Competitive inhibition of NMDA receptor-mediated currents by extracellular calcium chelators.
Calcium chelators have been widely used in electrophysiological recordings of N-methyl-D-aspartate (NMDA) receptor-mediated currents, as well as in studies of excitotoxicity. Intracellularly applied calcium chelators are known to inhibit, at least in part, such calcium-dependent processes as calmodulin-dependent inactivation, calcineurin-dependent desensitization, and rundown of NMDA receptors. On the other hand, the functional consequences and potential nonspecific effects of extracellularly applied chelators have not been extensively investigated. In whole-cell patch-clamp recordings from human embryonic kidney (HEK) 293 cells transiently transfected with recombinant NMDA receptors, we found that addition of calcium chelators such as EGTA shifted the glutamate dose-response curve to the right, from an EC(50) for NR1A/NR2A of 8 microM in 1.8 mM Ca(2+) to approximately 24 microM in a solution containing nominal 0 Ca(2+)/5 mM EGTA and further to approximately 80 microM in 20 mM EGTA. A similar shift in glutamate dose-response was observed for NR1A/NR2B currents. This dose-response shift was not due to a decrease in extracellular Ca(2+) concentration because there was no change in the glutamate EC(50) at Ca(2+) concentrations ranging from 10 mM to nominal 0/200 microM EGTA. Moreover, addition of 5 mM EGTA fully chelated with 6.8 mM Ca(2+) did not produce any shift in the glutamate dose-response curve. We propose that calcium chelators, containing four free carboxyl moieties, competitively inhibit glutamate binding to NMDA receptors.
Chen
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N
Kinsmen Laboratory of Neurological Research, Department of Psychiatry, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
Murphy
T H
TH
Raymond
L A
LA
D001667:Binding, Competitive / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000493:pharmacokinetics*; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology*; D004594:Electrophysiology; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D018698:Glutamic Acid / Q000494:pharmacology; D006801:Humans; D007668:Kidney / Q000166:cytology; D010957:Plasmids; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D002614:Chelating Agents; C409466:NR1 NMDA receptor; C120997:NR2A NMDA receptor; D016194:Receptors, N-Methyl-D-Aspartate; D018698:Glutamic Acid; D004533:Egtazic Acid; D002118:Calcium