Biomedical subjects

Ronald S Petralia

Publications and source records attributed to Ronald S Petralia.

At least 19 recordsLinked to original sources

PubMed2006

Arc/Arg3.1 interacts with the endocytic machinery to regulate AMPA receptor trafficking.

Arc/Arg3.1 is an immediate-early gene whose mRNA is rapidly transcribed and targeted to dendrites of neurons as they engage in information processing and storage. Moreover, Arc/Arg3.1 is known to be required for durable forms of synaptic plasticity and learning. Despite these intriguing links to plasticity, Arc/Arg3.1's molecular function remains enigmatic. Here, we demonstrate that Arc/Arg3.1 protein interacts with dynamin and specific isoforms of endophilin to enhance receptor endocytosis. Arc/Arg3.1 selectively modulates trafficking of AMPA-type glutamate receptors (AMPARs) in neurons by accelerating endocytosis and reducing surface expression. The Arc/Arg3.1-endocytosis pathway appears to regulate basal AMPAR levels since Arc/Arg3.1 KO neurons exhibit markedly reduced endocytosis and increased steady-state surface levels. These findings reveal a novel molecular pathway that is regulated by Arc/Arg3.1 and likely contributes to late-phase synaptic plasticity and memory consolidation.

Acyltransferases↗

PubMed2006

A novel family of adhesion-like molecules that interacts with the NMDA receptor.

We have identified a novel family of synaptic adhesion-like molecules (SALMs). The family members, SALM1-SALM4, have a single transmembrane (TM) domain and contain extracellular leucine-rich repeats, an Ig C2 type domain, a fibronectin type III domain, and an intracellular postsynaptic density-95 (PSD-95)/Discs large/zona occludens-1 (PDZ) binding domain, which is present on all members except SALM4. SALM1 interacts with PSD-95, synapse-associated protein 102 (SAP102), and SAP97 based on coimmunoprecipitation of detergent-solubilized brain. Distribution studies show that SALM1 is present in synaptic membrane and postsynaptic density fractions but is also distributed in axons and dendrites. Transfection of hippocampal neurons for 4 d in vitro (DIV) with SALM1 more than doubles the dendritic lengths of neurons after 48 h, whereas transfection of neurons 14 DIV has no significant effect on neurite outgrowth. Overexpression of SALM1 in 14 DIV neurons recruits NMDA receptors (NR) and PSD-95 to dendritic puncta. This effect is dependent on the PDZ-binding domain of SALM1. SALM1 also enhances surface expression of transfected NR2A subunit. Immunoprecipitation of detergent-solubilized brain membranes with anti-SALM1 antibodies shows coimmunoprecipitation of NR1 and NR2 subunits. After transfection of heterologous cells with NR1 and NR2 cDNAs, through coimmunoprecipitation analyses, we find that SALM1 also interacts with the NMDA receptor NR1 subunit through its extracellular or TM1 domains.

Amino Acid Sequence↗

PubMed2006

Partially overlapping distribution of epsin1 and HIP1 at the synapse: analysis by immunoelectron microscopy.

Synapses of neurons use clathrin-mediated endocytic pathways for recycling of synaptic vesicles and trafficking of neurotransmitter receptors. Epsin 1 and huntingtin-interacting protein 1 (HIP1) are endocytic accessory proteins. Both proteins interact with clathrin and the AP2 adaptor complex and also bind to the phosphoinositide-containing plasma membrane via an epsin/AP180 N-terminal homology (ENTH/ANTH) domain. Epsin1 and HIP1 are found in neurons; however, their precise roles in synapses remain largely unknown. Using immunogold electron microscopy, we examine and compare the synaptic distribution of epsin1 and HIP1 in rat CA1 hippocampal synapse. We find that epsin1 is located across both sides of the synapse, whereas HIP1 displays a preference for the postsynaptic compartment. Within the synaptic compartments, espin1 is distributed similarly throughout, whereas postsynaptic HIP1 is concentrated near the plasma membrane. Our results suggest a dual role for epsin1 and HIP1 in the synapse: as broadly required factors for promoting clathrin assembly and as adaptors for specific endocytic pathways.

Adaptor Proteins, Vesicular Transport↗

PubMed2005

mPins modulates PSD-95 and SAP102 trafficking and influences NMDA receptor surface expression.

Appropriate trafficking and targeting of glutamate receptors (GluRs) to the postsynaptic density is crucial for synaptic function. We show that mPins (mammalian homologue of Drosophila melanogaster partner of inscuteable) interacts with SAP102 and PSD-95 (two PDZ proteins present in neurons), and functions in the formation of the NMDAR-MAGUK (N-methyl-D-aspartate receptor-membrane-associated guanylate kinase) complex. mPins enhances trafficking of SAP102 and NMDARs to the plasma membrane in neurons. Expression of dominant-negative constructs and short-interfering RNA (siRNA)-mediated knockdown of mPins decreases SAP102 in dendrites and modifies surface expression of NMDARs. mPins changes the number and morphology of dendritic spines and these effects depend on its Galphai interaction domain, thus implicating G-protein signalling in the regulation of postsynaptic structure and trafficking of GluRs.

Animals↗

PubMed2005

TARP gamma-8 controls hippocampal AMPA receptor number, distribution and synaptic plasticity.

Synaptic plasticity involves activity-dependent trafficking of AMPA-type glutamate receptors. Numerous cytoplasmic scaffolding proteins are postulated to control AMPA receptor trafficking, but the detailed mechanisms remain unclear. Here, we show that the transmembrane AMPA receptor regulatory protein (TARP) gamma-8, which is preferentially expressed in the mouse hippocampus, is important for AMPA receptor protein levels and extrasynaptic surface expression. By controlling the number of AMPA receptors, gamma-8 is also important in long-term potentiation, but not long-term depression. This study establishes gamma-8 as a critical protein for basal AMPA receptor expression and localization at extrasynaptic sites in the hippocampus and raises the possibility that TARP-dependent control of AMPA receptors during synapse development and plasticity may be widespread.

Animals↗

PubMed2005

Persistent hippocampal CA1 LTP in mice lacking the C-terminal PDZ ligand of GluR1.

The C-terminal PDZ ligand of the AMPA receptor GluR1 subunit may be important for expression of CA1 hippocampal long-term potentiation. To test this directly in vivo, we generated a knock-in mouse lacking the last seven residues of GluR1, comprising the PDZ ligand. This deletion did not affect basal GluR1 synaptic localization, basal synaptic transmission, long-term potentiation or long-term depression, indicating that the ligand is not required for CA1 hippocampal synaptic plasticity.

Animals↗

PubMed2005

Synaptic distribution of the endocytic accessory proteins AP180 and CALM.

Clathrin-coated vesicles mediate a variety of endocytosis pathways in cells, including endocytic events at synapses. AP180 and clathrin assembly lymphoid myeloid leukemia protein (CALM) are clathrin accessory proteins that promote the formation of clathrin-coated vesicles. Both proteins bind to membrane lipids through their epsin N-terminal homology domains and interact with clathrin and related protein components through their carboxyl-terminal peptide motifs. We examine their neuronal expression and synaptic distribution. We show that both proteins are detected in synapses but demonstrate different distribution patterns. AP180 is located predominantly in presynaptic profiles, whereas CALM is found nonselectively in pre- and postsynaptic profiles and also in perisynaptic processes. These observations reveal an unexpected relationship between AP180 and the presumed non-neuronal homologue CALM. We propose that both AP180 and CALM function as endocytic accessory proteins at synapses, but each may regulate distinct clathrin pathways.

Animals↗

PubMed2005

Ontogeny of postsynaptic density proteins at glutamatergic synapses.

In glutamatergic synapses, glutamate receptors (GluRs) associate with many other proteins involved in scaffolding and signal transduction. The ontogeny of these postsynaptic density (PSD) proteins involves changes in their composition during development, paralleling changes in GluR type and function. In the CA1 region of the hippocampus, at postnatal day 2 (P2), many synapses already have a distinct PSD. We used immunoblot analysis, subcellular fractionation, and quantitative immunogold electron microscopy to examine the distribution of PSD proteins during development of the hippocampus. Synapses at P2 contained substantial levels of NR1 and NR2B and most GluR-associated proteins, including SAP102, SynGAP, the chain of proteins from GluRs/SAP102 through GKAP/Shank/Homer and metabotropic glutamate receptors, and the adhesion factors, cadherin, catenin, neuroligin, and Nr-CAM. Development was marked by substantial decreases in NR2B and SAP102 and increases in NR2A, PSD-95, AMPA receptors, and CaMKII. Other components showed more moderate changes.

Animals↗

PubMed2005

Roles of 3-deoxy-D-manno-2-octulosonic acid transferase from Moraxella catarrhalis in lipooligosaccharide biosynthesis and virulence.

Lipooligosaccharide (LOS), a major outer membrane component of Moraxella catarrhalis, is a possible virulence factor in the pathogenesis of human infections caused by the organism. However, information about the roles of the oligosaccharide chain from LOS in bacterial infection remains limited. Here, a kdtA gene encoding 3-deoxy-D-manno-2-octulosonic acid (Kdo) transferase, which is responsible for adding Kdo residues to the lipid A portion of the LOS, was identified by transposon mutagenesis and construction of an isogenic kdtA mutant in strain O35E. The resulting O35EkdtA mutant produced only lipid A without any core oligosaccharide, and it was viable. Physicochemical and biological analysis revealed that the mutant was susceptible to hydrophobic reagents and a hydrophilic glycopeptide and was sensitive to bactericidal activity of normal human serum. Importantly, the mutant showed decreased toxicity by the Limulus amebocyte lysate assay, reduced adherence to human epithelial cells, and enhanced clearance in lungs and nasopharynx in a mouse aerosol challenge model. These data suggest that the oligosaccharide moiety of the LOS is important for the biological activity of the LOS and the virulence capability of the bacteria in vitro and in vivo. This study may bring new insights into novel vaccines or therapeutic interventions against M. catarrhalis infections.

Bacterial Adhesion↗

PubMed2004

AMPA receptor subunit expression in chick vestibular nucleus neurons.

The principal cells of the chick tangential nucleus are vestibular nucleus neurons whose responses on vestibular nerve stimulation are abolished by glutamate receptor antagonists. Using confocal microscopy, we quantified immunolabeling for AMPA receptor subunits GluR1, GluR2, GluR2/3, and GluR4 in principal cells that were identified by the neuronal marker, microtubule-associated protein 2 (MAP2). This work was focused primarily on 9 days after hatching (H9) when the principal cells have acquired some important mature electrophysiologic properties. At H9, the principal cell bodies stained strongly with GluR2/3 and GluR4, whereas GluR1 and GluR2 produced weak signals. Moreover, GluR2/3 and GluR4 receptor subunit clusters in principal cell bodies and dendrites were localized at sites contacted by biocytin-labeled vestibular nerve terminals and synaptotagmin-labeled terminals. Developmental expression of AMPA receptor immunolabeling was studied in the principal cell bodies at embryonic day 16 (E16) and hatching (H1). At E16, labeling for GluR4 was already strong, and continued to increase at H1 and H9. In contrast, GluR2/3 labeling was weak at E16, but increased significantly at H1, and more so by H9. GluR1 and GluR2 were present at low levels at E16 and H1. From E16 to H9, overall AMPA receptor subunit expression increased steadily, with H9 showing the strongest labeling. Ultrastructural observations at E16 and H3 confirmed the presence of immunogold labeling for AMPA receptor subunits at the vestibular nerve and non-vestibular nerve synapses on the principal cell bodies. In summary, these results indicate that GluR3 and GluR4 are the major AMPA receptor subunits involved in excitatory synaptic transmission in principal cells during the perinatal period.

Age Factors↗

PubMed2004

Unique domain anchoring of Src to synaptic NMDA receptors via the mitochondrial protein NADH dehydrogenase subunit 2.

Src is the prototypic protein tyrosine kinase and is critical for controlling diverse cellular functions. Regions in Src define structural and functional domains conserved in many cell signaling proteins. Src also contains a region of low sequence conservation termed the unique domain, the function of which has until now remained enigmatic. Here, we show that the unique domain of Src is a protein-protein interaction region and we identify NADH dehydrogenase subunit 2 (ND2) as a Src unique domain-interacting protein. ND2 is a subunit of complex I in mitochondria, but we find that ND2 interacts with Src outside this organelle at excitatory synapses in the brain. ND2 acts as an adapter protein anchoring Src to the N-methyl-d-aspartate (NMDA) receptor complex, and is crucial for Src regulation of synaptic NMDA receptor activity. By showing an extramitochondrial action for a protein encoded in the mitochondrial genome, we identify a previously unsuspected means by which mitochondria regulate cellular function, suggesting a new paradigm that may be of general relevance for control of Src signaling.

Animals↗

PubMed2004

Differences in the expression of AMPA and NMDA receptors between axospinous perforated and nonperforated synapses are related to the configuration and size of postsynaptic densities.

Axospinous synapses are traditionally divided according to postsynaptic density (PSD) configuration into a perforated subtype characterized by a complex-shaped PSD and nonperforated subtype exhibiting a simple-shaped, disc-like PSD. It has been hypothesized that perforated synapses are especially important for synaptic plasticity because they have a higher efficacy of impulse transmission. The aim of the present study was to test this hypothesis. The number of postsynaptic AMPA receptors (AMPARs) is widely regarded as the major determinant of synaptic efficacy. Therefore, the expression of AMPARs was evaluated in the two synaptic subtypes and compared with that of NMDA receptors (NMDARs). Postembedding immunogold electron microscopy was used to quantify the immunoreactivity following single labeling of AMPARs or NMDARs in serial sections through the CA1 stratum radiatum of adult rats. The results showed that all perforated synapses examined were immunopositive for AMPARs. In contrast, only a proportion of nonperforated synapses (64% on average) contained immunogold particles for AMPARs. The number of immunogold particles for AMPARs was markedly and significantly higher in perforated synapses than in immunopositive nonperforated synapses. Although all synapses of both subtypes were NMDAR immunopositive perforated synapses contained significantly more immunogold particles for NMDARs than nonperforated ones. Multivariate analysis of variance revealed that the mode of AMPAR and NMDAR expression is related to the complexity of PSD configuration, not only to PSD size. These findings support the notion that perforated synapses may evoke larger postsynaptic responses relative to nonperforated synapses and, hence, contribute to an enhancement of synaptic transmission associated with some forms of synaptic plasticity.

Animals↗

PubMed2004

Loss of GLUR2 alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid receptor subunit differentially affects remaining synaptic glutamate receptors in cerebellum and cochlear nuclei.

The alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA) type of ionotropic glutamate receptor is the major mediator of fast neurotransmission in the brain and spinal cord. Most AMPA receptors are impermeable to calcium because they contain the GluR2 subunit. However, some AMPA receptors lack GluR2 and pass calcium which can mediate synaptic plasticity and, in excess, neurotoxicity. Previously, we showed a decrease in the density of synaptic AMPA receptors in the hippocampus of mice lacking GluR2. In this study, using these GluR2-lacking mice, we examined other areas of the brain that differ in the amount of GluR2 normally present. Like hippocampal spines, cerebellar Purkinje spines normally express AMPA receptors with high GluR2 and showed a decrease in synaptic AMPA receptors in mutant mice. In contrast, neurons that normally express AMPA receptors with little or no GluR2, such as in the anteroventral cochlear nucleus, showed no decrease in AMPA receptors and even showed an increase in one AMPA receptor subunit. These two different patterns may relate to preadaptations to prevent calcium neurotoxicity; such mechanisms might be absent in Purkinje and hippocampal spines so that these neurons must decrease their total expression of synaptic AMPA receptors (calcium permeable in mutant mice) to prevent calcium neurotoxicity. In addition, we found that another glutamate receptor, GluRdelta2, which is abundant only in parallel fibre synapses on Purkinje cells and in the dorsal cochlear nucleus, is up-regulated at these synapses in mutant mice; this probably reflects some change in GluRdelta2 targeting to these synapses.

Animals↗

PubMed2003

Activation of the TRPC1 cation channel by metabotropic glutamate receptor mGluR1.

Group I metabotropic glutamate receptors (consisting of mGluR1 and mGluR5) are G-protein-coupled neurotransmitter receptors that are found in the perisynaptic region of the postsynaptic membrane. These receptors are not activated by single synaptic volleys but rather require bursts of activity. They are implicated in many forms of neural plasticity including hippocampal long-term potentiation and depression, cerebellar long-term depression, associative learning, and cocaine addiction. When activated, group I mGluRs engage two G-protein-dependent signalling mechanisms: stimulation of phospholipase C and activation of an unidentified, mixed-cation excitatory postsynaptic conductance (EPSC), displaying slow activation, in the plasma membrane. Here we report that the mGluR1-evoked slow EPSC is mediated by the TRPC1 cation channel. TRPC1 is expressed in perisynaptic regions of the cerebellar parallel fibre-Purkinje cell synapse and is physically associated with mGluR1. Manipulations that interfere with TRPC1 block the mGluR1-evoked slow EPSC in Purkinje cells; however, fast transmission mediated by AMPA-type glutamate receptors remains unaffected. Furthermore, co-expression of mGluR1 and TRPC1 in a heterologous system reconstituted a mGluR1-evoked conductance that closely resembles the slow EPSC in Purkinje cells.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed2003

Aberrant formation of glutamate receptor complexes in hippocampal neurons of mice lacking the GluR2 AMPA receptor subunit.

The number and type of receptors present at the postsynaptic membrane determine the response to the neurotransmitter released from the presynaptic terminal. Because most neurons receive multiple and distinct synaptic inputs and contain several different subtypes of receptors stimulated by the same neurotransmitter, the assembly and trafficking of receptors in neurons is a complex process involving many levels of regulation. To investigate the mechanism that neurons use to regulate the assembly of receptor subunits, we studied a GluR2 knock-out mouse. GluR2 is a critical subunit that controls calcium permeability of AMPA receptors and is present in most native AMPA receptors. Our data indicate that in the absence of GluR2, aberrant receptor complexes composed of GluR1 and GluR3 are formed in the hippocampus, and that there is an increased number of homomeric GluR1 and GluR3 receptors. We also show that these homomeric and heteromeric receptors are less efficiently expressed at the synapse. Our results show that GluR2 plays a critical role in controlling the assembly of AMPA receptors, and that the assembly of subunits may reflect the affinity of one subunit for another or the stability of intermediates in the assembly process. Therefore, GluR1 may have a greater preference for GluR2 than it does for GluR3.

Animals↗

PubMed2003

Distribution of kainate receptor subunits at hippocampal mossy fiber synapses.

Kainate receptors function as mediators of postsynaptic currents and as presynaptic modulators of synaptic transmission at mossy fiber synapses. Despite intense research into the physiological properties of mossy fiber kainate receptors, their subunit composition in the presynaptic and postsynaptic compartments is unclear. Here we describe the distribution of kainate receptor subunits in mossy fiber synapses using subunit-selective antibodies and knock-out mice. We provide morphological evidence for the presynaptic localization of KA1 and KA2 receptor subunits at mossy fiber synapses. Immunogold staining for KA1 and KA2 was commonly seen at synaptic contacts and in vesicular structures. Postsynaptic labeling in dendritic spines was also observed. Although KA1 predominantly showed presynaptic localization, KA2 was concentrated to a greater degree on postsynaptic membranes. Both subunits coimmunoprecipitated from hippocampal membrane extracts with GluR6 but not GluR7 subunits. These results demonstrate that KA1 and KA2 subunits are localized presynaptically and postsynaptically at mossy fiber synapses where they most likely coassemble with GluR6 subunits to form functional heteromeric kainate receptor complexes.

Animals↗

PubMed2003

Narp and NP1 form heterocomplexes that function in developmental and activity-dependent synaptic plasticity.

Narp is a neuronal immediate early gene that plays a role in excitatory synaptogenesis. Here, we report that native Narp in brain is part of a pentraxin complex that includes NP1. These proteins are covalently linked by disulfide bonds into highly organized complexes, and their relative ratio in the complex is dynamically dependent upon the neuron's activity history and developmental stage. Complex formation is dependent on their distinct N-terminal coiled-coil domains, while their closely homologous C-terminal pentraxin domains mediate association with AMPA-type glutamate receptors. Narp is substantially more effective in assays of cell surface cluster formation, coclustering of AMPA receptors, and excitatory synaptogenesis, yet their combined expression results in supraadditive effects. These studies support a model in which Narp can regulate the latent synaptogenic activity of NP1 by forming mixed pentraxin assemblies. This mechanism appears to contribute to both activity-independent and activity-dependent excitatory synaptogenesis.

Amino Acid Sequence↗

PubMed2003

Impaired NMDA receptor-mediated postsynaptic function and blunted NMDA receptor-dependent persistent pain in mice lacking postsynaptic density-93 protein.

Modification of synaptic NMDA receptor (NMDAR) expression influences NMDAR-mediated synaptic function and associated persistent pain. NMDARs directly bind to a family of membrane-associated guanylate kinases (MAGUKs) that regulate surface and synaptic NMDAR trafficking in the CNS. We report here that postsynaptic density-93 protein (PSD-93), a postsynaptic neuronal MAGUK, is expressed abundantly in spinal dorsal horn and forebrain, where it colocalizes and interacts with NMDAR subunits NR2A and NR2B. Targeted disruption of the PSD-93 gene reduces not only surface NR2A and NR2B expression but also NMDAR-mediated excitatory postsynaptic currents and potentials, without affecting surface AMPA receptor expression or its synaptic function, in the regions mentioned above. Furthermore, mice lacking PSD-93 exhibit blunted NMDAR-dependent persistent pain induced by peripheral nerve injury or injection of Complete Freund's Adjuvant, although they display intact nociceptive responsiveness to acute pain. PSD-93 appears to be important for NMDAR synaptic targeting and function and to be a potential biochemical target for the treatment of persistent pain.

Animals↗