Synaptic Trafficking Research Articles

Using ProPeL to discover the binding specificity of human DYRK1a

Pairing kinases to their substrates is central to elucidating signal transduction pathways and understanding their functions in normal and diseased states. Kinases discriminate between potential substrates in part by linear sequence “motifs” surrounding the residue to be phosphorylated, yet motifs for the majority of the 518 human kinases remain unknown. One largely uncharacterized kinase is Dual specificity tyrosine‐phosphorylation‐regulated kinase 1a (DYRK1a), which is implicated in Down's Syndrome, Alzheimer's Disease, alternative splicing and synaptic trafficking. Using our new Proteomic Peptide Library (ProPeL) methodology, in which we express a human kinase in E. coli and analyze the resulting phosphopeptides with our pLogo and motif‐x software, we extracted a strong sequence motif for DYRK1a from 933 unique phosphorylation sites. We also detected 8 novel DYRK1a autophosphorylation sites. Currently, the only known tyrosine targets of DYRK1a are on DYRK1a itself. Strikingly, however, we observed 66 unique phosphotyrosine sites on substrates other than DYRK1a, suggesting that its ability to phosphorylate tyrosine residues may not be limited to autophosphorylation. Finally we predicted human DYRK1a substrates using the authors’ scan‐x tool. This study was funded in part by grants from the University of Connecticut Research Foundation (DS) and Genomes to Life from US DoE (GMC).

Progress in research of synaptic membrane-associated guanylate kinase scaffolding proteins in central nervous system involved in pain modulation

Background A family of membrane-associated guanylate kinases (MAGUKs) in central nervous system contains 4 kinds of synapses-associated proteins:PSD-93、PSD-95、SAP-102、SAP-97.They are involved in modulating synaptic trafficking of glutamate receptors and associate with pain,learning and memory.Objective To review the interactions between the synaptic MAGUK scaffolding proteins and the receptors、molecules involved in pain and the roles of synaptic MAGUK scaffolding proteins in pain signal modulation and transmission.Content Synaptic MAGUK scaffolding proteins are involved in modulating pain signal transduction in synapses by interacting with NMDAR,nNOS,AMPAR,SynCAM and other molecules.Trend Disruption of the interactions between the synaptic MAGUK scaffolding proteins and the receptors or molecules involved in pain can influence pain signal transduction in synapses,suggesting a new target for the treatment of pain. Key words: Membrane-associated guanylate kinase proteins; Glutamate receptors; Pain

Disrupted cortical function underlies behavior dysfunction due to social isolation

Stressful events during early childhood can have a profound lifelong influence on emotional and cognitive behaviors. However, the mechanisms by which stress affects neonatal brain circuit formation are poorly understood. Here, we show that neonatal social isolation disrupts molecular, cellular, and circuit developmental processes, leading to behavioral dysfunction. Neonatal isolation prevented long-term potentiation and experience-dependent synaptic trafficking of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors normally occurring during circuit formation in the rodent barrel cortex. This inhibition of AMPA receptor trafficking was mediated by an increase of the stress glucocorticoid hormone and was associated with reduced calcium/calmodulin-dependent protein kinase type II (CaMKII) signaling, resulting in attenuated whisker sensitivity at the cortex. These effects led to defects in whisker-dependent behavior in juvenile animals. These results indicate that neonatal social isolation alters neuronal plasticity mechanisms and perturbs the initial establishment of a normal cortical circuit, which potentially explains the long-lasting behavioral effects of neonatal stress.

Homeostatic Synaptic Scaling Is Regulated by Protein SUMOylation

Homeostatic scaling allows neurons to alter synaptic transmission to compensate for changes in network activity. Here, we show that suppression of network activity with tetrodotoxin, which increases surface expression of AMPA receptors (AMPARs), dramatically reduces levels of the deSUMOylating (where SUMO is small ubiquitin-like modifier) enzyme SENP1, leading to a consequent increase in protein SUMOylation. Overexpression of the catalytic domain of SENP1 prevents this scaling effect, and we identify Arc as a SUMO substrate involved in the tetrodotoxin-induced increase in AMPAR surface expression. Thus, protein SUMOylation plays an important and previously unsuspected role in synaptic trafficking of AMPARs that underlies homeostatic scaling.

Acute Cannabinoids Impair Working Memory through Astroglial CB1 Receptor Modulation of Hippocampal LTD

Impairment of working memory is one of the most important deleterious effects of marijuana intoxication in humans, but its underlying mechanisms are presently unknown. Here, we demonstrate that the impairment of spatial working memory (SWM) and in vivo long-term depression (LTD) of synaptic strength at hippocampal CA3-CA1 synapses, induced by an acute exposure of exogenous cannabinoids, is fully abolished in conditional mutant mice lacking type-1 cannabinoid receptors (CB(1)R) in brain astroglial cells but is conserved in mice lacking CB(1)R in glutamatergic or GABAergic neurons. Blockade of neuronal glutamate N-methyl-D-aspartate receptors (NMDAR) and of synaptic trafficking of glutamate α-amino-3-hydroxy-5-methyl-isoxazole propionic acid receptors (AMPAR) also abolishes cannabinoid effects on SWM and LTD induction and expression. We conclude that the impairment of working memory by marijuana and cannabinoids is due to the activation of astroglial CB(1)R and is associated with astroglia-dependent hippocampal LTD in vivo.

Aspects of Innate Immunity and Parkinson’s Disease

Genetic studies on PARK genes have identified dysfunction in proteasomal, lysosomal, and mitochondrial enzymes as pathogenic for Parkinson’s disease (PD). We review the role of these and similar enzymes in mediating innate immune signaling. In particular, we have identified that a number of PARK gene products as well as other enzymes have roles in innate immune signaling as well as DNA repair and regulation, ubiquitination, mitochondrial functioning, and synaptic trafficking. PD enzymatic dysfunction is likely to contribute to inadequate innate immune responses to a variety of extra- and intra-cellular stimuli, with a number of the innate immunity related enzymes found in the characteristic Lewy body pathology of PD. The decrease in innate immunity in PD is associated with an increase in markers of adaptive immunity, and recent GWAS studies have identified variants in human leukocyte antigen region as associated with late-onset sporadic PD (Hamza et al., 2010; Hill-Burns et al., 2011). Intriguing new data also suggest that peripheral immune responses may be involved, giving some potential to alleviate such peripheral dysfunction more directly in patients with PD. It is now important to identify the cell type specific immune responses contributing to the initial changes that occur in PD, as well as to the propagating immune responses important for the progression of PD pathology between cells and within the brain. Overall, a complex interplay between different types of immunity appear to be involved in the underlying pathology of PD.

Morphologic and Functional Correlates of Synaptic Pathology in the Cathepsin D Knockout Mouse Model of Congenital Neuronal Ceroid Lipofuscinosis

Mutations in the cathepsin D (CTSD) gene cause an aggressive neurodegenerative disease (congenital neuronal ceroid lipofuscinosis) that leads to early death. Recent evidence suggests that presynaptic abnormalities play a major role in the pathogenesis of CTSD deficiencies. To identify the early events that lead to synaptic alterations, we investigated synaptic ultrastructure and function in presymptomatic CTSD knockout (Ctsd) mice. Electron microscopy revealed that there were significantly greater numbers of readily releasable synaptic vesicles present in Ctsd mice than in wild-type control mice as early as postnatal day 16. The size of this synaptic vesicle pool continued to increase with disease progression in the hippocampus and thalamus of the Ctsd mice. Electrophysiology revealed a markedly decreased frequency of miniature excitatory postsynaptic currents (mEPSCs) with no effect on paired-pulse modulation of the evoked excitatory post synaptic potentials in the hippocampus of Ctsd mice. The reduced mEPSCs frequency was observed before the appearance of epilepsy or any morphologic sign of synaptic degeneration. Taken together, these data indicate that CTSD is required for normal synaptic function and that a failure in synaptic trafficking or recycling may bean early and important pathologic mechanism in Ctsd mice; these presynaptic abnormalities may initiate synaptic degeneration in advance of subsequent neuronal loss.

Glutamate receptor subunit GluA1 is necessary for long-term potentiation and synapse unsilencing, but not long-term depression in mouse hippocampus

Receptor subunit composition is believed to play a major role in the synaptic trafficking of AMPA receptors (AMPARs), and thus in activity-dependent synaptic plasticity. To isolate a physiological role of GluA1-containing AMPARs in area CA3 of the hippocampus, pair recordings were performed in organotypic hippocampal slices taken from genetically modified mice lacking the GluA1 subunit. We report here that long-term potentiation (LTP) is impaired not only at active but also at silent synapses when the GluA1 subunit is absent. The GluA1 knockout mice also exhibited reduced AMPAR-mediated evoked currents between pairs of CA3 pyramidal neurons under baseline conditions suggesting a significant role for GluA1-containing AMPARs in regulating basal synaptic transmission. In two independent measures, however, long-term depression (LTD) was unaffected in tissue from these mice. These data provide a further demonstration of the fundamental role that GluA1-containing AMPARs play in activity-dependent increases in synaptic strength but do not support a GluA1-dependent mechanism for reductions in synaptic strength.

Mechanisms underlying contextual fear learning

The hippocampus plays a central role in learning and memory. Although synaptic delivery of AMPA-type glutamate receptors (AMPARs) contributes to experience-dependent synaptic strengthening, its role in hippocampus-dependent learning remains elusive. In a recent study,we found that the inhibitory avoidance (IA) task, a hippocampus-dependent contextual fear learning paradigm, drives GluR1-containing AMPARs into CA3-CA1 synapses of the dorsal hippocampus.We expressed mutated membrane-proxymal region (14 amino acids) of the GluR1-cytoplasmic tail (serines mutated to phospho-mimicking aspartates:MPR-DD) in the dorsal hippocampus to block the synaptic delivery of endogenous AMPARs. Learning-driven synaptic AMPARs delivery in CA1 neurons was prevented by the expression of MPR-DD. Bilateral expression of MPR-DD in CA1 region of the dorsal hippocampusattenuated IA learning, indicating that synaptic GluR1 trafficking in the hippocampus is required for encoding contextual fear memories. Furthermore, fraction of CA1 neurons with synaptic strengthening positively correlated with the performance in the IA fear memory task. Thus, the robustness of a contextual fear memory may depend on thenumber of hipppocampal neurons that are involved in the encoding of a memory trace.

NR2B phosphorylation at tyrosine 1472 in spinal dorsal horn contributed to N‐methyl‐D‐aspartate‐induced pain hypersensitivity in mice

Calcium influx via N-methyl-D-aspartate (NMDA)-subtype glutamate receptors (NMDARs) regulates the intracellular trafficking of NMDARs, leading to long-lasting modification of NMDAR-mediated synaptic transmission that is involved in development, learning, and synaptic plasticity. The present study investigated the contribution of such NMDAR-dependent synaptic trafficking in spinal dorsal horn to the induction of pain hypersensitivity. Our data showed that direct activation of NMDARs by intrathecal NMDA application elicited pronounced mechanical allodynia in intact mice, which was concurrent with a specific increase in the abundance of NMDAR subunits NR1 and NR2B at the postsynaptic density (PSD)-enriched fraction. Selective inhibition of NR2B-containing NMDARs (NR2BR) by ifenprodil dose dependently attenuated the mechanical allodynia in NMDA-injected mice, suggesting the importance of NR2BR synaptic accumulation in NMDA-induced pain sensitization. The NR2BR redistribution at synapses after NMDA challenge was associated with a significant increase in NR2B phosphorylation at Tyr1472, a catalytic site by Src family protein tyrosine kinases (SFKs) that has been shown to prevent NR2B endocytosis. Intrathecal injection of a specific SFKs inhibitor, PP2, to block NR2B tyrosine phosphorylation eliminated NMDA-induced NR2BR synaptic expression and also attenuated the mechanical allodynia. These data suggested that activation of spinal NMDARs was able to accumulate NR2BR at synapses via SFK signaling, which might exaggerate NMDAR-dependent nociceptive transmission and contribute to NMDA-induced nociceptive behavioral hyperresponsiveness.

SAP97-mediated local trafficking is altered in Alzheimer disease patients' hippocampus

Synapse-asssociated protein-97 (SAP97) is responsible for the trafficking of both glutamate receptor subunits, GluR1 and NR2A, and α-secretase ADAM10 to the synaptic membrane. Here we evaluate the trafficking capability of SAP97 in Alzheimer disease (AD) patients' brain. We analyzed autoptic hippocampus and superior frontal gyrus, respectively as an affected and a less affected area, from 6 AD patients (Braak 4) and 6 healthy controls. In hippocampus, but not in superior frontal gyrus, of AD patients, ADAM10 and GluR1 synaptic membrane levels are altered while NR2A localization is not affected. Both immunoprecipitation and pull-down assays demonstrated that SAP97 failed to correctly couple to ADAM10 and GluR1, but not to NR2A. These findings not only indicate SAP97 as a point of convergence between amyloid cascade and synaptic failure in AD, but also allow a different interpretation of AD which can be now perceived as synaptic trafficking defect pathology.

Synaptic Vesicle Recycling: Genetic and Cell Biological Studies

We review mainly the work from our research group here. Our focus has been on the use of genetic methods to delineate the mechanisms of synaptic vesicle recycling and cellular trafficking. Acute temperature-sensitive paralytic mutants have been of particular value in this approach. We have primarily used screens for suppressor and enhancer mutations to identify genetic loci coding for proteins that interact with Dynamin in Drosophila. In addition, we have used reverse genetic approaches to investigate few other candidate molecules that may play a role in synaptic vesicle endocytosis. We have in particular discussed at some length the role of endocytic accessory proteins Stoned and Eps15 in vesicle recycling.

Regulation of stargazin synaptic trafficking by C‐terminal PDZ ligand phosphorylation in bidirectional synaptic plasticity

Stargazin is a transmembrane alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor regulatory protein that controls the surface and synaptic expression of AMPA-type glutamate receptors (AMPARs). Synaptic anchoring of AMPARs is influenced by the interaction between stargazin's C-terminal post-synaptic density-95 (PSD-95)/discs large/zona occludens-1 (PDZ) ligand and the synaptic scaffolding protein PSD-95. Phosphorylation of the stargazin PDZ ligand by protein kinase A (PKA) disrupts stargazin's interaction with PSD-95, but whether this phosphorylation plays a role in activity-dependent regulation of stargazin/AMPAR synaptic trafficking is unknown. Here, we show that stargazin is phosphorylated within the PDZ ligand at threonine residue 321 (T321) by mitogen-activated protein kinases (MAPKs) as well as PKA. By expressing constructs that selectively block T321 phosphorylation by either PKA or MAPKs, we show that stargazin T321 phosphorylation is required for activity-dependent changes in stargazin synaptic clustering in dissociated rat hippocampal neuron cultures. Specifically, we find that mutations that block stargazin T321 phosphorylation by PKA prevent activity-dependent increases in stargazin synaptic clustering, whereas a point mutant that blocks MAPK phosphorylation of T321 prevents activity-dependent decreases in stargazin synaptic clustering. Taken together, our studies implicate phosphorylation of stargazin T321 by PKA and MAPKs in bidirectional control of stargazin/AMPAR synaptic clustering during synaptic plasticity.

Analyzing NMDA and AMPA evoked spike-trains in CA1: GluA1 phosphorylation specific patterns in vivo

Event Abstract Back to Event Analyzing NMDA and AMPA evoked spike-trains in CA1: GluA1 phosphorylation specific patterns in vivo Balázs Barkóczi1*, Gábor Juhász2, Robert Averkin1, Imre Vörös3, Botond Penke1, 2 and Viktor Szegedi1 1 Bay Zoltán Foundation for Applied Research, BAYGEN, Hungary 2 University of Szeged, Department of Medical Chemistry, Faculty of General Medicine, Hungary 3 University of Oxford, St John's College, United Kingdom AMPA and NMDA type glutamate receptors convey fast synaptic transmission in the CNS and mediate various forms of hippocampal plasticity. AMPA receptors comprise four subunits – GluA1, GluA2, GluA3 and GluA4 (or GluR1-4) – that combine to form tetramers. GluA1 is central in synaptic plasticity: disruption of glutamate receptor type 1 causes synaptic alterations and learning/memory deficits in mice. Phosphorylation of serines 831 and 845 in the GluA1 subunit by CaMKII and PKA respectively regulates the ion channel properties and synaptic trafficking of GluR1-containing AMPARs during hippocampal LTP. Moreover, altered phosphorylation pattern of GluA1 was shown in various forms of mental disorders, like addiction, mania and major depressive disorder. Tianeptine, an atypical antidepressant, have been shown to increase the phosphorylation level of GluA1 on the CaMKII and PKA site, resulting in enhanced AMPA-evoked spiking activity in vivo. Here we investigated whether the increased level of GluA1 phosphorylation has a specific “fingerprint” on the AMPA-evoked spiking pattern of single CA1 cell in vivo. AMPA evoked spiking trains were recorded from the CA1 region of chloral-hydrate anaesthetized male rats before and after i.p. Tianeptine application. In another set of experiments, weak and strong AMPA and NMDA-stimulation was applied for evoking spike-trains. By comparing AMPA evoked spike-trains with similar firing rates, we show that the spike-trains recorded after Tianeptine application show characteristic features, distinguishing from spike-trains triggered by strong AMPA stimulation. Despite the similar firing rates, the interspike interval distributions are significantly different among the two groups, showing that neuronal output may differ when new receptors are activated compared to posttranscriptional regulation of previously activated receptors. Furthermore, we also show that NMDA evokes spiking activity with different pattern than AMPA induced spike-trains. Conference: IBRO International Workshop 2010, Pécs, Hungary, 21 Jan - 23 Jan, 2010. Presentation Type: Poster Presentation Topic: Disorders of the nervous system Citation: Barkóczi B, Juhász G, Averkin R, Vörös I, Penke B and Szegedi V (2010). Analyzing NMDA and AMPA evoked spike-trains in CA1: GluA1 phosphorylation specific patterns in vivo. Front. Neurosci. Conference Abstract: IBRO International Workshop 2010. doi: 10.3389/conf.fnins.2010.10.00025 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: 19 Apr 2010; Published Online: 19 Apr 2010. * Correspondence: Balázs Barkóczi, Bay Zoltán Foundation for Applied Research, BAYGEN, Szeged, Hungary, barkoczib@baygen.hu 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 Balázs Barkóczi Gábor Juhász Robert Averkin Imre Vörös Botond Penke Viktor Szegedi Google Balázs Barkóczi Gábor Juhász Robert Averkin Imre Vörös Botond Penke Viktor Szegedi Google Scholar Balázs Barkóczi Gábor Juhász Robert Averkin Imre Vörös Botond Penke Viktor Szegedi PubMed Balázs Barkóczi Gábor Juhász Robert Averkin Imre Vörös Botond Penke Viktor Szegedi 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.

The replacement of GluN2B with GluN2A increases synaptic trafficking of AMPA receptors in the neonatal mouse brain

The replacement of GluN2B with GluN2A increases synaptic trafficking of AMPA receptors in the neonatal mouse brain

Stress at Learning Facilitates Memory Formation by Regulating AMPA Receptor Trafficking Through a Glucocorticoid Action

Stress and glucocorticoids (GCs) can facilitate memory formation. However, the molecular mechanisms mediating their effects are largely unknown. Alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor (AMPAR) trafficking has been implicated in the changes in synaptic strength at central glutamatergic synapses associated with memory formation. In cell cultures, corticosterone has been shown to condition the synaptic trafficking of the AMPAR GluA2 subunit. In this study, we investigated the involvement of GluA2 trafficking in the facilitation of learning by stress. Using the water maze spatial task involving different stress levels, mice trained under more stressful conditions (water at 22 degrees C) showed better learning and memory, and higher post-training corticosterone levels, than mice trained under lower stress (water at 30 degrees C). Strikingly, this facilitated learning by stress was accompanied by enhanced synaptic expression of GluA2 AMPARs that was not observed in mice trained under less stressful conditions. Interfering with GC actions by injecting the GC synthesis inhibitor, metyrapone, blocked both the memory facilitation and the enhanced GluA2 trafficking induced by stressful learning. Intracerebroventricular infusion of the peptide, pep2m, that blocks GluA2 synaptic trafficking by interfering with the interaction between N-ethylmaleimide-sensitive factor and GluA2, impaired immediate performance at learning as well as long-term memory retrieval, supporting a causal role for GluA2 trafficking in stress-induced facilitation of spatial learning and memory. Evidence for the involvement of the neural cell adhesion molecule N-cadherin in interaction with GluA2 is also provided. These findings underscore a new mechanism whereby stress can improve memory function.

Metabotropic glutamate receptor 4 interacts with microtubule-associated protein 1B

The metabotropic glutamate receptor 4 (mGluR4) is a G-protein-coupled receptor that mediates inhibition of neurotransmitter release. Here, we used a proteomic approach to identify novel interaction partners of mGluR4 and report that the cytoplasmic C-terminal tail of mGluR4 interacts with microtubule-associated protein 1B (MAP1B). Binding of MAP1B to mGluR4 is inhibited by Ca 2+/calmodulin, and MAP1B and mGluR4 colocalize at excitatory synapses in cultured hippocampal neurons. Thus, MAP1B might be implicated in the synaptic trafficking and/or regulation of mGluR4.

Activity Level-Dependent Synapse-Specific AMPA Receptor Trafficking Regulates Transmission Kinetics

Central glutamatergic synapses may express AMPA-sensitive glutamate receptors (AMPA-Rs) with distinct gating properties and exhibit different transmission dynamics, which are important for computing various synaptic inputs received at different populations of synapses. However, how glutamatergic synapses acquire AMPA-Rs with distinct kinetics to influence synaptic integration remains poorly understood. Here I report synapse-specific trafficking of distinct AMPA-Rs in rat cortical layer 4 stellate and layer 5 pyramidal neurons. The analysis indicates that in single layer 4 stellate neurons thalamocortical synapses generate faster synaptic responses than intracortical synapses. Moreover, GluR1-containing AMPA-Rs traffic selectively into intracortical synapses, and this process requires sensory experience-dependent activity and slows down transmission kinetics. GluR4-containing AMPA-Rs traffic more heavily into thalamocortical synapses than intracortical synapses, and this process requires spontaneous synaptic activity and speeds up transmission kinetics. GluR2-containing AMPA-Rs traffic equally into both thalamocortical and intracortical synapses, and this process requires no synaptic activity and resets transmission kinetics. Notably, synaptic trafficking of distinct AMPA-Rs differentially regulates synaptic integration. Thus, synapse-specific AMPA-R trafficking coarsely sets and synaptic activity finely tunes transmission kinetics and integration properties at different synapses in central neurons.

Induction and expression of GluA1 (GluR‐A)‐independent LTP in the hippocampus

Long-term potentiation (LTP) at hippocampal CA3–CA1 synapses is thought to be mediated, at least in part, by an increase in the postsynaptic surface expression of α-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) receptors induced by N-methyl-d-aspartate (NMDA) receptor activation. While this process was originally attributed to the regulated synaptic insertion of GluA1 (GluR-A) subunit-containing AMPA receptors, recent evidence suggests that regulated synaptic trafficking of GluA2 subunits might also contribute to one or several phases of potentiation. However, it has so far been difficult to separate these two mechanisms experimentally. Here we used genetically modified mice lacking the GluA1 subunit (Gria1−/− mice) to investigate GluA1-independent mechanisms of LTP at CA3–CA1 synapses in transverse hippocampal slices. An extracellular, paired theta-burst stimulation paradigm induced a robust GluA1-independent form of LTP lacking the early, rapidly decaying component characteristic of LTP in wild-type mice. This GluA1-independent form of LTP was attenuated by inhibitors of neuronal nitric oxide synthase and protein kinase C (PKC), two enzymes known to regulate GluA2 surface expression. Furthermore, the induction of GluA1-independent potentiation required the activation of GluN2B (NR2B) subunit-containing NMDA receptors. Our findings support and extend the evidence that LTP at hippocampal CA3–CA1 synapses comprises a rapidly decaying, GluA1-dependent component and a more sustained, GluA1-independent component, induced and expressed via a separate mechanism involving GluN2B-containing NMDA receptors, neuronal nitric oxide synthase and PKC.

Dopamine D4 Receptors Regulate GABAA Receptor Trafficking via an Actin/Cofilin/Myosin-dependent Mechanism

The GABA(A) receptor-mediated inhibitory transmission in prefrontal cortex (PFC) is implicated in cognitive processes such as working memory. Our previous study has found that GABA(A)R current is subject to the regulation of dopamine D(4) receptors, a PFC-enriched neuromodulator critically involved in various mental disorders associated with PFC dysfunction. In this study, we have investigated the cellular mechanism underlying D(4) modulation of GABA(A)Rs. We found that the density of surface clusters of GABA(A)R beta2/3 subunits was reduced by D(4), suggesting that the D(4) reduction of GABA(A)R current is associated with a decrease in functional GABA(A)Rs at the plasma membrane. Moreover, the D(4) reduction of GABA(A)R current was blocked by the actin stabilizer phalloidin and was occluded by the actin destabilizer latrunculin, suggesting that D(4) regulates GABA(A)R trafficking via an actin-dependent mechanism. Cofilin, a major actin depolymerizing factor whose activity is strongly increased by dephosphorylation at Ser(3), provides the possible link between D(4) signaling and the actin dynamics. Because myosin motor proteins are important for the transport of vesicles along actin filaments, we also tested the potential involvement of myosin in D(4) regulation of GABA(A)R trafficking. We found that dialysis with a myosin peptide, which competes with endogenous myosin proteins for actin-binding sites, prevented the D(4) reduction of GABA(A)R current. These results suggest that D(4) receptor activation increases cofilin activity presumably via its dephosphorylation, resulting in actin depolymerization, thus causing a decrease in the myosin-based transport of GABA(A)R clusters to the surface.