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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of [125I]I-MK-801 to N-methyl-D-aspartate (NMDA) receptors on membranes prepared from cultured cerebral cortical neurons and from forebrain of rats of different ages was investigated. Specific binding of [125I]I-MK-801 was enhanced by glutamate, glycine, and polyamines and was inhibited by divalent cations and open-channel blockers of the NMDA receptor, indicating that [125I]I-MK-801 selectively labels a component of the NMDA receptor/ion channel complex. The ability of spermine to enhance the binding of [125I]I-MK-801 was lower in membranes prepared from cultured cerebral cortical neurons or from neonatal rat brain than in membranes prepared from adult rat brain. There was a progressive increase in the potency of spermine and in the magnitude of the stimulatory effect of spermine in rat forebrain between days 3 and 10 of postnatal life. In contrast, the apparent affinity of the NMDA receptor for spermine remained unchanged in cerebral cortical neurons maintained in culture for up to 5 weeks. Mg2+ also enhanced the binding of [125I]I-MK-801 and was more potent in membranes prepared from adult than from 3-day-old rat forebrain. The potency of glutamate for enhancing the binding of [125I]I-MK-801 was not altered in 3-day-old, compared with adult, brain tissue. The increase in the affinity of the polyamine recognition site on the NMDA receptor complex in rat forebrain during the first 2 weeks of postnatal life suggests that the macromolecular properties of the NMDA receptor are altered during development. This may suggest that the subunit composition of the NMDA receptor is under developmental control. Cultured cortical neurons may represent a useful system for investigating factors that regulate developmental changes in the properties of the NMDA receptor.
Mol Pharmacol 1991 Nov
PMID:Developmental changes in the sensitivity of the N-methyl-D-aspartate receptor to polyamines. 168 96

Acute chemical anoxic injury was produced in primary cerebellar granule cell cultures incubated with iodoacetate (IAA) alone or IAA combined with potassium cyanide (KCN). Cytotoxicity was assessed using Trypan blue exclusion or LDH release. Four millimolars of KCN induced approx 30% neuron death at 3 h, whereas greater than 50% cell death was produced by 0.2 mM IAA. No potentiation of cytotoxicity was observed by IAA + KCN. A total of 0.2 mM IAA produced an early major reduction of intracellular ATP prior to the onset of neuron injury or reduction in intracellular glutathione (GSH). Medium Na+ replacement by choline, K+, or methylglucamine protected against IAA-induced neuronal injury, reduced the rate of decline of intracellular ATP but had no effect on intracellular GSH. Some 80% neuronal survival was obtained when Na+ was deleted from the medium even after the intracellular ATP had been reduced to less than 10% of control. Removal of Ca2+ from the medium had no effect on control culture, Trypan blue exclusion, GSH, or ATP, but potentiated the onset and magnitude of IAA-induced cytotoxicity. ATP and GSH decline. Loading of granule cells with the Ca2+ chelator Fura-2 did not influence IAA-induced cytotoxicity in control or low Ca2+ media. Addition of 50 microM glutamate had a minimal cytotoxic effect over 3 h and the combined addition of 0.2 mM IAA plus 50 microM glutamate did not potentiate IAA-induced injury. The glutamate receptor antagonists, D-2-amino-5-phosphonovaleric acid (APV) or kynurenate did not block IAA-induced injury in control medium but inhibited the potentiation of toxicity seen in the low Ca2+ medium. This study suggests the use of IAA as a chemical anoxic agent in cerebellar granule cell culture. The early, dose-dependent decline in ATP may be dissociated from GSH change. Acute IAA-induced injury is Na+/Cl- dependent but paradoxically potentiated in low Ca2+ medium. The low Ca2+ potentiated component was sensitive to glutamate/NMDA receptor antagonists and associated with reduction of intracellular GSH.
Mol Chem Neuropathol 1991 Dec
PMID:Paradoxical potentiation by low extracellular Ca2+ of acute chemical anoxic neuronal injury in cerebellar granule cell culture. 168 39

Zinc noncompetitively antagonizes N-methyl-D-aspartate (NMDA) receptor-mediated responses in cultured neurons. We investigated the mechanism of this inhibition by examining the effect of zinc on ligand binding to three distinct sites on the NMDA receptor in rat hippocampal membranes. Zinc dose-dependently inhibited both the association and dissociation of the NMDA channel blocker [3H]N-(1-[thienyl]cyclohexyl)piperidine ([3H]TCP) but had no effect on steady state levels of [3H]TCP binding. This suggests that zinc inhibits the receptor-gated access of [3H]TCP to its site in the ion channel but has no effect on the binding site itself. Zinc inhibition of [3H]TCP association was not mediated by an action at the NMDA recognition site, because zinc had no effect on NMDA-displaceable L-[3H]glutamate binding. On the other hand, zinc dose-dependently inhibited [3H]glycine binding by a noncompetitive interaction. Stoichiometric analysis of equilibrium binding data indicated the presence of two [3H]glycine binding sites/[3H]TCP binding site. Comparison of the potencies of zinc in inhibiting glycine-dependent [3H]TCP association and [3H]glycine binding suggests that blockade of only one of the two glycine sites is sufficient to prevent [3H]TCP association. We hypothesize that synaptically released zinc inhibits NMDA receptor-mediated responses by binding to a site on the receptor/channel complex, reducing glycine binding, and thereby decreasing what would otherwise be a tonically present action of endogenous extracellular glycine.
Mol Pharmacol 1990 Jul
PMID:Evidence that zinc inhibits N-methyl-D-aspartate receptor-gated ion channel activation by noncompetitive antagonism of glycine binding. 169 16

Ethanol has been shown to inhibit N-methyl-D-aspartate (NMDA)-stimulated calcium influx into cerebellar granule cells grown in culture. Because NMDA-mediated responses are modulated by a number of substances, we investigated the effects of several of these agents on ethanol-induced inhibition of calcium flux. Ethanol (50 mM) inhibited NMDA-dependent Ca2+ influx by approximately 50%. The percentage of inhibition remained constant with increasing NMDA concentrations (5-250 microM). Increasing Mg2+ concentrations in the assay medium inhibited NMDA-stimulated calcium influx but the EC50 for Mg2+ was unchanged in the presence of ethanol. Glycine at concentrations of 0.3-100 microM potentiated the effects of NMDA. Glycine at concentrations in excess of 10 microM decreased ethanol-mediated inhibition of NMDA-stimulated calcium influx. D-Serine was shown to have effects similar to those of glycine, whereas L-serine was significantly less active in potentiating NMDA-stimulated activity and reversing the ethanol-induced inhibition of calcium influx. N-Methylglycine and L-leucine were ineffective in potentiating NMDA actions but high concentrations (1 mM) of N-methylglycine attenuated ethanol-induced inhibition, whereas L-leucine (1 mM) had no effect. High concentrations of N-methylglycine were shown to reduce glycine-induced enhancement at the NMDA receptor, whereas L-leucine did not affect the glycine response. Glycine did not affect kainate-stimulated calcium influx and did not alter the small amount of inhibition produced by ethanol in the response of the cells to kainate. The results demonstrate that the in vivo actions of ethanol on the NMDA systems of brain may be dependent on glycine concentrations at these receptor sites.
Mol Pharmacol 1990 Dec
PMID:Glycine site-directed agonists reverse the actions of ethanol at the N-methyl-D-aspartate receptor. 170 Dec 11

The action of tetrahydroaminoacridine (THA), a centrally active cholinesterase inhibitor that may provide symptomatic benefit in Alzheimer's disease, was studied on responses to the excitatory amino acid N-methyl-D-aspartate (NMDA) in cultured hippocampal neurons, using whole-cell voltage-clamp and single-channel recording techniques. THA produced a concentration-dependent block of NMDA-evoked inward current responses (IC50, 190 microM at -60 mV), without affecting responses to quisqualate or kainate. THA block of NMDA responses was voltage dependent and was nearly completely relieved at positive holding potentials. Analysis of the voltage dependency indicated that the THA binding site senses 56% of the transmembrane electrostatic field. In single-channel recordings from outside-out membrane patches, THA appeared to reduce the frequency and duration of NMDA-evoked single-channel currents, without affecting the single-channel amplitude. The effects of THA on NMDA responses occur at concentrations 1-2 orders of magnitude greater than the therapeutic serum concentrations and, therefore, blockade of NMDA receptor-mediated responses is unlikely to contribute to the putative therapeutic action of THA. However, because NMDA receptors may play a critical role in cognitive and memory function, THA has the potential to produce undesirable central nervous system side effects at high doses.
Mol Pharmacol 1991 May
PMID:Tetrahydroaminoacridine block of N-methyl-D-aspartate-activated cation channels in cultured hippocampal neurons. 170 20

The inhibition of N-methyl-D-aspartate (NMDA) receptor channels by the vasodilatory and anti-ischemic agent ifenprodil was examined on cultured rat hippocampal neurons. Whole-cell and single-channel patch recordings were used. Ifenprodil inhibition of NMDA currents could be separated into two components, with IC50 values of 0.75 and 161 microM. The high and low affinity components were both voltage independent but could be separated by their kinetics and dependence on extracellular calcium and glycine. The maximal inhibition of inward current by ifenprodil (approximately 90%) was equally divided between the two components in 0.3 mM extracellular calcium and 500 nM glycine. The low affinity action of ifenprodil had rapid kinetics and appeared to result from allosteric inhibition of the glycine modulatory site on the NMDA receptor. The macroscopic kinetics of the high affinity component were slow. The rate of onset was concentration dependent, and complete recovery required 1-2 min. Unlike open-channel blockers, ifenprodil block was not use dependent, and pre-exposure to ifenprodil also reduced subsequent NMDA responses. Low concentrations of ifenprodil were less effective after calcium-dependent inactivation of whole-cell currents, but the IC50 was unaffected, suggesting that calcium and ifenprodil act on a common set of channels. On outside-out membrane patches, ifenprodil reduced the frequency of channel opening without altering the single-channel conductance. Open time histograms of the large conductance events revealed two mean open times of approximately 2 and 8 msec, but only the duration of the long openings was decreased by ifenprodil. This effect was concentration dependent and revealed a blocking rate constant of 6 x 10(7) M-1sec-1. However, the proportion of current blocked by low concentrations of ifenprodil was larger in outside-out patches than in whole-cell recordings, suggesting that intracellular factors may influence ifenprodil efficacy. These results indicate that high affinity ifenprodil binding is extracellular and does not require agonist binding or channel opening. Because low concentrations of ifenprodil only partially inhibited the current and affected only the long openings, ifenprodil may promote a modal shift in channel gating.
Mol Pharmacol 1991 Aug
PMID:Ifenprodil blocks N-methyl-D-aspartate receptors by a two-component mechanism. 171 17

Desensitization of the N-methyl-D-aspartate (NMDA) receptor-channel complex was studied in isolated rat hippocampal neurons using a fast drug application system. 1) Desensitization rate was slower at more negative membrane potentials and when external [Ca2+] was lowered. 2) In the presence of 10 microM glycine, 2-amino-5-phosphonovalerate neither induced desensitization nor prevented recovery from it. 3) Preincubation in 500 microM aspartate or 10 microM glycine alone elicited desensitization only weakly or not at all. 4) Aspartate appeared to bind at its receptor site in the absence of glycine, and vice versa. It is proposed that, for the NMDA receptor, channel opening is necessary for the occurrence of desensitization and, thus, that desensitization involves structural changes in the channel-lining section of the protein rather than the glycine or NMDA binding sites.
Mol Pharmacol 1991 Sep
PMID:Desensitization of N-methyl-D-aspartate receptors in neurons dissociated from adult rat hippocampus. 171 28

In the presence of 1.2 mM Mg2+, glycine (30-100 microM) inhibited [3H]dopamine ([3H]DA) release stimulated by N-methyl-D-aspartate (NMDA), in fetal rat mesencephalic cell cultures. Strychnine (1 microM) blocked the inhibitory effect of 100 microM glycine, indicating an action via strychnine-sensitive inhibitory glycine receptors. A higher concentration of strychnine (100 microM), by itself, inhibited NMDA-evoked [3H]DA release in the presence or absence of Mg2+. Spontaneous [3H]DA release and [3H]DA release stimulated by kainate and quisqualate were unaffected by glycine (less than or equal to 100 microM) or strychnine (less than or equal to 100 microM), indicating that glycine and strychnine modulatory effects are only associated with the NMDA receptor subtype. [3H]DA release evoked by K+ (56 mM) was unaffected by glycine (less than or equal to 100 microM) but was attenuated by a high concentration of strychnine (100 microM). In the absence of exogenous Mg2+, glycine (30-100 microM) potentiated NMDA-evoked [3H]DA release by a strychnine-insensitive mechanism. A selective antagonist of the NMDA-associated glycine receptor, 7-chlorokynurenate (10 microM), attenuated NMDA-evoked [3H]DA release in the absence of Mg2+. The effect of 10 microM 7-chlorokynurenate was overcome by 1 microM glycine. Also, when tested in the presence of 1.2 nM Mg2+ and 1 microM strychnine, 100 microM 7-chlorokynurenate inhibited NMDA-evoked [3H]DA release, and this antagonism was overcome by 30 to 100 microM glycine. These results indicate that two distinct glycine receptors modulate NMDA-stimulated [3H]DA release from mesencephalic cells in culture. Manipulation of extracellular Mg2+ permits the differentiation of a strychnine-sensitive glycine response (inhibition of NMDA-evoked [3H]DA release) from a strychnine-insensitive glycine response (potentiation of NMDA-evoked [3H]DA release). It is suggested that voltage-dependent Mg2+ blockade of the NMDA response may allow for the expression of these opposing effects of glycine.
Mol Pharmacol 1991 Feb
PMID:Inhibitory and potentiating influences of glycine on N-methyl-D-aspartate-evoked dopamine release from cultured rat mesencephalic cells. 182 45

Whole-cell and single-channel patch-clamp recordings from hippocampal neurons in culture have been used to study the receptor channel selectivity of the glutamate analog quisqualate. The dose-response relationship of quisqualate acting at the N-methyl-D-aspartate (NMDA) receptor was measured as that portion of the whole-cell current activated by quisqualate that could be blocked by the addition of two NMDA antagonists, 5-fluoroindole-2-carboxylic acid, a competitive antagonist of the NMDA receptor-associated glycine site, and D-2-amino-5-phosphonovalerate, a competitive NMDA binding site antagonist. We found that quisqualate was 10-fold less potent than NMDA. In outside-out patches quisqualate activates single-channel events that range in conductance from 5 to 50 pS. The NMDA antagonists 5-fluoroindole-2-carboxylic acid and D-2-amino-5-phosphonovalerate completely blocked all of the 40-50-pS channel openings in the presence of quisqualate. These results indicate that quisqualate gates 40-50-pS events by activating NMDA receptor channels.
Mol Pharmacol 1990 Apr
PMID:Quisqualate activates N-methyl-D-aspartate receptor channels in hippocampal neurons maintained in culture. 197 Jan 15

In primary cultures of rat cerebellar neurons, a brief stimulation of glutamate receptors results in coordinated activation of a programmed early gene response involving increases in the amount of c-fos, c-jun, jun-B, and zif/268 mRNAs. Each of these genes was induced to a different extent and showed a temporal pattern characterized by either a monophasic "early" response, occurring within 30 min of glutamate addition, or a biphasic response (c-jun), lasting for up to 6 to 8 hr after the initial stimulus. The early phase of the glutamate-induced gene expression was prevented by 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid, a highly selective isosteric antagonist of the N-methyl-D-aspartate (NMDA)-sensitive glutamate receptor (NMDA receptor). The second phase of the c-jun response was not blocked when the NMDA receptors were completely inhibited after the initial pulse of agonist or when the quisqualate-kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione was added, suggesting that a brief NMDA receptor stimulation triggers a cascade of events critical for the manifestation of the delayed c-jun expression. Furthermore, gel retardation assays demonstrated that NMDA receptor activation results in a prolonged increase in nuclear DNA-binding activity specific for the AP-1 transcriptional regulatory element. Protein immunoblot analysis showed that the composition of this nucleoprotein complex changes as a function of time, reflecting a cascade that involves an increased translation of Fos and several Fos-related proteins. The coordinated induction of several different transcription factors and the variations in transcriptional complex formation initiated by NMDA receptor stimulation may be a key mechanism in the orchestration of specific target gene expression that underlies various aspects of neuronal function, including plasticity responses.
Mol Pharmacol 1990 Nov
PMID:Transcriptional program coordination by N-methyl-D-aspartate-sensitive glutamate receptor stimulation in primary cultures of cerebellar neurons. 197 42


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