The synthesis of nitric oxide (NO) has been demonstrated in the vascular endothelial cells and other tissues including in rat and bovine adrenal gland. To determine whether NO mediates aldosterone production in the adrenals, we evaluated the basal production of aldosterone, and the ACTH or angiotensin II (A II)-stimulated aldosterone in the presence of L-NG monomethyl arginine (L-NMMA) or L-arginine or L-NMMA + L-arginine in isolated rat adrenal capsular tissue by an ex vivo perfusion technique. ACTH increased the release of aldosterone by 40%. Such release of aldosterone was inhibited by L-NMMA, but not by a mixture of L-NMMA and L-arginine. A II stimulated aldosterone release by 40%. The increase in aldosterone release in response to A II was inhibited by L-NMMA or L-arginine. The increase in aldosterone release stimulated by A II was also inhibited by L-NMMA with L-arginine. L-glutamine did not inhibit the A II stimulated aldosterone release. In conclusion, the inhibition of NO synthesis prevented the release of aldosterone stimulated by ACTH, suggesting NO is required for the action of ACTH on aldosterone production.