Cerebellar Long-term Synaptic Depression Research Articles

A Late Phase of Long-Term Synaptic Depression in Cerebellar Purkinje Cells Requires Activation of MEF2

SUMMARYThe MEF2 family of transcription factors restricts excitatory synapse number in an activity-dependent fashion during development, yet MEF2 has not been implicated in long-term synaptic depression (LTD), which is thought to initiate synapse elimination. Mutations in MEF2 pathways are implicated in autism spectrum disorders, which include cerebellar dysfunction. Here, we test the hypothesis that cerebellar LTD requires postsynaptic activation of MEF2. Knockdown of MEF2D produces suppression of the transcription-dependent late phase of LTD in cultured Purkinje cells. The late phase of LTD is also completely blocked in Purkinje cells derived from MEF2A+MEF2D null mice and rescued with plasmids that drive expression of MEF2D but not phosphatase-resistant mutant MEF2D S444D. Wild-type Purkinje cells transfected with a constitutively active form of MEF2 show no alterations of synaptic strength. Thus, postsynaptic activation of MEF2 by S444 dephosphorylation is necessary, but not sufficient, for the late phase of cerebellar LTD.

DHHC8-Dependent PICK1 Palmitoylation is Required for Induction of Cerebellar Long-Term Synaptic Depression

The palmitoyl acyltransferase (PAT) DHHC8 is implicated in synaptic regulation but few DHHC8 substrates are known. Here we report that DHHC8 binds and palmitoylates the PDZ domain-containing protein PICK1 at a cysteine residue that is essential for long-term synaptic depression (LTD) in cultured mouse cerebellar Purkinje neurons. Cerebellar LTD is palmitoylation-dependent and induction of LTD requires DHHC8. Furthermore, PICK1 is a critical DHHC8 substrate whose palmitoylation is necessary for LTD. These results identify the first DHHC8 substrate required for a specific form of synaptic plasticity and provide new insights into synaptic roles of palmitoylation.

Raf Kinase Inhibitory Protein is Required for Cerebellar Long-Term Synaptic Depression by Mediating PKC-Dependent MAPK Activation

It was demonstrated previously that a positive feedback loop, including protein kinase C (PKC) and mitogen-activated protein kinase (MAPK), is required for the gradual expression of cerebellar long-term depression (LTD). PKC and MAPK are mutually activated in this loop. MAPK-dependent PKC activation is likely to be mediated by phospholipase A2. On the other hand, it is not clear how PKC activates MAPK. Therefore, the entire picture of this loop was not fully understood. We here test the hypothesis that this loop is completed by the PKC substrate, Raf kinase inhibitory protein (RKIP). To test this hypothesis, we used a mutant form of RKIP that is not phosphorylated by PKC and thus constitutively inhibits Raf-1 and MEK, upstream kinases of MAPK. When this RKIP mutant was introduced into Purkinje cells of mouse cerebellar slices through patch-clamp electrodes, LTD was blocked, while wild-type (WT) RKIP had no effect on LTD. Physiological epistasis experiments demonstrated that RKIP works downstream of PKC and upstream of MAPK during LTD induction. Furthermore, biochemical analyses demonstrated that endogenous RKIP dissociates from Raf-1 and MEK during LTD induction in a PKC-dependent manner, suggesting that RKIP binding-dependent inhibition of Raf-1 and MEK is removed upon LTD induction. We therefore conclude that PKC-dependent regulation of RKIP leads to MAPK activation, with RKIP completing the positive feedback loop that is required for LTD.

A late phase of LTD in cultured cerebellar Purkinje cells requires persistent dynamin-mediated endocytosis

Long-term synaptic depression (LTD) of cerebellar parallel fiber-Purkinje cell synapses is a form of use-dependent synaptic plasticity that may be studied in cell culture. One form of LTD is induced postsynaptically through an mGlu1/Ca influx/protein kinase Cα (PKCα) cascade, and its initial expression requires phosphorylation of ser-880 in the COOH-terminal PDZ-ligand region of GluA2 and consequent binding of PICK1. This triggers postsynaptic clathrin/dynamin-mediated endocytosis of GluA2-containing surface AMPA receptors. Cerebellar LTD also has a late phase beginning 45-60 min after induction that is blocked by transcription or translation inhibitors. Here, I have sought to determine the expression mechanism of this late phase of LTD by applying various drugs and peptides after the late phase has been established. Neither bath application of mGluR1 antagonists (JNJ-16259685, LY-456236) nor the PKC inhibitor GF-109203X starting 60-70 min after LTD induction attenuated the late phase. Similarly, achieving the whole cell configuration with a second pipette loaded with the peptide PKC inhibitor PKC(19-36) starting 60 min postinduction also failed to alter the late phase. Late internal perfusion with peptides designed to disrupt PICK1-GLUA2 interaction or PICK1 dimerization failed to impact late phase LTD expression. However, late internal perfusion with two different blockers of dynamin, the drug dynasore and a dynamin inhibitory peptide (QVPSRPNRAP), produced rapid and complete reversal of cerebellar LTD expression. These findings suggest that the protein synthesis-dependent late phase of LTD requires persistent dynamin-mediated endocytosis, but not persistent PICK1-GluA2 binding nor persistent activation of the upstream mGluR1/PKCα signaling cascade.

Calcium as a Trigger for Cerebellar Long-Term Synaptic Depression

Cerebellar long-term depression (LTD) is a form of long-term synaptic plasticity that is triggered by calcium(Ca2+) signals in the postsynaptic Purkinje cell. This Ca2+comes both from IP3-mediated release from intracellular Ca2+ stores, as well as from Ca2+ influx through voltage-gated Ca2+ channels. The Ca2+ signal that triggers LTD occurs locally within dendritic spines and is due to supralinear summation of signals coming from these two Ca2+ sources. The properties of this postsynaptic Ca2+signal can explain several features of LTD, such as its associativity, synapse specificity, and dependence on thetiming of synaptic activity, and can account for the slow kinetics of LTD expression. Thus, from a Ca2+ signaling perspective, LTD is one of the best understood forms of synaptic plasticity.

Is there a functional link between the cerebellum and the hippocampus relevant to spatial navigation?

Event Abstract Back to Event Is there a functional link between the cerebellum and the hippocampus relevant to spatial navigation? Arnaud Arabo1*, Christelle Rochefort1, Sylvain Quet1, Marion Andre2, Etienne Save2 and Laure Rondi-Reig1 1 Universite Pierre et Marie Curie, NPA, UMR7102, France 2 Universite de Provence, LNC, France Path integration requires the online estimation of a navigator position using self-motion cues. This process is necessary to the acquisition of an internal representation of the environment in the absence of allothetic cues. The hippocampal formation plays a key role in the development of spatial representations. The cerebellum is likely to mediate context-dependent optimization of navigation trajectories. Empirical experimental evidence exists that the cerebellum and the hippocampal formation are electrophysiologically coupled through a number of synaptic connections that remain unknown. We put forth the hypothesis that the cerebellum may contribute to the processing of self-motion cues, and therefore interact with the hippocampus for the elaboration of spatial representations.We adopt a multidisciplinary behavioural, electrophysiological, and neuroanatomical approach to study the spatial navigation capabilities of transgenic mice (L7-PKCI), which have impaired long-term synaptic depression (LTD) at cerebellar parallel fiber-Purkinje cell (PF-PC) synapses. Behavioural data show that in a path integration version of the Morris water maze, L7-PKCI animals present a drift in their trajectory toward a platform in the dark. Electrophysiological recordings of hippocampal place cells in L7-PKCI were performed as they explored a circular arena. Preliminary data may suggest an alteration of hippocampal place fields in the case of either an absence of allothetic cues or a conflict between external and self-motion cues. Using neuronal tracers, we are currently investigating if the connections between the hippocampus and the cerebellum are mono- or multi-synaptic. Our data seem to suggest that cerebellar PF-PC LTD might be important for the path integration process. A deficit in this process seems to induce impairment in the hippocampal place field representation. This may support the hypothesis of a functional link between the hippocampal formation and the cerebellum. Conference: 41st European Brain and Behaviour Society Meeting, Rhodes Island, Greece, 13 Sep - 18 Sep, 2009. Presentation Type: Poster Presentation Topic: Poster presentations Citation: Arabo A, Rochefort C, Quet S, Andre M, Save E and Rondi-Reig L (2009). Is there a functional link between the cerebellum and the hippocampus relevant to spatial navigation?. Conference Abstract: 41st European Brain and Behaviour Society Meeting. doi: 10.3389/conf.neuro.08.2009.09.360 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 16 Jun 2009; Published Online: 16 Jun 2009. * Correspondence: Arnaud Arabo, Universite Pierre et Marie Curie, NPA, UMR7102, Paris, France, christelle.rochefort@upmc.fr Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Arnaud Arabo Christelle Rochefort Sylvain Quet Marion Andre Etienne Save Laure Rondi-Reig Google Arnaud Arabo Christelle Rochefort Sylvain Quet Marion Andre Etienne Save Laure Rondi-Reig Google Scholar Arnaud Arabo Christelle Rochefort Sylvain Quet Marion Andre Etienne Save Laure Rondi-Reig PubMed Arnaud Arabo Christelle Rochefort Sylvain Quet Marion Andre Etienne Save Laure Rondi-Reig Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

A Positive Feedback Signal Transduction Loop Determines Timing of Cerebellar Long-Term Depression

Synaptic activity produces short-lived second messengers that ultimately yield a long-term depression (LTD) of cerebellar Purkinje cells. Here, we test the hypothesis that these brief second messenger signals are translated into long-lasting biochemical signals by a positive feedback loop that includes protein kinase C (PKC) and mitogen-activated protein kinase. Histochemical "epistasis" experiments demonstrate the reciprocal activation of these kinases, and physiological experiments--including the use of a light-activated protein kinase--demonstrate that such reciprocal activation is required for LTD. Timed application of enzyme inhibitors reveals that this positive feedback loop causes PKC to be active for more than 20 min, allowing sufficient time for LTD expression. Such regenerative mechanisms may sustain other long-lasting forms of synaptic plasticity and could be a general mechanism for prolonging signal transduction networks.

The Glutamate Receptor-Interacting Protein Family of GluR2-Binding Proteins Is Required for Long-Term Synaptic Depression Expression in Cerebellar Purkinje Cells

Glutamate receptor-interacting protein 1 (GRIP1) and GRIP2 are closely related proteins that bind GluR2-containing AMPA receptors and couple them to structural and signaling complexes in neurons. Cerebellar long-term synaptic depression (LTD) is a model system of synaptic plasticity that is expressed by persistent internalization of GluR2-containing AMPA receptors. Here, we show that genetic deletion of both GRIP1 and GRIP2 blocks LTD expression in primary cultures of mouse cerebellar neurons but that single deletion of either isoform allows LTD to occur. In GRIP1/2 double knock-out Purkinje cells, LTD can be fully rescued by a plasmid-driving expression of GRIP1 and partially rescued by a GRIP2 plasmid. These results indicate that the GRIP family comprises an essential molecular component for cerebellar LTD.

Ca2+ Requirements for Cerebellar Long-Term Synaptic Depression: Role for a Postsynaptic Leaky Integrator

Photolysis of a caged Ca(2+) compound was used to characterize the dependence of cerebellar long-term synaptic depression (LTD) on postsynaptic Ca(2+) concentration ([Ca(2+)](i)). Elevating [Ca(2+)](i) was sufficient to induce LTD without requiring any of the other signals produced by synaptic activity. A sigmoidal relationship between [Ca(2+)](i) and LTD indicated a highly cooperative triggering of LTD by Ca(2+). The duration of the rise in [Ca(2+)](i) influenced the apparent Ca(2+) affinity of LTD, and this time-dependent behavior could be described by a leaky integrator process with a time constant of 0.6 s. A computational model, based on a positive-feedback cycle that includes protein kinase C and MAP kinase, was capable of simulating these properties of Ca(2+)-triggered LTD. Disrupting this cycle experimentally also produced the predicted changes in the Ca(2+) dependence of LTD. We conclude that LTD arises from a mechanism that integrates postsynaptic Ca(2+) signals and that this integration may be produced by the positive-feedback cycle.

Arc is required for the late phase of cerebellar long-term synaptic depression

Arc is required for the late phase of cerebellar long-term synaptic depression

A Unique PDZ Ligand in PKCα Confers Induction of Cerebellar Long-Term Synaptic Depression

Induction of cerebellar long-term depression (LTD) requires a postsynaptic cascade involving activation of mGluR1 and protein kinase C (PKC). Our understanding of this process has been limited by the fact that PKC is a large family of molecules, many isoforms of which are expressed in the relevant postsynaptic compartment, the cerebellar Purkinje cell. Here, we report that LTD is absent in Purkinje cells in which the alpha isoform of PKC has been reduced by targeted RNA interference or in cells derived from PKCalpha null mice. In both of these cases, LTD could be rescued by expression of PKCalpha but not other PKC isoforms. The special role of PKCalpha in cerebellar LTD is likely to derive from its unique PDZ ligand (QSAV). When this motif is mutated, PKCalpha no longer supports LTD. Conversely, when this PDZ ligand is inserted in a nonpermissive isoform, PKCgamma, it confers the capacity for LTD induction.

Cerebellar Long-Term Synaptic Depression Requires PKC-Mediated Activation of CPI-17, a Myosin/Moesin Phosphatase Inhibitor

Cerebellar LTD requires brief activation of PKC and is expressed as a functional downregulation of AMPA receptors. Modulation of vascular smooth-muscle contraction by G protein-coupled receptors (called Ca 2+ sensitization) also involves PKC phosphorylation and activation of a specific inhibitor of myosin/moesin phosphatase (MMP). This inhibitor, called CPI-17, is also expressed in brain. Here, we tested the hypothesis that LTD, like Ca 2+ sensitization, employs a PKC/CPI-17 cascade. Introduction of activated recombinant CPI-17 into cells produced a use-dependent attenuation of glutamate-evoked responses and occluded subsequent LTD. Moreover, the requirement for endogenous CPI-17 in LTD was demonstrated with neutralizing antibodies plus gene silencing by siRNA. These interventions had no effect on basal synaptic strength but blocked LTD induction. Thus, a biochemical circuit that involves PKC-mediated activation of CPI-17 modulates the distinct physiological processes of vascular contractility and cerebellar LTD.

The expression of cerebellar LTD in culture is not associated with changes in AMPA-receptor kinetics, agonist affinity, or unitary conductance.

Cerebellar long-term synaptic depression (LTD) is a model system of neuronal information storage that is expressed postsynaptically as a functional down-regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. What properties of postsynaptic AMPA receptors are changed? Several lines of evidence argue against changes in AMPA-receptor kinetics. Neither LTD evoked in cultured granule-cell Purkinje cell (PC) pairs nor an LTD-like phenomenon evoked by phorbol ester application was associated with alterations in evoked AMPA receptor-mediated excitatory post-synaptic current (EPSC) or mEPSC kinetics. LTD produced by pairing glutamate pulses with depolarization was not altered by prior application of the desensitization-reducing compound cyclothiazide. Finally, rapid application of glutamate to lifted PCs revealed no significant alterations in AMPA-receptor kinetic properties after LTD induction. When this system was used to apply varying concentrations of glutamate, no alteration in AMPA-receptor glutamate affinity was seen after LTD induction. Finally, peak-scaled nonstationary fluctuation analysis was applied to estimate AMPA-receptor unitary conductance before and after LTD induction in a cultured cell pair, and this analysis too revealed no significant change. These results suggest that cerebellar LTD may be expressed solely as a reduction in the number of functional AMPA receptors in the postsynaptic density [Wang, Y.-T. & Linden, D. J. (2000) Neuron 25, 635-664].