle tends to be situated somewhat more ventrolaterally on the right than on the left side, but the difference is less clear than in the lateral cell group.
Comments The pontine projection to the lateral-intermediate part of the anterior lobe differs in some respect from that to the vermis. In the first place, the projection to the intermediate-lateral part is almost exclusively contralateral. Furthermore, the area lateral to the peduncle extends more rostrally when injections are placed in the intermediate part than when they are limited to the vermis (cp. Figs. 2 and 4). Finally, only the intermediate-lateral part receives a substantial projection from a region medial to the peduncle (Figs. 4 and 5). The distribution of labeled cells is very similar in cases with injections of the intermediate-lateral part of lobules IV and V, labeled cells being found within two well restricted areas, medial and lateral to the peduncle. Although certain differences exist between the results of injections in lobules IV and V, they do not exceed what may be due to individual differences, as appears from comparisons between cases with practically identical injections. However, following injection of lobules I I I - I V on one side and lobule V on the other within the same animal (cat C.Co.L. 198, Fig. 5), thus eliminating individual variations, it becomes evident that within the lateral region dorsal parts project to anterior lobuli, ventral parts to posterior ones, according to the same
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indicated with open (lobules I-IV) and filled (lobulus V) circles. The projection is bilateral, and cells projecting to rostral lobuli (hindlimb representation) are located more dorsally than those projecting to caudal lobuli (forelimb representation). The intermediate-lateral part receives fibres from more extensive, nearly ,8 exclusively controlateral pontine cell groups (hatching). One group of cells is found medial, another lateral to the peduncle. Caudally the lateral area coincides with that projecting to corresponding parts of the vermis. A somtoto
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concerning the anterior lobe vermis is present within the proiection from the lateral pontine region to the intermediate part. Within the medial pontine region the somatotopical pattern seems to be reversed, but is less clear than in the lateral region
CAUDAL
somatotopical principle as found within the projection to the vermis. The reverse pattern seems to be present within the medial region, but it is much less clear (Figs. 3, 5 and 6). Discussion By use of the retrograde axonal transport of HRP we have been able to map the pontocerebellar projection to the anterior lobe in more detail than has
.
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been possible with anterograde (Voogd, 1964) and retrograde degeneration techniques (Brodal and Jansen, 1946, and others). The advantages and limitations of the H R P method with regard to the study of cerebellar afferents have been documented and discussed (Walberg et al., 1976), and need not be repeated here. Suffice it to say that the H R P method is better suited for topographical than for quantitative purposes. Origin and Termination of Pontocerebellar Fibres to the.Anterior Lobe
Following injections of the anterior lobe, labeled cells in the pontine nuclei are confined to certain restricted parts, arranged as transverse bands or longitudinal columns. This arrangement resembles the one found previously with regard to the pontine projection to the paramedian lobule (Hoddevik, 1975) and the cerebellar visual areas (lobules VI-VIII, Hoddevik et al., 1977). It is also similar to the organization of terminal areas of corticopontine fibres from the frontal part of the cerebral cortex (P. Brodal, 1968a, b, 1971a, b). More specifically, the majority of pontine cells projecting to the anterior lobe are found lateral and medial close to the peduncle in the caudal part of the pons, in agreement with the findings of Voogd (1964) based upon anterograde degeneration technique. Furthermore, it is in agreement with the conclusion of certain of the older anatomists using normal or pathological human material (see Voogd, 1964, for references to the older literature). Brodal and Jansen (1946) using the modified Gudden method (Brodal, 1940) had no lesions restricted to the anterior lobe. However, in cases with partial involvement of the anterior lobe, retrograde changes or cell loss were found (among other places) in the areas found by us to project to the anterior lobe. Verrnis. We find that the anterior lobe vermis receives afferents almost exclusively from a well-restricted area in the caudal and lateral part of the pontine nuclei (Figs. 3 and 6). Although longitudinal zones of the anterior lobe vermis differ with regard to development (Korneliussen, 1968) and connections (Voogd, 1964; Voogd et al., 1969; Oscarsson, 1973; Armstrong et al., 1974; Brodal and Walberg, 1977), no such differences could be discerned in the present material. This can probably not be fully explained by the fact that our injections were not restricted to particular zones. Thus, using the same experimental material, Brodal and Walberg (1977) found different origins of the projections from the inferior olive to medial and lateral parts of the vermis. It is of interest in this connection that while Voogd et al. (1969) found spinocerebellar fibres to terminate in clear sagittal strips in the anterior lobe, this was not the case for pontocerebellar fibres. In harmony with this, Provini et al. (1968) with electrical stimulation of the sensorimotor cortex found evidence for a longitudinal subdivision of the anterior lobe vermis concerning climbing fibre responses (mediated at least mainly via the inferior olive; see especially Batini et al., 1976) but not concerning mossy fibre responses. A somatotopical pattern is present within the pontine projection to the anterior lobe vermis, since lobule V (representing the forelimb) receives fibres from more ventral parts of the pontine area than do lobules I-IV (hindlimb part). A pertinent question is then whether these pontine regions receive
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projections from cerebrocortical forelimb and hindlimb areas, respectively. When the present results are compared with those obtained by mapping the distribution of anterograde degeneration i n the pontine nuclei following small lesions of the sensorimotor cortex (P. Brodal, 1968a), it appears that most of the pontine area connected with the vermal part of lobulus V receives afferents from the motor forelimb area (lateral part of the anterior sigmoid gyrus), and to a lesser degree from the sensory forelimb region (lateral part of the posterior sigmoid gyrus). Parts of the pontine region projecting onto the hindlimb part of anterior lobe vermis (lobules I-IV) seem to receive afferents from the medial part of the posterior sigmoid gyrus, usually termed the sensory hindlimb area. The medial part of the anterior sigmoid gyrus, previously assumed to be the motor hindlimb area (Woolsey, 1958) does not appear to send fibres to those parts of the pons which project to the vermis of lobules I-IV. In agreement with this, stimulation of this part of the cortex evokes only slight mossy fibre potentials in lobules III-IV (Allen et al., 1974; Oka et al., 1976). It is interesting that recent data (see Wiesendanger et al., 1974) strongly suggest that the medial part of the anterior sigmoid gyrus represents the supplementary motor area in the cat, while the exact localization of the motor hindlimb area is still not clear. It may be speculated whether in the cat separation between sensory and motor subdivisions is less sharp in the hindlimb than in the forelimb region, so that the medial part of the posterior sigmoid gyrus is to be considered as a motor as well as a sensory hindlimb area. It should be noted that all parts of the pontine regions which project to the anterior lobe vermis do not seem to receive afferents from the primary sensorimotor area. These regions may be influenced from the second somatosensory area (P. Brodal, 1968b), the auditory cortex (P. Brodal, 1972c), and the superior and inferior colliculi (Kawamura and Brodal, 1973). However, in order to clarify these problems, it will be necessary to map in one experimental animal the terminal areas of the corticopontine fibres and the localization of the pontine cells projecting to the anterior lobe vermis. Such studies are now in progress in our laboratory with the combined use of the Fink and Heimer silver impregnation method and the HRP method. That this approach can be fruitful has recently been shown by Blomqvist and Westman (1975) in the dorsal column nuclei. Intermediate-Lateral Part. There is a main difference between the pontine projections to the vermis and the intermediate-lateral part of the anterior lobe: the latter receives a much stronger projection, and from more extensive parts of the pontine nuclei than does the former (Fig. 6). Thus, the intermediate-lateral part receives afferents from a longitudinal cell column medial to the peduncle, as well as from one lateral to it. It appears that the caudal part of the lateral cell column coincides with the one projecting to the vermis. Both the medial and the lateral columns projecting to the intermediate-lateral part receive afferents from the motor cortex, at least from its forelimb area. This is evident from experiments combining silver impregnation of degenerating fibres and retrograde labeling with HRP (P. Brodal, unpublished observations). By the use of bilateral HRP injections in the anterior lobe it has been possible to demonstrate a somatotopical pattern within the pontine projection to
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the intermediate part, at least within the lateral pontine region. The pattern is the same as that found within the proiection to the vermis (Fig. 6). Within the corticopontine projection likewise certain topographical differences became evident only with the use of bilateral cortical lesions (P. Brodal, 1968a, b, 1972a, b, c). While anatomically no clear difference in precision seems to exist between the pontocerebellar projection to the vermis and the intermediate part, physiologically the somatotopical organization of mossy fibre responses is sharper in the intermediate part than in the vermis (Provini et al., 1968). However, it should be recalled that mossy fibre potentials to the anterior lobe may be relayed in brain stem nuclei other than the pontine (nucleus reticularis lateralis, nucleus reticularis paramedianus, the perihypoglossal nuclei and the nucleus reticularis tegmenti pontis, for a review, see Brodal, 1972). It may be that some of these nuclei project more diffusely onto the vermis than to the intermediate part, and therefore blurr the somatotopical pattern of evoked potentials after cerebrocortical stimulation. It is of interest to consider our findings in relation to what is known of pontocerebellar projections to other parts of the cerebellum, when studied by the retrograde transport of HRP. The vermis of the anterior lobe receives afferents from largely other parts of the pons than does the vermis of lobules VI-VIII (Hoddevik et al., 1977). These lobules receive fibres from several longitudinal, sharply delimited cell columns in various parts of the pons, while the cells projecting to the anterior lobe vermis are distributed more transversally in the caudal part of the pons. The paramedian lobule likewise receives pontine afferents from several longitudinal cell columns (Hoddevik, 1975). Some of these appear to coincide closely with terminal areas of corticopontine fibres from the first and second somatosensory cortices (P. Brodal, 1968a, b), while there is less overlap with terminal areas of fibres from the motor cortex. As discussed above, the pontine regions projecting to the anterior lobe probably receive more cortical fibres from the motor than from the sensory cortex. It may therefore be suggested that the anterior lobe is chiefly (but not exclusively, see Allen et al., 1974; Oka et al., 1976) influenced from the motor cortex, while the paramedian lobule is functionally more closely related to the somatosensory cortical areas. The pattern of organization within the pontocerebellar projection appearing from the above mentioned studies (Hoddevik, 1975; Hoddevik et al., 1977) and from the present study, is indeed complex. A large number of often very discrete cell groups project onto particular parts of the cerebellum. A restricted part of the cerebellar cortex may receive fibres from several pontine cell groups, and one pontine cell group may project to different cerebellar parts. Thus, there is an extensive convergence as well as divergence within the pontocerebellar projection. The complexity in the cortico-ponto-cerebellar pathway is increased by the fact that each small part of the cerebral cortex as a rule projects onto several discrete parts of the pontine nuclei, while fibres from different cortical areas may converge onto one pontine cell group. This pattern represents a warning against regarding the cortico-ponto-cerebellar projection as being rather
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diffusely organized. It may well be that with the use of the present rather crude anatomical techniques, destroying thousands of functionally different cortical cells, or injecting HRP around thousands of possibly dissimilar pontocerebellar axon terminals, we can clarify only the most crude pattern of organization. When studied in detail, individual pontine cells are found to be functionally very specific with regard to their input (Baker et al., 1976), a fact which certainly reflects that their afferent fibres terminate very specifically. Many questions remain to be solved. For instance, when different pontine cell groups project to one cerebellar area, do they converge on identical Purkinje cells (via granule cells), or is there a very discrete mosaic pattern (see Eccles et al., 1972) not shown by our present method? Furthermore, when one pontine cell group projects to several different cerebellar areas, are the axons bifurcating, or are different, but closely adjoining cells projecting differently? The latter question may be solved by using different tracers for retrograde axonal transport as done, for example, by Steward and Scoville (1976) in the hippocampus.
References Allen, G.I., Tsukahara, N.: Cerebrocerebellar communication systems. Physiol. Rev. 54, 957-1006 (1974) Allen, G.I., Azzena, G.B., Ohno, T.: Somatotopically organized inputs from fore- and hindlimb areas of sensorimotor cortex to cerebellar Purkyn6 cells. Exp. Brain Res. 20, 255-272 (1974) Adrian, E. D.: Afferent areas in the cerebellum connected with the limbs. Brain 66, 289-315 (1943) Armstrong, D.M., Harvey, R.J., Schild, R.F.: Topographical localization in the olivocerebellar projection: An electrophysiologicval study in the cat. J. comp. Neurol. 154, 287-302 (1974) Baker, J., Gibson, A., Glickstein, M., Stein, J.: Visual cells in the pontine nuclei of the cat. J. Physiol. (Lond.) 255,415-434 (1976) Batini, C., Corvisier, J., Destombes, J., Gioauni, H., Everett, J.: The climbing fibers of the cerebellar cortex, their origin and pathways in the cat. Exp. Brain Res. 26, 407-422 (1976) Blomqvist, A., Westman, J.: Combined HRP and Fink-Heimer staining applied on the gracile nucleus in the cat. Brain Res. 99, 339-342 (1975) Brodal, A.: Modification of Gudden method for study of cerebral localization. Arch. Neurol. Psychiat. (Chic.) 43, 46-58 (1940) Brodal, A.: Cerebrocerebellar pathways. Anatomical data and some functional implications. Acta Neurol. Scand., Suppl. 51, 153-196 (1972) Brodal, A., Jansen, J.: The pontocerebellar projection in the rabbit and cat. Experimental investigations. J. comp. Neurol. 84, 31-118 (1946) Brodal, A., Walberg, F.: The olivocerebellar projection in the cat studied with the method of retrograde axonal transport with horseradish peroxidase. IV. The projection to the anterior lobe. J. comp. Neurol. 172, 85-108 (1977) Brodal, P.: The corticopontine projection in the cat. I. Demonstration of a somatotopically organized projection from the primary sensorimotor cortex. Exp. Brain Res. 5, 210-234 (1968a) Brodal, P.: The corticopontine projection in the cat. Demonstration of a somatotopically organized projection from the second somatosensory cortex. Arch. Ital. Biol. 106, 310-332. (1968b) Brodal, P.: The corticopontine projection in the cat. I. The projection from the proreate gyrus. J. comp. Neurol. 142, 127-140 (1971a) Brodal, P.: The corticopontine projection in the cat. II. The projection from the orbital gyrus. J. comp. Neurol. 142, 141-152 (1971b) Brodal, P.: The corticopontine projection from the visual cortex in the cat. I. The total projection and the projection from areas 17. Brain Res. 39, 297-317 (1972a)
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Brodal, P.: The corticopontine projection from the visual cortex in the cat. II. The projection from areas 18 and 19. Brain Res. 39, 319-335 (1972b) Brodal, P.: The corticopontine projection in the cat. The projection from the auditory cortex. Arch. Itai. Biol. 10, 119-144 (1972c) Eccles, J.C., Sabah, N.H., Schmidt, R.F., T~ibo~ikov~, H.: Mode of operation of the cerebellum in the dynamic loop control of movement. Brain Res. 40, 73-80 (1972) Evarts, E.V., Thach, W.T.: Motor mechanisms of the CNS: cerebrocerebellar interrelations. Ann. Rev. Physiol. 31, 451--498 (1969) Graham, R.C., Jr., Kamovsky, M.J.: The early stages of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney: ultrastructural cytochemistry.by a new technique. J. Histochem. Cytochem. 14, 291-302 (1966) Hampson, J.L.: Relationship between cerebral and cerebellar cortices in cat. J. Neurophysiol. 12, 37-50 (1949) Hoddevik, G.H.: The pontocerebellar projection onto the paramedian lobule in the cat: An experimental study with the use of horseradish peroxidase as a tracer. Brain Res. 95, 291-307 (1975) Hoddevik, G.H., Brodal, A., Kawamura, K., Hashikawa, T.: The pontine projection to the cerebellar vermal visual area studied by means of the retrograde axonal transport of horseradish per0xidase. Brain Res. 123, 209-227 (1977) Kawamura, K., Brodal, A.: The tectopontine projection in the cat: An experimental anatomical study with comments on pathways for teleceptive impulses to the cerebellum. J. comp. Neurol. 149, 371-390 (1973) Korneliussen, H.K.: Comments on the cerebellum and its division. Brain Res. 8, 229-236 (1968) Oka, H., Yasuda, T., Jinnai, K., Yoneda, Y.: Reexamination of cerebellar responses to stimulation of sensorimotor areas of the cerebral cortex. Brain Res. 118, 312-319 (1976) Oscarsson, O.: Functional organization of spinocerebellar paths. In: Handbook of sensory physiology, Vol. 2 (ed. A. Iggo), p. 339-380. Somatosensory system. Berlin-Heidelberg-New York: Springer 1973 Provini, L., Redman, S., Strata, P.: Mossy and climbing fibre organization on the anterior lobe of the cerebellum activated by forelimb and hindlimb areas of the sensorimotor cortex. Exp. Brain Res. 6, 216-233 (1968) Snider, R.S., Eldred, E.: Electro-anatomical studies on cerebrocerebellar connections in the cat. J. comp. Neurol. 95, 1-16 (1951) Steward, O., Scoville, S.A.: Retrograde labeling of central nervous pathways with tritiated or Evans blue-labeled bovine serum albumine. Neuroscience Letters 3, 191-196 (1976) Tomasch, J.: The overall information capacity of the major afferent and efferent cerebellar cell and fiber systems. Confin. neurol. (Basel) 30, 359-367 (1968) Voogd, J.: The cerebellum of the cat. Structure and fibre connections, pp. 215. Leiden: Van Gorcum & Comp. 1964 Voogd, J., Broere, G., van Rossum, J.: The medio-lateral distribution of the spinocerebellar projection in the anterior lobe and the simple lobule in the cat and a comparison with some other afferent fibre systems. Psychiat. Neurol. Neurochir. (Amst.) 72, 137-151 (1969) Walberg, F., Brodal, A., Hoddevik, G.H.: A note on the method of retrograde transport of horseradish peroxidase as a tool in studies of afferent cerebellar connections, particularly those from the inferior olive; with comments on the orthograde transport in Purkinje cell axons. Exp. Brain Res. 24, 383-401 (1976) Wiesendanger, M., Seguin, J.J., Kiinzte, H.: The supplementary motor area - a control system for posture? Advanc. Behav. Biol. 7, 331-346 (1974) Woolsey, C.N.: Organization of somatic sensory and motor areas of the cerebral cortex. In: Biological and biochemical bases of behavior (eds. Harlow and Woolsey), pp. 63-81. Madion: University of Wisconsin Press 1958
Received February 2, 1977
Experimental Brain Research
@ Springer-Verlag1977
The Pontine Projection to the Cerebellar Anterior Lobe. An Experimental Study in the Cat with Retrograde Transport of Horseradish Peroxidase P. Brodal and F. Walberg Anatomical Institute, University of Oslo, Karl Johansgt. 47, Oslo 1, Norway
Summary. By use of the retrograde axonal transport of horseradish peroxidase (HRP) the projection to the anterior lobe from the pontine nuclei was mapped in detail, In 18 cats 0.1-0.5 ~tl of a,50% suspension of Sigma VI or Serva H R P was injected in the anterior lobe under visual guidance or stereotaetically. The main findings are as follows: I. The projection to the vermis of the anterior lobe is bilateral with a contralateral preponderance (about 3/4). 2. The vermis of the anterior lobe receives afferents from a restricted, laterally located region in the caudal part of the pons. Dorsal parts of this region project anteriorly (lobules I-III), ventral parts posteriorly (lobulus
V). 3. The projection to the intermediate-lateral part is almost exclusively contralateral and is considerably heavier than the projection to the vermis. 4. The intermediate-lateral part receives afferents from two pontine regions. One is located laterally and coincides caudally with the region projecting to the vermis. The other is located medially and projects only sparsely to the vermis. 5. Within the projection from the lateral pontine region to the intermediate part there is a somatotopical pattern corresponding to that within the projection to the vermis. Within the medial pontine region the somatotopical pattern is less clear, but there is a tendency for cells projecting to lobules III-IV to be located more ventrolaterally than those sending their fibres to lobulus V. 6. The pontine regions projecting to the anterior lobe seem to coincide closely with those receiving fibres from the primary sensorimotor cortex, particularly the lateral part of the anterior sigmoid gyrus and medial part of the posterior sigmoid gyrus, as determined previously (P. Brodal, 1968a). Key words: P o n s - Cerebellum - Pontocerebellar projection - HRP method - Cat
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Introduction
Among the various cerebellar connections, the cortico-ponto-cerebeUar pathway is remarkable on account of its much higher number of fibres than the others (Tomasch, 1968). Its functional significance presumably increases during evolution in parallel with its quantitative increase. However, little is understood of the information brought to the cerebellum via the pontine nuclei (see Evarts and Thach, 1969; Allen and Tsukahara, 1974), and our knowledge of the anatomical organization of this large pathway is insufficient. The first link in the cerebro-ponto-cerebellar pathway, the corticopontine projection, has recently been studied in some detail using small cortical lesions and silver impregnation methods (P. Brodal, 1968a, b, 1971a, b, 1972a, b, c). It was shown that the pattern of organization is very complex, and that there is a higher degree of topographical order in this projection than previously found. In general, each small part of the cerebral cortex projects onto several discrete pontine regions, often arranged as longitudinal columns, while each of these receives fibres from several cortical spots. Some of the corticopontine projections show a somatotopical localization. Due to the utilization of axonal transport of macromolecules for mapping purposes, it has now been possible to study the pontocerebellar projection in corresponding detail (Hoddevik, 1975; Hoddevik et al., 1977). Using the retrograde axonal transport of horseradish peroxidase (HRP), Hoddevik (1975) found that the paramedian lobule receives afferents from several discrete parts of the pons, mainly arranged as longitudinal columns. The classical visual receptive area of the cerebellum (lobules VI-VIII) receives fibres from similarly arranged, but mainly differently located cell columns (Hoddevik et al., 1977). In the present study we have used the HRP method in order to map the pontocerebellar projection to the anterior lobe. Since there is no agreement in the literature as to which parts of the pons do project to the anterior lobe (see Brodal and Jansen, 1946; Voogd, 1964), it is hardly surprising that details concerning projections to subdivisions of the anterior lobe are lacking. Physiologically, however, the anterior lobe is one of the most completely studied parts of the cerebellum. Thus, the somatotopically organized relationship between the sensorimotor cortex and the anterior lobe is well documented (Adrian, 1943; Hampson, 1949; Snider and Eldred, 1951; Provini et al., 1968; Allen et al., 1974; Oka et al., 1976). Since the corticopontine projection from the sensorimotor cortex is somatotopically organized (P. Brodal, 1968a, b) there is strong reason to believe that the somatotopically organized mossy fibre potentials evoked in the anterior lobe after cortical stimulation are relayed, at least mainly, in the pontine nuclei (Allen and Tsukahara, 1974). The main aims of the present study are, therefore, to decide which parts of the pontine nuclei project to the anterior lobe, and to see whether there is a sumatotopical localization within this projection. If such a localization is present, it will be of interest to compare its pattern with that of the corticopontine projection determined previously. Since the anterior lobe exhibits a longitudinal organization in addition to the transverse (Voogd, 1964; Korneliussen, 1968; Oscarsson, 1973; Armstrong et al., 1974; Brodal and
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W a l b e r g , 1977), we tried to decide w h e t h e r this principle is e v i d e n t also in the pontocerebellar projection.
Material and Methods Altogether 18 young and adult cats have used in the present study. Most of them belong to a larger material prepared for the study of olivocerebellar connections (Brodal and Walberg, 1977). Under Nembutal anesthesia the HRP solution was injected slowly (about 30 min) with a Hamilton syringe. Usually from 0.1-0.5 ~tl of a 50% buffered solution of Sigma VI or Serva HRP was injected. The animals survived from one to 3 days, and were then perfused through the heart under deep Nembutal anesthesia with a buffered mixture of 0.4% paraformaldehyde and 1.25% glutaraldehyde. After the perfusion the brain and brain stem were dissected out, divided into appropriate blocks and stored overnight in the same fixative at 4 ~ C. Then the blocks were stored in a phosphate buffer solution containing 30% sucrose for another 24 hrs. Sagittal sections of the cerebellum and transverse sections of the pons were cut at 50 ~tm (in a few cases the pons was cut at 20 ~m) on a freezing microtome. The sections were collected into groups of five consecutive ones, and two from each group were treated according to Graham and Karnovsky (1966). One series was left unstained, the other weakly stained with cresyl violet. From one or both of the series the sections of the pons were drawn using an electronic pantograph, and the labeled cells (as identified with bright field and/or dark field microscopy) were plotted as dots. In some cases the plotted cells were counted, in order to determine the proportion of cells projecting contralaterally and ipsilaterally. The diagram of the pontine nuclei in Figure 2 shows the topography of the pons in a particular cat (B.St.L. 700) as seen in equidistant sections throughout its rostrocaudal extent. The findings made in the other cases were transferred to corresponding levels of this diagram (Figs. 3-6). The subdivision of the pontine grey into nuclei is that used by Brodal and Jansen (1946). This division is used here only to facilitate the description.
Results T h e e x t e n t of the s t a i n i n g following i n j e c t i o n s of H R P in the a n t e r i o r lobe varies c o n s i d e r a b l y from case to case. I n the p r e s e n t m a t e r i a l t h e r e was always s t a i n i n g of the m o l e c u l a r a n d g r a n u l a r layers a n d o f t e n also of the a d j a c e n t white m a t t e r . I n the diagrams the staining of the c e r e b e l l a r cortex as it a p p e a r s o n the surface is i n d i c a t e d w i t h o u t d i f f e r e n t i a t i n g b e t w e e n i n v o l v e m e n t of the m o l e c u l a r a n d g r a n u l a r layers. T h e i n t r a c e r e b e l l a r n u c l e i were u n s t a i n e d in all cases except o n e (B.St.L. 665). I n this case the fastigial n u c l e u s was slightly involved, b u t according to H o d d e v i k et al. (1977) this does n o t receive a p o n t i n e p r o j e c t i o n . O n e to t h r e e days after i n j e c t i o n of H R P in the a n t e r i o r lobe l a b e l e d cells c a n be f o u n d in the p o n t i n e n u c l e i (Fig. 1). T h e i r d i s t r i b u t i o n did n o t d e p e n d u p o n w h e t h e r Sigma V I or Serva H R P was used. N e i t h e r did the d i s t r i b u t i o n of l a b e l e d cells a p p e a r to d e p e n d u p o n w h e t h e r superficial or deep parts of the l o b u l i were stained, a l t h o u g h this q u e s t i o n was n o t studied systematically. T h e size a n d d e n s i t y of the H R P g r a n u l e s in the p o n t i n e cells m a y vary c o n s i d e r a b l y , u s u a l l y heavily l a b e l e d cells are f o u n d i n t e r m i n g l e d with m o r e lightly l a b e l e d ones. T h e n u m b e r of l a b e l e d cells seems to vary r o u g h l y i n p r o p o r t i o n to the e x t e n s i o n of the H R P s t a i n i n g in the c e r e b e l l a r cortex. A l t h o u g h m o s t l a b e l e d cells are f o u n d w i t h i n well-restricted areas (Fig. 1A), their d i s t r i b u t i o n does n o t respect the b o r d e r s b e t w e e n different p o n t i n e s u b n u c l e i o u t l i n e d o n a c y t o - a r c h i t e c t o n i c a l basis ( B r o d a l a n d J a n s e n , 1946).
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Fig. 1. Horseradish peroxidase labeled cells in the pontine nuclei. A A group of labeled cells in the lateral pontine nucleus 2 days following injection of HRP into lobules I and II of the vermis in cat B.St.L. 700 (cp Fig. 2). Dark field. X 112. B Labeled cells from the same group as in A as seen in the interference microscope. X 1200
In the following some representative cases showing the main features of the projection will be described in detail, while others with corresponding findings will be presented briefly. Due to the spread of staining along the cerebellar folia, the present material is not suited for a detailed analysis of possible differences in the projection between sagittal zones of cortex, while the more precise localization within the olivocerebellar projection permits an analysis of this kind for t~le projection from the oliva to the anterior lobe (Brodal and Watberg, 1977). Therefore, the cases will be separated into two main groups only: 1. those with injections mainly or exclusively restricted to the vermis, and 2. those with injections of the intermediate-lateral part of the anterior lobe.
Injections of the Vermis Lobules I and II. In cat B.St.L. 700 (injected stereotactically 0.5 ~1, survival time 2 days, Fig. 2) the staining involves the medial parts of these lobules on both sides. In the pontine nuclei labeled cells are found almost exclusively in a somewhat twisted band in the caudal fourth of the pons (sections 2 - 4 ) on both sides. It is located dorsally in the peduncular and lateral nuclei and extends from
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BSt.L700 IGHT
CAUDAL
Fig. 2. The pontine projection to the lobules I and II of the anterior lobe vermis. The extent of staining in the cerebellar cortex is indicated in black in a schematized drawing of the unfolded cerebellum. Labeled cells are indicated as dots in drawings of transverse sections through the ports. All labeled cells observed in the represented sections are indicated in this and the following figures. Note that labeled cells are mainly confined to a transversally oriented band in the caudal part of the pons lateral to the peduncle. In order to facilitate comparisons, this series of drawings of transverse sections is used in the following figures also
Abbreviations Br.p.: Brachium pontis L.m.: Medial lemniscus N.d.: Nucleus dorsolateralis N.I.: Nucleus lateralis N.p.: Nucleus peduncularis N.pm~: Nucleus paramedianus N.v.: Nucleus ventralis Ped.: The continuation through the ports of the cerebral peduncle
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B.St.L.725
\
B.St.L. 7 0 9
R0STRA"~ ,
C.Co.L.192
a
CAUDAL
Fig. 3. Organization of the pontine projection to the anterior lobe vermis. Four representative cases with injections in lobules I-V. The extent of the staining is shown in black in the cerebellar diagrams at the top of the figure. Below the distribution of labeled cells in the pontine nuclei is shown as dots in drawings of transverse sections through the pons. Since after vermal injections the distribution is symmetrical on the two sides of the pons, one side only is represented. Only the caudal third of the pons is represented, since very few labeled cells were found in more rostral parts (cp. Fig. 2). Rostral lobules (hindlimb representation) of the vermis are supplied from more dorsal parts of the pons than caudal lobules (forelimb representation)
the lateral aspect of the peduncle to the c a u d a l m o s t part of the dorsolateral nucleus (Fig. 2). O n l y a few labeled cells are f o u n d medial to the peduncle. Closely c o r r e s p o n d i n g findings are m a d e in two o t h e r animals (cats B.St.L. 665, Fig. 3, and 715, not illustrated, injected 0.25 and 0.15 ~tl, survival time 2 days). I n cat B.St.L. 715 the staining is restricted to lobulus II, mainly on the right side. T h e r e are s o m e w h a t m o r e labeled cells in the left pontine nuclei than in the right. Lobule III. W e have no case with staining restricted to lobule III. H o w e v e r , the only difference in distribution of labeled cells w h e n injections involve lobule I I I in addition to lobulus II is that the transverse b a n d tends to extend s o m e w h a t m o r e ventrally than following injections of lobules I and II (cats B.St.L. 701, not illustrated, and 725, Fig. 3, injected stereotactically 0.25 and 0.20 ~1, survival time 1 and 2 days, respectively). Lobule IV. I n cat B.St.L. 709 (injected stereotactically 0.2 ~tl, killed after 2 days, Fig. 3) the staining is mainly restricted to the right vermal part of lobulus IV, but extends s o m e w h a t over the midline. L a b e l e d cells are distributed symmetrically in the left and rightpontine nuclei, but the m a j o r i t y (approx. two thirds) is f o u n d
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on the left side. The labeled cells are restricted to the caudal fourth of the pons, and are distributed lateral to the peduncle in a transversally oriented band in the dorsal part (Fig. 3). Corresponding observations are made in two other cases with injections of lobulus IV (cats B.St.L. 720 and 723, not illustrated, 0.15 and 0.2 ~tl, survival time 2 and 3 days, respectively). In cat B.St.L. 723 with practically unilateral staining, three fourths of the labeled cells are found contralateral to the injection. The band of labeled cells extends somewhat more ventrally in these cases with injections of lobule IV than in those affecting lobules I - I I I (Fig. 3). Lobule V. In five cases the staining is restricted to the vermal part of lobulus V. Even if different folia are involved in different cases, the distribution of labeled cells in the pontine nuclei is not significantly different. In cat C.Co. L. 192 (injected 0.15 ~!, survival time 1 day, Fig. 3) the staining extends more laterally on the left than on the right side. It is restricted to the vermis. In the rightpontine nuclei labeled cells are found in an area lateral to the peduncle in the caudal third of pons. A few labeled cells are also found in the caudalmost part of the dorsolateral nucleus (section 4) and medial to the peduncle. In the left pontine nuclei a smaller number of cells show a closely similar distribution. In three other cases, (cats B.St.L. 648,654 and 689, injected 0.25, 0.075 and 0.4 ~1, survival times 2, 1 and 2 days, respectively, not illustrated), mainly the medial zone of the vermis of lobule V is stained, but the distribution of labeled cells is the same as in C.Co.L. 192 (Fig. 3). In the fifth case (cat C.Co.L. 193, injected 0.2 ~tl, survival time 1 day, not illustrated), the staining involves in addition to the vermis the medialmost strip of the intermediate region, but the findings in the pons do not differ from those made when the staining is restricted to the vermis. In this case the staining extends only slightly across the midline, and three fourths of the labeled cells are found in the contralateral pontine nuclei.
Comments Following injections restricted to the vermis of the anterior lobe, labeled cells are concentrated in a well restricted area lateral to the peduncle in the caudal third to fourth of the pons. The main difference between cases with injections of the rostral (lobules I, II and III) and the caudal (lobule V) parts of the anterior lobe is that the band of labeled cells is found more ventrally in the latter (Fig. 3, see also Fig. 6). Judging from cases with almost unilateral staining of the cortex, the pontocerebellar projection to the anterior lobe vermis appears to be mainly contralateral (about 3/4), however with a significant (1/4) ipsilateral contribution.
The Intermediate-Lateral Part of the Anterior Lobe Due to the small size of the intermediate-lateral part in the rostral part of the anterior lobe, injections without involvement of the vermis or the deep cerebellar nuclei have only been obtained in lobules V and III-IV.
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Fig. 4. The pontine projection to the intermediate part of lobules V. The extent of staining in the lobulus V is indicated with black in the diagram of the cerebellum (the staining involves to a lesser degree also lobulus VI). Labeled cells are represented with black dots in drawings of transverse sections through the pontine nuclei. One group of cells is found medial to the peduncle, another lateral to it. Both have the shape of longitudinal columns. The labeled cells within the circle are due to the involvement of lobulus VI by the injection (cfr. text)
Lobule V. I n cat C. Co. L. 190 (0.2 ~1, survival t i m e 1 day, Fig. 4) t h e c a u d a l folia o f t h e right i n t e r m e d i a t e p a r t o f l o b u l e V a r e stained, b u t in a d d i t i o n t h e r e is s o m e slight staining o f t h e a d j o i n i n g folia o f l o b u l e VI. In t h e contralateral pontine nuclei n u m e r o u s l a b e l e d cells (up to 80 cells in o n e 40 ~m section) a r e d i s t r i b u t e d in two c o l u m n - s h a p e d a r e a s in t h e c a u d a l 2/3 o f the p o n s , o n e m e d i a l , t h e o t h e r l a t e r a l to t h e p e d u n c l e . T h e m e d i a l c o l u m n is f o u n d d o r s a l l y close to t h e p e d u n c l e . In a d d i t i o n to t h e s e two c o l u m n s , a m i n o r g r o u p of l a b e l e d cells is l o c a t e d close to t h e v e n t r a l a s p e c t o f t h e p e d u n c l e (circles w i t h b r o k e n line in Fig. 4). This g r o u p is n e v e r f o u n d in cases with i n j e c t i o n s r e s t r i c t e d to t h e a n t e r i o r lobe, a n d c o i n c i d e s with a r e g i o n f o u n d b y H o d d e v i k et al. ( 1 9 7 7 ) to p r o j e c t to l o b u l u s VI. I n t h e ipsilateral pontine nuclei o n l y a few
Pontocerebellar Projeetion to the Anterior Lobe
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241
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Fig. 5. Organization of the pontine projection to the intermediate part of the vermis. No convincing differences are apparent between the distribution of pontine cells projecting to lobulus IV (hindlimb) and lobulus V (forelimb) judged from cases with unilateral injections (B.St.L. 623 and C.Co.L. 195). However, in C.Co.L. 198, with bilateral injections, a somatotopical pattern is evident. Dorsal parts of the lateral region project to lobules III-IV, while ventral parts supply lobulus V. Within the medial region there is a slight tendency for cells projecting to lubules III-IV to be located more ventrolaterally
labeled cells are f o u n d (about 1 % of the contralateral n u m b e r ) . In two o t h e r cases with injections of the intermediate-lateral part of the lobulus V (cats C.Col.L. 189, not illustrated, and 195, Fig. 5, 0.3 and 0.1 ~1, respectively, survival time 2 days) the distribution of labeled cells c o n f o r m s to the description given above, except that the v e n t r o m e d i a l c o l u m n is lacking. Lobule IV. I n cat B . S t . L . 623 (injected 0.5 ~1, survival time 3 days, Fig. 5) the staining is restricted to the left intermediate-lateral part o f lobulus IV. I n the contralateral pontine nuclei a considerable n u m b e r of labeled cells (up to 110 cells in one 50 [~m section) are distributed mainly into two regions in the caudal
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2/3 of the ports, extending caudo-rostrally, medial and lateral to the peduncle, respectively. In the ipsiIateral pontine nuclei only a few labeled cells are found. Corresponding observations are made in another case with a closely similar injection (cat B.St.L. 710, injected stereotactically 0.2 ~tl, survival time 3 days, not illustrated). However, the labeled cells are on the whole located somewhat more dorsally than in cat B.St.L. 623.
Bilateral Injection of Lobules I l l - I V and V In order to compare in more detail the projection to the intermediate part of the different lobules, bilateral injections were made in one case. In cat C.Co.L. 198, 0.1 ~tl H R P was injected in lobule V and 0.1 ~1 stereotactically in lobules I I I - I V (survival time 2 days, Fig. 5). On the left side the staining is found in the intermediate part of lobules III and IV. The intracerebellar nuclei are not involved. In addition, there is a minute area of staining in the intermediate part of lobule V where the cannula has entered. On the right side the staining is confined to the intermediate part of lobule V. In thepontine nuclei labeled cells are distributed on both sides in a pattern in agreement with the findings in cases with corresponding unilateral injections (Figs. 4 and 5). However, certain differences between the two sides are obvious. Thus, in most sections the group of labeled cells lateral to the peduncle is located more ventrally on the left side than on the right. The group of labeled cells medial to the peduncle tends to be situated somewhat more ventrolaterally on the right than on the left side, but the difference is less clear than in the lateral cell group.
Comments The pontine projection to the lateral-intermediate part of the anterior lobe differs in some respect from that to the vermis. In the first place, the projection to the intermediate-lateral part is almost exclusively contralateral. Furthermore, the area lateral to the peduncle extends more rostrally when injections are placed in the intermediate part than when they are limited to the vermis (cp. Figs. 2 and 4). Finally, only the intermediate-lateral part receives a substantial projection from a region medial to the peduncle (Figs. 4 and 5). The distribution of labeled cells is very similar in cases with injections of the intermediate-lateral part of lobules IV and V, labeled cells being found within two well restricted areas, medial and lateral to the peduncle. Although certain differences exist between the results of injections in lobules IV and V, they do not exceed what may be due to individual differences, as appears from comparisons between cases with practically identical injections. However, following injection of lobules I I I - I V on one side and lobule V on the other within the same animal (cat C.Co.L. 198, Fig. 5), thus eliminating individual variations, it becomes evident that within the lateral region dorsal parts project to anterior lobuli, ventral parts to posterior ones, according to the same
Pontocerebellar Projection to the Anterior Lobe
243
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indicated with open (lobules I-IV) and filled (lobulus V) circles. The projection is bilateral, and cells projecting to rostral lobuli (hindlimb representation) are located more dorsally than those projecting to caudal lobuli (forelimb representation). The intermediate-lateral part receives fibres from more extensive, nearly ,8 exclusively controlateral pontine cell groups (hatching). One group of cells is found medial, another lateral to the peduncle. Caudally the lateral area coincides with that projecting to corresponding parts of the vermis. A somtoto
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concerning the anterior lobe vermis is present within the proiection from the lateral pontine region to the intermediate part. Within the medial pontine region the somatotopical pattern seems to be reversed, but is less clear than in the lateral region
CAUDAL
somatotopical principle as found within the projection to the vermis. The reverse pattern seems to be present within the medial region, but it is much less clear (Figs. 3, 5 and 6). Discussion By use of the retrograde axonal transport of HRP we have been able to map the pontocerebellar projection to the anterior lobe in more detail than has
.
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been possible with anterograde (Voogd, 1964) and retrograde degeneration techniques (Brodal and Jansen, 1946, and others). The advantages and limitations of the H R P method with regard to the study of cerebellar afferents have been documented and discussed (Walberg et al., 1976), and need not be repeated here. Suffice it to say that the H R P method is better suited for topographical than for quantitative purposes. Origin and Termination of Pontocerebellar Fibres to the.Anterior Lobe
Following injections of the anterior lobe, labeled cells in the pontine nuclei are confined to certain restricted parts, arranged as transverse bands or longitudinal columns. This arrangement resembles the one found previously with regard to the pontine projection to the paramedian lobule (Hoddevik, 1975) and the cerebellar visual areas (lobules VI-VIII, Hoddevik et al., 1977). It is also similar to the organization of terminal areas of corticopontine fibres from the frontal part of the cerebral cortex (P. Brodal, 1968a, b, 1971a, b). More specifically, the majority of pontine cells projecting to the anterior lobe are found lateral and medial close to the peduncle in the caudal part of the pons, in agreement with the findings of Voogd (1964) based upon anterograde degeneration technique. Furthermore, it is in agreement with the conclusion of certain of the older anatomists using normal or pathological human material (see Voogd, 1964, for references to the older literature). Brodal and Jansen (1946) using the modified Gudden method (Brodal, 1940) had no lesions restricted to the anterior lobe. However, in cases with partial involvement of the anterior lobe, retrograde changes or cell loss were found (among other places) in the areas found by us to project to the anterior lobe. Verrnis. We find that the anterior lobe vermis receives afferents almost exclusively from a well-restricted area in the caudal and lateral part of the pontine nuclei (Figs. 3 and 6). Although longitudinal zones of the anterior lobe vermis differ with regard to development (Korneliussen, 1968) and connections (Voogd, 1964; Voogd et al., 1969; Oscarsson, 1973; Armstrong et al., 1974; Brodal and Walberg, 1977), no such differences could be discerned in the present material. This can probably not be fully explained by the fact that our injections were not restricted to particular zones. Thus, using the same experimental material, Brodal and Walberg (1977) found different origins of the projections from the inferior olive to medial and lateral parts of the vermis. It is of interest in this connection that while Voogd et al. (1969) found spinocerebellar fibres to terminate in clear sagittal strips in the anterior lobe, this was not the case for pontocerebellar fibres. In harmony with this, Provini et al. (1968) with electrical stimulation of the sensorimotor cortex found evidence for a longitudinal subdivision of the anterior lobe vermis concerning climbing fibre responses (mediated at least mainly via the inferior olive; see especially Batini et al., 1976) but not concerning mossy fibre responses. A somatotopical pattern is present within the pontine projection to the anterior lobe vermis, since lobule V (representing the forelimb) receives fibres from more ventral parts of the pontine area than do lobules I-IV (hindlimb part). A pertinent question is then whether these pontine regions receive
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projections from cerebrocortical forelimb and hindlimb areas, respectively. When the present results are compared with those obtained by mapping the distribution of anterograde degeneration i n the pontine nuclei following small lesions of the sensorimotor cortex (P. Brodal, 1968a), it appears that most of the pontine area connected with the vermal part of lobulus V receives afferents from the motor forelimb area (lateral part of the anterior sigmoid gyrus), and to a lesser degree from the sensory forelimb region (lateral part of the posterior sigmoid gyrus). Parts of the pontine region projecting onto the hindlimb part of anterior lobe vermis (lobules I-IV) seem to receive afferents from the medial part of the posterior sigmoid gyrus, usually termed the sensory hindlimb area. The medial part of the anterior sigmoid gyrus, previously assumed to be the motor hindlimb area (Woolsey, 1958) does not appear to send fibres to those parts of the pons which project to the vermis of lobules I-IV. In agreement with this, stimulation of this part of the cortex evokes only slight mossy fibre potentials in lobules III-IV (Allen et al., 1974; Oka et al., 1976). It is interesting that recent data (see Wiesendanger et al., 1974) strongly suggest that the medial part of the anterior sigmoid gyrus represents the supplementary motor area in the cat, while the exact localization of the motor hindlimb area is still not clear. It may be speculated whether in the cat separation between sensory and motor subdivisions is less sharp in the hindlimb than in the forelimb region, so that the medial part of the posterior sigmoid gyrus is to be considered as a motor as well as a sensory hindlimb area. It should be noted that all parts of the pontine regions which project to the anterior lobe vermis do not seem to receive afferents from the primary sensorimotor area. These regions may be influenced from the second somatosensory area (P. Brodal, 1968b), the auditory cortex (P. Brodal, 1972c), and the superior and inferior colliculi (Kawamura and Brodal, 1973). However, in order to clarify these problems, it will be necessary to map in one experimental animal the terminal areas of the corticopontine fibres and the localization of the pontine cells projecting to the anterior lobe vermis. Such studies are now in progress in our laboratory with the combined use of the Fink and Heimer silver impregnation method and the HRP method. That this approach can be fruitful has recently been shown by Blomqvist and Westman (1975) in the dorsal column nuclei. Intermediate-Lateral Part. There is a main difference between the pontine projections to the vermis and the intermediate-lateral part of the anterior lobe: the latter receives a much stronger projection, and from more extensive parts of the pontine nuclei than does the former (Fig. 6). Thus, the intermediate-lateral part receives afferents from a longitudinal cell column medial to the peduncle, as well as from one lateral to it. It appears that the caudal part of the lateral cell column coincides with the one projecting to the vermis. Both the medial and the lateral columns projecting to the intermediate-lateral part receive afferents from the motor cortex, at least from its forelimb area. This is evident from experiments combining silver impregnation of degenerating fibres and retrograde labeling with HRP (P. Brodal, unpublished observations). By the use of bilateral HRP injections in the anterior lobe it has been possible to demonstrate a somatotopical pattern within the pontine projection to
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the intermediate part, at least within the lateral pontine region. The pattern is the same as that found within the proiection to the vermis (Fig. 6). Within the corticopontine projection likewise certain topographical differences became evident only with the use of bilateral cortical lesions (P. Brodal, 1968a, b, 1972a, b, c). While anatomically no clear difference in precision seems to exist between the pontocerebellar projection to the vermis and the intermediate part, physiologically the somatotopical organization of mossy fibre responses is sharper in the intermediate part than in the vermis (Provini et al., 1968). However, it should be recalled that mossy fibre potentials to the anterior lobe may be relayed in brain stem nuclei other than the pontine (nucleus reticularis lateralis, nucleus reticularis paramedianus, the perihypoglossal nuclei and the nucleus reticularis tegmenti pontis, for a review, see Brodal, 1972). It may be that some of these nuclei project more diffusely onto the vermis than to the intermediate part, and therefore blurr the somatotopical pattern of evoked potentials after cerebrocortical stimulation. It is of interest to consider our findings in relation to what is known of pontocerebellar projections to other parts of the cerebellum, when studied by the retrograde transport of HRP. The vermis of the anterior lobe receives afferents from largely other parts of the pons than does the vermis of lobules VI-VIII (Hoddevik et al., 1977). These lobules receive fibres from several longitudinal, sharply delimited cell columns in various parts of the pons, while the cells projecting to the anterior lobe vermis are distributed more transversally in the caudal part of the pons. The paramedian lobule likewise receives pontine afferents from several longitudinal cell columns (Hoddevik, 1975). Some of these appear to coincide closely with terminal areas of corticopontine fibres from the first and second somatosensory cortices (P. Brodal, 1968a, b), while there is less overlap with terminal areas of fibres from the motor cortex. As discussed above, the pontine regions projecting to the anterior lobe probably receive more cortical fibres from the motor than from the sensory cortex. It may therefore be suggested that the anterior lobe is chiefly (but not exclusively, see Allen et al., 1974; Oka et al., 1976) influenced from the motor cortex, while the paramedian lobule is functionally more closely related to the somatosensory cortical areas. The pattern of organization within the pontocerebellar projection appearing from the above mentioned studies (Hoddevik, 1975; Hoddevik et al., 1977) and from the present study, is indeed complex. A large number of often very discrete cell groups project onto particular parts of the cerebellum. A restricted part of the cerebellar cortex may receive fibres from several pontine cell groups, and one pontine cell group may project to different cerebellar parts. Thus, there is an extensive convergence as well as divergence within the pontocerebellar projection. The complexity in the cortico-ponto-cerebellar pathway is increased by the fact that each small part of the cerebral cortex as a rule projects onto several discrete parts of the pontine nuclei, while fibres from different cortical areas may converge onto one pontine cell group. This pattern represents a warning against regarding the cortico-ponto-cerebellar projection as being rather
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diffusely organized. It may well be that with the use of the present rather crude anatomical techniques, destroying thousands of functionally different cortical cells, or injecting HRP around thousands of possibly dissimilar pontocerebellar axon terminals, we can clarify only the most crude pattern of organization. When studied in detail, individual pontine cells are found to be functionally very specific with regard to their input (Baker et al., 1976), a fact which certainly reflects that their afferent fibres terminate very specifically. Many questions remain to be solved. For instance, when different pontine cell groups project to one cerebellar area, do they converge on identical Purkinje cells (via granule cells), or is there a very discrete mosaic pattern (see Eccles et al., 1972) not shown by our present method? Furthermore, when one pontine cell group projects to several different cerebellar areas, are the axons bifurcating, or are different, but closely adjoining cells projecting differently? The latter question may be solved by using different tracers for retrograde axonal transport as done, for example, by Steward and Scoville (1976) in the hippocampus.
References Allen, G.I., Tsukahara, N.: Cerebrocerebellar communication systems. Physiol. Rev. 54, 957-1006 (1974) Allen, G.I., Azzena, G.B., Ohno, T.: Somatotopically organized inputs from fore- and hindlimb areas of sensorimotor cortex to cerebellar Purkyn6 cells. Exp. Brain Res. 20, 255-272 (1974) Adrian, E. D.: Afferent areas in the cerebellum connected with the limbs. Brain 66, 289-315 (1943) Armstrong, D.M., Harvey, R.J., Schild, R.F.: Topographical localization in the olivocerebellar projection: An electrophysiologicval study in the cat. J. comp. Neurol. 154, 287-302 (1974) Baker, J., Gibson, A., Glickstein, M., Stein, J.: Visual cells in the pontine nuclei of the cat. J. Physiol. (Lond.) 255,415-434 (1976) Batini, C., Corvisier, J., Destombes, J., Gioauni, H., Everett, J.: The climbing fibers of the cerebellar cortex, their origin and pathways in the cat. Exp. Brain Res. 26, 407-422 (1976) Blomqvist, A., Westman, J.: Combined HRP and Fink-Heimer staining applied on the gracile nucleus in the cat. Brain Res. 99, 339-342 (1975) Brodal, A.: Modification of Gudden method for study of cerebral localization. Arch. Neurol. Psychiat. (Chic.) 43, 46-58 (1940) Brodal, A.: Cerebrocerebellar pathways. Anatomical data and some functional implications. Acta Neurol. Scand., Suppl. 51, 153-196 (1972) Brodal, A., Jansen, J.: The pontocerebellar projection in the rabbit and cat. Experimental investigations. J. comp. Neurol. 84, 31-118 (1946) Brodal, A., Walberg, F.: The olivocerebellar projection in the cat studied with the method of retrograde axonal transport with horseradish peroxidase. IV. The projection to the anterior lobe. J. comp. Neurol. 172, 85-108 (1977) Brodal, P.: The corticopontine projection in the cat. I. Demonstration of a somatotopically organized projection from the primary sensorimotor cortex. Exp. Brain Res. 5, 210-234 (1968a) Brodal, P.: The corticopontine projection in the cat. Demonstration of a somatotopically organized projection from the second somatosensory cortex. Arch. Ital. Biol. 106, 310-332. (1968b) Brodal, P.: The corticopontine projection in the cat. I. The projection from the proreate gyrus. J. comp. Neurol. 142, 127-140 (1971a) Brodal, P.: The corticopontine projection in the cat. II. The projection from the orbital gyrus. J. comp. Neurol. 142, 141-152 (1971b) Brodal, P.: The corticopontine projection from the visual cortex in the cat. I. The total projection and the projection from areas 17. Brain Res. 39, 297-317 (1972a)
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Brodal, P.: The corticopontine projection from the visual cortex in the cat. II. The projection from areas 18 and 19. Brain Res. 39, 319-335 (1972b) Brodal, P.: The corticopontine projection in the cat. The projection from the auditory cortex. Arch. Itai. Biol. 10, 119-144 (1972c) Eccles, J.C., Sabah, N.H., Schmidt, R.F., T~ibo~ikov~, H.: Mode of operation of the cerebellum in the dynamic loop control of movement. Brain Res. 40, 73-80 (1972) Evarts, E.V., Thach, W.T.: Motor mechanisms of the CNS: cerebrocerebellar interrelations. Ann. Rev. Physiol. 31, 451--498 (1969) Graham, R.C., Jr., Kamovsky, M.J.: The early stages of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney: ultrastructural cytochemistry.by a new technique. J. Histochem. Cytochem. 14, 291-302 (1966) Hampson, J.L.: Relationship between cerebral and cerebellar cortices in cat. J. Neurophysiol. 12, 37-50 (1949) Hoddevik, G.H.: The pontocerebellar projection onto the paramedian lobule in the cat: An experimental study with the use of horseradish peroxidase as a tracer. Brain Res. 95, 291-307 (1975) Hoddevik, G.H., Brodal, A., Kawamura, K., Hashikawa, T.: The pontine projection to the cerebellar vermal visual area studied by means of the retrograde axonal transport of horseradish per0xidase. Brain Res. 123, 209-227 (1977) Kawamura, K., Brodal, A.: The tectopontine projection in the cat: An experimental anatomical study with comments on pathways for teleceptive impulses to the cerebellum. J. comp. Neurol. 149, 371-390 (1973) Korneliussen, H.K.: Comments on the cerebellum and its division. Brain Res. 8, 229-236 (1968) Oka, H., Yasuda, T., Jinnai, K., Yoneda, Y.: Reexamination of cerebellar responses to stimulation of sensorimotor areas of the cerebral cortex. Brain Res. 118, 312-319 (1976) Oscarsson, O.: Functional organization of spinocerebellar paths. In: Handbook of sensory physiology, Vol. 2 (ed. A. Iggo), p. 339-380. Somatosensory system. Berlin-Heidelberg-New York: Springer 1973 Provini, L., Redman, S., Strata, P.: Mossy and climbing fibre organization on the anterior lobe of the cerebellum activated by forelimb and hindlimb areas of the sensorimotor cortex. Exp. Brain Res. 6, 216-233 (1968) Snider, R.S., Eldred, E.: Electro-anatomical studies on cerebrocerebellar connections in the cat. J. comp. Neurol. 95, 1-16 (1951) Steward, O., Scoville, S.A.: Retrograde labeling of central nervous pathways with tritiated or Evans blue-labeled bovine serum albumine. Neuroscience Letters 3, 191-196 (1976) Tomasch, J.: The overall information capacity of the major afferent and efferent cerebellar cell and fiber systems. Confin. neurol. (Basel) 30, 359-367 (1968) Voogd, J.: The cerebellum of the cat. Structure and fibre connections, pp. 215. Leiden: Van Gorcum & Comp. 1964 Voogd, J., Broere, G., van Rossum, J.: The medio-lateral distribution of the spinocerebellar projection in the anterior lobe and the simple lobule in the cat and a comparison with some other afferent fibre systems. Psychiat. Neurol. Neurochir. (Amst.) 72, 137-151 (1969) Walberg, F., Brodal, A., Hoddevik, G.H.: A note on the method of retrograde transport of horseradish peroxidase as a tool in studies of afferent cerebellar connections, particularly those from the inferior olive; with comments on the orthograde transport in Purkinje cell axons. Exp. Brain Res. 24, 383-401 (1976) Wiesendanger, M., Seguin, J.J., Kiinzte, H.: The supplementary motor area - a control system for posture? Advanc. Behav. Biol. 7, 331-346 (1974) Woolsey, C.N.: Organization of somatic sensory and motor areas of the cerebral cortex. In: Biological and biochemical bases of behavior (eds. Harlow and Woolsey), pp. 63-81. Madion: University of Wisconsin Press 1958
Received February 2, 1977
