Previously, we demonstrated that human serotonin (5-HT) 5-HT(7) receptors display marked constitutive activity. Here, we tested if the constitutive activation of adenylyl cyclase by 5-HT(7) receptors influenced both the desensitization properties of transfected 5-HT(7) receptors and the ability of endogenous G(s)-coupled receptors to activate adenylyl cyclase. Using membranes from stably transfected HEK293 cells expressing the recombinant human 5-HT(7) receptor splice variants (5-HT(7(a)), 5-HT(7(b)) and 5-HT(7(d))), we compared the effects of 1-h or 24-h preincubation of the agonist 5-HT, partial inverse agonists mesulergine and SB269970, and full inverse agonists clozapine and methiothepin on subsequent activation of adenylyl cyclase by both 5-HT through transfected 5-HT(7) receptors and the endogenous G(s)-coupled beta-adrenoceptors and prostaglandin receptors of HEK293 cells. The data show that stable expression of 5-HT(7) receptors is sufficient to attenuate adenylyl cyclase activation by endogenous G(s)-coupled receptors. Interestingly, preincubation with inverse agonists not only failed to result in the predicted resensitization of all receptor mediated adenylyl cyclase activation, but some inverse agonists further attenuated (desensitized) beta-adrenoceptor and prostaglandin-stimulated adenylyl cyclase activation similar to long-term agonist exposure by 5-HT. These effects were not correlated with inverse agonist efficacy, were not accompanied by receptor down-regulation and appear to be mediated by a protein kinase A (PKA) independent mechanism. It is concluded that the human 5-HT(7) receptor mediates heterologous desensitization of endogenous G(s)-coupled receptors through an unknown and potentially novel mechanism.