Enhancement of glutamate uptake mediates the neuroprotection exerted by activating group II or III metabotropic glutamate receptors on astrocytes

We investigated whether the activation of astroglial group II and III metabotropic glutamate receptors (mGluRs) could exert neuroprotective effects and whether the neuroprotection was related to glutamate uptake. Our results showed that the activation of astroglial group II or III mGluRs exerted neuroprotection against 1-methyl-4-phenylpyridinium (MPP+) astroglial conditioned medium-induced neurotoxicity in midbrain neuron cultures. Furthermore, MPP+ decreased glutamate uptake of primary astrocytes and C6 glioma cells, which was recovered by activating group II or III mGluRs. Specific group II or III mGluRs antagonists completely abolished the neuroprotective effects and the enhancement of glutamate uptake of their respective agonists. Our results showed that the primary cultured rat astrocytes and C6 glioma cells expressed receptor proteins for group II mGluR2/3, group III mGluR4, mGluR6 and mGluR7. C6 glioma cells expressed mRNA for group II mGluR3, group III mGluR4, mGluR6, mGluR7 and mGluR8. In conclusion, we confirmed that the activation of astroglial mGluRs exerted neuroprotection, and demonstrated that the mechanism underlying this protective role was at least partially related to the enhancement of glutamate uptake.     

Phosphorylation of Mitogen-Activated Protein Kinase in Cultured Rat Cortical Glia by Stimulation of Metabotropic Glutamate Receptors

Abstract: Activation of metabotropic glutamate receptors (mGluRs) in glia results in significant physiological effects for both the glia and the neighboring neurons; but in many cases, the mGluR subtypes and signal transduction mechanisms mediating these effects have not been determined. In this study, we report that mGluR activation in primary cultures of rat cortical glia results in tyrosine phosphorylation of several proteins, including p44/p42 mitogen-activated protein kinases, also referred to as extracellular signal-regulated kinases (ERK1/2). Incubation of glial cultures with the general mGluR agonist 1-aminocyclopentane-1 S ,3 R -dicarboxylate and the mGluR group I-selective agonists ( RS )-3,5-dihydroxyphenylglycine (DHPG) and l -quisqualate resulted in increased tyrosine phosphorylation of ERK1/2. The group II-selective agonist (2 S ,2' R ,3' R )-2-(2',3'-dicarboxycyclopropyl)glycine and group III-selective agonist l (+)-2-amino-4-phosphonobutyric acid had no effect on tyrosine phosphorylation. DHPG-induced ERK1/2 phosphorylation could be inhibited by an antagonist that acts at group I or group II mGluRs but not by antagonists for group II and group III mGluRs. Protein kinase C (PKC) activators also induced ERK1/2 phosphorylation, but the PKC inhibitor bisindolylmaleimide I did not inhibit DHPG-induced ERK1/2 phosphorylation at a concentration that inhibited the response to phorbol 12,13-dibutyrate. These data suggest that mGluR activation of ERK1/2 in cultured glia is mediated by group I mGluRs and that this effect is independent of PKC activation. Furthermore, immunoblots with antibodies against various mGluR subtypes show expression of mGluR5, but no other mGluRs in our cultures. Taken together, these results suggest that mGluR5 stimulation results in tyrosine phosphorylation of ERK1/2 and other glial proteins.     

Group I metabotropic glutamate receptors mediate phospholipase D stimulation in rat cultured astrocytes

We have studied the activation of phospholipase D (PLD) by glutamate in rat cultured astrocytes by measuring the PLD-catalyzed formation of [32P]phosphatidylbutanol in [32P]Pi-prelabeled cells, stimulated in the presence of butanol. Glutamate elicited the activation of PLD in cortical astrocytes but not in cortical neurons, whereas similar glutamate activation of phosphoinositide phospholipase C was found in both astrocytes and neurons. The extent of PLD stimulation by glutamate was similar in astrocytes from brain cortex and hippocampus, but no effect was found in cerebellar astrocytes. In cortical astrocytes, the glutamate response was insensitive to antagonists of ionotropic glutamate receptors and was reproduced by agonists of metabotropic glutamate receptors (mGluRs) with a rank order of agonist potency similar to that reported for group I mGluR-mediated phosphoinositide phospholipase activation [quisqualate > (S)-3,5-dihydroxyphenylglycine > (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid]. The response to (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid was inhibited by the mGluR antagonist (S)-alpha-methyl-4-carboxyphenylglycine and, less potently, by 1-aminoindan-1,5-dicarboxylic acid and 4-carboxyphenylglycine, two antagonists of group I mGluRs that display higher potency on mGluR1 than on mGluR5. The mGluR5-selective agonist (RS)-2-chloro-5-hydroxyphenylglycine also activated PLD in astrocytes. These findings indicate the involvement of group I mGluRs, most likely mGluR5, in the glutamate activation of PLD in cultured rat cortical astrocytes.     

MGluRs regulate the expression of neuronal calcium sensor proteins NCS-1 and VILIP-1 and the immediate early gene arg3.1/arc in the hippocampus in vivo

The metabotropic glutamate receptor (mGluR) agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) is involved in several forms of hippocampal synaptic plasticity. DHPG application can induce slow-onset potentiation, a form of long-term potentiation (LTP), in the dentate gyrus and in the CA1 region in vivo. The induction of LTP correlates with increased expression levels of neuronal calcium sensor (NCS), considered as key elements for plasticity. In this study we investigated mGluR- and time-dependent changes in the expression of two different NCS proteins. Following DHPG application in vivo NCS-1 and VILIP-1 expression increased, with significant levels reached after 8 and 24h. The effect was attenuated by treatment with the group I mGluR specific antagonist S-4-carboxyphenylglycine. The immediate early gene (IEG) arg3.1/arc showed highest expression levels 2h after DHPG-treatment. Therefore, mGluRs at concentrations which induce synaptic plasticity regulate the expression of IEGs and NCS proteins in different time frames and thus contribute to late phases of synaptic plasticity.     

The mechanism of presynaptic long-term depression mediated by group I metabotropic glutamate receptors

1. Metabotropic glutamate receptors (mGluRs) are known to play a role in synaptic plasticity. In a study of rat hippocampal brain slices, we find that a brief perfusion of a group I mGluR agonist, (S)-3,5-dihydroxyphenylglycine (DHPG), induced a robust long-term depression (DHPG-LTD) in area CA1.2. The action was accompanied by an enhancement of the paired-pulse facilitation (PPF) ratio.3. At the same time DHPG enhanced ionophoretic responses to alpha-amino-3- hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA), kainic acid (KA), and N-methyl-D-aspartate (NMDA) in CA1 pyramidal neurons. This was only partially reversed by washing.4. These observations indicate that DHPG exerts two opposing actions, suppression of the synaptic transmission and facilitation of postsynaptic responses. However, the presynaptic action dominates, since the net effect of monosynaptic activation is a reduction of response.5. Perfusion of DHPG reduced three calcium-dependent responses in CA3 pyramidal neurons, which are presynaptic to CA1 neurons. These are calcium spike width and amplitude, after-hyperpolarization (AHP), and spike frequency adaptation (SFA).6. These results suggest that the DHPG-LTD results from modulation of the presynaptic calcium currents by group l mGluRs.     

Localization and function of metabotropic glutamate receptor 8 in the enteric nervous system

The enteric nervous system (ENS) contains glutamatergic neurons, transporters, and functional ionotropic and groups I and II metabotropic glutamate receptors (mGluRs). The aim of this study was to determine whether the ENS contains functional group III mGluRs. RT-PCR demonstrated the expression of mGluR7 and mGluR8 mRNA in rat myenteric ganglia. Western blot analysis confirmed the presence of mGluR8 protein. Immunocytochemistry, in conjunction with confocal microscopy, demonstrated mGluR8 immunoreactivity in the ENS of several species, including humans. mGluR8 immunoreactivity was localized to the membrane of nerve cell bodies that received glutamatergic input. Significant receptor internalization of mGluR8 was observed on activation, and localization to membrane was observed on blocking with the mGluR III antagonist (RS)-cyclopropyl-4-phosphonophenylglycine (CPPG). mGluR8-positive myenteric neurons contained glutamate or nitric oxide synthase (NOS), a marker of inhibitory motorneurons. Enteric group III mGluRs are functional because mGluR8 agonists inhibited forskolin-induced accumulation of cAMP in isolated myenteric ganglia, and CPPG reduced this effect. In addition, an accelerating effect on guinea pig colonic motility was observed after the application of mGluR8 agonists. Increase in motility was specific, because CPPG inhibited it. Moreover, in the presence of hexamethonium or Nomega-nitro-l-arginine methyl ester, an inhibitor of NOS, responses caused by mGluR8 agonists were abolished. mGluR8 agonists also increased longitudinal muscle contractions. These findings suggest that mGluR8 agonists increase motility by inhibiting nitrergic relaxation and possibly by facilitating cholinergic contractions.     

Nicotine has a permissive role on the activation of metabotropic glutamate 5 receptors coexisting with nicotinic receptors on rat hippocampal noradrenergic nerve terminals

The existence of metabotropic glutamate receptors (mGluRs) on hippocampal noradrenergic nerve terminals and their interaction with coexisting nicotinic acetylcholine receptors (nAChRs) were investigated in superfused rat synaptosomes using [(3)H]-noradrenaline ([(3)H]-NA) release as a readout. The selective agonist of group I mGluRs, (S)-3,5-dihydroxyphenylglycine (DHPG), inactive on its own, acquired ability to release [(3)H]-NA when added together with (-)-nicotine. The effect of DHPG was prevented by 2-methyl-6-(phenylethynyl)-pyridine (MPEP), a selective antagonist of mGluR5, but not by 7-(hydroxyimino)cyclopropane[b]chromen-1-carboxylate ethyl ester (CPCCOEt), selective antagonist of mGluR1. The [(3)H]-NA release evoked by (-)-nicotine plus DHPG was totally abrogated by the nAChR antagonist mecamylamine. Veratrine mimicked the permissive role of (-)-nicotine on the activation of mGluR5 mediating [(3)H]-NA release. The mGluR5-mediated component of the [(3)H]-NA release provoked by DHPG plus (-)-nicotine was blocked by xestospongin C, a selective antagonist of inositoltrisphosphate (IP(3)) receptors. It can be concluded that (i) release-enhancing mGluRs of subtype 5 exist on hippocampal noradrenergic axon terminals; (ii) activation of mGluR5 to mediate IP(3)-dependent NA release requires activation of depolarizing nAChRs coexisting on the same terminals.     

NMDA receptor, PKC and ERK prevent fos expression induced by the activation of group I metabotropic glutamate receptors in the spinal trigeminal subnucleus oralis

Fos, a protein product of immediate early gene c-fos, has been used as a marker for activation of nociceptive neurons in central nervous system including spinal trigeminal nucleus (Vsp). By noxious stimulation applied to orofacial area, the expression of Fos occurred in the Vsp pars oralis (Vo), the subnucleus receiving inputs from trigeminal primary afferents that predominantly innervate intraoral receptive fields. The present study demonstrates that the in vitro activation of group I metabotropic glutamate receptors (mGluRs; mGluR1 and 5) by bath-application of their well-known agonist (S)-3,5-dihydroxyphenylglycine (DHPG) increased the number of Fos-expressing neurons in the Vo area. In addition, bath application of DHPG caused inward currents, a parameter of neuronal excitation, in the Vo neurons held at −70 mV in voltage-clamp mode of whole-cell recordings. In further experiments characterizing two phenomena, the increased Fos expression in the Vo was mediated by an additive activation of both mGluR1 and mGluR5, which required the activation of N-methyl-D-aspartate (NMDA) receptors, protein kinase C (PKC) and extracellular signal-regulated kinase (ERK). In contrast, the inward currents were mediated only by mGluR1, but not by others. The data resulting from this in vitro study indicate that the DHPG-induced membrane depolarisation or neuronal excitation may be upstream to, or skip, the NMDA receptor, PKC and ERK pathways for the DHPG-induced Fos expression.     

Dopamine-induced plasticity, phospholipase D (PLD) activity and cocaine-cue behavior depend on PLD-linked metabotropic glutamate receptors in amygdala

Cocaine-cue associations induce synaptic plasticity with long lasting molecular and cellular changes in the amygdala, a site crucial for cue-associated memory mechanisms. The underlying neuroadaptations can include marked alterations in signaling via dopamine (DA) receptors (DRs) and metabotropic glutamate (Glu) receptors (mGluRs). Previously, we reported that DR antagonists blocked forms of synaptic plasticity in amygdala slices of Sprague-Dawley rats withdrawn from repeated cocaine administration. In the present study, we investigated synaptic plasticity induced by exogenous DA and its dependence on mGluR signaling and a potential role for phospholipase D (PLD) as a downstream element linked to mGluR and DR signaling. Utilizing a modified conditioned place preference (CPP) paradigm as a functional behavioral measure, we studied the neurophysiological effects after two-weeks to the last cocaine conditioning. We recorded, electrophysiologically, a DR-induced synaptic potentiation in the basolateral to lateral capsula central amygdala (BLA-lcCeA) synaptic pathway that was blocked by antagonists of group I mGluRs, particularly, the PLD-linked mGluR. In addition, we observed 2-2.5 fold increase in PLD expression and 3.7-fold increase in basal PLD enzyme activity. The enhanced PLD activity could be further stimulated (9.3 fold) by a DA D1-like (D1/5R) receptor agonist, and decreased to control levels by mGluR1 and PLD-linked mGluR antagonists. Diminished CPP was observed by infusion of a PLD-linked mGluR antagonist, PCCG-13, in the amygdala 15 minutes prior to testing, two weeks after the last cocaine injection. These results imply a functional interaction between D1/5Rs, group I mGluRs via PLD in the amygdala synaptic plasticity associated with cocaine-cues.     

Developmental regulation of group I metabotropic glutamate receptors in the premature brain and their protective role in a rodent model of periventricular leukomalacia

Cerebral white matter injury in premature infants, known as periventricular leukomalacia (PVL), is common after hypoxia-ischemia (HI). While ionotropic glutamate receptors (iGluRs) can mediate immature white matter injury, we have previously shown that excitotoxic injury to premyelinating oligodendrocytes (preOLs) in vitro can be attenuated by group I metabotropic glutamate receptor (mGluR) agonists. Thus, we evaluated mGluR expression in developing white matter in rat and human brain, and tested the protective efficacy of a central nervous system (CNS)-penetrating mGluR agonist on injury to developing oligodendrocytes (OLs) in vivo. Group I mGluRs (mGluR1 and mGluR5) were strongly expressed on OLs in neonatal rodent cerebral white matter throughout normal development, with highest expression early in development on preOLs. Specifically at P6, mGluR1 and mGLuR5 were most highly expressed on GalC-positive OLs compared to neurons, axons, astrocytes and microglia. Systemic administration of (1S,3R) 1-aminocyclopentane-trans-1,3,-dicarboxylic acid (ACPD) significantly attenuated the loss of myelin basic protein in the white matter following HI in P6 rats. Assessment of postmortem human tissue showed both mGluR1 and mGluR5 localized on immature OLs in white matter throughout development, with mGluR5 highest in the preterm period. These data indicate group I mGluRs are highly expressed on OLs during the peak period of vulnerability to HI and modulation of mGluRs is protective in a rodent model of PVL. Group I mGluRs may represent important therapeutic targets for protection from HI-mediated white matter injury.     

Antagonists of group I metabotropic glutamate receptors do not inhibit induction of ischemic tolerance in gerbil hippocampus

In this study we tested the effect of antagonists of two subtypes of the group I metabotropic glutamate receptors (mGluRs GI) on the induction of ischemic tolerance in relation to brain temperature. These experiments were prompted by indications that glutamate receptors may participate in the mechanisms of ischemic preconditioning. The role of NMDA receptors in the induction of ischemic tolerance has been debated while there is lack of information concerning the involvement of mGluRs GI in this phenomenon. The tolerance to injurious 3 min forebrain ischemia in Mongolian gerbils was induced 48 h earlier by 2 min preconditioning ischemia. Brain temperature was measured using telemetry equipment. EMQMCM and MTEP, antagonists of mGluR1 and mGluR5, respectively, were injected i.p. at a dose of 5 mg/kg. They were administered either before preconditioning ischemia in a single dose or after 2 min ischemia three times every 2 h. Both antagonists did not inhibit the induction of ischemic tolerance. Thus, our data indicate that group I metabotropic glutamate receptors do not play an essential role in the induction of ischemic tolerance.     

Mechanism of regulation of casein kinase I activity by group I metabotropic glutamate receptors

Previously, we reported that (S)-3,5-dihydroxypenylglycine (DHPG), an agonist for group I metabotropic glutamate receptors (mGluRs), stimulates CK1 and Cdk5 kinase activities in neostriatal neurons, leading to enhanced phosphorylation, respectively, of Ser-137 and Thr-75 of DARPP-32 (dopamine and cAMP-regulated phosphoprotein, 32 kDa). We have now investigated the signaling pathway that leads from mGluRs to casein kinase 1 (CK1) activation. In mouse neostriatal slices, the effect of DHPG on phosphorylation of Ser-137 or Thr-75 of DARPP-32 was blocked by the phospholipase Cbeta inhibitor, the Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA/AM), and the calcineurin inhibitor cyclosporin A. In neuroblastoma N2a cells, the effect of DHPG on the activity of transfected HA-tagged CK1(epsilon) was blocked by BAPTA/AM and cyclosporin A. In neostriatal slices, the effect of DHPG on Cdk5 activity was also abolished by BAPTA/AM and cyclosporin A, presumably through blocking activation of CK1. Metabolic labeling studies and phosphopeptide mapping revealed that a set of C-terminal sites in HA-CK1epsilon were transiently dephosphorylated in N2a cells upon treatment with DHPG, and this was blocked by cyclosporin A. A mutant CK1epsilon with a nonphosphorylatable C-terminal domain was not activated by DHPG. Together, these studies suggest that DHPG activates CK1(epsilon) via Ca(2+)-dependent stimulation of calcineurin and subsequent dephosphorylation of inhibitory C-terminal autophosphorylation sites.     

Metabotropic glutamate receptor expression in olfactory receptor neurons from the channel catfish,Ictalurus punctatus

Metabotropic glutamate receptors (mGluRs) were identified in olfactory receptor neurons of the channel catfish, Ictalurus punctatus, by polymerase chain reaction. DNA sequence analysis confirmed the presence of two subtypes, mGluR1 and mGluR3, that were coexpressed with each other and with the putative odorant receptors within single olfactory receptor neurons. Immunocytochemical data showed that both mGluR subtypes were expressed in the apical dendrites and some cilia of olfactory neurons. Pharmacological analysis showed that antagonists to each mGluR subtype significantly decreased the electrophysiological response to odorant amino acids. α-Methyl-L -CCG1/(2S,3S,4S)-2-methyl-2-(carboxycyclopropyl)glycine (MCCG), a known antagonist to mGluR3, and (S)-4-carboxyphenylglycine (S-4CPG), a specific antagonist to mGluR1, each significantly reduced olfactory receptor responses to L -glutamate. S-4CPG and MCCG reduced the glutamate response to 54% and 56% of control, respectively, which was significantly greater than their effect on a neutral amino acid odorant, methionine. These significant reductions of odorant response by the antagonists, taken with the expression of these receptors throughout the dendritic and ciliated portions of some olfactory receptor neurons, suggest that these mGluRs may be involved in olfactory reception and signal transduction. © 1998 John Wiley & Sons, Inc. J Neurobiol 35: 94–104, 1998     

Myelin-induced microglial neurotoxicity can be controlled by microglial metabotropic glutamate receptors

Microglia are present in an activated state in multiple sclerosis lesions. Incubation of primary cultured rat microglia with rat-brain derived myelin (0.1–1 μg/mL) for 24 h induced microglial activation; cells displayed enhanced ED1 staining, expression of inducible nitric oxide synthase, production and release of the cytokine tumour necrosis factor-α and glutamate release. Exposure of microglia to myelin induced the expression of neuronal caspases and ultimately neuronal death in cultured cerebellar granule cell neurons; neurotoxicity was directly because of microglial-derived soluble toxins. Co-incubation of microglia with agonists or antagonists of different metabotropic glutamate receptor (mGluR) subtypes ameliorated microglial neurotoxicity by inhibiting soluble neurotoxin production. Activation of microglial mGluR2 exacerbated myelin-evoked neurotoxicity whilst activation of mGluR3 was protective as was activation of group III mGluRs. These data show that myelin-induced microglial neurotoxicity can be prevented by regulation of mGluRs and suggest these receptors on microglia may be promising targets for therapeutic intervention in multiple sclerosis.     

Novel mGluR- and CB1R-Independent Suppression of GABA Release Caused by a Contaminant of the Group I Metabotropic Glutamate Receptor Agonist,DHPG

Background

Metabotropic glutamate receptors (mGluRs) are ubiquitous throughout the body, especially in brain, where they mediate numerous effects. MGluRs are classified into groups of which group I, comprising mGluRs 1 and 5, is especially important in neuronal communication. Group I actions are often investigated with the selective agonist, S-3,5-dihydroxyphenylglycine (DHPG). Despite the selectivity of DHPG, its use has often led to contradictory findings. We now report that a particular commercial preparation of DHPG can produce mGluR-independent effects. These findings may help reconcile some discrepant reports.

Methods

We carried out electrophysiological recordings in the rat in vitro hippocampal slice preparation, focusing mainly on pharmacologically isolated GABA_A-receptor-mediated synaptic currents. Principal Findings: While preparations of DHPG from three companies suppressed GABAergic transmission in an mGluR-dependent way, one batch had an additional, unusual effect. Even in the presence of antagonists of mGluRs, it caused a reversible, profound suppression of inhibitory transmission. This mGluR - independent action was not due to a higher potency of the compound, or its ability to cause endocannabinoid-dependent responses. Field potential recordings revealed that glutamatergic transmission was not affected, and quantal analysis of GABA transmission confirmed the unusual effect was on GABA release, and not GABA_A receptors. We have not identified the responsible factor in the DHPG preparation, but the samples were 99% pure as determined by HPLC and NMR analyses.

Conclusions

In certain respects our observations with the anomalous batch strikingly resemble some published reports of unusual DHPG effects. The present findings could therefore contribute to explaining discrepancies in the literature. DHPG is widely employed to study mGluRs in different systems, hence rigorous controls should be performed before conclusions based on its use are drawn.     

Cyclic ADP ribose-dependent Ca2+ release by group I metabotropic glutamate receptors in acutely dissociated rat hippocampal neurons

Group I metabotropic glutamate receptors (group I mGluRs; mGluR1 and mGluR5) exert diverse effects on neuronal and synaptic functions, many of which are regulated by intracellular Ca²⁺. In this study, we characterized the cellular mechanisms underlying Ca²⁺ mobilization induced by (RS)-3,5-dihydroxyphenylglycine (DHPG; a specific group I mGluR agonist) in the somata of acutely dissociated rat hippocampal neurons using microfluorometry. We found that DHPG activates mGluR5 to mobilize intracellular Ca²⁺ from ryanodine-sensitive stores via cyclic adenosine diphosphate ribose (cADPR), while the PLC/IP₃ signaling pathway was not involved in Ca²⁺ mobilization. The application of glutamate, which depolarized the membrane potential by 28.5±4.9 mV (n = 4), led to transient Ca²⁺ mobilization by mGluR5 and Ca²⁺ influx through L-type Ca²⁺ channels. We found no evidence that mGluR5-mediated Ca²⁺ release and Ca²⁺ influx through L-type Ca²⁺ channels interact to generate supralinear Ca²⁺ transients. Our study provides novel insights into the mechanisms of intracellular Ca²⁺ mobilization by mGluR5 in the somata of hippocampal neurons.     

Co-stimulation of mGluR5 and N-methyl-D-aspartate receptors is required for potentiation of excitatory synaptic transmission in hippocampal neurons

In the central nervous system, excitatory synaptic transmission is mediated by the neurotransmitter glutamate and its receptors. Interestingly, stimulation of group I metabotropic glutamate receptors (mGluRs) can either enhance or depress synaptic transmission at CA1 hippocampal synapses. Here we report that co-activation of mGluR5, a member of the group I mGluR family, and N-methyl-d-aspartate receptors (NMDARs) potentiates NMDAR currents and induces a long lasting enhancement of excitatory synaptic transmission in primary cultured hippocampal neurons. Unexpectedly, activation of mGluR5 alone fails to enhance evoked NMDAR currents and synaptic alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor (AMPAR) AMPAR currents. The observed potentiation requires an mGluR5-induced, inositol 1,4,5-trisphosphate receptor-mediated mobilization of intracellular Ca2+, which acts in concert with a protein kinase C, calcium-activated tyrosine kinase cascade to induce a long lasting enhancement of NMDAR and AMPAR currents.     

Activation of Metabotropic Glutamate Receptors Inhibits Synapsin I Phosphorylation in Visceral Sensory Neurons

Activation of glutamate metabotropic receptors (mGluRs) in nodose ganglia neurons has previously been shown to inhibit voltage-gated Ca⁺⁺ currents and synaptic vesicle exocytosis. The present study describes the effects of mGluRs on depolarization-induced phosphorylation of the synaptic-vesicle-associated protein synapsin I. Depolarization of cultured nodose ganglia neurons with 60 mm KCl resulted in an increase in synapsin I phosphorylation. Application of mGluR agonists 1-aminocyclopentane-1s-3r-dicarboxylic acid (t-ACPD) and L(+)-2-Amino-4-phosphonobutyric acid (L-AP4) either in combination or independently inhibited the depolarization induced phosphorylation of synapsin I. Application of the mGluR antagonist (RS)-α-Methyl-4-carboxyphenylglycine (MCPG) blocked t-ACPD-induced inhibition of synapsin phosphorylation but not the effects of L-AP4. In addition, application of either t-ACPD or L-AP4 in the absence of KCl induced depolarization had no effect on resting synapsin I phosphorylation. RT-PCR analysis of mGluR subtypes in these nodose ganglia neurons revealed that these cells only express group III mGluR subtypes 7 and 8. These results suggest that activation of mGluRs modulates depolarization-induced synapsin I phosphorylation via activation of mGluR7 and/or mGluR8 and that this process may be involved in mGluR inhibition of synaptic vesicle exocytosis in visceral sensory neurons of the nodose ganglia. Received 28 June 2000/Revised: 11 September 2000