Abstract
Short-term activation of Gαi/o-coupled receptors inhibits adenylyl cyclase, whereas persistent activation of Gαi/o-coupled receptors results in a compensatory sensitization of adenylyl cyclase activity after subsequent activation by Gαs or forskolin. Several indirect observations have suggested the involvement of increased Gαs-adenylyl cyclase interactions in the expression of sensitization; however, evidence supporting a direct role for Gαs has not been well established. In the present report, we used two genetic approaches to further examine the role of Gαs in heterologous sensitization of Ca2+-sensitive type 1 adenylyl cyclase (AC1). In the first approach, we constructed Gαs-insensitive mutants of AC1 (F293L and Y973S) that retained sensitivity to Ca2+ and forskolin activation. Persistent (2 h) activation of the D2 dopamine receptor resulted in a significant augmentation of basal or Ca2+- and forskolin-stimulated AC1 activity; however, sensitization of Gαs-insensitive mutants of AC1 was markedly reduced compared with wild-type AC1. In the second strategy, we examined the requirement of an intact receptor-Gαs signaling pathway for the expression of sensitization using dominant-negative Gαs mutants (α3β5 G226A/A366S or α3β5 G226A/E268A/A366S) to disrupt D1 dopamine receptor activation of recombinant AC1. D1 dopamine receptor-Gαs signaling was attenuated in the presence of α3β5 G226A/A366S or α3β5 G226A/E268A/A366S, but D2 agonist-induced sensitization of Ca2+-stimulated AC1 activity was not altered. Together, the present findings directly support the hypothesis that the expression of sensitization of AC1 involves Gαs-adenylyl cyclase interactions.
- Received March 5, 2004.
- Accepted September 10, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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