Granule cell ascending axon excitatory synapses onto Golgi cells implement a potent feedback circuit in the cerebellar granular layer.

The function of inhibitory interneurons within brain microcircuits depends critically on the nature and properties of their excitatory synaptic drive. Golgi cells (GoCs) of the cerebellum inhibit cerebellar granule cells (GrCs) and are driven both by feedforward mossy fiber (mf) and feedback GrC excitation. Here, we have characterized GrC inputs to GoCs in rats and mice. We show that, during sustained mf discharge, synapses from local GrCs contribute equivalent charge to GoCs as mf synapses, arguing for the importance of the feedback inhibition. Previous studies predicted that GrC-GoC synapses occur predominantly between parallel fibers (pfs) and apical GoC dendrites in the molecular layer (ML). By combining EM and Ca(2+) imaging, we now demonstrate the presence of functional synaptic contacts between ascending axons (aa) of GrCs and basolateral dendrites of GoCs in the granular layer (GL). Immunohistochemical quantification estimates these contacts to be ∼400 per GoC. Using Ca(2+) imaging to identify synaptic inputs, we show that EPSCs from aa and mf contacts in basolateral dendrites display similarly fast kinetics, whereas pf inputs in the ML exhibit markedly slower kinetics as they undergo strong filtering by apical dendrites. We estimate that approximately half of the local GrC contacts generate fast EPSCs, indicating their basolateral location in the GL. We conclude that GrCs, through their aa contacts onto proximal GoC dendrites, define a powerful feedback inhibitory circuit in the GL.journal articleresearch support, non-u.s. gov't2013 Jul 24importe

Pre and Post Synaptic NMDA Effects Targeting Purkinje Cells in the Mouse Cerebellar Cortex

N-methyl-D-aspartate (NMDA) receptors are associated with many forms of synaptic plasticity. Their expression level and subunit composition undergo developmental changes in several brain regions. In the mouse cerebellum, beside a developmental switch between NR2B and NR2A/C subunits in granule cells, functional postsynaptic NMDA receptors are seen in Purkinje cells of neonate and adult but not juvenile rat and mice. A presynaptic effect of NMDA on GABA release by cerebellar interneurons was identified recently. Nevertheless whereas NMDA receptor subunits are detected on parallel fiber terminals, a presynaptic effect of NMDA on spontaneous release of glutamate has not been demonstrated. Using mouse cerebellar cultures and patch-clamp recordings we show that NMDA facilitates glutamate release onto Purkinje cells in young cultures via a presynaptic mechanism, whereas NMDA activates extrasynaptic receptors in Purkinje cells recorded in old cultures. The presynaptic effect of NMDA on glutamate release is also observed in Purkinje cells recorded in acute slices prepared from juvenile but not from adult mice and requires a specific protocol of NMDA application

Current and Calcium Responses to Local Activation of Axonal NMDA Receptors in Developing Cerebellar Molecular Layer Interneurons

In developing cerebellar molecular layer interneurons (MLIs), NMDA increases spontaneous GABA release. This effect had been attributed to either direct activation of presynaptic NMDA receptors (preNMDARs) or an indirect pathway involving activation of somato-dendritic NMDARs followed by passive spread of somatic depolarization along the axon and activation of axonal voltage dependent Ca2+ channels (VDCCs). Using Ca2+ imaging and electrophysiology, we searched for preNMDARs by uncaging NMDAR agonists either broadly throughout the whole field or locally at specific axonal locations. Releasing either NMDA or glutamate in the presence of NBQX using short laser pulses elicited current transients that were highly sensitive to the location of the spot and restricted to a small number of varicosities. The signal was abolished in the presence of high Mg2+ or by the addition of APV. Similar paradigms yielded restricted Ca2+ transients in interneurons loaded with a Ca2+ indicator. We found that the synaptic effects of NMDA were not inhibited by blocking VDCCs but were impaired in the presence of the ryanodine receptor antagonist dantrolene. Furthermore, in voltage clamped cells, bath applied NMDA triggers Ca2+ elevations and induces neurotransmitter release in the axonal compartment. Our results suggest the existence of preNMDARs in developing MLIs and propose their involvement in the NMDA-evoked increase in GABA release by triggering a Ca2+-induced Ca2+ release process mediated by presynaptic Ca2+ stores. Such a mechanism is likely to exert a crucial role in various forms of Ca2+-mediated synaptic plasticity

A family of photoswitchable NMDA receptors

NMDA receptors, which regulate synaptic strength and are implicated in learning and memory, consist of several subtypes with distinct subunit compositions and functional properties. To enable spatiotemporally defined, rapid and reproducible manipulation of function of specific subtypes, we engineered a set of photoswitchable GIuN subunits ('LiGluNs'). Photo-agonism of GIuN2A or GIuN2B elicits an excitatory drive to hippocampal neurons that can be shaped in time to mimic synaptic activation. Photo-agonism of GIuN2A at single dendritic spines evokes spine specific calcium elevation and expansion, the morphological correlate of LTP. Photo-antagonism of GIuN2A alone, or in combination with photo-antagonism of GluN1a, reversibly blocks excitatory synaptic currents, prevents the induction of long-term potentiation and prevents spine expansion. In addition, photo-antagonism in vivo disrupts synaptic pruning of developing retino-tectal projections in larval zebrafish. By providing precise and rapidly reversible optical control of NMDA receptor subtypes, LiGluNs should help unravel the contribution of specific NMDA receptors to synaptic transmission, integration and plasticity

Sensory Stimulation-Dependent Plasticity in the Cerebellar Cortex of Alert Mice

In vitro studies have supported the occurrence of cerebellar long-term depression (LTD), an interaction between the parallel fibers and Purkinje cells (PCs) that requires the combined activation of the parallel and climbing fibers. To demonstrate the existence of LTD in alert animals, we investigated the plasticity of local field potentials (LFPs) evoked by electrical stimulation of the whisker pad. The recorded LFP showed two major negative waves corresponding to trigeminal (broken into the N2 and N3 components) and cortical responses. PC unitary extracellular recording showed that N2 and N3 occurred concurrently with PC evoked simple spikes, followed by an evoked complex spike. Polarity inversion of the N3 component at the PC level and N3 amplitude reduction after electrical stimulation of the parallel fiber volley applied on the surface of the cerebellum 2 ms earlier strongly suggest that N3 was related to the parallel fiber–PC synapse activity. LFP measurements elicited by single whisker pad stimulus were performed before and after trains of electrical stimuli given at a frequency of 8 Hz for 10 min. We demonstrated that during this later situation, the stimulation of the PC by parallel and climbing fibers was reinforced. After 8-Hz stimulation, we observed long-term modifications (lasting at least 30 min) characterized by a specific decrease of the N3 amplitude accompanied by an increase of the N2 and N3 latency peaks. These plastic modifications indicated the existence of cerebellar LTD in alert animals involving both timing and synaptic modulations. These results corroborate the idea that LTD may underlie basic physiological functions related to calcium-dependent synaptic plasticity in the cerebellum

Target-specific expression of presynaptic NMDA receptors in neocortical microcircuits.

Traditionally, NMDA receptors are located postsynaptically; yet, putatively presynaptic NMDA receptors (preNMDARs) have been reported. Although implicated in controlling synaptic plasticity, their function is not well understood and their expression patterns are debated. We demonstrate that, in layer 5 of developing mouse visual cortex, preNMDARs specifically control synaptic transmission at pyramidal cell inputs to other pyramidal cells and to Martinotti cells, while leaving those to basket cells unaffected. We also reveal a type of interneuron that mediates ascending inhibition. In agreement with synapse-specific expression, we find preNMDAR-mediated calcium signals in a subset of pyramidal cell terminals. A tuned network model predicts that preNMDARs specifically reroute information flow in local circuits during high-frequency firing, in particular by impacting frequency-dependent disynaptic inhibition mediated by Martinotti cells, a finding that we experimentally verify. We conclude that postsynaptic cell type determines presynaptic terminal molecular identity and that preNMDARs govern information processing in neocortical columns

Fonction des récepteurs NMDA présynaptiques dans la plasticité du cervelet

La caractérisation détaillée de la plasticité synaptique a conduit à préciser les règles hebbiennes simples. Des règles plus complexes sont basées sur l ordre relatif des potentiels d action pré- et postsynaptiques. Dans ce mémoire de thèse, nous décrivons un mécanisme sous-tendant une nouvelle règle de plasticité basée sur le patron des potentiels d action présynaptiques. La règle hebbienne classique repose sur la détection des activités simultanées pré- et postsynaptiques par les récepteurs NMDA postsynaptiques. Désormais, l existence de récepteurs NMDA présynaptiques est établie dans plusieurs structures cérébrales. Nous proposons que les récepteurs NMDA présynaptiques définissent la structure temporelle de la règle d induction de la dépression à long terme (LTD) des synapses entre fibres parallèles et cellules de Purkinje dans le cervelet. Nous avons montré que plusieurs potentiels d action présynaptiques à des fréquences entre 40 Hz et 1 kHz sont nécessaires pour l induction de la LTD. Nous avons caractérisé le sous-type de récepteurs NMDA impliqué dans l induction de la LTD ainsi que ses cinétiques. Nous montrons comment les cinétiques des courants issus de récepteurs contenant les sous-unités NR2A, exprimées par les fibres parallèles, génèrent un filtre passe-haut pour permettre la dépression sélective des synapses présentant une activité en bouffées de haute fréquence. Nous proposons que ce mécanisme soit un principe général. En fonction du type de sous-unité présent, les synapses exprimant des autorécepteurs NMDA génèrent des filtres passe-haut de différentes fréquences.PARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

ACTIVATION OF CEREBELLAR INHIBITORY LOOPS: EVIDENCE FOR NOVEL GRANULE CELL-GOLGI CELL CONTACTS IN THE GRANULAR LAYER

The granular layer, the input stage of the cerebellar cortex, is an important target of EtOH that is populated by two main neuronal types, excitatory GrCs and inhibitory Golgi cells (GoCs). The function of these neurons is altered by acute EtOH exposure. Mossy fibers (mfs) provide excitatory inputs to both neuron types; GoCs are also excited by GrCs, and their output inhibits GrCs. Thus mfs activate local feed-forward and feed-back inhibitory loops. Sensory input can either activate a GoC with high temporal precision, or evoke less precisely timed responses. While direct input from mfs may trigger GoC firing with millisecond precision, the functional impact of GrC input on GoC firing is unknown. GrC-GoC contacts are believed to occur in the molecular layer, between parallel fibers (pfs) and GoC apical dendrites, although morphological data also propose the presence of contacts between the GrC ascending axon (aa) and GoC basolateral (bl) and apical dendrites. Using a combination of electrophysiological, 2-photon Ca2+-imaging and EM approaches, we have obtained evidence for functional aa-GoC contacts occurring in the rat GL onto bl dendrites. EPSCs from these contacts (aaEPSCs) have fast kinetics, similar to mf-evoked EPSCs (mf-EPSCs), in contrast with the slow kinetics of pf-evoked EPSCs. mf spikes evoke both mono-synaptic mf- and di-synaptic aa-EPSCs, which may summate to evoke a GoC spike within milliseconds. Hence, the impact of novel aaEPSCs on GoC firing may parallel the one of mf inputs, contributing to generate fast GoC responses, which may allow for precise control of the time window for information flow through GrCs. The pf\u2013GoC input is more complex, involving weak excitatory and inhibitory metabotropic components, and this may implement spatial contrast enhancement upon intense pf excitation. The potential impact of EtOH on aa and pf GoC inputs will be discussed

Granule Cell Ascending Axon Excitatory Synapses onto Golgi Cells Implement a Potent Feedback Circuit in the Cerebellar Granular Layer

The function of inhibitory interneurons within brain microcircuits depends critically on the nature and properties of their excitatory synaptic drive. Golgi cells (GoCs) of the cerebellum inhibit cerebellar granule cells (GrCs) and are driven both by feedforward mossy fiber (mf) and feedback GrC excitation. Here, we have characterized GrC inputs to GoCs in rats and mice. We show that, during sustained mf discharge, synapses from local GrCs contribute equivalent charge to GoCs as mf synapses, arguing for the importance of the feedback inhibition. Previous studies predicted that GrC-GoC synapses occur predominantly between parallel fibers (pfs) and apical GoC dendrites in the molecular layer (ML). By combining EM and Ca2+ imaging, we now demonstrate the presence of functional synaptic contacts between ascending axons (aa) of GrCs and basolateral dendrites of GoCs in the granular layer (GL). Immunohistochemical quantification estimates these contacts to be 3c400 per GoC. Using Ca2+ imaging to identify synaptic inputs, we show that EPSCs from aa and mf contacts in basolateral dendrites display similarly fast kinetics, whereas pf inputs in the ML exhibit markedly slower kinetics as they undergo strong filtering by apical dendrites. We estimate that approximately half of the local GrC contacts generate fast EPSCs, indicating their basolateral location in the GL. We conclude that GrCs, through their aa contacts onto proximal GoC dendrites, define a powerful feedback inhibitory circuit in the GL