The activity of the neuroprotective agent clomethiazole at glutamate and ion channel sites has been investigated. Dizocilpine (3.25 mg/kg intraperitoneally) provided almost total protection against the damage produced by infusion of N-methyl-DL-aspartate (NMDLA; 75 micrograms) into the right hippocampus. In contrast, clomethiazole (96 mg/kg intraperitoneally) was without effect. Using ligand binding techniques, no evidence was found for clomethiazole interacting with NMDA, AMPA or sigma binding sites. Clomethiazole did inhibit the stimulatory effect of the metabotropic glutamate receptor agonist 1S3R-aminocyclopentone-1,3-dicarboxylic acid (ACPD) on phosphoinositol hydrolysis, but only at a concentration of 10(-3) M, which is unlikely to have functional relevance. Clomethiazole was also without effect on ligand binding to Ca2+ channels (N- or L- type), Na+ channels or ATP-sensitive K+ channels. Potentiation of GABA function therefore remains the most plausible explanation for the neuroprotective activity of clomethiazole.