3D electron microscopic reconstruction of segments of rat cerebellar purkinje cell dendrites receiving ascending and parallel fiber granule cell synaptic inputs

Growing physiological evidence suggests that there are functional differences between synapses made by the ascending and parallel fiber segments of the granule axon on cerebellar Purkinje cells. Supporting this view, our previous electron microscopic studies suggested that these synapses also contacted different regions of the Purkinje cell dendrite, and in particular that ascending segment synapses are made exclusively on the smallest diameter Purkinje cell dendrites. In the current study we used serial electron microscopic techniques to reconstruct Purkinje cell dendritic segments up to almost 10 μm in length. Using a combination of anatomical and immunological labeling techniques we identified the ascending or parallel fiber origins of the excitatory synaptic inputs onto dendritic spines, as well as the location of inhibitory synapses made directly on the dendritic shaft. The results confirmed that there are regions of the Purkinje cell dendrite receiving exclusively ascending or parallel fiber synapses and that ascending segment synapses are only found on small‐diameter dendrites. In addition, we describe for the first time small‐diameter dendritic regions contacted by both types of excitatory synapses. While our data suggest that the majority of inhibitory inputs to the Purkinje cell tree are associated with parallel fiber synaptic inputs, we also found inhibitory inputs on dendritic regions with mixed ascending and parallel fiber inputs, or exclusively parallel fiber inputs. The finding that ascending and parallel fiber inputs can be segregated on the Purkinje cell dendritic tree provides further evidence that these excitatory granule cell synaptic inputs may be functionally distinct. J. Comp. Neurol. 514:583–594, 2009. © 2009 Wiley‐Liss, Inc.     

Depression of parallel and climbing fiber transmission to Bergmann glia is input specific and correlates with increased precision of synaptic transmission

In the cerebellar cortex, Bergmann glia enclose the synapses of both parallel and climbing fiber inputs to the Purkinje neuron. The glia express Ca(2+)-permeable AMPA receptors, and the GLAST and GLT-1 classes of glutamate transporter, which are activated by glutamate released during synaptic transmission. We have previously reported that parallel fiber to Bergmann glial transmission in rat cerebellar slices exhibits a form of frequency-dependent plasticity, namely long-term depression, following repetitive stimulation at 0.1-1 Hz. Here, we report that this form of plasticity is also present at the climbing fiber input, that climbing and parallel fibers can be depressed independently, that discrete parallel fiber inputs can also be depressed independently, and that depression is maintained when a distributed array of parallel fibers are stimulated (in contrast to several forms of synaptic plasticity at the Purkinje neuron). Depression of glutamate transporter currents does not correlate with a decrease in the stringency with which Purkinje neuron synapses are isolated. Rather, postsynaptic currents in Purkinje neurons decay more rapidly and perisynaptic metabotropic glutamate receptors are activated less effectively after stimulation at 0.2 and 1 Hz, suggesting that depression arises from a decrease in extrasynaptic glutamate concentration and not from impairment of glutamate clearance in and around the synapse. These results indicate that neuron-glial plasticity is activity dependent, input specific and does not require spillover between adjacent synapses to manifest. They also argue against a withdrawal of the glial sheath from synaptic regions as the putative mechanism of plasticity.     

Retrograde endocannabinoid signaling in the cerebellar cortex

The regulation of Purkinje cell activity is important for motor behavior and motor learning. As the sole output cell of the cerebellar cortex, Purkinje cell firing is controlled by parallel fibers and climbing fiber synapses, and by inhibitory interneurons. Depolarization of Purkinje cells evokes endocannabinoid release that activates cannabinoid CB1 receptors expressed on boutons of its synaptic inputs to transiently decrease neurotransmitter release. In addition, associative activation of the excitatory inputs can liberate endocannabinoids to decrease synaptic strength for a prolonged duration. Here we review the different mechanisms of evoking endocannabinoid release and discuss the physiological role of endocannabinoids in mediating global modulation of synaptic strength, localized short-term associative plasticity and cerebellar long term depression.     

Bidirectional plasticity at developing climbing fiber–Purkinje neuron synapses

Climbing fibers provide one of the two major excitatory inputs to the cerebellar cortex. In an immature animal, several climbing fibers form synapses with one Purkinje neuron. During postnatal development most climbing fiber innervations with a Purkinje neuron are eliminated and only one strong fiber remains. Previous studies suggested that this pruning of surplus climbing fiber innervations depends on the neuronal activity. We hypothesized that synaptic plasticity might play a role in the maturation and refinement of such a climbing fiber projection pattern, and examined the plasticity properties of synapses between postnatal days 5 and 9 in mice. We found that a 5 Hz conditioning stimulation of climbing fibers forming relatively strong synapses with a Purkinje neuron induced long-term potentiation of the transmission accompanied by a decrease in the paired-pulse ratio of excitatory postsynaptic current amplitudes. This was suggestive of an increased probability of presynaptic release. However, the conditioning stimulation of climbing fibers forming relatively weak synapses induced long-term depression and tended to increase the paired-pulse ratio. Thus, the direction of plasticity appears to be determined by the strength of synaptic connection. Long-term depression occurred only in the conditioned climbing fiber, whereas long-term potentiation spread to unconditioned climbing fibers. A postsynaptic increase in the intracellular Ca²⁺ concentration was required for long-term potentiation but not for long-term depression. These results reveal the existence of novel presynaptic plasticity at immature climbing fiber–Purkinje cell synapses, which may contribute to the maturation and refinement of the climbing fiber projection pattern.     

Ascending granule cell axon: an important component of cerebellar cortical circuitry.

Physiologic evidence suggests that local activation of the cerebellar granule cell layer produces a much more restricted spatial activation of overlying Purkinje cells than would be expected from the parallel fiber system. These results have led to the suggestion that synapses associated with the ascending granule cell axon may provide a large, direct, excitatory input to Purkinje cells, whereas parallel fiber synapses may be more modulatory in nature. In the current experiments, serial electron microscopy was used to reconstruct synapses associated with these two segments of the granule cell axons in the cerebellar cortex of albino rats. The results indicate that there are significantly more presynaptic vesicles in ascending segment synapses than in parallel fiber synapses. Furthermore, a first-order linear regression analysis revealed positive correlations between all measures of pre- and postsynaptic morphology for parallel fibers, but not for ascending segment synapses. Perhaps most surprisingly, serial reconstructions of postsynaptic spines and their associated dendrites demonstrated that spines contacted by ascending segment synapses are located exclusively on the smallest diameter distal regions of the Purkinje cell dendrites, whereas parallel fiber synapses are found exclusively on intermediate- and large-diameter regions of the spiny branchlets. Based on two independent calculations, we estimate that 20% of the granule cell synapses onto a Purkinje cell are actually made by the ascending segment. By using computer simulations of a single Purkinje cell dendrite, we have also demonstrated that synchronous activation of these distal ascending segment inputs could produce a substantial somatic response. Taken together, these results suggest that the two different regions of granule cell axons may play very different physiologic roles in cerebellar cortex.     

Activity-dependent axonal and synaptic plasticity in the cerebellum

The cerebellum is a brain region endowed with a high degree of plasticity also in adulthood. After damage or alteration in the patterns of activity, it is able to undergo remarkable changes in its architecture and to form new connections based upon a process of synaptic reorganization. This review addresses cellular and molecular mechanisms that regulate the competition between two inputs belonging to different neuronal populations in innervating two contiguous but separate domains of the same target cell. The two inputs are the parallel fibers, the axon of the cerebellar granule cells, and the olivocerebellar neurons, that terminate as climbing fibers in the cerebellar cortex. The target is the Purkinje cell characterized by two dendritic domains that are different in size and number of spines, upon which the two afferent inputs impinge. Both inputs express several genes related to plasticity throughout the life span conferring the ability to remodel their synapses. In addition, we provided evidence that climbing fibers and Purkinje cells show remarkable reciprocal trophic interactions that are required for the maintenance of the correct synaptic connectivity. Through their activity, climbing fibers sustain the competition with parallel fibers by displacing this input to the distal territory of the Purkinje cell dendrite. In addition, they operate on the Purkinje cells through AMPA receptor suppressing spines in the territory surrounding their synapses. In this way, climbing fibers are able to optimize spine distribution and functional connectivity.     

The role of calcium in synaptic plasticity and motor learning in the cerebellar cortex

The cerebellum is important for motor coordination, as well as motor learning and memories. Learning is believed to occur in the cerebellar cortex, in the form of synaptic plasticity. Central to motor learning theory are Purkinje cells (PCs), which are the sole output neurons of the cerebellar cortex. Motor memories are postulated to be stored in the form of long-term depression (LTD) at parallel fiber synapses with PCs, once thought to be the only plastic synapse in the cerebellar cortex. However, in the past few decades many studies have demonstrated that several other synapses in the cerebellar cortex are indeed plastic, and that LTD or long-term potentiation at these various synapses could affect the overall output signal of PCs from the cerebellar cortex. Almost all of these forms of synaptic plasticity are dependent on calcium to some extent. In the current review we discuss various types of synaptic plasticity in the cerebellar cortex and the role of calcium in these forms of plasticity.     

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.     

Diversity and Complexity of Roles of Granule Cells in the Cerebellar Cortex. Editorial

The cerebellar granule cell, the most numerous neurons in the brain, forms the main excitatory neuron of the cerebellar cortical circuitry. Granule cells are synaptically connected with both mossy fibers and Golgi cells inside specialized structures called glomeruli, and thereby, they are subject to both feed-forward and feed-back inhibition. Their unique architecture with about four dendrites and a single axon ascending in the cerebellar cortex to bifurcate into two parallel fibers making synapses with Purkinje neurons has attracted numerous scientists. Recent advances show that they are much more than just relays of mossy fibers. They perform diverse and complex transformations in the spatiotemporal domain. This special issue highlights novel avenues in our understanding of the roles of this key neuronal population of the cerebellar cortex, ranging from developmental up to physiological and pathological points of view.     

Frequency-dependent recruitment of inhibition mediated by stellate cells in the rat cerebellar cortex

In the cerebellum, dendritic inhibition of Purkinje cells (PCs) is mediated by stellate cells (SCs). These inhibitory interneurons are critically involved in the cerebellar network; they control the timing and firing frequency of PCs, the only output cells of the cerebellar cortex. However, the underlying properties of parallel fiber (PF) to SC excitatory synapses have not been fully determined. To characterize the conditions favoring the recruitment of SCs in the cerebellum, we analyzed evoked and spontaneous excitatory postsynaptic currents (EPSCs) recorded from SCs of rat cerebellar slices. We found that SC EPSCs evoked with single suprathreshold-intensity stimulations were mostly unitary, with a large amplitude and variable latencies, and failed with a high rate. Increasing the frequency of stimulation above 60 Hz significantly reduced failures, whereas mean SC EPSC amplitude was increased by less than 20%. Decreasing failures at PF-SC synapses experimentally enhanced the number of asynchronous SC EPSCs per stimulation but, again, moderately increased the mean SC EPSC amplitude. Finally, brief presynaptic bursts transiently depressed synaptic transmission. This depression resulted from the release of endocannabinoids and might act as a negative-feedback mechanism. Thus, we conclude that SC EPSCs evoked with single suprathreshold-intensity stimulations are mostly unitary and that PF-SC synapse efficacy is highly regulated by the presynaptic temporal pattern of activity and the frequency of afferent inputs. Such synaptic properties may control the responsiveness of SC synapses to the frequency of PF stimulations, which may control the spatial extent and duration of the recruitment of inhibition in the cerebellar cortex.     

Synaptic 5'-nucleotidase is transient and indicative of climbing fiber plasticity during the postnatal development of rat cerebellum

The transient appearance of 5'-nucleotidase, an adenosine-producing ecto-enzyme, was studied during specific stages of postnatal synaptogenesis in the rat cerebellum. For ultrastructural detection of 5'-nucleotidase activity, an enzyme-cytochemical technique was used. Between postnatal days 4 and 6, enzymatic reaction product was present in the synaptic clefts of climbing fibers containing the perisomatic spines, apical cones and emerging dendrites of Purkinje cells (CF-PC synapses). Labeled parallel fiber synapses were observed on dendritic shafts of cerebellar interneurons. At postnatal days 9 and 12, enzyme-positive parallel fiber terminals were in addition numerous on the spines of peripheral Purkinje branchlets, and gradually disappeared thereafter. Between postnatal days 8 and 15, labeling of perisomatic CF-PC contacts persisted. In contrast, climbing fiber synapses on Purkinje dendrites were only occasionally labeled. Between postnatal days 18 and 21, synaptic reaction product was restricted to mossy fibers. At the same time, association of 5'-nucleotidase with glial profiles was prominent throughout the cerebellar layers. In adult cerebellum (from 24 days onwards) all synapses were devoid of enzymatic activity. Throughout development, basket, stellate and Golgi cell synapses were devoid of enzymatic activity. We conclude that 5'-nucleotidase is present in excitatory cerebellar synapses during part of their generation period. The transient nature of this phenomenon suggests that 5'-nucleotidase may serve as a novel, cytochemical marker for a specific state of synaptic maturation, and in particular for climbing fiber plasticity. A role of 5'-nucleotidase in purinergic neuromodulation and cellular contact formation could be significant in these processes.     

The Purkinje neuron: II. Electron microscopic analysis of the mature Purkinje neuron in organotypic culture

Purkinje neurons of organotypic cultures were investigated electron microscopically following their analysis with the Golgi technique. The purpose of this study was to critically examine the issue of synaptic specificity in CNS cultures. The unique finding was the synaptic cluster, a terminal which engulfs many Purkinje spines. In the neuropil and on the major dendrites, this synaptic arrangement was interpreted to be a hypertrophic parallel fiber, representing a type of synaptic modulation. The terminals on the somatic spines are also in the form of clusters; some or all of these spines were thought to have developed to form synapses with the climbing fiber. In the absence of this afferent, the parallel fiber—a competing system—takes over the site. This represents a form of synaptic plasticity in these cultures. The inhibitory synaptic relationships were maintained on the soma and dendrites, but it was found that the basket synapses did not quantitatively encase the soma as is seen in the intact animal. Mossy-type terminals were found occasionally synapsing with Purkinje dendritic spines, as has been seen in agranular cerebellum in animals. These mossy terminals are presently thought to originate from the deep cerebellar nuclei within these cultures. Synaptic errors were rarely encountered. It is concluded that this preparation develops in accordance with established neurobiological principles, that the Purkinje neuron reaches a mature state in culture, and that this model has a sound anatomical basis for further experimental work.     

Number of parallel fiber synapses on an individual Purkinje cell in the cerebellum of the rat

In the present study, stereological techniques applied to electron micrographs of the molecular layer of the rat cerebellum have been used to estimate the number of parallel fiber synapses on the dendritic tree of a single Purkinje cell. Quantitative features of the parallel fiber to Purkinje cell dendritic spine synapses and of the parallel fibers were investigated as a preliminary to estimating the number of synapses. Parallel fiber to Purkinje cell synapses are flattened disclike structures with a mean axial ratio of 14.7 and a mean diameter of 319 microns in fixed tissue. The density of synapses in our fixed material was 8.17 x 10(8) per microliters of molecular layer. Determination of the length density of the synapses per unit area of micrograph indicated a synapse density of 8.03 x 10(8) per microliters. These densities give a total number of synapses per Purkinje cell of 1.74 x 10(5) and 1.71 x 10(5), respectively. Estimation of the number of parallel fiber varicosities and of varicosity length gave a density of 9.31 x 10(8) varicosities per microliters of molecular layer and determining the mean number of parallel fiber to Purkinje cell synapses per varicosity gave a synapse density of 9.82 x 10(8) per microliters, equivalent to 2.09 x 10(5) per Purkinje cell. The reasons why this estimate is likely to be too high are discussed. We conclude that there are some 175,000 parallel fiber synapses on an individual Purkinje cell dendritic tree in the cerebellar cortex of the rat, considerably more than previously reported.     

THE EFFECTS OF CARBON MONOXIDE ON NEURONS OF THE CEREBELLUM*

The effects of carbon monoxide on neurons of the cerebellum Acute CO exposure in rats produces multifocal edema and neuroglial swelling in cerebellar cortex and nuclei. Chronic multiple daily exposures to CO produce degenerative changes in neurons–shrinkage of cell somata and dendrites, enlargement of nuclei and nucleoli, vacuolation in neuronal cytoplasm, pyknosis and ultimate engulfment by microglia. Focal areas of Purkinje cell loss are typical; individual basket cells and their axons, stellate cells and the deep nuclear neurons are severely affected. Counts of Purkinje cells and their dendritic thorns show reductions of 30% and 31% respectively in affected regions. In foci of severe Purkinje cell loss, dendritic thorns of the neighboring cells which remain assume a greater number of synapses with unusual presynaptic partners, e. g. both parallel fibers and climbing fibers in a single thorn. An increase of 4.2% more synapses per thorn was measured when compared to the normal. Thus, the stress of chronic CO exposure stimulates synaptic plasticity or a compensatory remodelling of neuronal elements in the central nervous system. A scheme is proposed to summarize these changing relationships caused by the loss of some dendritic thorns and the migration of the presynaptic partners to other surviving thorns.     

The unipolar brush cell: a remarkable neuron finally receiving deserved attention

Unipolar brush cells (UBC) are small, glutamatergic neurons residing in the granular layer of the cerebellar cortex and the granule cell domain of the cochlear nuclear complex. Recent studies indicate that this neuronal class consists of three or more subsets characterized by distinct chemical phenotypes, as well as by intrinsic properties that may shape their synaptic responses and firing patterns. Yet, all UBCs have a unique morphology, as both the dendritic brush and the large endings of the axonal branches participate in the formation of glomeruli. Although UBCs and granule cells may share the same excitatory and inhibitory inputs, the two cell types are distinctively differentiated. Typically, whereas the granule cell has 4-5 dendrites that are innervated by different mossy fibers, and an axon that divides only once to form parallel fibers after ascending to the molecular layer, the UBC has but one short dendrite whose brush engages in synaptic contact with a single mossy fiber terminal, and an axon that branches locally in the granular layer; branches of UBC axons form a non-canonical, cortex-intrinsic category of mossy fibers synapsing with granule cells and other UBCs. This is thought to generate a feed-forward amplification of single mossy fiber afferent signals that would reach the overlying Purkinje cells via ascending granule cell axons and their parallel fibers. In sharp contrast to other classes of cerebellar neurons, UBCs are not distributed homogeneously across cerebellar lobules, and subsets of UBCs also show different, albeit overlapping, distributions. UBCs are conspicuously rare in the expansive lateral cerebellar areas targeted by the cortico-ponto-cerebellar pathway, while they are a constant component of the vermis and the flocculonodular lobe. The presence of UBCs in cerebellar regions involved in the sensorimotor processes that regulate body, head and eye position, as well as in regions of the cochlear nucleus that process sensorimotor information suggests a key role in these critical functions; it also invites further efforts to clarify the cellular biology of the UBCs and their specific functions in the neuronal microcircuits in which they are embedded. High density of UBCs in specific regions of the cerebellar cortex is a feature largely conserved across mammals and suggests an involvement of these neurons in fundamental aspects of the input/output organization as well as in clinical manifestation of focal cerebellar disease.     

Organizational principles and microcircuitry of the cerebellum

The cerebellum is known to influence motor behavior and to enable smooth, coordinated movements. Recent evidence also suggests that the cerebellum contributes to non-motor behavior, including components of cognition and regulation of affective state.This review summarizes the organization and circuitry of the cerebellum as a basis for understanding newly emerging concepts about the function of this neuronal system. The cerebellum consists of several divisions with separate functions. One region is associated with the vestibular system and another with brainstem and spinal cord. A third region, the cerebrocerebellum, has extensive interconnections with cerebral cortex and is likely to be involved in motor coordination and regulation of non-motor behavior. The cerebellar cortex is made up of radial modules of interconnected neurons. The Purkinje cell is the principle integrating neuron and focal point of each module. Other neuron types include the granule cell and three inhibitory interneurons. The Purkinje cell integrates excitatory inputs from climbing and parallel fibers, while its axon modulates activity of neurons in the deep nuclei, which represents the final outflow from cerebellum to other parts of the brain. Cerebellar circuitry exhibits a strong parasagittal organization based on climbing fiber input and the distributions of neuronal proteins and neuronal vulnerability to insults. The combination of this parasagittal circuitry with the mediolateral course of parallel fibers results in a Cartesian coordinate system which is likely to be a crucial factor in its signal processing function. Although numerous details of cerebellar microcircuitry, synaptic transmission and signal transduction have been determined, the functional contribution of cerebellar signalling to brain function remains highly enigmatic.     

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.     

Development of spontaneous and glutamate-evoked activity is altered by chronic ethanol in cultured cerebellar Purkinje neurons

The effects of continuous exposure to ethanol on the cytological and physiological development of a central nervous system (CNS) neuron were studied using the cultured Purkinje neuron of the rat cerebellar cortex. Purkinje neurons in fetal rat brain cultures which are established at one day before birth show development comparable to that described in vivo in other studies. In culture, Purkinje neurons progress from immature rounded cells with fine neurites to mature neurons with a branched dendritic structure. These structural changes are accompanied by an increase in the duration and complexity of the excitatory response to glutamate, by transitions in the patterns of spontaneous activity, and by an increase in mean firing rate. Our results demonstrate that chronic exposure to a low concentration of ethanol (90 mg%; 19.5 mM) during development selectively alters the electrophysiological but not the morphological properties of Purkinje neurons. Specifically, ethanol treatment reduces the responsiveness of these neurons to glutamate, delays the expected developmental transitions in patterns of spontaneous activity, and induces increased spontaneous bursting activity, particularly at the stage of dendritic formation. Impairment of responsiveness to glutamate is significant in that it may reflect the compromise by ethanol of a major excitatory pathway in the cerebellar cortex, resulting from the decreased efficacy of glutamatergic input from parallel fibers. In contrast to the results of other studies using adult neurons as a model for the effects of ethanol, our work suggests that the developing CNS neurons does not become tolerant; that is, in the continuing presence of ethanol, it does not express physiological function equivalent to that of the control.     

Differential neuronal and glial expression of GluR1 AMPA receptor subunit and the scaffolding proteins SAP97 and 4.1N during rat cerebellar development

In neurons, AMPA glutamate receptors are developmentally regulated and selectively targeted to synaptic sites. Astroglial cells also express AMPA receptors, but their developmental pattern of expression and targeting mechanisms are unknown. In this study we investigated by immunocytochemistry at the light and electron microscopy level the expression of GluR1 and its scaffolding proteins SAP97 (synapse-associated protein) and 4.1N during cerebellar development. In cerebellar cortex the GluR1 AMPA receptor subunit is expressed exclusively in Bergmann glia in the adult rodent. Interestingly, we observed that GluR1 was expressed postsynaptically at the climbing fibers (CF) synapse at early ages during Purkinje cell dendritic growth and before the complete ensheathment of CF/Purkinje cell synapses by Bergmann glia. However, its expression changed from neurons to Bergmann glia once these glial cells had completed their enwrapping process. In contrast, GluR2/3 and GluR4 AMPAR subunits were stably expressed in both Purkinje cells (GluR2/3) and Bergmann glia (GluR4) throughout postnatal development. Our data indicate that GluR1 expression undergoes a developmental switch from neurons to glia and that this appears to correlate with the degree of Purkinje cell dendritic growth and their enwrapping by Bergmann glia. SAP97 and 4.1N were developmentally regulated in the same pattern as GluR1. Therefore, SAP97 and 4.1N may play a role in the transport and insertion of GluR1 at CF/Purkinje cell synapses during early ages and at Bergmann glia plasma membrane in the adult. The parallel fiber (PF)/Purkinje cell synapse contained GluR2/3 but lacked GluR1, SAP97, and 4.1N at the time of PF synaptogenesis.     

Intraventricular administration of substance p increases the dendritic arborisation and the synaptic surfaces of Purkinje cells in rat's cerebellum

Substance P was infused in the lateral ventricles of twenty Lewis rats for twenty days. On the twentieth day the animals were sacrificed and the cerebellar cortex was processed for electron microscopy. The ultrastructural morphometric analysis revealed that the Purkinje cell dendritic arborisation and the number of the synapses between the parallel fibres and the Purkinje cell dendritic spines were much higher than in control animals. Numerous unattached spines of the secondary and tertiary dendritic branches of the Purkinje cells were also seen in the molecular layer either free or surrounded by astrocytic sheath. The increased number of synapses between the Purkinje cell dendrites and the parallel fibres in the animals, which received substance P intraventricularly, in correlation to control animals, supports a neurotrophine-like activity of the substance P in the mammalian cerebellum, enforcing the pre-programmed capability of the Purkinje cells to develop new synaptic surfaces.