WAY-100635 is the first selective, silent 5-HT1A (5-hydroxytryptamine1A, serotonin-1A) receptor antagonist. We have investigated the use of [3H]WAY-100635 as a quantitative autoradiographic ligand in post-mortem human hippocampus, raphe and four cortical regions, and compared it with the 5-HT1A receptor agonist, [3H]8-OH-DPAT. Saturation studies showed an average Kd for [3H]WAY-100635 binding in hippocampus of 1.1 nM. The regional and laminar distributions of [3H]WAY-100635 binding and [3H]8-OH-DPAT binding were similar. The density of [3H]WAY-100635 binding sites was 60-70% more than that of [3H]8-OH-DPAT in all areas examined except the cingulate gyrus where it was 165% higher. [3H]WAY-100635 binding was robust and was not affected by the post-mortem interval, freezer storage time or brain pH (agonal state). Using [3H]WAY-100635, we confirmed an increase of 5-HT1A receptor binding sites in the frontal cortex in schizophrenia, previously demonstrated with [3H]8-OH-DPAT. Compared to [3H]8-OH-DPAT, [3H]WAY-100635 has two advantages: it has a higher selectivity and affinity for the 5-HT1A receptor, and it recognizes 5-HT1A receptors whether or not they are coupled to a G-protein, whereas [3H]8-OH-DPAT primarily detects coupled receptors. Given these considerations, the [3H]WAY-100635 binding data in schizophrenia clarify two points. First, they indicate that the elevated [3H]8-OH-DPAT binding seen in the same cases is attributable to an increase of 5-HT1A receptors rather than any other binding site. Second, the enhanced [3H]8-OH-DPAT binding in schizophrenia reflects an increased density of 5-HT1A receptors, not an increased percentage of 5-HT1A receptors which are G-protein-coupled. We conclude that [3H]WAY-100635 is a valuable autoradiographic ligand for the qualitative and quantitative study of 5-HT1A receptors in the human brain.