Nitric oxide inhibits aldosterone synthesis by a guanylyl cyclase-independent effect

To investigate the mechanism of nitric oxide (NO) inhibition of aldosterone release, this study compared the effects of type A natriuretic peptide and heat-stable enterotoxin to a nitric oxide donor, deta nonoate, on cGMP production and angiotensin II-stimulated aldosterone synthesis ill primary cultures of bovine adrenal zona glomerulosa cells. Type A natriuretic peptide (10(-10)-10(-6) M) and deta nonoate (10(-6)-10(-3) M) stimulated concentration-related increases in cGMP production. Heat-stable enterotoxin (10(-6) M) failed to stimulate cGMP synthesis in zona glomerulosa cells. Type A natriuretic peptide and deta nonoate attenuated angiotensin II-stimulated aldosterone production over the same concentration range that stimulated cGMP production. Heat-stable enterotoxin (10(-6) M) was without effect on aldosterone release. To further test the hypothesis that cGMP mediated the inhibition of aldosterone synthesis, the selective inhibitor of soluble guanylyl cyclase, 1H-(1,2,4)oxadiazolo [4,3-a]quinoxalin-1-one (ODQ) was used. ODQ pretreatment (10(-5) M) completely prevented deta nonoate-stimulated cGMP production without altering the inhibitory effect of deta nonoate on angiotensin II-stimulated steroidogenesis. Consistent with its selectivity for inhibiting soluble guanylyl cyclase, ODQ did not block type A natriuretic peptide-stimulated cGMP synthesis or type A natriuretic peptide inhibition of steroidogenesis. Deta nonoate completely blocked 25-hydroxycholesterol- and progesterone-stimulated aldosterone synthesis in zona glomerulosa cells and inhibited the conversion of 25-hydroxycholesterol to pregnenolone in mitochondrial fractions from bovine adrenal cortex. Deta nonoate-derived NO gave an absorbance maximum of the mitochondrial cytochrome P450 of 453 nm and inhibited the absorbance at 450 nm caused by carbon monoxide binding to the enzyme. These results suggest that deta nonoate reduces steroidogenesis independent of guanylyl cyclase activation and that NO has a direct effect to inhibit the activity of cytochrome P450, probably by binding to the heme groups of the cytochrome.