In an attempt to elucidate early biochemical events in the action of aldosterone on rat kidney in situ, the response of the pyridine nucleotide oxidation-reduction state of surface cells was directly and continuously recorded with an organ-fluorometer. Intravenous administration of aldosterone induced a rapid (maximum response in 10 min) and dose-related (2.5-25 microng/100 g rat) oxidative response that was specific to aldosterone and to the kidney. The oxidative response was 1) detectable with minute dise (approximately 0.2 microng/100 g rat) of hormone: 2) reproduced by other mineralocorticoids; 3) enhanced by a maneuver for expanding the extracellular fluid compartment or by adrenalectomy; and 4) prevented by spironolactone, progesterone, actinomycin D, and cycloheximide. All of these data argue that the redox response is related to the subsequent changes in aldosteronemediated ion transport. Experiments with an uncoupler and with redox substrates showed that mitochondrial NADH was the major nucleotide pool responding to hormone. The oxidation was not accompanied by changes in the adenylate energy charge level of the whole organ. These observations support the view that aldosterone acts on energy metabolism of tubular cells before developing apparent cation transport effects.