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.
Kinsmen Laboratory of Neurological Research, Department of Psychiatry, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
N
N
Chen
T H
TH
Murphy
L A
LA
Raymond
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