G-protein-coupled receptor agonists augment adenylyl cyclase activity induced by forskolin in human corpus cavernosum smooth muscle cells.

Abstract

The goal of this study was to investigate the synergistic effects between G-protein-coupled receptor agonists and forskolin-induced activation of adenylyl cyclases, in cultured human corpus cavernosum smooth-muscle cells. Treatment of human corpus cavernosum smooth-muscle cells with forskolin (0.1-10 microM) produced an increase in cAMP synthesis in a concentration-dependent manner. Forskolin-induced adenylyl cyclase activity was markedly augmented by prostaglandin E1 (PGE1) and its metabolite, PGE0, isoproterenol, carbachol, and phenylephrine. Augmentation of forskolin-induced cAMP by PGE1, and PGE0 is probably mediated by prostaglandin E receptors (EP). Enhancement of forskolin-induced cAMP synthesis by isoproterenol is mediated by beta-adrenergic receptors (beta-AR), since this activity was inhibited by propranolol. Stimulation of forskolin-induced cAMP synthesis by carbachol is attributed to activation of muscarinic acetylcholine receptors (mAChR), as demonstrated by inhibition with atropine. The augmentation of forskolin-induced cAMP synthesis by phenylephrine, an alpha1-adrenergic receptor (AR) agonist, however, was unexpected and cannot be attributed to increased intracellular Ca2+, since treatment of cells with either the Ca2+ ionophore, A23187, or 80 mM KCl did not affect forskolin-induced cAMP synthesis. Stimulation of forskolin-induced cAMP synthesis by phenylephrine is explained by its binding to beta-AR and activation of Gs protein, since this augmentation was inhibited by the beta-AR antagonist, propranolol. This observation was further supported by physiological studies in organ bath chambers, in which forskolin-induced relaxation of precontracted corpus cavernosum strips was enhanced by phenylephrine. These studies suggest that synergism between agonist-induced cAMP synthesis and forskolin is attributed to increased conformational stabilization of activated adenylyl cyclase catalytic domains by forskolin and the Gs(alpha)-subunit of activated Gs proteins.