Cellular localization of the Ca2+-binding protein parvalbumin in the developing avian cerebellum

Summary The appearance and distribution of the calciumbinding protein parvalbumin was investigated immunocytochemically at different postnatal developmental stages of the zebra finch cerebellum. Purkinje, basket and stellate, but not granule neurons or glial cells were labeled by an antiserum against chicken parvalbumin. At all developmental stages investigated immunostained Purkinje cells were found in clusters separated by spaces containing unstained large cells, probably Purkinje and Golgi type-II cells, and unstained smaller cells resembling granule neurons. Perisomatic processes, dendrites and spines of Purkinje cells were heavily immunoreactive. Axons of Purkinje cells were observed to be parvalbumin-positive throughout their entire length until developmental stage D 24, i.e., 10 days after hatching. Their immunoreactivity gradually decreased up to adulthood, when only their proximal portions, in addition to a few punctate structures in the internal granular layer and in the deep cerebellar nuclei presumably representing the synaptic terminals, remained immunoreactive. This decrease in immunoreactivity might be related to progressive maturation and/or degree of myelination. The developmental expression of parvalbumin immunoreactivity and its ultrastructural localization in spines, postsynaptic densities and on microtubular elements leads to several suggestions concerning the possible function of parvalbumin in neurons. In outgrowing dendrites and axons the protein might be involved in the regulation of the synthesis of membrane components, their intracellular transport and fusion of new membrane components into the plasmalemma, events that are Ca- and/or Mg-dependent. In spines and postsynaptic densities parvalbumin might be involved in the development and regulation of synaptic activities in Ca-spiking elements such as the inhibitory Purkinje cells, and possibly also in stellate and basket cells. Furthermore, in developing and adult neurons parvalbumin might be involved in the Ca-/Mg-regulation of a variety of enzymatic activities and hence influence the alteration of the intracellular metabolic potential in response to extracellular signals.This work was supported by the Deutsche Forschungsgemeinschaft, SPP Verhaltensontogenie and by the Swiss National Science Foundation, Grant No. 3.185-0.82     

Norepinephrine-dependent and independent mechanisms of persistent effects of amphetamine in rat cerebellum

Previous studies of the effects of chronic low-dose amphetamine (2 mg/kg per day X 21 days) on the spontaneous discharge rate of cerebellar Purkinje neurons have shown persistent depressant effects for up to 50 days after cessation of drug administration. The depression of spontaneous discharge observed was only partially reversible by various pharmacological agents which disrupt noradrenergic neurotransmission in cerebellum. In the present study, several additional approaches were used to investigate further this persistent effect. Rats were treated, either before or after chronic treatment with amphetamine, with intracisternal 6-hydroxydopamine at doses which destroy most noradrenergic fibers in cerebellum. In either case Purkinje neurons were still significantly slowed after cessation of amphetamine treatment, although the depression was not as great as previously observed. In another experiment, cerebellar cortical levels of 3-methoxy, 4-hydroxy phenyl glycol (MHPG) were measured after cessation of amphetamine administration, to determine if there was biochemical evidence for increased noradrenergic neurotransmission. At ten days, MHPG levels were elevated by 36%, and they returned to control values by 30 days. The evidence obtained in these studies suggests that chronic amphetamine treatment causes a persistent increase in noradrenergic neurotransmission, but non-noradrenergic mechanisms may also be important mechanisms in the long-lasting depression of activity of cerebellar Purkinje neurons.     

ATM, the Mre11/Rad50/Nbs1 complex, and topoisomerase I are concentrated in the nucleus of Purkinje neurons in the juvenile human brain

The genetic disease ataxia telangiectasia (AT) results from mutations in the ataxia telangiectasia mutated (ATM) gene. AT patients develop a progressive degeneration of cerebellar Purkinje neurons. Surprisingly, while ATM plays a criticial role in the cellular reponse to DNA damage, previous studies have localized ATM to the cytoplasm of rodent and human Purkinje neurons. Here we show that ATM is primarily localized to the nucleus in cerebellar Purkinje neurons in postmortem human brain tissue samples, although some light cytoplasmic ATM staining was also observed. No ATM staining was observed in brain tissue samples from AT patients, verifying the specificity of the antibody. We also found that antibodies against components of the Mre11/Rad50/Nbs1 (MRN) complex showed strong staining in Purkinje cell nuclei. However, while ATM is present in both the nucleoplasm and nucleolus, MRN proteins are excluded from the nucleolus. We also observed very high levels of topoisomerase 1 (TOP1) in the nucleus, and specifically the nucleolus, of human Purkinje neurons. Our results have direct implications for understanding the mechanisms of neurodegeneration in AT and AT-like disorder.     

Electrophysiological analysis of the functional organization of cortico-cerebellar connections

The effect of stimulation of cortical association (orbito-frontal, parietal) and projection (auditory, sensomotor) areas on the activity of Purkinje neurons of the cerebellar cortex was studied in adult cats anesthetized with pentobarbital, with or without chloralose. These responses were compared with those to peripheral stimuli. Definite similarity was found between the responses of Purkinje cells to different cortical (association and projection) stimuli as regards both the types of responses of the neurons and their ability to respond. No similarity was observed in the responses of Purkinje cells to peripheral (visual, auditory, electrodermal) stimulation. Whereas almost identical numbers of neurons (over 50%) were excited in response to the different forms of cortical stimulation, the ability of the neurons to respond to peripheral stimuli differed considerably: 44.6% of neurons responded to electrodermal stimulation, 34.2% to auditory, and 18.8% to visual.Medical Institute, Kemerovo. Translated from Neirofiziologiya, Vol. 8, No. 5, pp. 483–489, September–October, 1976.     

Increase in the efficacy of synapses between parallel fibers and Purkinje cells after joint stimulation of climbing and parallel fibers in the frog

A study was made of the susceptibility of Purkinje cells to long-term plasticity changes produced by joint stimulation of two inputs: the parallel and the climbing fibers. Experiments were conducted on a preparation of isolated frog cerebellum, joined to the medulla by one peduncle. A total of 18 neurons were investigated which showed a monosynaptic response to stimulation of the parallel fibers and maintained stable background activity over a 2 h period. Curves were plotted throughout this time for the likelihood of a reaction occurring in Purkinje cells in response to stimulation of the parallel fibers. Level of current required to stimulate a Purkinje cell firing index of 0.5 (I_0.5) was calculated. Neurons in which compound response to the "climber" type had been produced by stimulating the medulla showed a I_0.5 of 0.7 (less than one unit) at the start and finish of experiments, which would suggest an increase in the efficacy of the synapses of parallel fibers in Purkinje cells when parallel and climbing fibers are stimulated simultaneously.Institute of Problems in Information Transmission, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 19, No. 2, pp. 156–164, March–April, 1987.     

Effect of alkylating drugs on rat cerebellum

Ultrastructural and histochemical studies were performed on Wistar rats treated during 6 days with Cyclophosphamide 50 mg/kg/day or Lomustine 20 mg/kg/day. Changes in cerebellum after both drugs were similar. Most severe ultrastructural lesions were found in glia, mainly astroglia cells localized in perivascular area. Cytoplasm of these cells was on the large area devoid of organelles. Neurons were less affected but in Purkinje cells mild ultrastructural lesions were found. Increased activity of glucose-6-phosphatase and decreased of succinate dehydrogenase activity in these cells suggest that cytostatic treatment disturbed metabolism in the neurons.     

Neurophysiological mechanisms in the genetics of ethanol sensitivity

Abstract

Cerebellar Purkinje neurons of long‐sleep (LS) mice express a higher sensitivity than do those of short‐sleep (SS) mice to the depressant effects of ethanol in situ, in vitro, and in intraocular cerebellar brain grafts. The ethanol sensitivity of Purkinje neurons is intrinsic to the cerebellum, may be associated with only certain brain areas, and shows a high genetic correlation with the behavioral sensitivity of mice to ethanol‐induced ataxia. Tolerance develops to the depressant effects of ethanol on cerebellar neurons in both lines of mice. However, ethanol‐tolerant LS mice are more sensitive to the electrophysiological effects of ethanol on Purkinje neurons than are ethanol‐tolerant SS mice. In addition, the behavioral sensitivity to this drug probably also involves noncerebellar neurons since neonatally cerebellectomized LS and SS mice retain a different sensitivity to the ataxic effects of ethanol.     

Ankyrin-based subcellular gradient of neurofascin, an immunoglobulin family protein, directs GABAergic innervation at purkinje axon initial segment

Distinct classes of GABAergic synapses are segregated into subcellular domains (i.e., dendrite, soma, and axon initial segment-AIS), thereby differentially regulating the input, integration, and output of principal neurons. In cerebellum, for example, basket interneurons make exquisitely precise "pinceau synapses" on AIS of Purkinje neurons, but the underlying mechanism is unknown. Using BAC transgenic reporter mice, we found that basket axons always contacted Purkinje soma before innervating AIS. This synapse targeting process followed the establishment of a subcellular gradient of neurofascin186 (NF186), an L1 family immunoglobulin cell adhesion molecule (L1CAM), along the Purkinje AIS-soma axis. This gradient was dependent on ankyrinG, an AIS-restricted membrane adaptor protein that recruits NF186. In the absence of neurofascin gradient, basket axons lost directional growth along Purkinje neurons and precisely followed NF186 to ectopic locations. Disruption of NF186-ankyrinG interactions at AIS reduced pinceau synapse formation. These results implicate ankyrin-based localization of L1CAMs in subcellular organization of GABAergic synapses.     

DNA CONTENT OF PURIFIED PREPARATIONS OF MOUSE PURKINJE NEURONS ISOLATED BY A VELOCITY SEDIMENTATION TECHNIQUE

Abstract— The DNA content of mouse Purkinje neurons was investigated employing a biochemical approach. Material for the biochemical assay was provided by means of a sedimentation velocity separation technique which yields bulk quantities of well-preserved Purkinje perikarya in a high degree of purity. The same amount of DNA/cell was recorded for mixed cerebellar cell somata (7·6 ± 0±2 pg/cell), as for the Purkinje perikarya enriched fractions (7±2 & 0·2 pg/cell). No evidence could be found for the existence of a tetraploid DNA complement in mouse Purkinje neurons despite indications to the contrary from a parallel cytophotometric study.     

Chromatin organization in frog Purkinje neurons during the annual cycle: cytochemical and ultrastructural studies

Feulgen-DNA microdensitometric and propidium Iodide-DNA (PI-DNA) microfluorometric evaluations were made in Purkinje cells of cerebella from hibernating frogs and from active frogs. Higher Feulgen-DNA/distribution area ratios and lower PI-DNA values were found in the hibernating frog (both at high and low fluorochrome concentrations). These data indicate a higher degree of chromatin compactness. During the activity period, the Feulgen-DNA/distribution area ratios are lower and the Feulgen-DNA content is higher; the corresponding higher PI-DNA values indicate a lower degree of chromatin condensation. Histochemical and ultrastructural data show clearly different distribution patterns of chromatin and ribosomes in the two periods. The overall results underscore lower activity of neurons and a greater homogeneity within the Purkinje cell population during hibernation.     

Resurgent current and voltage sensor trapping enhanced activation by a beta-scorpion toxin solely in Nav1.6 channel. Significance in mice Purkinje neurons

Resurgent currents are functionally crucial in sustaining the high frequency firing of cerebellar Purkinje neurons expressing Na(v)1.6 channels. Beta-scorpion toxins, such as CssIV, induce a left shift in the voltage-dependent activation of Na(v)1.2 channels by "trapping" the IIS4 voltage sensor segment. We found that the dangerous Cn2 beta-scorpion peptide induces both the left shift voltage-dependent activation and a transient resurgent current only in human Na(v)1.6 channels (among 1.1-1.7), whereas CssIV did not induce the resurgent current. Cn2 also produced both actions in mouse Purkinje cells. These findings suggest that only distinct beta-toxins produce resurgent currents. We suggest that the novel and unique selectivity of Cn2 could make it a model drug to replace deep brain stimulation of the subthalamic nucleus in patients with Parkinson disease.     

Characterization of the Responses of Purkinje Cells to Neurotrophin Treatment

Abstract: The ability of the neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4/5 (NT-4/5) to promote neuronal survival and phenotypic differentiation was examined in dissociated cultures from embryonic day 16 rat cerebellum. BDNF treatment increased the survival of neuron-specific enolase-immunopositive cells by 250 and 400% after 8 and 10 days in culture, respectively. A subpopulation of these neurons, the Purkinje cells, identified by calbindin staining, was increased to an equivalent extent, ∼200%, following BDNF, NT-4/5, or NT-3 treatment. The number of GABAergic neurons, identified by GABA immunoreactivity, was greatly increased by treatment with BDNF (470%) and moderately by NT-4/5 (46%), whereas NT-3 was without effect. NGF failed to increase the number of either Purkinje cells or GABAergic neurons. Addition of BDNF within 48 h of cell plating was required to obtain a maximal increase in Purkinje cell number after 8 days. In contrast, the NT-3 responses were nearly equivalent even if treatment was delayed for 96 h after plating. BDNF, NT-4/5, and NT-3, but not NGF, induced the rapid expression of the immediate early gene c- fos . Immunocytochemical double-labeling with antibodies to c-fos and calbindin was used to identify Purkinje cells that responded to neurotrophin treatment by induction of c-fos. After 4 days in vitro, both BDNF and NT-3 induced the formation of c-fos protein in calbindin-immunopositive neurons, whereas NT-4/5 did not. The latter results suggest that although BDNF and NT-4/5 have been shown to act through a common receptor, TrkB, it appears that the effects of BDNF and NT-4/5 are not identical.     

Localization of 4.1 related proteins in cerebellar neurons

Localization of 4.1 related proteins in neurons was studied with immunofluorescence microscopy and with immunoelectron microscopy on ultrathin cryosections. In rat cerebellum, 4.1 immunoreactive proteins were demonstrated in Purkinje cell bodies, dendrites and other neurons in the cerebellar cortex. Some glial cells showed staining, but no labeling was found in myelinated axons of the white matter and of the glomeruli in the granule cell layer. At the ultrastructural level, the 4.1 related proteins were localized mainly in the cytoplasmic matrix, while some labeling was found underneath the plasma membrane. To determine whether 4.1 related proteins in neuronal cytoplasm exist as part of the cytoskeleton or not, PC12 cells cultured in the presence of nerve growth factor were stained with the anti-4.1 antibody. Since cytoplasmic staining was retained after detergent treatment, the 4.1 related proteins seem to exist as a component of the neural cell cytoskeleton. Localization of 4.1 related proteins during the postnatal development of the cerebellum was also studied. In Purkinje cells, localization of 4.1 related proteins changed according to the stages of the postnatal development. The present data suggest that 4.1 related proteins in neurons localized mainly in the cytoplasm and may play some role in organizing cytoskeletal networks in the cytomatrix. Their distribution is developmentally regulated in some neurons, possibly in relationship to their maturation in the cytoskeleton.     

Rapid Ca2+-dependent increase in oxygen consumption by mitochondria in single mammalian central neurons

Oxygen consumption increases within a fraction of a second after the onset of neuronal activity, a phenomenon referred to as the "initial dip" in functional imaging studies of the living brain. The cellular mechanism that underlies this rapid increase in oxygen consumption has remained unclear, however. We have now used two-photon excitation imaging to characterize rapid activity-dependent mitochondrial responses in single neurons. This approach allowed simultaneous multicolor imaging of individual mitochondria in single mouse Purkinje neurons in culture. Mitochondrial depolarization was induced immediately when the cytosolic free Ca(2+) concentration ([Ca(2+)](i)) exceeded 15 microM and was associated with oxidation of mitochondrial NAD(P)H, suggesting that Ca(2+)-induced mitochondrial depolarization mediated by the Ca(2+) uniporter directly facilitated oxidation of NAD(P)H. With the use of a miniature oxygen electrode, we detected a burst of oxygen consumption within 0.2s after the onset of cell depolarization in single Purkinje neurons, and this rapid increase in oxygen consumption was dependent on the increase in [Ca(2+)](i). We have thus demonstrated a rapid Ca(2+)-dependent consumption of oxygen that is mediated by mitochondrial depolarization in mammalian central neurons. This process might function as a rapid feed-forward mechanism in homeostatic control of the cytosolic ATP concentration.     

Increased number of nitric oxide synthase immunoreactive Purkinje cells and dentate nucleus neurons in schizophrenia

There is growing interest in the cerebellum as a site of neuropathological changes in schizophrenia. Reports showing that schizophrenics have higher nitric oxide synthase (NOS) activity and MAPKinase levels in the vermis, point to possible aberrations in the cerebellar signal transduction of schizophrenics. It has been speculated that Ca²⁺-dependent extracellular to intracellular signal transduction may be disrupted in the cerebellum of schizophrenics. We decided to test this hypothesis by studying the nitrergic system and markers of the Ca²⁺-triggered signal cascade in the cerebellum of schizophrenics, depressives and controls. The cellular distribution of two calcium sensor proteins (VILIP-1 and VILIP-3) and of neuronal NOS immunoreactivity was studied morphometrically in the flocculonodulus, the inferior vermis and the dentate nucleus of 9 schizophrenics, 7 depressive patients and 9 matched controls. In comparison to controls and depressed patients there were fewer Nissl-stained neurons in the dentate nucleus of schizophrenics. The number of NOS-expressing Purkinje neurons was however strongly increased. In the flocculonodulus and the vermis no differences between the groups were found with regard to the density of Nissl-stained Purkinje cells. The number of NOS-expressing Purkinje neurons was increased in schizophrenics, however. No differences between schizophrenics, depressives and controls were found in the number of VILIP-1 immunoreactive dentate nucleus neurons and VILIP-3 immunoreactive vermal and flocculonodular Purkinje cells. Our data provide further histochemical evidence in favor of structural abnormalities in discrete cerebellar regions of schizophrenics. They confirm and extend earlier reports of increased cerebellar NOS immunoreactivity in schizophrenia and point to possible neurodevelopmental disturbances. Our failure to show an altered expression of two calcium sensor proteins possibly points to a less important role of calcium signaling in cerebellar pathology of the disease.     

Cell-type specific proximity of centromeric domains of one homologue each of chromosomes 2 and 11 in nuclei of cerebellar Purkinje neurons

In Purkinje neurons of the mouse cerebellum, the centromeres of several chromosomes are placed in close proximity to form a distinct pattern of clusters and exhibit reproducible spatial redistributions during development. In granule neurons, an adjacent cell type in the cerebellum, the pattern, size, and number of centromeric aggregations are different from those of Purkinje neurons. The present work was undertaken to test the hypothesis that the same chromosomes form part of one aggregate in a cell-type-specific manner. Fluorescence in situ hybridization (FISH) with chromosome-specific paracentromeric probes was used to identify centromeric regions of individual chromosomes in cerebellar Purkinje and granule neurons of the adult mouse. When pairs of centromeric probes were used in two-color FISH, one homologue each of chromosomes 2 and 11 were routinely found close to each other in Purkinje neurons but not in granule neurons. This finding of specific proximity was limited to the pair 2 and 11, out of the ten chromosome pairs that were randomly selected and studied. Our results indicate that, in adult Purkinje neurons, a cell-type-specific spatial proximity is present between centromeric domains of one homologue each of chromosomes 2 and 11.     

Chronic ethanol exposure enhances AMPA-elicited Ca2+ signals in the somatic and dendritic regions of cerebellar Purkinje neurons.

Intracellular Ca2+ signals produced by the glutamate receptor agonist alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA; 5 microM) were measured in the somatic and dendritic regions of cerebellar Purkinje neurons in mature cerebellar control cultures (> or = 20 days in vitro) and cultures chronically treated with 32 mM ethanol (146 mg%; 8-11 days). Recordings were made in physiological saline without ethanol. The mean peak amplitude of the Ca2+ signal elicited by AMPA (applied by brief 1-s microperfusion) in the somatic region was enhanced 38% in chronic ethanol-treated Purkinje neurons compared with control neurons. In contrast, Ca2+ signals evoked by AMPA in the dendritic region were similar in magnitude between control and chronic ethanol-treated Purkinje neurons. When tetrodotoxin (TTX; 500 nM) was included in the bath saline to block spike activity and synaptically-generated events, the mean peak amplitude of the Ca2+ signal elicited by AMPA was enhanced 60% in both the somatic and dendritic regions of chronic ethanol-treated Purkinje neurons compared with control neurons. Thus, TTX-sensitive mechanisms (i.e., spike or synaptic activity) appear to play a role in normalizing neuronal functions involved in Ca2+ signaling in the chronic ethanol-treated neurons. In parallel current clamp experiments, the resting membrane potential of chronic ethanol-treated neurons was slightly depolarized compared with control neurons. However, no differences were found between control and chronic ethanol-treated Purkinje neurons in input resistance or the peak amplitude or duration of the depolarizations or hyperpolarizations elicited by AMPA. AMPA receptors mediate fast excitatory neurotransmission in the majority of neurons in the central nervous system (CNS) and Ca2+ signals in response to AMPA receptor activation contribute to synaptic function. Thus, our results suggest that modulation of Ca2+ signals to AMPA receptor activation (or other cellular inputs) may provide an important mechanism contributing to the actions of prolonged ethanol exposure in the CNS.     

Activity of cat cerebellar neurons in penicillin epilepsy and amphetamine treatment.

The discharge pattern of cerebellar Purkinje cells and fastigial neurons was studied after acute amphetamine treatment in immobilized cats, as well as during generalized penicillin-induced epilepsy. There was a marked reversible decrease in spontaneous firing rate of Purkinje cells and an increase in spontaneous firing rate of fastigial neurons after acute d-1 amphetamine administration (5 mg/kg, s.c.). The discharge pattern of Purkinje cells showed tendency towards inhibition, while the fastigial neurons showed less clear tendency towards disinhibition in the course of epilepsy induced by parenteral administration of penicillin (400.000-500.000 I.U./kg, i.m.). Moreover, acute amphetamine treatment (5 mg/kg, s.c.) performed after the development of penicillin-induced epileptic episodes elicited a prominent suppression of Purkinje cell discharges associated with a parallel increase in discharges of fastigial neurons. These results suggest that the changes in discharge rate of cerebellar corticonuclear neurons induced by amphetamine contribute to suppression of seizural activity in the feline model of generalized epilepsy.     

The distribution of GABA-like-immunoreactive neurons in the brain of the newt,Triturus cristatus carnifex,and the green frog,Rana esculenta

Summary The distribution of -aminobutyric acid (GABA) immunoreactivity was studied in the brain of two amphibian species (Triturus cristatus carnifex, Urodela; Rana esculenta, Anura) by employing a specific GABA antiserum. A noteworthy immunoreactive neuronal system was found in the telencephalic dorsal and medial pallium (primordium pallii dorsalis and primordium hippocampi) and in the olfactory bulbs. In the diencephalic habenular nuclei there was a rich GABAergic innervation, and immunoreactive neurons were observed in the dorsal thalamus. In the hypothalamus the GABA immunoreactivity was found in the preoptic area, the paraventricular organ and in the hypothalamo-hypophysial complex. In the preoptic area of the frog some GABA-immunoreactive CSF-contacting cells were shown. In the optic tectum immunolabeled neurons were present in all the cellular layers. A rich GABAergic innervation characterized both the fibrous layers of the tectum and the neuropil of the tegmentum and interpeduncular nucleus. In the cerebellum, in addition to the Purkinje cells showing a variable immunopositivity, some immunoreactive cell bodies appeared in the central grey. Abundant immunolabeled nerve fibers in the acoustico-lateral area and some immunopositive neurons in the region of the raphe nucleus were observed. In conclusion, the GABAergic central systems, well-developed in the amphibian species studied, were generally characterized by close similarities to the pattern described in mammals.Dedicated to Professor Valdo Mazzi (Dipartimento di Biologia Animale, Università di Torino), in honor of his 70th birthday     

Formation of new synaptic contacts by Purkinje axon collaterals in the granular layer of deafferented cerebellar cortex of adult rat

In addition to (i) mossy terminals, (ii) Golgi axons, (iii) granule cell dendrites and (iv), occasionally, Golgi cell dendrites, a third axonal profile identified by morphological criteria as the collateral of Purkinje axons, has been found in 2% of all cerebellar glomeruli. These infrequent components of a few glomeruli, however, were never seen in normal cerebellar cortex to establish specialized synaptic contact with glomerular dendrites. Two to four weeks after surgical isolation of the cerebellar cortex, i.e. following the destruction of both efferent and afferent fibres, the number of glomeruli containing (hypertrophic) axonal branches of Purkinje cells has increased to 13% of all surveyed glomeruli. In addition, the Purkinje axon terminals in the mossy fibre-deprived glomeruli were observed to establish numerous Gray II-type synaptic contacts with surrounding granule cell dendrites. It is suggested that the development of heterologous synapses between hypertrophic, or even intact, Purkinje axon collaterals on the one hand and the mossy fibre-vacated granule cell dendrites on the other, is a compensatory, reactive process to the synaptic "desaturation" of granule neurons, which demonstrate a dormant potential of Purkinje cells to form new synaptic contacts in the adult cerebellum.