Contractile responses to serotonin were examined in vitro in the longitudinal portal vein to determine whether such responses were mediated by the interaction of serotonin with 5HT1 receptors (those that preferentially bind [3H]serotonin) or 5HT2 receptors (those that preferentially bind [3H]spiperone). Using eight serotonin receptor antagonists (spiperone, metergoline, LY53857, ketanserin, trazodone, benzoctamine, 1-(1-naphthyl)piperazine, and 1-meta-methoxyphenylpiperazine), we found a significant correlation between the affinity for serotonin receptors in the rat portal vein and the ability to bind to 5HT2, but not 5HT1 receptors in rat frontal cortical membranes. Thus, the receptors mediating vascular contraction to serotonin in the rat portal vein were similar to those receptors defined in other vascular beds from the rat (aorta, jugular vein,and caudal artery). Furthermore, contraction resulting from the cumulative addition of serotonin in the rat portal vein was associated with desensitization (higher ED50 value) relative to contractions produced by the non-cumulative addition of serotonin. Affinities of serotonin receptor antagonists were also lower when determined by antagonism of cumulative serotonin concentration-response curves compared to affinities obtained by antagonism of non-cumulative concentration-response curves. Thus, 5HT2 receptor affinities of antagonists in the rat portal vein are best determined by the shift of non-cumulative responses to serotonin.