In the present study, we examined whether chronic exposure of C6BU-1 cells to 100 nM of several different types of antidepressants directly influences serotonin2A (5-HT2A) receptor-stimulated intracellular Ca2+ mobilization. Imipramine, desipramine, clomipramine, and maprotiline amplified the 5-HT response at 48, but not at 2, h. Imipramine increased the maximum response to 5-HT without altering the EC50 of the dose-response curve. This effect was time dependent and cycloheximide blocked the maximal induction, suggesting an essential role for protein synthesis in this process. Previous exposure of the cells to thrombin or isoproterenol did not influence 5-HT2A receptor function and pretreatment with imipramine did not alter the thrombin- or bradykinin-induced Ca2+ mobilization, which indicates that the effects of imipramine appear to be specific to the 5-HT2A receptor. The effect of imipramine was potently suppressed by a calmodulin antagonist, W-13, in a dose-dependent manner. Furthermore, this amplified 5-HT response was blocked by KN-93, but not by H-7. Taken together, these results suggest that imipramine has a modulatory effect on the 5-HT2A receptor-coupled intracellular Ca2+ in C6 cells through a calmodulin-dependent pathway, possibly involving Ca2+/calmodulin kinase activation.