CHARACTERIZATION OF GLUTAMATE BINDING-SITES IN RECEPTORS ASSEMBLED FROM TRANSFECTED NMDA RECEPTOR SUBUNITS

Previous studies in brain and recombinant NMDA receptors have observed heterogeneity in NMDA-sensitive glutamate binding site, We further ch aracterized the glutamate site assembled from NR1a, NR2A, and NR2B NMD A receptor subunits using L-[H-3]glutamate and [H-3] CGP 39653 binding assays, In contrast to earlier reports, we demonstrate a unique pharm acology for the NR2A subunit alone, which has high affinity for agonis ts but low affinity for competitive antagonists compared with heterome ric combinations of NR1a + NR2A and NR1a; NR2B, Similar to previous re ports, we find unequal antagonist affinity between heteromeric combina tions of NR1a + NR2A and NR1a + NR2B, However, unlike earlier reports, we describe two binding components within each heteromeric transfecti on that more closely resemble data obtained for binding to brain membr anes, In addition, we show Mg2+ can alter [H-3]CGP 39653 binding in bo th the NR1a + NR2A and the NR1a + NR2B combination, thus allowing comp arison of the [H-3]CGP 39653-labeled site between the two heteromeric combinations, Agonist inhibition of [H-3]CGP 39653 binding revealed di fferences between the heteromeric combinations as well as within each heteromeric combination, the latter of which more closely resembled re sults from brain, These results further determine components of the ag onist and antagonist binding sites of the NMDA receptor as well as sug gest additional possible mechanisms of heterogeneity of the glutamate site in the brain.