CALCIUM-DEPENDENT INACTIVATION OF RECOMBINANT N-METHYL-D-ASPARTATE RECEPTORS IS NR2 SUBUNIT-SPECIFIC

Intracellular Ca2+ can reversibly reduce the activity of native N-meth yl-D-aspartate (NMDA) receptors in hippocampal neurons, a phenomenon t ermed Ca2+-dependent inactivation. We examined inactivation in heterom eric NMDA receptors expressed in human embryonic kidney (HEK) 293 cell s using whole-cell recording. NR1-1a/2A heteromers showed reversible i nactivation that was very similar to native NMDA receptors in cultured hippocampal neurons. Inactivation was dependent on the extracellular Ca2+ concentration and the degree of intracellular Ca2+ buffering. In 2 mM extracellular Ca2+, inactivation resulted in a 46.1 +/- 12.6% red uction in the whole-cell current during a 5-sec agonist application. I nactivation of NR1-1a/2A heteromers was unaffected by calcineurin inhi bitors, staurosporine, or phalloidin. NR1-1a/2D heteromers also showed a similar degree of inactivation. In contrast, NR1-1a/2B and NR1-1a/2 C heteromers showed no significant inactivation. At saturating concent rations of NMDA(I mM), NR1-1a/2A heteromers also showed Ca- and glycin e-independent desensitization, as seen in native hippocampal neurons. Ca2+- and glycine-independent desensitization Was less pronounced in N R1-1a/2B heteromers and absent in NR1-1a/2C heteromers. Activation of NR1-1a/2C heteromers triggered intracellular Ca2+ transients similar t o NR1-1a/2A heteromers as verified by combined Ca2+ imaging and whole- cell recording. Thus differences in Ca2+ permeability were not respons ible for the lack of inactivation in NR1-1a/2C heteromers. Our results show that inactivation of recombinant NMDA receptors requires either the NR2A or NR2D subunit, whereas both inactivation and desensitizatio n were absent in NR2C-containing receptors. The gating of inactivating NMDA receptors is more likely to be influenced by ongoing NMDA recept or activity and Ca2+ transients, perhaps consistent with the prominent expression of NR2A in hippocampus and cerebral cortex.