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

The synthesis of nerve growth factor (NGF) and nerve growth factor receptor (NGFR) were studied in a C6 glioma cell line by Northern blot hybridization. In response to a glutamate agonist N-methyl-D-aspartic acid (NMDA), NGF mRNA increased by up to 2-fold after 4-12 h of culture. The non-NMDA receptor agonists, quisqualate and kainate, did not induce any increase of NGF mRNA, and kainate actually produced a decrease. The increase in NGF mRNA in response to NMDA was dose-dependent at 1, 5 and 10 microM. NGF receptor (NGFR) mRNA showed changes in expression which were similar to those for NGF mRNA, but were less marked. The specific glutamate antagonist 2-aminophosphonovaleric acid (APV) blocked the increase of NGF mRNA produced by NMDA. In the absence of Ca2+, an increase of NGF mRNA was still observed but in the presence of 1 mM ethylglycol-bis-(beta-aminoethyl ether) N,N'-tetraacetic acid (EGTA), NGF mRNA production abolished. The mechanism producing an increase in NGF mRNA by NMDA may be mediated by cyclic AMP since intracellular cyclic AMP and NGF mRNA levels both increased following treatment with NMDA or dibutyryl cyclic AMP.
Brain Res Mol Brain Res 1992 Jun
PMID:Regulation of nerve growth factor and nerve growth factor receptor production by NMDA in C6 glioma cells. 135 54

Exposure of cultured cerebellar granule cells to glutamate results in a concentration-dependent (EC50 = 22.7 +/- 0.4 microM) and delayed (24-72 hr) neurotoxicity, which is blocked by the specific N-methyl-D-aspartate (NMDA) receptor antagonists 2-amino-5-phosphovalerate and MK-801 but is unaffected by the non-NMDA receptor antagonists 6-cyano-7-nitroquinoxaline-2,3-dione and 6,7-dinitroquinoxaline-2,3-dione. Although glutamate toxicity in these cells is mediated by the NMDA subtype of glutamate receptor, pretreatment of cerebellar granule cells with subtoxic concentrations of NMDA markedly antagonizes the neurotoxic actions of glutamate, with an IC50 of 55 +/- 4 microM. The neuroprotective effect of NMDA requires a preincubation time of approximately 120 min to be fully manifested and does not require the presence of NMDA during glutamate exposure. These data demonstrate that NMDA receptors mediate both neurotoxicity and neuroprotection in cerebellar granule cells. Among four glutamate receptor agonists tested (NMDA, quisqualate, ibotenate, and kainate), only NMDA was able to provide a robust neuroprotection against glutamate toxicity. Quisqualate was neither neurotoxic nor neuroprotective, whereas ibotenate, which was nontoxic by itself, induced a small degree of neuroprotection. In contrast, kainate, which was neurotoxic to cerebellar granule cells, also provided considerable neuroprotection against glutamate toxicity. Because preincubation of cerebellar granule cells with NMDA fails to alter NMDA receptor-mediated phosphoinositide hydrolysis or the specific binding of [3H]MK-801 to NMDA receptors, it appears that the neuroprotective effects of NMDA are not due to NMDA receptor desensitization.
Mol Pharmacol 1992 Aug
PMID:N-methyl-D-aspartate exposure blocks glutamate toxicity in cultured cerebellar granule cells. 135 59

We have recently shown that a high-affinity AMPA receptor labelled with the antagonist [3H]CNQX can be regulated in a 'living' cortical slice preparation by agonist stimulation or changes in electrical activity (Lanius, R.A. and Shaw, C. (1992) Anat. Rec., in press). Based on a study of GABAA receptors (Shaw, C. and Scarth, B.A. (1992) Mol. Brain Res., in press), which showed age-dependent changes in regulation, we have now investigated the regulation of high-affinity AMPA receptors in neocortex at different stages in postnatal development. The results show that regulation by agonist stimulation and increases in bioelectric activity are age-dependent in amount and, in the latter case, in direction. Agonist stimulation using quisqualate resulted in a significant receptor down-regulation of approximately 7% at ages less than 20 days postnatal; in adult rats quisqualate led to a significant 23% decrease. Changes in bioelectric activity induced by a combination of veratridine and glutamate showed a significant increase in AMPA receptor number of 16% at ages less than 20 days, whereas such treatment resulted in a significant 18% decrease in adult rats. The present data reveal a near mirror-image to the effects of veratridine and glutamate and agonist on GABAA receptors in the same preparation, but with a temporal mismatch in the amount and direction of regulation. We speculate that the age-dependent differences in direction of regulation for the receptor populations which serve key excitatory and inhibitory functions in cortex may provide a molecular basis for the gradual decline of neuronal plasticity during the critical period.
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PMID:Cortical AMPA receptors: age-dependent regulation by cellular depolarization and agonist stimulation. 135 59

We have shown previously that GABAA receptors labelled with the antagonist [3H]SR95531 can be regulated in a living cortical slice preparation by agonists or changes in electrical activity. Due to the important role that receptor regulation may play in controlling neural activity, we have now investigated the regulation of GABAA receptors in neocortex at different stages in postnatal life. We found that regulation by agonist stimulation and increases in bioelectric activity is age-dependent in amount and, in the latter case, in direction. Using muscimol as an agonist we observed a GABAA receptor down-regulation of between 60 and 70% at 20-30 days of age; in adults muscimol gave an 11% down-regulation. A combination of veratridine and glutamate gave a peak down-regulation of 39% at 20 days postnatal, but an up-regulation of 58% in adults. These age-dependent effects may signal a role for receptor regulation in cortical neural critical period plasticity.
Brain Res Mol Brain Res 1992 Jul
PMID:Age-dependent regulation of GABAA receptors in neocortex. 135 70

We have characterized a high affinity site of the alpha-amino-3 hydroxy-5-methyl-isoxazole-4-propionate (AMPA) receptor in in vitro living slices of adult rat neocortex using [3H]CNQX, and AMPA antagonist. [3H]CNQX labelled multiple binding sites with a Bmax of a high affinity site of approximately 470 fmol/mg protein and an apparent Kd of 11.3 nM. The high affinity site of the AMPA receptor could be down-regulated (36%) by 2 h preincubations in quisqualate, an AMPA agonist. Increases in electrical activity induced by a combination of veratridine and glutamate also led to an average decrease of the high affinity AMPA receptor number of 17%. In addition, preincubations with muscimol, a GABAA agonist, as well as glutamate agonists kainate and N-methyl-D-aspartate (NMDA) led to an average increase in high affinity AMPA receptor number of 17%, 14%, and 37%, respectively. The present results show that a ligand-gated high affinity AMPA receptor can be regulated by agonist stimulation as well as changes in neural activity.
Brain Res Mol Brain Res 1992 Oct
PMID:Characterization and regulation of a high affinity [3H]CNQX labelled AMPA receptor in rat neocortex. 135 75

The genes encoding the Na+/H+/L-glutamate symport proteins of the thermophilic organisms Bacillus stearothermophilus (gltTBs) and Bacillus caldotenax (gltTBc) were cloned by complementation of Escherichia coli JC5412 for growth on glutamate as sole source of carbon, energy and nitrogen. The nucleotide sequences of the gltTBs and gltTBc genes were determined. In both cases the translated sequences corresponded with proteins of 421 amino acid residues (96.7% amino acid identity between GltTBs and GltTBc). Putative promoter, terminator and ribosome-binding-site sequences were found in the flanking regions. These expression signals were functional in E. coli. The hydropathy profiles indicate that the proteins are hydrophobic and could form 12 membrane-spanning regions. The Na+/H+ coupled L-glutamate symport proteins GltTBs and GltTBc are homologous to the strictly H+ coupled L-glutamate transport protein of E. coli K-12 (overall 57.2% identity). Functional expression of glutamate transport activity was demonstrated by uptake of glutamate in whole cells and membrane vesicles. In accordance with previous observations (de Vrij et al., 1989; Heyne et al., 1991), glutamate uptake was driven by the electrochemical gradients of sodium ions and protons.
Mol Microbiol 1992 Oct
PMID:Characterization and functional expression in Escherichia coli of the sodium/proton/glutamate symport proteins of Bacillus stearothermophilus and Bacillus caldotenax. 135 85

The assembly of the insulin hexamer brings the six B13 glutamate side-chains at the centre into close proximity. Their mutual repulsion is unfavourable and zinc co-ordination to B10 histidine is necessary to stabilize the well known zinc-containing hexamers. Since B13 is always a carboxylic acid in all known sequences of hexamer forming insulins, it is likely to be important in the hormone's biology. The mutation of B13 Glu-->Gln leads to a stable zinc-free hexamer with somewhat reduced potency. The structures of the zinc-free B13 Gln hexamer and the 2Zn B13 insulin hexamer have been determined by X-ray analysis and refined with 2.5 A and 2.0 A diffraction data, respectively. Comparisons show that in 2Zn B13 Gln insulin, the hexamer structure (T6) is very like that of the native hormone. On the other hand, the zinc-free hexamer assumes a quaternary structure (T3/R3) seen in the native 4Zn insulin hexamer, and normally associated only with high chloride ion concentrations in the medium. The crystal structures show the B13 Gln side-chains only contact water in contrast to the B13 glutamate in 2Zn insulin. The solvation of the B13 Gln may be associated with this residue favouring helix at B1 to B8. The low potency of the B13 Gln insulin also suggests the residue influences the hormone's conformation.
J Mol Biol 1992 Dec 20
PMID:Role of B13 Glu in insulin assembly. The hexamer structure of recombinant mutant (B13 Glu-->Gln) insulin. 136 49

[3H]Dextrorphan recognition sites were characterized in rat brain membranes. The pharmacological profile and regional distribution of [3H]dextrorphan binding sites appear to distinguish these sites from those labeled either by [3H]dextromethorphan or by putative sigma receptor radioligands. Data from thoroughly washed forebrain membranes suggest that [3H]dextrorphan predominantly labels a high affinity site defined by the activated state of the N-methyl-D-aspartate (NMDA) receptor-channel complex. Regulation of [3H]dextrorphan binding by specific modulators of NMDA receptor function suggests that [3H]dextrorphan binding is predominantly localized to a domain of the receptor-channel complex also recognized by the prototypical noncompetitive antagonist radioligands (+)-[3H]5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imi ne (MK-801) and [3H]1-[1-(2-thienyl)cyclohexyl]piperidine (TCP). The critical relationship between [3H]dextrorphan binding and activation of the NMDA receptor-complex is suggested by the profound dependence of [3H]dextrorphan binding on glutamate in well washed membranes. Basal specific [3H]dextrorphan binding is nearly totally suppressed by the specific competitive NMDA antagonist D(-)-2-amino-5-phosphonopentanoic acid (D-AP5), in a glutamate- but not glycine-surmountable manner. Glutamate and glycine each stimulate [3H]dextrorphan binding in a concentration-dependent manner, effecting maximal increases from control of up to 30- and 14-fold, respectively. The NMDA receptor specificity of the modulation of [3H]dextrorphan binding by glutamate and glycine is indicated by the sensitivity of their effects to competitive antagonism by D-AP5 and 3-amino-1-hydroxy-2-pyrrolidone (HA-966), respectively, and by the accordant rank orders of potency of glycine analogs as modulators of [3H]dextrorphan binding and as ligands at the strychnine-insensitive glycine site. The divalent cations Mg2+ and Zn2+ and the polyamines spermine and spermidine regulate [3H]dextrorphan binding in a manner consistent with radioligand interaction at the noncompetitive NMDA antagonist domain. Mg2+ and spermidine regulate [3H]dextrorphan binding biphasically in well washed forebrain membranes, whereas Zn2+ monotonically inhibits [3H]dextrorphan binding. Mg2+ and spermidine regulate [3H]dextrorphan binding with qualitative similarity and in a contrasting fashion to their regulation of [3H]MK-801 and [3H]TCP binding. First, spermidine and Mg2+ are significantly more potent modulators of [3H]dextrorphan binding than of [3H]MK-801 and [3H]TCP binding in well washed membranes; second, whereas the potencies of spermidine and Mg2+ as modulators of [3H]MK-801 and [3H]TCP binding are significantly increased by glutamate and glycine in well washed membranes, their potencies as regulators of [3H]dextrorphan binding appear to be unaffected by glutamate and glycine.(ABSTRACT TRUNCATED AT 400 WORDS)
Mol Pharmacol 1992 Jan
PMID:High affinity [3H]dextrorphan binding in rat brain is localized to a noncompetitive antagonist site of the activated N-methyl-D-aspartate receptor-cation channel. 137 Jul 4

The endogenous neurotransmitter candidates L-aspartate, L-cysteine sulfinate (CSA), L-glutamate, L-homocysteate (HCA), and the endogenously occurring analogue quinolinate were compared in terms of potency, maximal activity, and selectivity for steady state activation of N-methyl-D-aspartate (NMDA) and non-NMDA [(RS)-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)] types of glutamate receptors expressed in Xenopus oocytes injected with mRNA isolated from rat brain (minus cerebellum). Selective activation of NMDA receptors was achieved by deleting Mg2+ and including 3-10 microM glycine in the perfusion medium and by applying ligands in the presence of 30 microM quisqualate, which blocks the AMPA receptor and desensitizes the oocyte's own Ca(2+)-dependent Cl- current. Oocytes were voltage clamped, and steady state inward currents were measured in response to perfusion with agonists at known concentrations. Under the NMDA receptor-preferring condition, the potency rank order was L-glutamate (EC50 = 2.2 microM, 95% confidence interval = 1.4-3.6 microM) greater than L-aspartate (13 microM) = HCA (13 microM) greater than CSA (59 microM) greater than quinolinate (greater than or equal to 7200 microM). All amino acids tested evoked similar maximal currents, which were 120-159% that of NMDA itself. The Hill coefficient was greater than 1 for all agonists except L-HCA (0.6), which might reflect heterogeneity of NMDA receptors expressed. This was supported by the finding that glycine was more potent in combination with HCA than NMDA, in activating NMDA receptors. To study the activity of agonists at AMPA receptors, glycine and quisqualate were omitted and 1 mM Mg2+ was included to block NMDA receptors. Ca(2+)-dependent Cl- currents activated by L-glutamate were prevented by inclusion of 0.4 M ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid in the recording electrode. All amino acids were less potent at AMPA receptors than at NMDA receptors; the potency rank order for steady state activation of AMPA receptors was L-glutamate (EC50 = 11 microM, 95% confidence interval = 7.3-18 microM) greater than HCA (430 microM) greater than CSA (3300 microM). L-Aspartate and quinolinate produced little or no inward current even up to 10 mM, i.e., were inactive at forebrain AMPA receptors. The maximal currents activated by all amino acids at steady state were 5-10% that of kainate, presumably due to severe desensitization of the AMPA receptor by the natural agonists. These results are consistent with L-glutamate acting as a mixed agonist at both AMPA and NMDA synaptic receptors and L-aspartate being involved exclusively in NMDA receptor-mediated synapses.
Mol Pharmacol 1992 Mar
PMID:Selectivity of amino acid transmitters acting at N-methyl-D-aspartate and amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors. 137 86

Escherichia coli RNA polymerase can terminate transcription efficiently at rho-independent terminators in a purified transcription system in the absence of accessory factors. This process of "intrinsic termination" involves direct recognition of the terminator by the core RNA polymerase, and provides an important model system for the study of the molecular interactions involved in the switch between elongation and termination. We have analyzed the intrinsic termination efficiency (%T) of 13 rho-independent terminators, under a variety of in vitro reaction conditions. Although all of these sites share the general sequence features of typical rho-independent terminators, we find a wide range of %T (2% to 90%) for the different sites under our standard transcription conditions. While %T for a particular site is characteristic of that site, the efficiency can be altered considerably by the nature and concentration of salts in the reaction, by alteration of the concentrations of the nucleoside triphosphate substrates, or by transcription from supercoiled rather than linear templates. Surprisingly, different conditions can alter %T to a different extent for different terminators. For neutral salts such as potassium chloride or potassium glutamate, changes in the range from 0.1 to 1 M affect %T for different terminators in a distinct manner, depending on the terminator and the anion involved. At some sites, %T is greatly increased by Cl- concentrations up to 1 M, while at other sites %T is reduced or unaffected by these conditions. At some sites K+ concentrations up to 1 M give a modest increase in %T, while at other sites %T is slightly reduced under the same conditions. Thus the actual values of %T, as well as the order of terminator sites ranked according to %T, can be altered greatly according to the choice of reaction conditions. Reduction of the Mg2+ concentration below 1 mM has a dramatic and quite different effect, enhancing termination to approximately 100% for all terminators tested. Transcription of supercoiled DNA templates gives somewhat reduced %T as compared with linear DNA templates. However, the effect is no greater than twofold. Our results are not consistent with those expected for models in which %T is determined by the differential stability of DNA, RNA and hybrid duplex structures at the melted region in the transcription complex. Thus, the Cl anion does not affect the stability of nucleic acid duplexes even at 1 M concentrations, but can enhance termination tenfold. Also, the alterations of monovalent cation concentration that affect %T are not expected to have a differential effect on Tm for DNA, RNA and hybrid duplexes.(ABSTRACT TRUNCATED AT 400 WORDS)
J Mol Biol 1992 Mar 05
PMID:Parameters affecting transcription termination by Escherichia coli RNA polymerase. I. Analysis of 13 rho-independent terminators. 137 65


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