Atrial natriuretic factor (ANF) inhibits basal and stimulated aldosterone production in vitro and in vivo. Infusions of ANF have been reported to inhibit renin release in several species, including humans. A 4-h infusion of ANF into 12 patients with mild hypertension reduced plasma aldosterone at 2 and 4 h, with a rebound increase 2 h after the end of the infusion. Plasma renin activity also showed a mild but significant reduction at 2 h with a rebound increase at 6 h. The effect of ANF on renin release in vitro is controversial. Using a superfusion system of rat renal cortical slices and of dispersed juxtaglomerular (JG) cells, we found that ANF did not affect basal or isoproterenol-stimulated renin release by the kidney slices or dispersed JG cells. These studies suggest that ANF does not inhibit renin release by a direct effect on the JG cells and that the inhibition of renin release found in vivo studies may be mediated through a macula densa mechanism. Since calcium plays an important role in aldosterone secretion, we investigated the effect of ANF on calcium fluxes in rat adrenal glomerulosa cells. ANF did not affect either calcium influx or calcium efflux in rat adrenal glomerulosa cells. These studies suggest that the inhibitory effect of ANF on aldosterone production is distal to the calcium messenger system.