NMDA channel gating is influenced by a tryptophan residue in the M2 domainbut calcium permeation is not altered

N-Methyl-D-aspartate (NMDA) receptors are susceptible to open-channel block by dizolcipine (MK-801), ketamine and Mg2+ and are permeable to Ca2+. It i s thought that a tryptophan residue in the second membrane-associated domai n (M2) may form part of the binding site for open-channel blockers and cont ribute to Ca2+ permeability. We tested this hypothesis using recombinant wi ld-type and mutant NMDA receptors expressed in HEK-293 cells. The tryptopha n was mutated to a leucine (W-5L) in both the NMDAR1 and NMDAR2A subunits. MK-801 and ketamine progressively inhibited currents evoked by glutamate, a nd the rate of inhibition was increased by the W-5L mutation. An increase i n open channel probability accounted for the acceleration. Fluctuation anal ysis of the glutamate-evoked current revealed that the NMDAR1 W-5L mutation increased channel mean open time, providing further evidence for an altera tion in gating. However, the equilibrium affinities of Mg2+ and ketamine we re largely unaffected by the W-5L mutation, and Ca2+ permeability was not d ecreased. Therefore, the M2 tryptophan residue of the NMDA channel is not i nvolved in Ca2+ permeation or the binding of open-channel blockers, but pla ys an important role in channel gating.