K+ occupancy of the N-methyl-D-aspartate receptor channel probed by Mg2+ block

The single-channel kinetics of extracellular Mg2+ block was used to probe K+ binding sites in the permeation pathway of rat recombinant NR1/NR2B NMDA receptor channels. K+ binds to three sites: two that are external and one that is internal to the site of Mg2+ block. The internal site is similar to0.84 through the electric field from the extracellular surface. The equilibrium dissociation constant for this site for K+ is 304 mM at 0 mV and with Mg2+ in the pore. The occupancy of any one of the three sites by K+ effectively prevents the association of extracellular Mg2+. Occupancy of the internal site also prevents Mg2+ permeation and increases (by approximately sevenfold) the rate constant for Mg2+ dissociation back to the extracellular solution. Under physiological intracellular ionic conditions and at -60 mV, there is similar to1,400-fold apparent decrease in the affinity of the channel for extracellular Mg2+ and similar to2-fold enhancement of the apparent voltage dependence of Mg2+ block caused by the voltage dependence of K+ occupancy of the external and internal sites.