Most biological actions of prostaglandins and thromboxane A2 are thought to be mediated by alterations of tissue cyclic nucleotide concentrations. We have shown that endogenous prostaglandins regulate basal and homone-stimulated cyclic AMP concentrations in cultured renal tubular cells retaining and homone-stimulated cyclic AMP concentrations in cultured renal tubular cells retaining differentiated properties. In short-term (2 min) incubations endogenous prostaglandins appeared to be 5 to 10 times more effective than exogenous prostaglandins in increasing intracellulr cyclic AMP concentrations. We have also shown that cyclic AMP and its analogs inhibit prostaglandin biosynthesis by decreasing acylhdrolase (phospholipase) and fatty acid cyclo-oxygenase activity. This applicant proposes to identify the specific endogenous prostaglandins which regulate cyclic AMP concentrations and the reasons for the increased efficacy of endogenous relative to exogenous prostaglandins in cultured renal tubular (MDCK), aortic endothelial and smooth mcuscle cells. Emphasis in aortic cultures, will be placed on prostacyclin (prostaglandin I(2)), a potent vasodilator and anti-thrombotic agent. The regulation of cyclic AMP and cyclic GMP concentrations in the aforementioned cultured cells by platelet-derived arachidonate metabolites, namely thromboxane A(2) and 12-hydroperoxy-5,8,10,14-eicosatetraenoic acid will also be investigated. I also propose to investigate, in detail, the mechanism by which cAMP may inhibit acylhydrolase activity by determining cAMP-induced changes of phospholipid and particularly, of phosphatidylinositol metabolism as well as the cAMP-induced changes of cyclo-oxygenase activity. The potential for cyclic GMP to stimulate prostaglandin biosynthesis will also be investigated. Among the many biochemical methods which will be used are radioimmunoassay of prostaglandins and cyclic nucleotides, as well as radiometric thin-layer chromatography of prostaglandins and phospholipids. Increased understanding of prostaglandin-cyclic nucleotide interactions in renal and vascular cells will be of potential benefit to our understanding of renal water and salt metabolism, regulation of vascular tone, thrombosis, and athersclerosis.
