Dopamine D2 receptor-induced heterologous sensitization of adenylyl cyclase requires Gαs:: Characterization Gαs-insensitive mutants of adenylyl cyclase V

Whereas acute stimulation of G alpha (i/o)-coupled receptors inhibits the activity of adenylyl cyclase, a delayed consequence of persistent activation of the receptors is heterologous sensitization, an enhanced responsiveness of adenylyl cyclase to activators such as forskolin or agonists of G alpha (s)-coupled receptors. G alpha (s)-insensitive mutants of adenylyl cyclase type V were used to test the hypothesis that heterologous sensitization requires Ga.-dependent activation of adenylyl cyclase. When adenylyl cyclase was stably expressed in human embryonic kidney (HEK) 293 cells with the D-2L dopamine receptor, basal, forskolin-stimulated, and isoproterenol-stimulated cyclic AMP accumulation were all enhanced by 2-h pretreatment with the D-2 receptor agonist quinpirole. Transient expression of wild-type adenylyl cyclase and three G alpha (s)-insensitive mutants (F379L, R1021Q, and F1093S) in HEK293 cells stably expressing the D-2L receptor demonstrated that all three mutants had little or no responsiveness to beta -adrenergic receptor-mediated activation of Ga. but that the mutants retained sensitivity to forskolin and to D-2L receptor-mediated inhibition. Transiently expressed adenylyl cyclase V was robustly sensitized by 2-h pretreatment with quinpirole. In contrast, the G alpha (s)-insensitive mutants displayed no sensitization of forskolin-stimulated cyclic AMP accumulation, indicating that responsiveness to G alpha (s) is required for the expression of heterologous sensitization.