Selectivity of amino acid transmitters acting at N-methyl-D-aspartate and amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors.
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.
Department of Pharmacology, University of North Carolina, Chapel Hill 27599.
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D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism; D007051:Ibotenic Acid / Q000031:analogs & derivatives* / Q000378:metabolism; D008297:Male; D018377:Neurotransmitter Agents / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism*; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid
D000596:Amino Acids; D002712:Chlorides; D018377:Neurotransmitter Agents; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D007051:Ibotenic Acid; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; D002118:Calcium