Zonal expression of endothelial nitric oxide synthase in sheep and rhesus adrenal cortex

There is mounting evidence that nitric oxide (NO) may inhibit adrenal stero idogenesis by binding to the heme group of P450 enzymes, particularly the r ate-limiting steps cholesterol side-change cleavage P450, aldosterone synth ase P450, and 17 alpha -hydroxylase/C-17/20-lyase P450. Using immunohistoch emistry, nitrotyrosine was detectable throughout the ovine adrenal cortex, and endothelial NO synthase (eNOS) was further identified in zona glomerulo sa (ZG) and at a higher level throughout the zona fasciculata, increasing t oward the medulla. Caveolin-1, 90-kDa heat shock protein, ERK-1/2, and Akt, all known and proposed regulators of eNOS activity, were detected througho ut the ovine adrenal cortex. Western immunoblotting confirmed the identity of these proteins as well as the absence of neuronal NOS, inducible NOS, ca veolin-2, and caveolin-3. Through dual immunostaining we further identified for the first time a zona intermedia without strong staining for 17 alpha -hydroxylase/C-17/20-lyase P450 or angiotensin II type 1 receptor, but posi tive for eNOS. Rhesus adrenals also stained positively for eNOS, but staini ng was seen only in the ZG and zona reticularis. We conclude that eNOS may play a role in controlling zone-specific aldosterone synthase vs. 11 beta - hydroxylase activities of the single CYP11B gene in sheep. In the rhesus mo nkey, NO may modulate ZG aldosterone synthase, but it is not needed for con trol of the distinct 11 beta -hydroxylase in the zona fasciculata. In the z ona reticularis, however, eNOS may control C-19 steroid production at the l evel of 17 alpha -hydroxylase vs. 17,20-lyase activity otherwise unopposed by 3 beta -hydroxysteroid dehydrogenase.