Adenylyl cyclase isoforms and vasopressin enhancement of agonist-stimulated cAMP in vascular smooth muscle cells

The influence of arginine vasopressin (AVP) on agonist-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) accumulation was investigated in vascular smooth muscle cells (VSMC) cultured from rat thoracic aorta. Incubation of VSMC with AVP for 60 s produced a 2- to 2.5-fold enhancement of isoproterenol-induced cAMP formation. AVP also increased cAMP stimulation by the prostaglandin I-2 analogue iloprost. The effect of AVP to enhance agonist-stimulated cAMP formation was completely inhibited in cells pretreated with a selective antagonist of V-1 vasopressin receptors but was not affected by blockade of V-2 receptors. Inhibition of protein kinase C activation failed to alter the action of AVP to potentiate cAMP stimulation, but treatment of cells with calmodulin antagonists significantly attenuated this effect of the peptide. Moreover, depletion of Ca2+ stores with thapsigargin decreased AVP enhancement of isoproterenol-stimulated cAMP by >70%. The action of AVP to increase cAMP stimulation was also demonstrated in freshly isolated strips of rat aorta where treatment with peptide produced a twofold increase in isoproterenol-stimulated cAMP formation. RNA blot analysis indicated expression in VSMC of mRNA encoding type III adenylyl cyclase, a Ca2+-calmodulin-sensitive isoform of the effector. Furthermore, when detergent-solubilized membrane extract was subjected to calmodulin affinity chromatography, a peak of adenylyl cyclase activity was identified which had affinity for calmodulin matrix in the presence of Ca2+. The results indicate that AVP activates V-1 receptors in VSMC to enhance agonist-stimulated cAMP formation by a Ca2+-calmodulin-dependent mechanism and suggest that type III adenylyl cyclase may provide a focal point in the VSMC for cross talk between constrictor and dilator pathways.