The production of aldosterone by isolated canine zona glomerulosa cells was measured after the incubation of cell suspensions with angiotensin II and ACTH, and during changes in extracellular potassium concentration. Adrenal cell suspensions were prepared by collagenase digestion and physical dispersion of the capsular layer of the dog adrenal cortex, and aldosterone production was determined by direct radioimmunoassay of cell incubation media. The isolated dog adrenal cells were highly responsive to angiotensin II, with aldosterone production significantly stimulated by concentrations of the octapeptide as low as 10(-11)M. Thus, the steroidogenic response of zona glomerulosa cells was consistently observed at peptide concentrations within the physiological range of angiotensin II in dog plasma, i.e., 2.0-5.0 X 5.0 X 10(-11)M. The maximum aldosterone response of 3-8 times the basal level of steroid production was induced by 3 X 10(-10)M angiotensin II, and a decrease in aldosterone production occurred at peptide concentrations above 10(-9)M. The aldosterone response of isolated adrenal cells to ACTH was consistently less sensitive than their response to angiotensin II, by a factor of 10-20 fold. Aldosterone production was significantly increased by 10(-10)M ACTH, and reached a maximum at 10(-8)M ACTH. By contrast with angiotensin II, ACTH usually evoked a higher maximal level of aldosterone production, and did not produce a decline in steroidogenesis at peptide concentrations above the level which caused maximum stimulation of aldosterone formation. Changes in the potassium concentration of cell incubation media were also accompanied by marked effects upon aldosterone synthesis which was abolished in the absence of potassium and became detectable in the presence of 0.5 mM K+. After remaining constant between 2.5 and 4.0 mM K+, aldosterone production rose sharply above 4.5 mM K+ and reached a maximum at 8 mM K+. These observations provide direct evidence that aldosterone production by zona glomerulosa cells is influenced by changes in angiotensin II levels within the normal plasma range.