Serotonin (5-HT)-induced changes in the levels of intracellular Ca++ were analyzed in human platelets, using the Ca+(+)-sensitive dye 1-(2-(5'-carboxyoxazol-2'-yl)-6-aminobenzofuran-5-oxy)-2-(2' -amino-5'- methylphenox)-ethane-N,N,N',N'-tetraacetic acid, pentaacetoxymethyl ester, to investigate the regulation of 5-HT2 receptor function. Serotonin mobilized intracellular Ca++ in a dose-dependent fashion from basal level of 98 +/- 2.7 and up to 211 +/- 5.8 nM with an EC50 value for 5-HT of 0.2 microM. Ketanserin, a 5-HT2 antagonist, reversed the 5-HT (10 microM)-induced Ca++ increase in a dose-dependent manner with an IC50 value of 2 nM. An initial treatment with 10 microM 5-HT abolished the response to a second treatment with 100 microM 5-HT, suggesting that 5-HT evoked an acute desensitization of 5-HT2 receptors in human platelets. Mezerein and phorbol 12-myristate 13-acetate, activators of protein kinase C, inhibited 5-HT-stimulated inositol monophosphate accumulation with IC50 values of 3 and 10 nM, respectively. Furthermore, a protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride prevented the protein kinase C activator-induced inhibition against 5-HT-mediated inositol monophosphate accumulation. Mezerein also inhibited 5-HT (10 microM)-mediated Ca++ release with an IC50 value of 3 nM. 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride prevented the inhibition by mezerein of the 5-HT-stimulated Ca++ increase. Moreover, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride by itself enhanced the Ca++ spike induced by 100 microM 5-HT, the plateau phase induced by 10 microM 5-HT and the second response to 5-HT. These findings suggest that 5-HT2 receptor activation mobilizes intracellular Ca++ in human platelets and that this receptor may be desensitized acutely by a protein kinase C mediated feedback system.