Physiological activation of presynaptic metabotropic glutamate receptors increases intracellular calcium and glutamate release.
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Activation of metabotropic glutamate receptors (mGluRs) has diverse effects on the functioning of vertebrate synapses. The cellular mechanisms that underlie these changes, however, are largely unknown. The role of presynaptic mGluRs in modulating Ca(2+) dynamics and regulating neurotransmitter release was investigated at the vestibulospinal-reticulospinal (VS-RS) synapse in the lamprey brain stem. Application of the specific Group I mGluRs antagonist 7-(hydroxyimino) cyclopropa[b]chromen-1a-carboxylate ethyl ester (CPCCOEt) reduced the amplitude of consecutive high-frequency evoked excitatory postsynaptic currents (EPSCs). A series of experiments using techniques of electrophysiology and calcium imaging were carried out to determine the cellular mechanisms by which this phenomenon occurs. Concentration-dependent increases in the pre- and postsynaptic [Ca(2+)](i) were seen with the application of mGluR agonists. Similarly, high-frequency stimulation of axons caused a Group I mGluR-dependent enhancement in presynaptic Ca(2+) transients. Application of mGluR agonist caused a depolarization of the presynaptic elements, while thapsigargin decreased the high-frequency stimulus- and agonist-induced rises in [Ca(2+)](i). These data suggest that both membrane depolarization and the release of Ca(2+) from intracellular stores potentially play a role in mGluR-induced Ca(2+) signaling. To determine the effect of this modulation of Ca(2+) dynamics on spontaneous glutamate release, miniature EPSCs were recorded from postsynaptic reticulospinal neurons. A potent Group I mGluR agonist, (S)-homoquisqualic acid, caused a large increase in the frequency of events. These results demonstrate the presence of presynaptic Group I mGluRs at the VS-RS synapse. Activation of these receptors leads to a rise in [Ca(2+)](i) and enhances the spontaneous and evoked release of glutamate. Taken together, these studies highlight the importance of synaptic activation of these facilitatory autoreceptors in both short-term plasticity and synaptic transmission. (+info)
Expression of functional metabotropic glutamate receptors in primary cultured rat osteoblasts. Cross-talk with N-methyl-D-aspartate receptors.
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Osteoblasts and osteoclasts express functional N-methyl-d-aspartate (NMDA) receptors, which participate in regulation of bone matrix. In rat femoral osteoblasts held in whole cell clamp there is a robust NMDA current but little if any response to l-glutamate. We have investigated expression of metabotropic glutamate receptors (mGluRs) in these cells. By reverse transcription polymerase chain reaction, we have detected expression of mGluR1b (but not mGluR1a, 2, 3, 4, 5, or 6). Blockade of mGluRs with (+/-)-alpha-methyl-carboxyphenyl-glycine resulted in an enlarged l-glutamate-induced current that resembled the response to NMDA. Conversely, prior stimulation of mGluRs with trans-(+/-)-1-amino-1, 3-cyclopentanedicarboxylic acid (1S,3R-ACPD; mGluR agonist) reduced the NMDA-induced current by 77%. Monitoring of [Ca(2+)](i) showed that NMDA induced a sustained elevation of [Ca(2+)](i), which was dependent upon [Ca(2+)](o). Treatment with 1S,3R-ACPD generated an initial transient that was independent of [Ca(2+)](o), followed by a sustained, [Ca(2+)](o)-dependent phase, a response consistent with phospholipase C-mediated mobilization of stored Ca(2+). Investigations of the interaction between the two receptors confirmed inhibitory modulation of the NMDA receptor-induced rise in [Ca(2+)](i) by mGluRs. Parathyroid hormone, which also activates phospholipase C in osteoblasts, had a similar inhibitory effect on the NMDA receptor-induced [Ca(2+)](i) response. Elevation of [Ca(2+)](i) mediated by mGluR activation was reduced by subsequent stimulation of NMDA receptors. This is the first description of mGluRs in bone and shows that complex glutamatergic signaling can occur in this tissue. (+info)
Control of dendritic outputs of inhibitory interneurons in the lateral geniculate nucleus.
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The thalamic relay to neocortex is dynamically gated. The inhibitory interneuron, which we have studied in the lateral geniculate nucleus, is important to this process. In addition to axonal outputs, these cells have dendritic terminals that are both presynaptic and postsynaptic. Even with action potentials blocked, activation of ionotropic and metabotropic glutamate receptors on these terminals increases their output, whereas activation of metabotropic (M2 muscarinic) but not nicotinic cholinergic receptors decreases their output. These actions can strongly affect retinogeniculate transmission. (+info)
Dendritic release of glutamate suppresses synaptic inhibition of pyramidal neurons in rat neocortex.
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Dual whole-cell recordings were made in layer 2/3 of the rat neocortex in synaptically connected pyramidal cells and fast-spiking non-accommodating (FSN) interneurons. In 75% of cell pairs (n = 80), the cells formed reciprocal synaptic connections. Trains of backpropagating action potentials in pyramidal cells induced Ca2+ transients in dendrites followed by inhibition of unitary IPSPs. IPSP depression was prevented by loading pyramidal cells with 5 mM BAPTA or EGTA. IPSP depression was mimicked by the metabotropic glutamate receptor (mGluR) agonist ACPD and was prevented by a mixture of the mGluR antagonists CPCCOEt and EGLU.IPSP depression was prevented by loading pyramidal cells with the antagonists of vesicular exocytosis botulinum toxin D (light chain) and GDP-beta-S. It is concluded that Ca2+-dependent release of a retrograde messenger, most probably glutamate, from pyramidal cell dendrites suppresses the inhibition of pyramidal neurons via activation of mGluRs located in FSN interneuron nerve terminals. (+info)
Slow synaptic inhibition mediated by metabotropic glutamate receptor activation of GIRK channels.
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Glutamate is the predominant excitatory neurotransmitter in the vertebrate CNS. Ionotropic glutamate receptors mediate fast excitatory actions whereas metabotropic glutamate receptors (mGluRs) mediate a variety of slower effects. For example, mGluRs can mediate presynaptic inhibition, postsynaptic excitation, or, more rarely, postsynaptic inhibition. We previously described an unusually slow form of postsynaptic inhibition in one class of projection neuron in the song-control nucleus HVc of the songbird forebrain. These neurons, which participate in a circuit that is essential for vocal learning, exhibit an inhibitory postsynaptic potential (IPSP) that lasts several seconds. Only a portion of this slow IPSP is mediated by GABA(B) receptors. Since these cells are strongly hyperpolarized by agonists of mGluRs, we used intracellular recording from brain slices to investigate the mechanism of this hyperpolarization and to determine whether mGluRs contribute to the slow synaptic inhibition. We report that mGluRs hyperpolarize these HVc neurons by activating G protein-coupled, inwardly-rectifying potassium (GIRK) channels. MGluR antagonists blocked this response and the slow synaptic inhibition. Thus, glutamate can combine with GABA to mediate slow synaptic inhibition by activating GIRK channels in the CNS. (+info)
Inositol 1,4,5-trisphosphate (IP3)-mediated Ca2+ release evoked by metabotropic agonists and backpropagating action potentials in hippocampal CA1 pyramidal neurons.
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We examined the properties of [Ca(2+)](i) changes that were evoked by backpropagating action potentials in pyramidal neurons in hippocampal slices from the rat. In the presence of the metabotropic glutamate receptor (mGluR) agonists t-ACPD, DHPG, or CHPG, spikes caused Ca(2+) waves that initiated in the proximal apical dendrites and spread over this region and in the soma. Consistent with previously described synaptic responses (Nakamura et al., 1999a), pharmacological experiments established that the waves were attributable to Ca(2+) release from internal stores mediated by the synergistic effect of receptor-mobilized inositol 1,4, 5-trisphosphate (IP(3)) and spike-evoked Ca(2+). The amplitude of the changes reached several micromoles per liter when detected with the low-affinity indicators fura-6F, fura-2-FF, or furaptra. Repetitive brief spike trains at 30-60 sec intervals generated increases of constant amplitude. However, trains at intervals of 10-20 sec evoked smaller increases, suggesting that the stores take 20-30 sec to refill. Release evoked by mGluR agonists was blocked by MCPG, AIDA, 4-CPG, MPEP, and LY367385, a profile consistent with the primacy of group I receptors. At threshold agonist concentrations the release was evoked only in the dendrites; threshold antagonist concentrations were effective only in the soma. Carbachol and 5-HT evoked release with the same spatial distribution as t-ACPD, suggesting that the distribution of neurotransmitter receptors was not responsible for the restricted range of regenerative release. Intracellular BAPTA and EGTA were approximately equally effective in blocking release. Extracellular Cd(2+) blocked release, but no single selective Ca(2+) channel blocker prevented release. These results suggest that IP(3) receptors are not associated closely with specific Ca(2+) channels and are not close to each other. (+info)
Protein kinase C activation modulates alpha-calmodulin kinase II binding to NR2A subunit of N-methyl-D-aspartate receptor complex.
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The N-methyl-d-aspartate (NMDA) receptor subunits NR2 possess extended intracellular C-terminal domains by which they can directly interact with a large number of postsynaptic density (PSD) proteins involved in synaptic clustering and signaling. We have previously shown that PSD-associated alpha-calmodulin kinase II (alphaCaMKII) binds with high affinity to the C-terminal domain of the NR2A subunit. Here, we show that residues 1412-1419 of the cytosolic tail of NR2A are critical for alphaCaMKII binding, and we identify, by site directed mutagenesis, PKC-dependent phosphorylation of NR2A(Ser(1416)) as a key mechanism in inhibiting alphaCaMKII-binding and promoting dissociation of alphaCaMKII.NR2A complex. In addition, we show that stimulation of PKC activity in hippocampal slices either with phorbol esters or with the mGluRs specific agonist trans-1-amino-1,3- cyclopentanedicarboxylic acid (t-ACPD) decreases alphaCaMKII binding to NMDA receptor complex. Thus, our data provide clues on understanding the molecular basis of a direct cross-talk between alphaCaMKII and PKC pathways in the postsynaptic compartment. (+info)
Relative contribution by GABA or glycine to Cl(-)-mediated synaptic transmission on rat hypoglossal motoneurons in vitro.
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The relative contribution by GABA and glycine to synaptic transmission of motoneurons was investigated using an hypoglossus nucleus slice preparation from neonatal rats. Spontaneous, miniature, or electrically evoked postsynaptic currents (sPSCs, mPSCs, ePSCs, respectively) mediated by glycine or GABA were recorded under whole cell voltage clamp after blocking excitatory glutamatergic transmission with kynurenic acid. The overall majority of Cl(-)-mediated sPSCs was glycinergic, while only one-third was GABAergic; 70 +/- 10% of mPSCs were glycinergic while 22 +/- 8% were GABAergic. Tetrodotoxin (TTX) application dramatically reduced the frequency (and slightly the amplitude) of GABAergic events without changing frequency or amplitude of glycinergic sPSCs. These results indicate that, unlike spontaneous GABAergic transmission, glycine-mediated neurotransmission was essentially independent of network activity. There was a consistent difference in the kinetics of GABAergic and glycinergic responses as GABAergic events had significantly slower rise and decay times than glycinergic ones. Such a difference was always present whenever sPSCs, mPSCs, or ePSCs were measured. Finally, GABAergic and glycinergic mPSCs were differentially modulated by activation of glutamate metabotropic receptors (mGluRs), which are abundant in the hypoglossus nucleus. In fact, the broad-spectrum mGluR agonist (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (50 microM), which in control solution increased the frequency of both GABAergic and glycinergic sPSCs, enhanced the frequency of glycinergic mPSCs only. These results indicate that on brain stem motoneurons, Cl(-)-mediated synaptic transmission is mainly due to glycine rather than GABA and that GABAergic and glycinergic events differ in terms of kinetics and pharmacological sensitivity to mGluR activation or TTX. (+info)