Distribution of metabotropic glutamate receptor type 1a in Purkinje cell dendritic spines is independent of the presence of presynaptic parallel fibers

The metabotropic glutamate receptor type 1a (mGluR1a) is expressed at a high level in the molecular layer of the cerebellar cortex, where it is localized mostly in dendritic spines of Purkinje cells, innervated by parallel fibers. Treatment with methylazoxymethanol (MAM) of mouse pups at postnatal days (PND) 0 + 1 or 5 + 6 results in the partial loss of granule cells, the extent of which depends on the age of the animal at the time of injection. As a consequence of hypogranularity, the number of parallel fibers is decreased to such an amount that many of the postsynaptic Purkinje cell dendritic spines are devoid of axonal input, and only a limited number of spines participate in the formation of parallel fiber synapses, or, infrequently, in heterologous or heterotopic synapses with other presynaptic partners. At PND 30, 50% of the spines in the cerebella of mice treated with MAM at PND 0 + 1 was not contacted by any presynaptic element, compared to 5% in controls or 15% in the cerebella of mice treated with MAM at PND 5 + 6. The localization of mGluR1a was visualized by immunocytochemistry on ultrathin sections: approximately 80% of all Purkinje cell dendritic spines were immunopositive in controls and in both groups of MAM-treated mice, indicating that mGluR1a was present in Purkinje dendritic spines even when the corresponding synaptic input was absent. This observation indicates that the expression and subcellular distribution of mGluR1a are inherent, genetically determined properties of Purkinje cells. J. Neurosci. Res. 50:433–442, 1997. © 1997 Wiley-Liss, Inc.     

Morphological development of dendritic spines on rat cerebellar Purkinje cells

The posterior cerebellum is strongly involved in motor coordination and its maturation parallels the development of motor control. Climbing and mossy fibers from the spinal cord and inferior olivary complex, respectively, provide excitatory afferents to cerebellar Purkinje neurons. From post-natal day 19 climbing fibers form synapses with thorn-like spines located on the lower primary and secondary dendrites of Purkinje cells. By contrast, mossy fibers transmit synaptic information to Purkinje cells trans-synaptically through granule cells. This communication occurs via excitatory synapses between the parallel fibers of granule cells and spines on the upper dendritic branchlets of Purkinje neurons that are first evident at post-natal day 21. Dendritic spines influence the transmission of synaptic information through plastic changes in their distribution, density and geometric shape, which may be related to cerebellar maturation. Thus, spine density and shape was studied in the upper dendritic branchlets of rat Purkinje cells, at post-natal days 21, 30 and 90. At 90 days the number of thin, mushroom and thorn-like spines was greater than at 21 and 30 days, while the filopodia, stubby and wide spines diminished. Thin and mushroom spines are associated with increased synaptic strength, suggesting more efficient transmission of synaptic impulses than stubby or wide spines. Hence, the changes found suggest that the development of motor control may be closely linked to the distinct developmental patterns of dendritic spines on Purkinje cells, which has important implications for future studies of cerebellar dysfunctions.     

Immunocytochemical localization of the alpha 1A subunit of the P/Q-type calcium channel in the rat cerebellum

Among various types of low- and high-threshold calcium channels, the high voltage-activated P/Q-type channel is the most abundant in the cerebellum. These P/Q-type channels are involved in the regulation of neurotransmitter release and in the integration of dendritic inputs. We used an antibody specific for the alpha1A subunit of the P/Q-type channel in quantitative pre-embedding immunogold labelling combined with three-dimensional reconstruction to reveal the subcellular distribution of pre- and postsynaptic P/Q-type channels in the rat cerebellum. At the light microscopic level, immunoreactivity for the alpha1A protein was prevalent in the molecular layer, whereas immunostaining was moderate in the somata of Purkinje cells and weak in the granule cell layer. At the electron microscopic level, the most intense immunoreactivity for the alpha1A subunit was found in the presynaptic active zone of parallel fibre varicosities. The dendritic spines of Purkinje cells were also strongly labelled with the highest density of immunoparticles detected within 180 nm from the edge of the asymmetrical parallel fibre-Purkinje cell synapses. By contrast, the immunolabelling was sparse in climbing fibre varicosities and axon terminals of GABAergic cells, and weak and diffuse in dendritic shafts of Purkinje cells. The association of the alpha1A subunit with the glutamatergic parallel fibre-Purkinje cell synapses suggests that presynaptic channels have a major role in the mediation of excitatory neurotransmission, whereas postsynaptic channels are likely to be involved in depolarization-induced generation of local calcium transients in Purkinje cells.     

F3/F11 cell surface molecule expression in the developing mouse cerebellum is polarized at synaptic sites and within granule cells.

The distribution of the F3/F11 neuronal cell surface molecule was investigated in the developing and adult mouse cerebellum by immunocytochemistry at the light and electron microscopic levels. F3/F11 was confined to subsets of neuronal types, since the Purkinje cell body and dendritic arborization as well as the stellate cells were not immunoreactive. In the young developing cerebellum, the granule cell axons strongly express F3/F11 as soon as they begin to grow, consistent with a functional role in promoting directional outgrowth of neuronal processes. In 10-d-old and adult cerebella, the granule cell bodies and dendrites were not immunoreactive whereas the parallel fibers, which are the granule cell axons, were labeled including in their presynaptic varicosities. By contrast, dendrites, cell bodies, and axons of Golgi cells were labeled by anti-F3 antibodies. Hence, F3/F11 can either be expressed throughout the cell or be polarized to the axons. This raises the question of how segregation of the glypiated F3/F11 molecule between different subcellular compartments depending on the type of neuron is achieved. F3/F11 was found to be present at three types of synaptic sites, suggesting that it might play a role in the formation and maintenance of synapses. However, in each type of synpase, F3/F11 was present at only the pre- or postsynaptic site, never at both: the parallel fiber varicosities contained F3/F11 whereas the postsynaptic compartment in contact, that is, the Purkinje cell dendritic spines, did not. The granule cell dendrites were unlabeled while the mossy fiber terminals contacting them were immunoreactive, and finally, the Golgi cell dendrites and dendritic spines were labeled while the presynaptic compartment contacting them was not. If F3/F11 functions as an adhesion molecule in vivo as indicated by in vitro assays, F3/F11-mediated adhesion is likely to be heterophilic.     

The development of synaptic contacts in the cerebellum of Macaca mulatta

The maturation of various cerebellar cortical cells, the appearance of afferent fibers to the cerebellum, and the development of synaptic contacts in the cerebellar cortex and deep nuclei was investigated in the fetal macaque. Ultrastructural studies were done on cerebellum obtained from fetuses at 75, 100, 125 and 150 days after conception to interrelate the temporal development of these three systems. At 75 days, synaptic contacts were seen on somas and axons of neurons in the deep cerebellar nuclei, and climbing fibers formed pericellular baskets around Purkinje cells. By 100 days the climbing fibers synapsed with somatic spines of the Purkinje cells, and mossy fiber endings were present in the internal granule cell layer. Synaptic contacts were also seen on dendritic processes of neurons in the deep cerebellar nuclei at this time. In the 125 day cerebellum, Golgi cells were identified for the first time and climbing fibers and parallel fibers made synaptic contact with both Purkinje and Golgi cells. At 150 days parallel fibers made synaptic contact with superficial stellate cells and mature cerebellar glomeruli had appeared. At this stage, axosomatic contacts of climbing fibers on the soma of Purkinje cells had disappeared. The relationship of these anatomical observations to possible functional activity is discussed.     

Purkinje cell abnormalities and synaptogenesis in genetically jaundiced rats (Gunn rats)

The course of cytological abnormalities and synaptogenesis of Purkinje cells were investigated in the culmen of cerebella from homozygous Gunn rats with hereditary hyperbilirubinemia from postnatal day 7 to adulthood (5-10 months old). The affected Purkinje cells were abundant at day 7. A large number of Purkinje cells reached the fully advanced stage of degeneration during the ensuring 16 days and disappeared between days 12 and 30. The Purkinje cells remaining at day 30 were less affected and recovered by the adult stage. Various abnormalities in Purkinje cell synaptogenesis with the parallel fibers, climbing fibers, and basket and stellate cell axons were observed. Primitive junctions between parallel fibers and Purkinje dendritic shafts were often found in adulthood. The parallel fiber boutons lacking postsynaptic partners and facing astrocytic processes were often noted from day 18 to adulthood. The persistence of such presynaptic elements suggests some mechanisms for stabilizing the synaptic elements once they have been formed. Many of the parallel fiber synaptic boutons with or without their postsynaptic partners were enlarged and were assumed to be transsynaptically affected by Purkinje cell damage. A number of climbing fiber synapses with perisomatic process of Purkinje cells, which are transient in normal synaptogenesis, were present at day 30 and a few of them were still found even in adulthood. Basket and stellate cell synapses were often found in abundance on the remaining Purkinje cells in adulthood, though they were not frequently encountered during the development period.     

Characterization of various nervous tissues of the chick embryos through responses to chronic application and immunocytochemistry of beta-bungarotoxin.

beta-Bungarotoxin (beta-BT) was applied to chick embryos at 3-day intervals beginning on the fourth day of incubation to investigate ultrastructurally the effects of chronically and massively applied beta-BT on various nervous tissues and muscles. On the twenty-first day of incubation, spinal cords of beta-BT treated embryos were conspicuously decreased in size. Ventral root fibers dorsal root fibers, white matter, and motor neurons disappeared. Although spinal ganglia and sympathetic trunk ganglia were completely absent, Auerbach's and Meissner's ganglia nerve cells in the small intestine and adrenal medullary cells were not affected. In retinas of beta-BT treated animals ganglion cells and optic nerve fibers disappeared, but photoreceptor cells, bipolar cells and horizontal cells remained intact. Furthermore, olfactory nerve cells and their unmyelinated nerve fibers ensheathed by Schwann cells were quite undamaged. Skeletal muscles degenerated, whereas cardiac muscles were unaffected. In the present study various nervous tissues of the twenty-first day normal chick embryos were incubated with beta-BT and target cells of beta-BT were detected directly by the reaction with horseradish peroxidase labelled anti beta-BT guinea pig IgG. Motor nerve cells in the spinal cords, spinal and sympathetic ganglia nerve cells, ganglion cells and some nerve cells at the inner part of the inner nuclear layer in the retinas were positively stained, whereas Auerbach's and Meissner's ganglia nerve cells in the small intestine, adrenal medullary cells, photoreceptor cells, bipolar cells and horizontal cells in the retina and olfactory nerve cells were negative. Thus the present study shows the beta-BT has extensive destructive effects on various nerve cells which were revealed to be target neurons of beta-BT by immunocytochemistry. Those nerve cells, affected by beta-BT and positively stained with immunocytochemical reaction were supposed to have different characteristics from unaffected cells. One of the differences between these affected cells and unaffected cells may be whether there exist binding sites for beta-BT on the plasma membrane or not. The possibility of the use of beta-BT to characterize various nervous tissues is presented in the present study.     

Plasticity of the parallel Fiber-Purkinje cell synapse by spine takeover and new synapse formation in the adult rat

Alteration in synaptic connectivity between Purkinje cell spines and parallel fibers of the cerebellum were studied following partial deafferentation of Purkinje cells in the adult rat. Transection of parallel fibers by two lesions placed at a 1 mm interval on the folial crest were used to produce degeneration of these afferents. Ultrastructural analysis of synapses on Purkinje cell spines revealed degeneration with vacating of postsynaptic sites within 6 h. Reactive synaptogenesis as takeover of Purkinje cell spines by formation of new synapses from remaining parallel fibers occurred even before degenerating parallel fibers had vacated postsynaptic sites. This was accompanied by a marked increase in the number of dual innervations by reactive parallel fibers within one day. Some vacated postsynaptic sites were lost as indicated by a reduction in the number of synapses and others may have been taken over by newly formed synapses on spines. In addition, new synapses formed between the shafts of Purkinje cell branchlets and parallel fibers. Sprouting of parallel fibers occurred as small extensions without tubules while Purkinje cell spines reacted by forming elongated and multiple heads which contacted different parallel fibers. After 5 days degenerating boutons were rarely found. Enlarged spine heads were each capped by a proportionally enlarged parallel fiber bouton and joined by an elongated synaptic junction to parallel fibers. Some parallel fiber boutons were greatly enlarged and capped numerous profiles of spines.

This study shows that formation of new pre- and postsynaptic sites takes precedence over reoccupation of original contacts and that multiple synapses on individual spines are being eliminated to give rise to single contacts with boutons. This elimination resulted in enlargement of synaptic contact areas between Purkinje cell spines and parallel fibers by taking over postsynaptic sites from some vacated and eliminated boutons.     

Electron microscopic analysis of postnatal histogenesis in the cerebellar cortex of staggerer mutant mice

Postnatal development of the cerebellar cortex has been compared in staggerer mutant and unaffected littermate mice. From postnatal day 3 to about day 21 the external granular layer in staggerer mice is decreased in thickness and area, and the number of postmitotic granule cell neurons is reduced. Those granule cells that are generated seem to differentiate normally, with the remarkable exception that they form only primitive junctions with Purkinje cell dendritic shafts. These specialized junctions are not superseded by the normal parallel fiber:Purkinje spine synapses and disappear by the third week. Purkinje cell somata and dendrites are smaller than normal at all stages examined. The dendrites are not confined to the sagittal plane as in the normal and, unique among mutant or other animals described to date, they exhibit virtually no branchlet spines. All other cortical synaptic relations of granule and Purkinje cells, including climbing fiber:Purkinje spine synapses, appear qualitatively normal. However, by 28 days virtually all staggerer granule cells have degenerated. While the primary genetic defect remains unknown, we postulate that the morphological abnormalities may be attributable to a block in the normal developmental relationship between granule cells and Purkinje cells. The small cell size and failure to form branchlet spines suggest that the Purkinje cell abnormality may be closer to the primary effect of the mutant gene than the more flagrant hypoplasia and degeneration of granule cell neurons.     

Altered intracellular localization of the glutamate receptor channel δ2 subunit in weaver and reeler Purkinje cells

The glutamate receptor (GluR) channel δ2 subunit is expressed abundantly and specifically in cerebellar Purkinje cells. Our previous study demonstrated that the GluR δ2 mRNA is expressed as early as embryonic day 15 prior to Purkinje cell synaptogenesis, and its protein product accumulates in dendritic spines during normal Purkinje cell maturation. In this study, we examined expression and distribution of the GluR δ2 in the weaver and reeler mutant cerebelli, which show abnormal cytoarchitecture and neural circuitry. In situ hybridization analysis showed that GluR δ2 mRNA was expressed in entire Purkinje cells in both mutant mice. Immunohistochemical analysis revealed that intracellular localization of GluR δ2 was altered in some region of mutant cerebelli. In the cortical surface where Purkinje cells form synapses with parallel fibers, GluR δ2-immunoreactivity was restricted to dendritic spines of Purkinje cells as observed in normal mice. In contrast, in the subcortical region where granule cells and parallel fibers are absent, the immunoreactivity was found widely in Purkinje dendrites. Thus, the GluR δ2 protein did not accumulate to the dendritic spines of Purkinje cells lacking synaptic contact with parallel fibers. These results suggest that the expression of both GluR δ2 mRNA and protein is independent of abnormalities in the mutant cerebelli, but relocalization of the GluR δ2 protein might depend on the formation of synapses between Purkinje cells and parallel fibers.     

Organization of cerebellar cortex secondary to deficit of granule cells in weaver mutant mice

This study deals with some consequences of the early postnatal abnormalities of cerebellar Bergmann glial fibers and granule cell neurons. (1) Cerebellar size is mildly reduced in heterozygous weaver (+/wv) mice and markedly reduced in homozygotes (wv/wv), but the pattern of fissures is essentially normal. Comparison with other mutants displaying small cerebella suggests that cell proliferation rate in the external granular layer is a key deteminant of cerebellar cortical folding. (2) Mossy fiber terminals differentiate on schedule despite the reduced number and abnormal positions of granule cells. However, many of them enter the modified molecular layer, and as noted especially in noninbred wv/wv mice one to two years old, form synapses with dendrites of aberrant granule cells. Where granule cells are absent, mossy fibers form more than the normal number of synapses with dendrites of Golgi type II neurons. (3) Purkinje cells are only mildly affected by the disorder of neighbouring cells. Their dendrites grow abnormally into the territory occupied by external granule cells, reach the external surface, and may turn inward. They form few tertiary branches. Dendritic spines are present in profusion and show membrane thickenings akin to normal postsynaptic elements. Although they receive no axonal contacts, the spines persist, enveloped by glial processes, for at least two years. Apart from the absence of parallel fiber contacts, afferent and intrinsic axons form the normal classes of synaptic connections with Purkinje cells. (4) Interneurons of the molecular layer are generated on schedule. At the time of their earliest recognition, they reside in the external granular layer, where they receive synaptic contacts from climbing fibers and other interneurons. In the absence of parallel fibers, interneurons differentiate in situ but their dendrites are abortive and randomly oriented. Growth of their dendrites, in contrast to that of Purkinje cell dendrites, appears to be markedly influenced by the organization of the local cellular milieu.     

The cerebellum of the frog Rana ridibunda. An electron microscopic study

An electron microscopic study of neuronal types and different synaptic contacts has been made in the cerebellum of the frog Rana ridibunda. The Purkinje cells have a pear-shaped cell body and in their cytoplasm the organelles show a special arrangement because of the great amount of microtubules they contain. The granule cells are small, rounded neurons with a large nucleus surrounded by a thin rim of cytoplasm. The stellate cells are interneurons of the molecular layer whose large nuclei show a single finger-like invagination of its nuclear envelope. The afferent tracts to the cerebellum end either as climbing fibers or mossy fibers. The axon terminals of climbing fibers are large and the synaptic complexes exhibit all the features of a type-I Gray synapse. The mossy fibers reach the granular layer and synapses between them and granule cell dendrites are by far the most abundant. The parallel fibers establish synaptic contacts on the spines arising from the spiny branchlet units of the Purkinje cells and with the perikaryon and dendrites of stellate cells. The stellate cell axons cross the molecular layer and establish type-II Gray synapses on the Purkinje cells.     

Granule cells migrate within raphes in the developing cerebellum: an evolutionarily conserved morphogenic event.

The early phase of granule cell migration in the developing chick cerebellum occurs within ribbons of cells moving through parasagittally arrayed gaps between Purkinje cell clusters. These parasagittal arrays of migrating granule cells, termed "granule cell raphes," also have been reported in rabbit and cat, but recent publications variously report that granule cell raphes are absent or present in rodents. By using Nissl counterstaining and Pax6 immunohistochemistry, we confirm that granule cells do migrate in raphes in the developing mouse cerebellum, and also in the primate cerebellum during a period of development that coincides with Purkinje cell compartmentation. In mouse and primate cerebellum, as in chick cerebellum, granule cell migratory streams occur at the borders of Purkinje cell clusters. GFAP immunostaining of Bergmann glial fibers shows no parasagittally localized pattern of distribution, indicating that the formation of granule cell ribbons is not prepatterned by heterogeneous distribution of radial glia. The conservation of the ribboned pattern of granule cell migration from bird to primate and the timing of this event suggest a possible role for granule cell raphes in parasagittal compartmentation of Purkinje cells. A potential mechanism for such an interaction is discussed.     

Developmental profile of inositol 1,4,5-trisphosphate 3-kinase in rat cerebellar cortex: light and electron microscopic immunohistochemical studies

Developmental expression and intracellular distribution of inositol 1,4,5-trisphosphate 3-kinase in the rat cerebellar cortex were studied immunohistochemically. Immunoreactivity appeared first at postnatal day 1 in the rostral region of the cerebellum and by day 15 had extended throughout the whole cerebellum, being localized in the Purkinje cell layer. Shortly after the expression of the enzyme in each Purkinje cell, the labelling showed a tendency to accumulate in the dendrites in a fine granular pattern. Electron microscopy revealed that immunoreactivity was present in the Purkinje dendritic trunks with accentuation in the distal segments during the early postnatal period, thereafter becoming concentrated in the dendritic spines at later developmental stages. Labelling was associated mainly with the plasmalemma, including the postsynaptic densities and open coated vesicles, and the subplasmalemmal vesicles of the smooth endoplasmic reticulum. Immunoreactivity was also evident in the perisomatic processes of immature Purkinje cells, which are transient projections synapsing with climbing fibers. In developing Purkinje axons, immunoreactivity was accentuated in the distal segments, associated with the plasmalemma and synaptic vesicles. These results suggest that inositol 1,4,5-trisphosphate 3-kinase is involved in the dendritic arborization and subsequent spine synaptogenesis of Purkinje cells, and that the developing presynaptic nerve endings of these cells are another functional site for the enzyme.     

Experimental reorganization of the cerebellar cortex. II. Effects of elimination of most microneurons with prolonged x-irradiation started at four days

The heads of Long-Evans rats were irradiated from the fourth day after birth with a schedule of repeated doses of low-level x-ray which essentially prevented the formation of basket, stellate and granule cells in all except the earliest-forming lobules (nodulus and uvula). The morphogenic and synaptogenic effects of this treatment were examined with light and electron microscopy in 30 day old animals, with particular attention paid to the pyramis. Although when irradiation was started the Purkinje cells formed a monolayer and had upward oriented apical poles, they became scattered later and had randomly oriented dendrites. This secondary disorientation was attributed to insufficient space available in the arrested cerebellum for the rapidly expanding Purkinje cells. Although basket cell terminals were scarce, basket cell-like terminals were formed on the somata of Purkinje cells, apparently by recurrent axon collaterals of these cells. The most common synapses with the thorns of Purkinje dendrites were formed by climbing fibers but other elements, including glial processes, were also in contact with postsynaptic loci on the thorns. Many mossy fiber terminals reached the surface. Where parallel fibers were present they were often thicker than in unirradiated animals and contained neurofilaments. No pathological changes were seen in these cerebella, with the possible exception of excessive lobulation of the nuclei of many Purkinje cells.     

Primary degeneration of the granular layer of the cerebellum (Norman type)

Summary Purkinje cells, impregnated with the rapid Golgi method, in a patient with primary degeneration of the granular layer showed abnormal orientation of the perikaryon and dendrites, reduction in size of the dendritic arbor, absence of spiny branchlets, and large numbers of stubby spines and hypertrophic spines on secondary dendritic branches; stubby spines and thorn-like formations were seldom observed on the primary dendrites and perikaryon of some Purkinje cells. These findings are similar to those described in the cerebellum of the homozygous weaver mutant mouse and in the cerebella of experimentally induced agranular phenocopies, thus suggesting that similar plastic changes occur in human and animal Purkinje cells as a result of the absence of parallel fibres input in early developmental stages. In addition, Purkinje cells in this patient showed club-shaped deformities in the distal region of primary dendrites, which were filled with radially oriented, short dendrites covered with stubby spines and hypertrophic spines. These latter structures appear to be fully impregnated asteroid bodies observed in paraffin sections.     

Anatomical, physiological and biochemical studies of the cerebellum from mutant mice. II. Morphological study of cerebellar cortical neurons and circuits in the weaver mouse.

The vermis of the homozygous weaver mice has been examined with Golgi and electron microscopic techniques. In addition to the findings already reported by previous authors 12, 29, new cytological features concerning all the cerebellar neuronal types and the synaptic reorganization of the cerebellar circuitry are described. As in other agranular cerebella, Purkinje cells do not develop spiny branchlets and have a randomly oriented dendritic tree. By contrast, their thick dendrites are studded with spines; according to their size and shape these were classified into: (a) small stubby spines which are the normal postsynaptic targets for climbing fibers; (b) tertiary-like spines, most of which are free of axonal contacts; (c) dolichoderus spines; (d) branching spines; and (e) hypertrophic spines. The last 3 types do not exist in normal cerebellum. Postsynaptic-like differentiations are frequently undercoating the smooth surface of the Purkinje dendrites. As it happens in the case of the free spines, free postsynaptic sites in the shafts of the dendrites develop an extracellular material similar to the material present in synaptic clefts. Basket and stellate cells also develop postsynaptic-like differentiations undercoating the somatic and dendritic plasma membranes. These free postsynaptic sites can reach a gigantic size, being longer than 3 mum in length. The rare postmigrative granule cells which persist in wv exhibit claw-endings not only at the dendritc terminal segments, but at the proximal dendritic stems as well. Some of these granule cells, besides having fully achieved migration, undergo a degenerative process indicating that they are probably directly affected by the mutation. Concerning the cerebellar circuitry, and despite the great number of free postsynaptic sites, the large majority of the synaptic contacts keep their specificity. However, some quantitative variations have been disclosed. The surface density of climbing varicosities is increased, whereas that of mossy rosettes is decreased. Stellate and basket fibers are present and their density also decreased. Furthermore, the pinceau formation around the initial segment of the Purkinje cell axon is missing. In addition to all normal synapt iccontacts (with the exception of the'parallel fiber-omnicellularsystem') present in weaver, heterologous synapses have also been encountered, mainly concerning the Purkinje dendritic spines, which can be contacted by mossy rosettes, granule cell bodies and/or dendrites. Morphological signs of partial innervation of the free postsynaptic sites on the smooth surface of Purknje dendrites and the perikarya and dendrites of interneurons have also been observed. These results confirm the existence of synaptic remodeling in wv cerebellum     

Autoradiographic visualization of a calcium channel antagonist, [125I]omega-conotoxin GVIA, binding site in the brains of normal and cerebellar mutant mice (pcd and weaver)

An in vitro autoradiographic technique has been used to localize [125I]omega-conotoxin GVIA binding sites in the brains of normal and cerebellar mutant mice. In the brains of normal mice, the highest densities of binding sites were observed at glomeruli of the olfactory bulb, cerebral cortex, caudate nucleus-putamen, hippocampus, and the nucleus of the solitary tract. Moderate densities of the silver grains occurred on the granular layer of the olfactory bulb, the molecular layer of the dentate gyrus, the molecular layer of the cerebellum, and the cochlear nucleus. No specific binding appeared in the white matter or the deep nucleus of the cerebellum, the corpus callosum, the internal capsule and the external plexiform layer of the olfactory bulb. Autoradiographic studies of the cerebella of Purkinje cell degeneration (pcd) mice showed that the distribution of binding sites on the molecular layer of the cerebellum are not affected by the degeneration of Purkinje cells. However, only background levels of the silver grains occurred on the cerebella of agranular weaver mutant mice, suggesting that the receptors for omega-conotoxin GVIA in the cerebellum are predominantly distributed on the parallel fibers of granule cells.     

Postnatal maturation of rat Purkinje cells cultivated in the absence of two afferent systems: an ultrastructural study.

Organized cultures of newborn rat cerebellum were established in Maximow chambers in order to study the maturation of Purkinje cells in absence of afferent systems. In the first model, standard cultures were devoid of extracerebellar afferents mossy and climbing fibers. Despite this absence, somatic spines appeared upon Purkinje cells during the first week in vitro and maturation proceeded normally except for the almost absence of spiny branchlets. Large dendritic trunks were studded with numerous spines, some of which were naked, a few bearing isolated post-synaptic densities and others occupied by boutons of parallel fibers. Stellate and basket axons made synapses upon the smooth portions of dendrites and soma. In a second model, the cultures were fed the antimitotic drug methylazoxymethanol (MAM) to prevent multiplication of granule cell precursors. Despite the absence of climbing and parallel fibers, the elongation of Purkinje dendrites was not prevented, but again the dendritic arbor consisted of large trunks studded with spines; somatic as well as dendritic spines were contacted by large boutons identified as Purkinje recurrent collaterals (PRC). It is concluded that the Purkinje cell possesses a large autonomy from afferent systems as to the growth of soma and dendrites. Conversely, the geometry of the dendrite and especially the spiny branchlets depend on the presence of both climbing and parallel fibers. One may conclude from the above experiments that specificity of synaptic contacts is maintained as long as postsynaptic sites are not devoid of their normal afferents. Heterologous synapses are formed when postsynaptic sites are present, their normal afferents absent and aberrant ones increasing by collateral sprouting. Such is probably the case in the second model of this study.     

Presynaptic plasticity at cerebellar parallel fiber terminals

The cerebellum plays a role in the control of sensorimotor functions and possibly also of higher cognitive processing. The granule cells, which are abundant and unique in their characteristic dendritic morphology, allow the cerebellum to combine the advantages of sparse coding with a high sensitivity for individual afferents at the input stage. Plastic changes in the granular layer circuitry may thus control instant transformation of inputs as well as long-term modifications so as to support procedural memory formation. Over recent decades, substantial research has been done to explore the mechanisms of postsynaptic changes that may sustain learning processes in the cerebellum, especially bidirectional plasticity at the parallel fiber to Purkinje cell synapse. In contrast, the presynaptic occurrence of synaptic plasticity has been relatively neglected. Here we review the current models of granular layer processing in the framework of cerebellar functioning with special emphasis on the presynaptic modulations of operations at the parallel fiber to Purkinje cell synapse. We argue that the wide range of possible mechanisms that can strengthen the parallel fiber to Purkinje cell synapse at the presynaptic level endows the cerebellar cortex with optimal computational capacities to potentiate both spatial and temporal cues that are relevant for fine-regulating memory formation.