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PubMed1988

Ultrastructure of synapses in the cerebellar cortex after long-term activation of climbing fibres.

Adult male rats were treated for 24 or 48 h with harmaline which selectively activates neurones in the inferior olivary nucleus which give rise to climbing fibres projecting to the cerebellar vermis. Electrophysiological studies have shown that harmaline-induced climbing fibre activity completely blocks the responses of the Purkinje cell to parallel fibre input. Morphometric analysis of the ultrastructure of climbing and parallel fibre synapses revealed no significant differences in morphology between vermis (experimental) and hemisphere (control). These findings indicate that the decreased responsiveness of Purkinje cells to parallel fibre inputs induced by increased climbing fibre activity over 24 or 48 h is not accompanied by any observable structural changes in the cerebellar cortex.

Action Potentials↗
PubMed2004

Localization of synapsin-I and PSD-95 in developing postnatal rat cerebellar cortex.

The comparative localization of two prominent synaptic proteins, synapsin-I (Syn-I) and PSD-95, was investigated in slices of developing (P3-P21) rat cerebellar cortex using double- or triple-label fluorescence immunohistochemistry and confocal microscopy. During the first postnatal week, Syn-I and PSD-95 immunoreactive (IR) puncta were strongly concentrated in the Purkinje cell layer (PCL) where they circumscribed irregularly shaped PC somata, forming pericellular nests that likely correspond to early climbing fiber synapses. PSD-95 and Syn-I puncta also were found along the shafts and at the tips of growing PC dendrite branches labeled with calbindin. During the second postnatal week, synaptic puncta were lost from the PC layer, while many new puncta were added to the molecular layer (ML). At P10, about half of the PCs were circumscribed by PSD-95 or Syn-I puncta, whereas at P14 no PCs were circumscribed. By P14, PSD-95 and Syn-I became most strongly localized to many small puncta in the ML and to large clusters at mossy fiber rosettes in the glomerular layer (GL) where PSD-95 often encircled Syn-I clusters. Some large clusters in the GL contained only PSD-95 or Syn-I, but not both, suggesting differential growth or remodeling of pre- and post-synaptic structures. No PSD-95 staining of pre-synaptic terminal pinceau was observed during the first 3 weeks of postnatal development. Thus, in relation to PCs, there is a developmental shift in PSD-95 localization whereby, first, it is concentrated on PC cell bodies and short dendrites (P3-P7), then it is lost on PC cell bodies (P7-14) and becomes localized almost exclusively to PC dendrites (P14-P21).

Animals↗
PubMed1979

Lipofuscin in the cerebellar cortex of albino rats: an electron microscopic study.

The ultrastructure of autofluorescent, PAS-positive lipofuscin in Purkinje, granule, Golgi epithelial, basket and stellate, microglial and perivascular cells in the cerebellar cortex of senescent rats is described. The membrane-bounded pigment is composed of three elements: 1) electron-lucent homogeneous droplets, 2) a granular matrix and 3) intensely osmiophilic patches. The proportions of these three components vary between cell types and one can grossly differentiate a neuronal and a glial lipofuscin. The lipofuscin granules of stellate and perivascular cells are different from lipofuscin of other cerebellar neurons and glia. It can be concluded from these morphological observations that each cerebellar cell type has its distinct lipofuscin.

Aging↗
PubMed1993

Neuronal differentiation rescued by implantation of Weaver granule cell precursors into wild-type cerebellar cortex.

The migration of postmitotic neurons away from compact, germinal zones is a critical step in neuronal differentiation in the developing brain. To study the molecular signals necessary for cerebellar granule cell migration in situ, precursor cells from the neurological mutant mouse weaver, an animal with phenotypic defects in migration, were implanted into the external germinal layer (EGL) of wild-type cerebellar cortex. In this region, labeled weaver precursor cells of the EGL progressed through all stages of granule neuron differentiation, including the extension of parallel fibers, migration through the molecular and Purkinje cell layers, positioning in the internal granule cell layer, and extension of dendrites. Thus, the weaver gene acts nonautonomously in vivo, and local cell interactions may induce early steps in neuronal differentiation that are required for granule cell migration.

Animals↗
PubMed1989

Physiological and anatomical studies of the interactions between Purkinje cells and basket cells in the cat's cerebellar cortex: evidence for a unitary relationship.

Intracellular recordings have been obtained from neurons in lobule V of the cat's vermis, which were identified as basket cells following intracellular injections of HRP. Stimulation of the inferior cerebellar peduncle or peripheral nerves elicited an initial depolarizing and subsequent hyperpolarizing response. Neither potential could be graded with changes in stimulus intensity; both displayed all-or-none properties at threshold levels of stimulation. The depolarization and hyperpolarization were confirmed as an excitatory postsynaptic potential and an inhibitory postsynaptic potential (IPSP), respectively, on the basis of their response to intracellular injections of hyperpolarizing and depolarizing currents into the cell body. A possible source of the unitary IPSP is the Purkinje cell, via its recurrent axonal collaterals. To test this hypothesis, an electron microscopic analysis was carried out to define the synaptic relationships between the recurrent collaterals of an HRP-filled Purkinje cell and 3 basket cells. Serial section analysis reveals that collaterals from a single Purkinje cell contact several basket cells, but each basket cell received somatic input from only one Purkinje cell. These data provide an anatomical substrate for the unitary IPSP observed during intracellular recording from basket cells. The unitary nature of the Purkinje cell-basket cell interaction indicates that a very limited population of cortical neurons may be involved in local circuits that integrate afferent information in the cerebellar cortex.

Animals↗
PubMed1987

[Chromatin matrix activity in Purkinje cells and granular cells of the rat cerebellar cortex during postnatal differentiation].

Moore's method used for the examination of chromatin template activity in Purkinje and granule cells of 7, 14, 30 days and 3 months old rate cerebellar cortex has shown the age-dependent changes during differentiation period. The histograms for Purkinje cells have demonstrated that all neurons were distributed into 3 groups of activity according to their nuclear labelling. The cell percentage in each group varied during ontogenesis.

Aging↗
PubMed1979

Projections from the spinal and the principal sensory nuclei of the trigeminal nerve to the cerebellar cortex in the cat, as studied by retrograde transport of horseradish peroxidase.

Projections from the spinal (Vsp) and the principle sensory (Vp) nuclei of the trigeminal nerve to the cerebellar cortex were studied by means of retrograde transport of horseradish peroxidase in the cat. Neurons projecting to the simple lobule and the dorsal part of the paramedian lobule (PMD) were located mainly in the dorsal part of the nucleus interpolaris (Vi) and of the caudal one third of the nucleus oralis (Vo) and in the rostralmost part of the Vo. Neurons projecting to the medial part of the posterior folia of crus II (crus IIp) were located in the dorsal to ventral parts of the Vi and of the caudal one third of the Vo and in the rostralmost part of the Vo, while those projecting to the lateral part of crus IIp were confined to the ventral part of the Vi and of the caudal one third of the Vo. Neurons of the Vp also projected to all of these cortical areas. They were relatively confined to the ventral part of this nucleus. Thest trigeminocerebellar projections were exclusively ipsilateral to the cell origin. There were sparse projections from the Vi and Vo to lobules V to VIIIa. In addition, a small group of neurons in the subnucleus magnocellularis of the nucleus caudalis of the Vsp also projected to the above cortical areas. No projections were, however, observed to the anterior portion of the anterior lobe, crus I, the anterior folia of crus II, paraflocculus, flocculus and the ventral part of the PMD. The majority of these cerebellar projection neurons were medium-sized and triangular, fusiform or ovoid in shape. There were small neurons of similar types and large multipolar neurons as well.

Animals↗
PubMed1995

Cerebellar cortex and eyeblink conditioning: bilateral regulation of conditioned responses.

We examined the role of the cerebellum in classical conditioning of the nictitating membrane response (NMR) of rabbits by comparing the effects of unilateral and bilateral cerebellar cortical lesions. Using extended preoperative conditioning to ensure high levels of learning, we confirmed that unilateral lesions of lobules HVI and ansiform lobe impaired conditioned responses (CRs) previously established to an auditory conditioned stimulus, but did not prevent some relearning with post-operative retraining. Bilateral lesions of HVI and ansiform lobe produced similar impairments of CRs, but also prevented subsequent relearning. Unilateral cortical lesions produced significant enhancement of unconditioned response (UR) amplitudes to periorbital electrical stimulation. Bilateral cortical lesions enhanced UR amplitudes to a lesser extent. Because there was no correlation between the degree of CR impairment and UR enhancement across the unilateral and bilateral lesion groups, the suggestion that the lesions impaired CRs due to general effects upon performance, rather than due to losses of learning, is not supported. Both sides of the cerebellar cortex contribute towards learning a unilaterally trained CR. This finding is important for the re-interpretation of unilateral, reversible inactivation studies that have found no involvement of the cerebellar deep nuclei in the acquisition of NMR conditioning. In addition, we found conditioning-dependent modifications of unconditioned responses that were particularly apparent at low intensities of periorbital electrical stimulation. This finding is important for the re-interpretation of studies that have found apparent changes in the UR of conditioned subjects after cerebellar lesions.

Animals↗
PubMed1993

Effects of recurrent collateral inhibition on Purkinje cell activity in the immature rat cerebellar cortex--an in vivo electrophysiological study.

We present here a study on the effects of inhibitory recurrent collaterals of Purkinje cell (PC) axons on the activity of the immature rat cerebellar cortex. Simultaneous extracellular recordings of pairs of PCs were performed in rat pups aged 5-8 days postnatal. Bicuculline was applied to the surface of the cortex in order to functionally antagonize PC recurrent collaterals. At this early developmental stage these are the only inhibitory links in the network. Dye marks from the microelectrode tips and 3D serial-section reconstruction of the structure allowed the exact determination of the distance separating recorded cells and of their respective orientation in the cortex. Standard statistical tests and an informational entropy index were used to calculate levels of cooperativity. By comparing PC activity under control conditions and after bicuculline superfusion it is shown that recurrent collateral inhibition has a structurating effect on the PC activity and that it increases the informational content of the network. Inhibition decreases the activity of the cells by 35% and drastically changes the interspike interval histograms. This leads to a more constrained state of the system. Three types of coupling via recurrent collaterals are present: symmetrical, asymmetrical or non existent. The exact type of coupling follows a simple vicinity rule and strongly influences the cooperativity level between the recorded cells. This cooperativity was also found to be spatially compartmentalized. Several pairs were driven by common inputs via climbing fibers or parallel fibers. Using the predictive value of a theoretical model of this immature structure we propose a complementary explanation of the role of the recurrent collaterals at this stage of development: that of a spatial and temporal filter, specific to each different microzone.

Action Potentials↗
PubMed1976

Aspartate aminotransferase activity and glutamic dehydrogenase in the cerebellar cortex in several species of animals. A histochemical study.

The histochemical study of the consumption of glutamic acid by way of the aspartate aminotransferase and the glutamic dehydrogenase in the cerebellar cortex of several species of animals have demonstrated that in that nerve centre exists some structures in which the mentioned consumption is specially or exclusively realized by means of one way and not for other different one. Is observed, as well, that in the rats, chicken and lizard, the baskets that surround the Purkinje cells are constituted by basket cells axons and by recurrent collaterals of Purkinje axons and that those structures have an intense aspartate aminotransferase activity, but not glutamic dehydrogenase. The aspartate aminotransferase activity was not observed on the other side, in the perikarya of the Purkinje cells of the related animals. However, there exists intense glutamic dehydrogenase activity. On the other hand, in the toad was not observed baskets with aspartate aminotransferase activity but this enzyme was presented on the other side in the perikarya of the Purkinje cells. All these observations have suggested the possibility that this special utilization of the glutamic acid is in some way concerned with the transmission phenomenons of the nerve impulse.

Animals↗
PubMed1997

Therapeutic motor training increases parallel fiber synapse number per Purkinje neuron in cerebellar cortex of rats given postnatal binge alcohol exposure: preliminary report.

Because therapeutic approaches to fetal alcohol effects in humans have been rare, this study explored the rehabilitative effect of complex motor training on an animal model of binge drinking in the third trimester of human pregnancy. Neonatal alcohol exposure induces significant and permanent reductions in Purkinje and granule cell number accompanied by impaired motor behavior in rats. The purpose of this study was to determine: (1) whether the motor skill impairment caused by exposure to alcohol in the early postnatal period could be ameliorated by the learning of a set of complex motor tasks that had been demonstrated to cause synaptogenesis in the cerebellar cortex; and (2) the extent to which cerebellar neurons in alcohol-exposed (AE) rats exhibit synaptic plasticity. The AE group was given 4.5 g/kg/day of ethanol from postnatal days 4 to 9 via an artificial rearing procedure producing a mean peak blood alcohol level of 257 mg/dl. Control groups consisted of a gastrostomy control (GC) group, that received an isocaloric mixture of maltose/dextrin instead of ethanol, and a suckle control (SC) group, that was reared normally by dams. At approximately 6 months of age, animals from the three groups were assigned either to a rehabilitation condition (RC; that received 10 days of training on the motor tasks) or to an inactive condition (IC; where rats stayed in isolation in their cages). Although SC rats were significantly faster to complete the course in the first 5 days of training, there were no differences in ability to perform among animals from all three groups-SC, GC, and AE--at the end of the training period. Unbiased stereological techniques were used to obtain estimates of the number of parallel fiber synapses/Purkinje cell within the cerebellar paramedian lobule. Results showed that the RC rats from the SC and AE groups had significantly more synapses/Purkinje cell than corresponding IC animals. These data demonstrate that rehabilitative intervention (complex motor training) can improve motor performance impaired by postnatal alcohol exposure and that surviving Purkinje neurons retain the capacity for synaptic plasticity.

Alcoholic Intoxication↗
PubMed1996

Morphology of the Golgi-impregnated Lugaro cell in the rat cerebellar cortex: a reappraisal with a description of its axon.

We present a detailed description of the somatodendritic and axonal features of the Golgi-impregnated Lugaro cell in the rat cerebellar cortex. This neuron, present throughout the cerebellum, is characterized by a fusiform cell body located at the border between the granular and Purkinje cell layers, horizontal bipolar dendrites extending in the parasagittal plane and an axon projecting exclusively to the molecular and granular layers. A quantitative analysis of 77 Lugaro cells confirms the somatodendritic homogeneity of this cell type. Computer reconstructions showed that the bipolar dendrites spread like an horizontal X centered by the cell body, implying that Lugaro cells are arranged in a flat lattice just beneath the Purkinje cell layer. Fifty-seven Lugaro cells with sufficiently impregnated axons displayed a common pattern of organization with, nevertheless, a certain variability of the axonal trajectory. Some axons projected directly into the molecular layer with a profuse plexus spreading just above the parent cell. Other axons first traveled downwards into the granular layer, sometimes even reaching and passing into the white matter. They always hooked back to reascend through the granular layer and also finally terminate in the molecular layer, near and sometimes at a distance from the parent cell. In a few samples, one or two remote axon collaterals were seen to extend longitudinally in the lower molecular layer for a few hundred microns, in the same direction as the parallel fibers. In all cases a few collaterals projected into the granular layer. In view of its dense afferentation by Purkinje cell recurrent collaterals and its profuse inhibitory projection in the molecular layer, the Lugaro cell could act as a feedback interneuron on the corticocerebellar output.

Animals↗
PubMed1994

Quantitative ultrastructural localization of glutamate dehydrogenase in the rat cerebellar cortex.

Glutamate dehydrogenase is one of the main enzymes involved in the formation and metabolism of the neurotransmitter glutamate. In the present study we investigated the enzyme ultrastructurally in the cerebellar cortex, a region rich in well defined glutamatergic neurons, by pre-embedding immunocytochemical staining (peroxidase-antiperoxidase), as well as by post-embedding immunogold labelling employing a new system for quantitation and for specificity testing under the conditions of the immunocytochemical procedure. A new antiserum against immunologically purified bovine liver glutamate dehydrogenase or antibodies isolated from this by affinity chromatography were used in rats fixed by perfusion with aldehydes. The pre-embedding method displayed peroxidase reaction preferentially in mitochondria of astroglial cells (including the Bergmann glia). Mitochondria of neuronal tissue elements were usually free of peroxidase-reaction product. Extra-mitochondrial staining was not observed. The post-embedding immunogold method was employed to overcome penetration problems and allow semiquantitative analysis of localization and specificity. The highest densities of gold particles were found over the mitochondria in astroglial cell elements (including the Bergmann glia). Mitochondria in cell bodies of Bergmann glia had a lower particle density than those in astrocytic processes. In the latter, analysis of frequency distribution revealed no evidence of a population of mitochondria lacking glutamate dehydrogenase, but suggested the presence of populations with different levels of immunoreactivity. Comparison with the labelling of embedded bovine liver glutamate dehydrogenase indicated that the enzyme constitutes a high proportion (10%) of the total matrix protein of these mitochondria. A weaker but significant labelling was found in oligodendrocytes of the white matter. The labelling of mitochondria in neuronal elements including glutamatergic mossy fibre terminals was of the order of 15% of that in astroglial mitochondria. No difference was detected between glutamatergic neurons (mossy and parallel fibres, granular cells) and non-glutamatergic neurons (Purkinje cells). The particle density over non-mitochondrial areas was very close to background over empty resin. The results, obtained with different methods of tissue and antibody preparation, agree to show that the present form of glutamate dehydrogenase is restricted to mitochondria and preferentially localized in astrocytes.

Animals↗
PubMed2001

Developmental expression of heterotrimeric G-proteins in the murine cerebellar cortex.

We utilized the murine cerebellum to analyze the expression of G-proteins during vertebrate neural differentiation. Combining reverse transcription-polymerase chain reaction and immunocytochemistry, we monitored the expression and cellular localization, within the nascent cerebellar cortex, of G-proteins subunits known to mediate signal transduction in the adult cerebellum. The mRNAs encoding subunits Galphaq, Galphao, Galphai-2, and Galphaz are expressed in the cerebellar anlage at least from embryonic day 14 onward, and relative levels of these mRNAs do not change appreciably from E14 to adulthood. Galphao, Galphaz, and Galphai-2 could be localized to granule cell neuroblasts and postmigratory, mature granule cells, but not to early postmitotic, premigratory, and migrating granule neurons. All of the Galpha subunits analyzed could also be localized to the cell somata of postmitotic Purkinje neurons, irrespective of age. In contrast, Purkinje cell dendrites stained for Galphao only up to postnatal day 8, dendritic immunoreactivity for Galphaz increased during dendritogenesis, and appreciable levels of Galphai-2 and Galphaq were seen in Purkinje cell dendrites only transiently during the 2nd and 3rd postnatal week. Of the G-beta and -gamma subunits analyzed (beta1, beta2, gamma2, gamma3, gamma5, and gamma7), only expression of gamma3 varied with development. It could be localized to Purkinje cell somata and dendrites in early postnatal, but not in adult animals. These changes in the cellular distribution and subcellular segregation of G-proteins are correlated to tangible aspects of cerebellar cortical histogenesis and suggest a role for G-protein-mediated signaling in their mechanistic implementation.

Age Factors↗