Both glucocorticoids and mineralcorticoids stimulate the renal Na-K-ATPase. However, the exact site of their respective action is not precisely determined and it is still unknown whether these effects are cumulative or not. We studied the effects of dexamethasone and aldosterone on Na-K-ATPase activity in microdissected nephron segments from adrenalectomized rabbits. In proximal convoluted tubule (PCT) the enzyme activity was altered neither by adrenalectomy nor by any steroid replacement. In the medullary thick ascending limb of the loop of Henle (MAL) and the distal convoluted tubule (DCT), Na-K-ATPase activity decreased by 40% after adrenalectomy, and was restored to control level three hours after administration of dexamethasone (100 micrograms/kg) but not by aldosterone (up to 10 micrograms/kg). In the cortical (CCT) and medullary (MCT) collecting tubule the enzyme activity decreased by 75% after adrenalectomy but in contrast with the MAL and the DCT, these two segments were sensitive to both dexamethasone (100 micrograms/kg) and aldosterone (10 micrograms/kg) and recovered their activities within 3 h after the hormone injection. These effects were not additive. Spironolactone (100 micrograms/kg) abolished the action of each of the two hormones on the CCT and MCT. In contrast, spironolactone did not curtail the effect of dexamethasone on MAL and DCT. These results indicate that whereas glucocorticoid action is localized in MAL, DCT, CCT and MCT, the mineralocorticoid effect is restricted to the CCT and MCT exclusively. They also suggest that, in the CCT and MCT, the two types of hormones share the same receptors.