The objectives of the proposed research are to develop additional support for the role of cyclic GMP in the vasodilation of isolated bovine intrapulmonary vessels and to use this system to reconstruct the physiological regulation of guanylate cyclase(GC). Several different classes of vasodilator agents, which potentially possess secondary actions, will be compared for the relationship between the degree of vascular relaxation and the magnitude of tissue level increases in cyclic GMP. These studies will be performed in isolated normal intra-pulmonary arteries and veins using phenylephrine for a contractile agent. The types of vasodilators to be studied include: acetylcholine or related endothelial cell dependent dilators, metabolites of oxygen, arachidonic acid(AA) and related non-prostanoid metabolites, and hydrazine derivatives including phenyl-hydrazine and hydralazine. These agents will be compared to the well characterized nitrogen oxide type vasodilators that are thought to act through cyclic GMP. The mechanism of vasodilator action will be probed by measurement of changes in cyclic nucleotide levels, the effects of some inhibitors of AA metabolism and determination of the endothelial cell dependence. Agents that directly activate purified GC will be compared for the maximal degree of enzyme activation and tissue level increases of cyclic GMP, and for responses to GC activation inhibitors. The mechanism of indirect activators of GC will be probed with additional inhibitors and by measurement of metabolites related to intra-cellular chemical redox pontential and hydroperoxide metabolism. Based on the results obtained, an attempt will be made to reconstruct the mechanism of the indirect activation of purified GC. Since the endothelial cell dependent activation of arterial smooth muscle GC by acetylcholine currently appears to be the best suited system for the study of the physiological regulation of GC, the mechanism of this process will be the primary focus of the proposal. In addition, these studies will contribute to the understanding of physiological and pharmacological mechanisms of vasodilation in the intra-pulmonary vessels.
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