Effects of Mg2+ on recovery of NMDA receptors from inhibition by memantine and ketamine reveal properties of a second site

Memantine and ketamine are NMDA receptor (NMDAR) open channel blockers that are thought to act via similar mechanisms at NMDARs, but exhibit divergent clinical effects. Both drugs act by entering open NMDARs and binding at a site deep within the ion channel (the deep site) at which the endogenous NMDAR channel blocker Mg2+ also binds. Under physiological conditions, Mg2+ increases the IC50s of memantine and ketamine through competition for binding at the deep site. Memantine also can inhibit NMDARs after associating with a second site accessible in the absence of agonist, a process termed second site inhibition (SSI) that is not observed with ketamine. Here we investigated the effects of 1 mM Mg2+ on recovery from inhibition by memantine and ketamine, and on memantine SSI, of the four main diheteromeric NMDAR subtypes. We found that: recovery from memantine inhibition depended strongly on the concentration of memantine used to inhibit the NMDAR response; Mg2+ accelerated recovery from memantine and ketamine inhibition through distinct mechanisms and in an NMDAR subtype-dependent manner; and Mg2+ occupation of the deep site disrupted memantine SSI in a subtype-dependent manner. Our results support the hypothesis that memantine associates with, but does not inhibit at the second site. After associating with the second site, memantine can either slowly dissociate directly to the extracellular solution, or transit to the deep site, resulting in typical channel block. Memantine's relatively slow dissociation from the second site underlies the dependence of NMDAR recovery from inhibition on both memantine concentration and on Mg2+.