The effects of ethanol on the binding of [3H]MK-801, [3H]L-glutamate, [3H]glycine and [3H]CGS 19755 to the N-methyl-D-aspartate receptor were determined in membranes from mouse cortex and hippocampus. Under equilibrium conditions, ethanol in vitro (100 mM) did not alter the apparent affinity or binding site density for any of these ligands. However, in the presence of glutamate and the selective glycine antagonist, 5,7-dichlorokynurenic acid, ethanol inhibited the non-equilibrium binding of [3H]MK-801. This inhibition could be reversed in a time- and concentration-dependent manner by addition of glycine. These data suggest that ethanol may inhibit N-methyl-D-aspartate receptor-mediated responses by altering the kinetics of channel activation. Chronic in vivo ethanol ingestion by mice, that resulted in tolerance to and physical dependence on ethanol, produced an increased density of hippocampal [3H]MK-801 and [3H]L-glutamate binding sites, but not [3H]glycine or [3H]CGS 19755 binding sites. It is possible that chronic ethanol ingestion may influence the subunit composition of the NMDA receptor complex.