The renin-angiotensin system produces the potent vasoconstrictor angiotensin II, which is involved in i regulation of vascular resistance, blood pressure, and virtually every form of hypertension. Renin is the rate-i limilmg enzymatic step in angiotensin formation. In the kidney, the vasoconstriction by angiotensin is buffered by the vasodilator endothelium-derived nitric oxide (NO). We propose that NO both counteracts angiotensin-induced vasoconstriction and regulates its formation by differentially regulating renin secretion. NO has been reported to both inhibit and stimulate renin secretion, but it is unclear how these different effects could occur. While the cyclic nucleotide which stimulates renin secretion is cAMP, we hypothesize that cGMP, the second messenger of NO, inhibits renin release when cAMP levels are low by activating protein kinase GI! (PKGII) and increasing JG cell intracellular calcium. In contrast, when cAMP is increased by secretagogues such as prostaglandin (PG)E, cGMP stimulates renin release indirectly by inhibiting phosphodiesterase (PDE)-3 and exaggerating cAMPs effects by reducing its metabolism. In aim 1, we propose that with low cAMP levels, NO inhibits renin by stimulating cGMP production and increasing PKGII activity in the JG cells. We will study effects of NO and cGMP on renin without stimulating cAMP both in vitro (primary cultures of isolated JG cells and isolated glomeruli) and in vivo using eNOS knockout mice. In aim 2, we propose that if cAMP levels are elevated, NO enhances renin release by cGMP inhibition of PDE-3 and potentiation of cAMP-mediated renin stimulation. We will study cAMP and macula densa stimulation of renin, blocking PDEs that break down either cGMP or CAMP, using primary JG cultures, isolated glomeruli (without the macula densa) and renal cortical slices (with the macula densa). We will use nNOS knockout mice to see if NO from the macula densa mediates renin stimulation. In aim 3 we propose that prostanoids, particularly PGE2 produced by COX-2 in the macule dense, regulate cAMP levels in JG cells. Co-expression of COX-2 and nNOS in the macula densa suggests NO's effect on renin depends on COX-2 expression and activity. We will manipulate COX-2 expression and PG synthesis in the macula densa and study the effects of NO and cGMP, using renal cortical slices and in vivo experiments stimulating macula densa-mediated renin secretion in rats and nNOS knockout mice. These studies should reveal the dual pathways by which NO can either inhibit or stimulate renin secretion.