C3 Zone Research Articles

Single Purkinje cell can innervate multiple classes of projection neurons in the cerebellar nuclei of the rat: a light microscopic and ultrastructural triple-tracer study in the rat.

Two different populations of projection neurons are intermingled in the cerebellar nuclei. One group consists of small, gamma-aminobutyric acid-containing (GABAergic) neurons that project to the inferior olive, and the other group consists of larger, non-GABAergic neurons that provide an input to one or more, usually premotor, centers in the brainstem, such as the red nucleus, the thalamus, and the superior colliculus. All cerebellar nuclear neurons are innervated by GABAergic Purkinje cells. In this study, we investigated whether individual Purkinje cells of the C1 zone of the paramedian lobe of the rat innervate both groups of projection neurons in the anterior interposed nucleus. Two different, retrogradely transported tracers, either cholera toxin beta subunit (CTb) or wheat germ agglutinin coupled to horseradish peroxidase (WGA-HRP) and a gold lectin tracer were injected into the red nucleus and the inferior olive, respectively, whereas Purkinje cell axons were anterogradely labeled with biotinylated dextran amine (BDA) injected into the paramedian lobule. Cerebellar nuclear sections studied with the light microscope demonstrated a close relation of varicosities from BDA-labeled Purkinje cell axons with both gold lectin- and CTb-labeled neurons. Branches of individual axons could be traced to both retrogradely labeled cell populations. At the ultrastructural level, synapses of labeled Purkinje cell terminals with profiles of WGA-HRP-labeled projection neurons predominated over contacts with gold lectin-containing neurons. Nine out of 367 investigated BDA-labeled terminals were observed to be presynaptic to a WGA-HRP-labeled profile as well as to a gold lectin-labeled profile. This indicates that nuclear cells that project to the inferior olive as well as those that project to premotor centers are under the influence of the same Purkinje cells. Such an arrangement would suggest an in-phase cortical modulation of the activation patterns of the inhibitory cells that project to the inferior olive and excitatory cells that project to premotor nuclei, which could explain why olivary neurons, especially those of the rostral part of the dorsal accessory olive, appear to be unresponsive to stimuli generated during active movement.

Zonal organization of cortico-nuclear and nucleo-cortical projections of the paramedian lobule of the cat cerebellum. 1. the C1 zone.

The cortico-nuclear (C-N) and nucleo-cortical (N-C) projections of the C1 cortical zone in pars anterior (pa) and pars copularis (pc) of the paramedian lobule (PML) in the posterior lobe of the cat cerebellum were investigated with a combined electrophysiological and neuroanatomical technique. In each experiment the mediolateral boundaries of the zone were located on the cortical surface by recording field potentials mediated via climbing fibres and evoked in the zone by activity elicited in spino-olivocerebellar paths through percutaneous stimulation of fore- and hindlimbs; a small (15-30 nl) injection of WGA-HRP was then made into the zone. The distributions in the deep cerebellar nuclei were determined (with light microscopy) both for terminal labelling due to anterograde axonal transport by Purkinje cells and for cell bodies labelled due to retrograde transport in N-C axons. The extent to which injection sites were confined to the C1 zone was assessed both by comparing injection site and zone widths and by determining the distributions of retrogradely labelled neurones within the contralateral inferior olive. The C-N projection from the part of the zone in PML pa (a forelimb part) terminates almost exclusively (perhaps exclusively) in nucleus interpositus anterior (NIA), primarily in caudal and dorsal parts, where it overlaps heavily with the C-N projections from the lobule V parts (also forelimb parts) of the C1 and C3 zones as previously defined. The C-N projection from the part of the zone in PML pc (a hindlimb part) also terminates virtually exclusively in NIA but primarily in almost all parts of the medial third of the nucleus. There is, nevertheless, sufficient overlap between the PML pa and PML pc projections that approximately one third of the termination territory of each projection overlaps that of the other. The PML pc part of the zone is almost entirely lacking in a N-C projection, as previously found for the lobule V part of the C1 zone (and C3 zone). However, the PML pa part of the zone receives N-C projections that arise, in descending order of size, from nucleus interpositus posterior (NIP), from NIA, from the NIA/nucleus lateralis (NL) fusion area and (perhaps) NL. The projection from NIP is similar in size to that provided by the nucleus to the C2 zone in lobule V of the anterior lobe. The findings are discussed, with particular emphasis on their implications for the hypothesis that the cerebellum is divisible into a number of olivo-cortico-nuclear complexes or compartments.

Zonal organization of cortico-nuclear and nucleo-cortical projections of the paramedian lobule of the cat cerebellum. 2. The C 2 zone

The cortico-nuclear (C-N) and nucleo-cortical (N-C) projections of the C2 cortical zone in pars anterior (pa) and pars posterior (pp) of the paramedian lobule (PML) in the posterior lobe of the cat cerebellum were investigated with a combined electrophysiological and neuroanatomical technique. In each experiment the mediolateral boundaries of the zone were localized on the cortical surface by recording field potentials mediated via climbing fibres and evoked in the zone by activity elicited in spino-olivocerebellar paths through percutaneous stimulation of the fore- and hindlimbs; a small (15-30 nl) injection of 1-2% WGA-HRP was then made into the zone. Distributions in the deep cerebellar nuclei were determined with light microscopy both for C-N terminal labelling due to anterograde axonal transport by Purkinje cells and for cell bodies labelled due to retrograde transport in N-C axons. The extent to which retrogradely labelled olivary neurones were confined to the part of the rostral medial accessory olive that innervates the C2 zone was estimated to provide an indication of the degree to which the injected tracer might have spread beyond the boundaries of the zone. The C-N projection from the part of the C2 zone in PML pa terminates almost exclusively (probably exclusively) in nucleus interpositus posterior (NIP) at all medio-lateral levels of the nucleus but most extensively at middle and lateral levels. At most levels the C-N termination territory forms a crescent with its outer curve following the caudal, dorsal and rostral borders of the nucleus and as a result it is mainly in the dorsal half of the nucleus. There is heavy overlap with the projection from the lobule V part of the C2 zone previously studied by us. The projection from the C2 zone in PML pp terminates entirely in NIP, but although at middle medio-lateral levels in the nucleus there is substantial overlap with the PML pa and lobule V projections, the projection territory is confined to the medial half of the nucleus. Evidence was obtained compatible with the view that throughout the C2 zone its lateral and medial parts project to different parts of NIP. In both PML pa and pp the C2 zone receives N-C projections from NIP. Most of the N-C cells concerned are in the dorsal half of NIP and the great majority lie within the corresponding C-N projection territory. However, the N-C projection to PML pa appears c. 6 times heavier than that to PML pp and the PML pa part of the zone also receives a minor additional projection from nucleus lateralis (NL). The findings are discussed in relation to the hypothesis of olivo-cortico-nuclear complexes or compartments, with particular reference to the internal organization of the C2 complex.

Numerical aspects of pontine, lateral reticular, and inferior olivary projections to two paravermal cortical zones of the cat cerebellum

Two different olivo-cortico-nuclear zones in the cat cerebellum have been compared quantitatively as regards the numbers of cells projecting to them from within several sources of mossy fibres (MFs), namely the basal pontine nuclei (BPN), nucleus reticularis tegmenti pontis (NRTP), and the ipsilateral lateral reticular nucleus (LRN). The zones chosen were the C3 zone in lobule V of the anterior lobe and the C1 zone in pars copularis of the paramedian lobule (PMLpc), localised by recording climbing fibre-mediated potentials evoked on their surface as a result of volleys set up in their spino-olivocerebellar paths. The zones were injected with fluorescent-labelled latex microspheres and cell bodies, retrogradely labelled in the MF source nuclei and in the contralateral inferior olive, were counted and mapped. Evidence was obtained that tracer efficiency was very high in both the MF projections and the olivo-cerebellar projection and that each olivocerebellar axon may provide only one climbing fibre to the upper part of a lobule V folium but an average of nearly two to the same part of a PML folium. When the numbers of labelled cells in each MF source nucleus were expressed as a percentage of the total number of labelled pontine cells, the biggest source for lobule V was the contralateral BPN, followed by LRN, contralateral NRTP, ipsilateral BPN, and ipsilateral NRTP. For PMLpc, the order was similar except that ipsilateral BPNs exceeded contralateral NRTPs, but the dominance of contralateral BPN as a source was much greater. Cell totals were converted into projection densities (i.e., numbers of cells labelled per square millimetre of cortical sheet involved in the injection site); densities for PMLpc were found to be almost three times greater than those for lobule V for contralateral BPN but the two densities were not significantly different for ipsilateral BPN. The three other MF sources projected at higher densities to lobule V than to PML. These findings indicate that two cortical zones, both of which receive climbing fibres from the rostral part of the dorsal accessory olive and project to nucleus interpositus anterior, nevertheless differ markedly in regard to both the relative and the absolute sizes of the projections they receive from several of their most important sources of MFs.

Somatotopical organisation within the climbing fibre projection to the paramedian lobule and copula pyramidis of the rat cerebellum

The climbing fibre projection to the paramedian lobule (lobule VII) and the copula pyramidis (lobule VIII) in the posterior lobe of the rat cerebellum was investigated in pentobarbitone-anaesthetised animals. Percutaneous electrical stimulation generated climbing fibre field potentials on the cerebellar surface that indicated a somatotopical organisation into five distinct cortical areas. Each area was identified by the site(s) of peripheral stimulation that evoked the largest response within that area, and from medial to lateral the cortex was subdivided as follows [principal site(s) of stimulation in parentheses with corresponding range of onset latencies]: in the paramedian lobule, area 1 (contralateral face, 17-18 ms), area 2 (ipsilateral forelimb, 10-15 ms) and area 3 (ipsi- and contralateral forelimbs, 16-26 ms and 15-30 ms, respectively); and in the copula pyramidis, area 4 (tail, 17-21 ms) and area 5 (ipsilateral hindlimb, 13-19 ms). In additional retrograde tracer experiments, small volumes of fluorescent-tagged beads were injected into each of the different areas and the location of retrogradely labelled olive cells was mapped. By comparison with other species, the results indicate that in the paramedian lobule area 1 corresponds to zone A2; area 2 corresponds, at least in part, to medial C1; and area 3 corresponds to C2; in addition, farther laterally, a C3 zone may be present. In the copula pyramidis, areas 4 and 5 correspond to subzones of medial C1. Overall, the results support the view that a general principle of cerebellar cortical organisation is a division into parasagittal zones, each characterised by its somatotopically organised climbing fibre input that arises from a specific, rostrocaudally oriented column of cells within the inferior olivary nucleus.

Somatotopical organisation within the climbing fibre projection to the paramedian lobule and copula pyramidis of the rat cerebellum

The climbing fibre projection to the paramedian lobule (lobule VII) and the copula pyramidis (lobule VIII) in the posterior lobe of the rat cerebellum was investigated in pentobarbitone-anaesthetised animals. Percutaneous electrical stimulation generated climbing fibre field potentials on the cerebellar surface that indicated a somatotopical organisation into five distinct cortical areas. Each area was identified by the site(s) of peripheral stimulation that evoked the largest response within that area, and from medial to lateral the cortex was subdivided as follows [principal site(s) of stimulation in parentheses with corresponding range of onset latencies]: in the paramedian lobule, area 1 (contralateral face, 17–18 ms), area 2 (ipsilateral forelimb, 10–15 ms) and area 3 (ipsi- and contralateral forelimbs, 16–26 ms and 15–30 ms, respectively); and in the copula pyramidis, area 4 (tail, 17–21 ms) and area 5 (ipsilateral hindlimb, 13–19 ms). In additional retrograde tracer experiments, small volumes of fluorescent-tagged beads were injected into each of the different areas and the location of retrogradely labelled olive cells was mapped. By comparison with other species, the results indicate that in the paramedian lobule area 1 corresponds to zone A2; area 2 corresponds, at least in part, to medial C1; and area 3 corresponds to C2; in addition, farther laterally, a C3 zone may be present. In the copula pyramidis, areas 4 and 5 correspond to subzones of medial C1. Overall, the results support the view that a general principle of cerebellar cortical organisation is a division into parasagittal zones, each characterised by its somatotopically organised climbing fibre input that arises from a specific, rostrocaudally oriented column of cells within the inferior olivary nucleus. J. Comp. Neurol. 389:249–263, 1997. © 1997 Wiley-Liss, Inc.

Maastrichtian dinocyst floras from Maud Rise and Georgia Basin (Southern Ocean): Their stratigraphic and paleoenvironmental implications

Dinocyst assemblages from Ocean Drilling Project (ODP) Leg 113, sites 689 and 690, located at Maud Rise, and Leg 114, Site 698 at South Georgia Basin, are of medium diversity with close to 40 taxa. The floras are dominated by Isabelidinium pellucidum and Odontochitina porifera; common species are Batiacasphaera reticulata, Microdinium cassiculum, Leberidocysta sp. A, Nematosphaeropsis sp. and Valensiella reticulata. The age of these dinocyst‐bearing sections has been determined, using paleomagnetics and biostratigraphy, as early Maastrichtian (Chron 32N), correlative to the uppermost part of the B. coronum to the lower part of the B. magnum nannofossil zones. Comparison with equally well‐dated Kerguelen Plateau palynofloras indicates an age equivalent to the dinoflagellate Zone C2, and perhaps also Zone D of Mao and Mohr (1992). The precise age model for these dinocyst floras allows better calibration of other Antarctic shallow marine sections from the James Ross Basin. Decreasing species diversity and increasing endemism of these floras can be used to reconstruct environmental conditions. Decreasing diversity is interpreted as response to cooling of the Antarctic water masses, probably caused by the onset of a distinct proto‐circum‐Antarctic current during the latest Campanian and early Maastrichtian.

Gating of cutaneous input to cerebellar climbing fibres during a reaching task in the cat.

1. Task-dependent modulation of cutaneous input to climbing fibres projecting to the C1, C2 and C3 zones in the cerebellar paravermal lobule V was investigated in awake cats during performance of a reaching task. 2. Climbing fibre responses resulting from low intensity (non-noxious) electrical stimulation of the ipsilateral superficial radial nerve were recorded as extracellular field potentials in the cerebellar cortex using chronically implanted microwires. 3. Response size, measured as the time-voltage integral of the evoked field potential, was assessed during three phases of the reaching movement, reaction, reach and grasp, and compared with the response size at rest. 4. At C1 and C3 zone recording sites response size was usually reduced during the task (7/10 sites). The reduction was most pronounced in the grasp phase, occasionally accompanied by a smaller reduction in the reach and reaction phases. In one case an enhancement was found in the reach phase. 5. Response size was also modulated during the task at four of six C2 zone recording sites. However, the results were mixed. In three cases the modulation resembled the pattern at C1/C3 sites with the responses being reduced in the grasp phase accompanied on occasion by a lesser reduction in the reach phase. In the remaining case there was an enhancement during grasp. In this case and one other there was also an enhancement during the reaction phase. 6. The findings indicate significant gating of cutaneous input to climbing fibres projecting to the C1, C2 and C3 zones during reaching movements, while the variability between recording sites suggests functional differences, both between and within zones.

Topographical organisation within the lateral reticular nucleus mossy fibre projection to the c1 and c2 zones in the rostral paramedian lobule of the cat cerebellum

The organisation of the projection from the principal lateral reticular nucleus (LRN) to the electrophysiologically defined cerebellar cortical c1 and c2 zones within apical folia of the forelimb-receiving area of the rostral paramedian lobule was investigated in cats. In individual experiments, small injections of either of two retrograde tracers, wheat germ agglutinin-horseradish peroxidase or fluorescent-tagged beads, were made into one or the other of the two zones. The mossy fibre projection from the LRN was found to arise almost entirely from the ipsilateral nucleus, primarily from its magnocellular region. In the pars anterior (folia 1-3) of the paramedian lobule, the projection to the c1 zone arose from cells located throughout most of the rostrocaudal extent of the magnocellular LRN, whereas the projection to the c2 zone was from cells in the rostral half of this region. There was also a small projection from cells in the dorsolateral parvicellular LRN to the c2 zone, but there was none to the c1 zone. Double-label experiments showed that, in territories within LRN where overlap occurred, cells that target either zone were intermingled, and, at most, about 9% of cells had axons that branched to innervate both zones. In the pars posterior (folia 4-6), cells targeting the two zones arose from largely overlapping areas throughout both the magnocellular and the dorsolateral parvicellular LRN, although cells targeting the c2 zone appeared to be more restricted in distribution. In addition to these interzonal differences, it was found that the projection to the pars posterior from cells in the parvicellular LRN was stronger than that to the pars anterior, irrespective of the cortical zone injected. Overall, the present study provides evidence that there is at least some topographical organisation within the LRN mossy fibre projection to certain cerebellar cortical zones.

Topographical organisation within the lateral reticular nucleus mossy fibre projection to the c1 and c2 zones in the rostral paramedian lobule of the cat cerebellum

Topographical organisation within the lateral reticular nucleus mossy fibre projection to the c1 and c2 zones in the rostral paramedian lobule of the cat cerebellum

Chapter 17 Signal processing in the C2 module of the flocculus and its role in head movement control

The major novel findings described and reviewed in the present study have all been demonstrated in the C2 module, which is formed by the rostral medial accessory olive, posterior interposed nucleus of the cerebellum, and zone C2. We show (1) that expression of dendritic lamellar bodies and dendrodendritic gap junctions in the rostral medial accessory olive are both down regulated by removal of the GABAergic input from the posterior interposed nucleus of the cerebellum to electrotonically coupled olivary dendrites; (2) that the high density of dendritic lamellar bodies in the rostral medial accessory olive can be correlated with a relatively high level of CS synchrony in the C2 zone of the flocculus; and (3) that the C2 zone of the flocculus is involved in head movements and probably gaze control. These results support the hypothesis that dendritic lamellar bodies are associated with dendrodentritic gap junctions, and they suggest that appropriate executions of compensatory head and eye movements require particular levels of complex spike synchrony in the flocculus.

Micro-organization of olivocerebellar and corticonuclear connections of the paravermal cerebellum in the cat.

The olivocerebellar and corticonuclear connections of the forelimb area of the paravermal medial C3 zone were studied in the cat using a combined electrophysiological and fluorescent tracer technique. During an initial operation under barbiturate anaesthesia, lobules IV/V of the cerebellar anterior lobe were exposed and small injections of dextran amines tagged with rhodamine or fluorescein were made into areas selected from four different electrophysiologically defined parts of the zone. The inferior olive and the deep cerebellar nuclei were then scrutinized for retrogradely labelled cells and anterogradely labelled axon terminals respectively. The findings demonstrate a detailed topographical organization within the olivocerebellar projection to the medial C3 zone and provide some evidence for a topographical organization of its projection to nucleus interpositus anterior. Both projections are described at a level of resolution not previously attained in neuroanatomical studies and the results strongly support the notion of a micro-compartmentalization of cerebellar olivo-corticonuclear circuits.

Inhibition of the inferior olive during conditioned responses in the decerebrate ferret.

Output from the interpositus nucleus can inhibit the inferior olive, probably via the GABA-ergic nucleo-olivary pathway. It has been suggested that the function of this inhibition might be to regulate synaptic plasticity resulting from parallel fibre/climbing fibre interaction in cerebellar Purkinje cells, by providing negative feedback information to the olive. Thus, when a learned response, generated by the interpositus nucleus, reaches a sufficient amplitude, the olive would be inhibited and further learning blocked. This suggestion was tested in a classical conditioning paradigm. Decerebrate ferrets were trained using electrical skin stimulation of the forelimb as the conditioned stimulus (CS) and periorbital stimulation as the unconditioned stimulus (US). Climbing fibre responses evoked in Purkinje cells by the US were recorded as surface field potentials in the part of the c3 zone controlling eyeblink. It was found that the CS did not inhibit the olive at the beginning of training, but when conditioned responses were large, the olive was inhibited by the CS in some animals. After a number of unpaired CS presentations, which caused extinction of the conditioned response, the inhibition disappeared. The size of individual conditioned responses correlated negatively with the size of the climbing fibre responses evoked by the US. Climbing fibre responses evoked by direct stimulation of the olive were also inhibited. It was concluded that cerebellar output during performance of a conditioned response inhibits the inferior olive. The results thus support the hypothesis of a cerebellar locus of conditioning and are consistent with the proposed role of cerebello-olivary inhibition.

Zonal organization of the flocculovestibular nucleus projection in the rabbit: A combined axonal tracing and acetylcholinesterase histochemical study

With the use of retrograde transport of horseradish peroxidase we confirmed the observation of Yamamoto and Shimoyama ([1977] Neurosci Lett. 5:279-283) that Purkinje cells of the rabbit flocculus projecting to the medial vestibular nucleus are located in two discrete zones, FZII and FZIV, that alternate with two other Purkinje cell zones, FZI and FZIII, projecting to the superior vestibular nucleus. The retrogradely labeled axons of these Purkinje cells collect in four bundles that occupy the corresponding floccular white matter compartments, FC1-4, that can be delineated with acetylcholinesterase histochemistry (Tan et al. [1995a] J. Comp. Neurol., this issue). Anterograde tracing from small injections of wheat germ agglutin-horseradish peroxidase in single Purkinje cell zones of the flocculus showed that Purkinje cell axons of FZII travel in FC2 to terminate in the medial vestibular nucleus. Purkinje cell axons from FZI and FZIII occupy the FC1 and FC3 compartments, respectively, and terminate in the superior vestibular nucleus. Purkinje cell axons from all three compartments pass through the floccular peduncle and dorsal group y. In addition, some fibers from FZI and FZII, but not from FZIII, arch through the cerebellar nuclei to join the floccular peduncle more medially. No anterograde tracing experiments were available to determine the projections of the FZIV and C2 zones. The functional implications of these results are discussed.

Zonal organization of the climbing fiber projection to the flocculus and nodulus of the rabbit: a combined axonal tracing and acetylcholinesterase histochemical study.

The localization and termination of olivocerebellar fibers in the flocculus and nodulus of the rabbit were studied with anterograde axonal transport methods [wheatgerm agglutinin-horseradish peroxidase (WGA-HRP) and tritiated leucine] and correlated with the compartments in the white matter of these lobules delineated with acetylcholinesterase histochemistry (Tan et al. J. Comp. Neurol., 1995, this issue). Olivocerebellar fibers originating from the caudal dorsal cap travel through floccular compartments FC2 and FC4 to terminate as climbing fibers in floccular zones FZII and FZIV. Fibers from the rostral dorsal cap and the ventrolateral outgrowth traverse compartments FC1 and FC3, which are interleaved with compartments FC2 and FC4, and terminate in zones FZI and FZIII. Fibers from the rostral pole of the medial accessory olive traverse the C2 compartment and terminate in the C2 zone. FZI-III extend into the adjoining folium (folium p) of the ventral paraflocculus. The C2 zone continues across folium p into other folia of the ventral paraflocculus and into the dorsal paraflocculus. Four compartments and five zones were distinguished in the nodulus. Medial compartment XC1 contains olivocerebellar fibers from the caudal dorsal cap and subnucleus beta that terminate in the XZI zone. Olivocerebellar fibers from the rostral dorsal cap and the ventrolateral outgrowth occupy XC2 and terminate in XZII. The XC4 compartment contains fibers from both the caudal dorsal cap and from the rostral dorsal cap and the ventrolateral outgrowth. The latter terminate in a central portion of the XZIV zone. The dorsomedial cell column projects to the XZIII zone, which is present only in the dorsal part of the nodulus. The rostral medial accessory olive projects to the XZV zone, which occupies the lateral border of the nodulus. These results confirm and extend the conclusions of Katayama and Nisimaru ([1988] Neurosci. Res. 5:424-438) on the zonal pattern in the olivo-nodular projection in the rabbit. Additional observations were made on the presence of a lateral A zone (Buisseret-Delmas [1988] Neurosci. Res. 5:475-493) in the hemisphere of lobules VI and VII. Retrograde labeling of the nucleo-olivary tract of Legendre and Courville ([1987] Neuroscience 21:877-891) was observed after WGA-HRP injections into the inferior olive including the rostral dorsal cap and the ventrolateral outgrowth. The anatomical and functional implications of these observations are discussed.

A Combined Retrograde Tracer and GABA‐immunocytochemical Study of the Projection from Nucleus Interpositus Posterior to the Posterior Lobe c2 Zone of the Cat Cerebellum

The extent to which the cells of origin of the cerebellar nucleocortical pathway are immunopositive for gamma-aminobutyric acid (GABA) was investigated in four cats using retrograde labelling of nucleocortical neurons in combination with immunocytochemistry. Neurons were retrogradely labelled by injection of fluorescent (coumarin)-tagged latex microspheres into the c2 zone in the rostral part of the paramedian lobule. The zone was identified electrophysiologically by the characteristics of the climbing fibre responses evoked on the cerebellar surface by percutaneous stimulation applied to the left and right forepaws in pentobarbitone-anaesthetized animals. Sections of the cerebellum containing the retrogradely labelled neurons were processed for GABA immunocytochemistry using a fluorescent (rhodamine)-tagged immunoglobulin. When viewed with epifluorescence microscopy and appropriate filter blocks the retrogradely labelled nucleocortical neurons could be visualized in the same sections as the GABA-immunopositive neurons. Almost all of a total of 254 labelled nucleocortical neurons were located in nucleus interpositus posterior, where a total of 711 GABAergic neurons were also found. None of these cells contained coumarin-tagged beads and displayed immunoreactivity for GABA (i.e. none was double-labelled). When compared by area of their cell body, the nucleocortical and GABA-immunopositive neurons appeared to form two partially overlapping populations. The mean cell area of the nucleocortical neurons was 620 +/- 233 microns2 (SD), whereas the GABA-immunopositive neurons were much smaller, with a mean cell area of 220 +/- 115 microns2. The results suggest that GABA does not play a major role in the nucleocortical pathway to the c2 zone of the rostral paramedian lobule of the cat cerebellum.

Functional relation between corticonuclear input and movements evoked on microstimulation in cerebellar nucleus interpositus anterior in the cat.

The functional relation between receptive fields of climbing fibres projecting to the C1, C3 and Y zones and forelimb movements controlled by nucleus interpositus anterior via the rubrospinal tract were studied in cats decerebrated at the pre-collicular level. Microelectrode tracks were made through the caudal half of nucleus interpositus anterior. This part of the nucleus receives its cerebellar cortical projection from the forelimb areas of these three sagittal zones. The C3 zone has been demonstrated to consist of smaller functional units called microzones. Natural stimulation of the forelimb skin evoked positive field potentials in the nucleus. These potentials have previously been shown to be generated by climbing fibre-activated Purkinje cells and were mapped at each nuclear site, to establish the climbing fibre receptive fields of the afferent microzones. The forelimb movement evoked by microstimulation at the same site was then studied. The movement usually involved more than one limb segment. Shoulder retraction and elbow flexion were frequently evoked, whereas elbow extension was rare and shoulder protraction never observed. In total, movements at the shoulder and/or elbow occurred for 96% of the interpositus sites. At the wrist, flexion and extension movements caused by muscles with radial, central or ulnar insertions on the paw were all relatively common. Pure supination and pronation movements were also observed. Movements of the digits consisted mainly of dorsal flexion of central or ulnar digits. A comparison of climbing fibre receptive fields and associated movements for a total of 110 nuclear sites indicated a general specificity of the input-output relationship of this cerebellar control system. Several findings suggested that the movement evoked from a particular site would act to withdraw the area of the skin corresponding to the climbing fibre receptive field of the afferent microzones. For example, sites with receptive fields on the dorsum of the paw were frequently associated with palmar flexion at the wrist, whereas sites with receptive fields on the ventral side of the paw and forearm were associated with dorsiflexion at the wrist. Correspondingly, receptive fields on the lateral side of the forearm and paw were often associated with flexion at the elbow, whereas sites with receptive fields on the radial side of the forearm were associated with elbow extension. The proximal movements that were frequently observed also for distal receptive fields may serve to produce a general shortening of the limb to enhance efficiency of the withdrawal. It has previously been suggested that the cerebellar control of forelimb movements via the rubrospinal tract has a modular organisation. Each module would consist of a cell group in the nucleus interpositus anterior and its afferent microzones in the C1, C3 and Y zones, characterised by a homogenous set of climbing fibre receptive fields. The results of the present study support this organisational principle, and suggest that the efferent action of a module is to withdraw the receptive field from an external stimulus. Possible functional interpretations of the action of this system during explorative and reaching movements are discussed.

Non-Reciprocal Connections between the Perihypoglossal Nuclei and the Cerebellar Paramedian Lobule in Rabbits and Their Relationship to the Climbing Fiber Zones

Retrogradely transported horseradish peroxidase (HRP) was used to investigate the organization of projections from the perihypoglossal nuclei to the cerebellar paramedian lobule, and possible reciprocal corticoperihypoglossal connections in rabbits. The absence of labeled Purkinje cells in the paramedian lobule after HRP injections of the perihypoglossal nuclei, compared with labeling in the nodulus, ipsilateral flocculus and adjacent ventral paraflocculus, strongly suggests that corticoperihypoglossal projections do not originate in the paramedian lobule. Comparison of labeling patterns in the perihypoglossal nuclei following HRP injections into various sublobules of the paramedian lobule suggests that projections from the perihypoglossal nuclei to the paramedian lobule exist and that some degree of topographic organization is present exclusively in bilateral projections from the caudal part of nucleus prepositus hypoglossi. All perihypoglossal projections end in deep parts of sublobules e-b of zones C2 and D1 (as defined by inferior olivary labeling). It is significant that in the ipsilateral nucleus prepositus hypoglossi, two groups of cells projecting to zone C2 are arranged rostrocaudally between three other groups projecting to zone D1.

Projections of individual purkinje cells of identified zones in the flocculus to the vestibular and cerebellar nuclei in the rabbit

The rabbit flocculus can be divided into five zones (zones 1, 2, 3, 4, and C2) with the use of acetylcholinesterase histochemistry. The projections of individual Purkinje cells in these zones to the vestibular and cerebellar nuclei were studied by using biocytin as an anterograde tracer. The zones were physiologically identified in terms of the Purkinje cell complex spike modulation occurring in response to optokinetic stimulation. In zones 1 and 3 neurons respond best to rotation about a horizontal axis that is close to perpendicular to the ipsilateral anterior semicircular canal, whereas in zones 2 and 4 neurons respond best to rotation about the vertical axis. Complex spike activity in zone C2 is unresponsive to optokinetic stimulation. Collectively, Purkinje cells of zone 1 projected to the ventral dentate nucleus, dorsal group y, and superior vestibular nucleus; Purkinje cells of zones 2 and 4 projected to the magnocellular and parvicellular parts of the medial vestibular nucleus; Purkinje cells of zone 3 projected to dorsal group y, ventral group y, and the superior vestibular nucleus; and Purkinje cells of zone C2 projected to the interposed posterior nucleus and dorsal group y. Some of the labeled Purkinje cell axons branched and innervated two nuclei. Branching axons from zone 1 either innervated both the ventral dentate nucleus and the superior vestibular nucleus or both dorsal group y and the superior vestibular nucleus. Branching axons from zones 2 and 4 innervated both the magnocellular and the parvicellular parts of the medial vestibular nucleus. Branching axons from zone 3 innervated both dorsal group y and the superior vestibular nucleus, or both ventral group y and the superior vestibular nucleus. Branching axons from zone C2 innervated both the interposed posterior nucleus and dorsal group y. Some of the target nuclei of the floccular Purkinje cell axons (e.g., dorsal group y and interposed posterior nucleus) project to the part of the inferior olive that, in turn, projects to the corresponding floccular zone, thus completing a closed pathway consisting of the inferior olive, the cerebellar cortex, and the cerebellar and vestibular nuclei. Other target nuclei (e.g., superior vestibular nucleus and medial vestibular nucleus) do not project back to the olivary subnuclei that innervate the flocculus and are part of an open olivofloccular pathway. An individual Purkinje cell thus can innervate a nucleus in the closed pathway as well as a nucleus in the open pathway.(ABSTRACT TRUNCATED AT 400 WORDS)

Correspondence between climbing fibre input and motor output in eyeblink-related areas in cat cerebellar cortex.

The purpose of the present work was to identify sites in the cerebellar cortex which are likely to control eyeblink. This work was motivated by findings suggesting that the cerebellum is involved in the learning and/or performance of the classically conditioned eyeblink response. The identification was based on climbing fibre input to the cortex and on the effects of electrical stimulation of the cerebellar cortex in cats decerebrated rostral to the red nucleus. The cerebellar surface was searched for areas receiving short latency climbing fibre input on periorbital electrical stimulation. Four such areas were found in the c1 and c3 zones of lobules VI and VII in the anterior lobe of the cerebellum and in the c3 zone in the paramedian lobule. Electrical stimulation of the cerebellar cortex with trains (150-400 Hz) of at least 10 ms duration evoked two types of EMG response in the orbicularis oculi muscle. An early response, time-locked to the onset of the stimulation, was unrelated to climbing fibre input and a delayed response, time-locked to the termination of the stimulation, could only be evoked from areas which received short latency climbing fibre input from the eye, that is, the c1 and c3 zones. The delayed responses had long latencies (up to 50 ms) after the termination of the stimulus train and could be delayed further by prolonging the stimulation. Both types of response were abolished by injections of small amounts of lignocaine into the brachium conjunctivum. A number of characteristics of the delayed responses are described. They could be inhibited by a further shock to the same area of the cerebellar cortex. Their latency could be increased by increasing the stimulation frequency. The period between stimulation and appearance of the response often showed a decrease in spontaneous EMG activity. There was a close topographical correspondence between input and output. Delayed responses could be evoked from all four of the areas in the c1 and c3 zones which have climbing fibre input from the periorbital area. They could not be evoked from other areas. In contrast, early responses were only evoked from areas without such climbing fibre input. It is proposed that the delayed responses were generated by activation of Purkinje cell axons leading to hyperpolarization and a subsequent rebound depolarization and activation of cells in the interpositus nucleus. The cortical areas are therefore probably involved in the control of the orbicularis oculi muscle.(ABSTRACT TRUNCATED AT 400 WORDS)