Evidence for a tyrosine kinase-dependent activation of the adenylyl cyclase/PKA cascade downstream from the G-protein-linked endothelin ETA receptor in vascular smooth muscle

Endothelin (ET-1), a contractor and mitogen in the vasculature, enhanced cAMP production (t(1/2), 2.2 min; EC50, 89 +/- 6.3 nM) and stimulated activity of the cAMP-dependent protein kinase (PKA) in pig coronary arteries. These responses were blunted by the protein tyrosine kinase (PTK) inhibitors genistein and herbimycin-A, but not by inhibitors of protein kinase C or cyclooxygenase. In contrast, forskolin-stimulated cAMP production was unaffected by PTK inhibition. Immunoblot analysis revealed that ET-1 induced a concentration-dependent protein tyrosine (PT) phosphorylation. Sarafotoxin-c, a selective ETB receptor agonist, had no effect on either cAMP levels or PT phosphorylation. Moreover, pervanadate (PV), a potent inhibitor of PT phosphatases, enhanced both cAMP formation and PT phosphorylation, both of which were blocked by PTK inhibitors. The effects of ET-1 and PV were not additive, suggesting a similar mode of activation, whereas responses to ET-1 and forskolin were synergistic. These findings indicate that AC and PKA are activatable via a nonreceptor PTK-dependent pathway downstream from the G-protein-linked ETA receptor. Because cAMP is a dilator and antimitogen in smooth muscle, stimulation of AC activity may be a negative feedback mechanism regulating ET-1-induced vasoconstriction and/or mitogenesis, (C) 1998 Academic Press.