To test the proposal that the enzyme 11β-hydroxysteroid dehydrogenase (11β-HSD) confers aldosterone specificity on mineralocorticoid receptors in the distal nephron by inactivating glucocorticoids, we performed a free-flow micropuncture study of distal tubular function in adrenalectomized rats infused with high-dose corticosterone. One-half of the rats were additionally given intravenous carbenoxolone (CBX; 6 mg/h) to inhibit renal 11β-HSD activity. Although this maneuver lowered fractional Na⁺ excretion (1.1 ± 0.2 vs. 1.9 ± 0.2%,P < 0.01), Na⁺ reabsorption within the accessible distal tubule was found to be similar in the two groups of animals. In contrast, distal tubular K⁺ secretion was enhanced in CBX-treated rats: fractional K⁺ deliveries to the early and late distal collection sites in the corticosterone-alone group were 13 ± 1 and 20 ± 3%, respectively (not significant), whereas corresponding data in the CBX-treated group were 9 ± 1 and 24 ± 2% (P < 0.01). This stimulation of distal K⁺ secretion provides the first direct in vivo evidence that 11β-HSD normally prevents corticosterone from exerting a mineralocorticoid-like effect in the distal tubule. The reduction in fractional Na⁺ excretion during inhibition of 11β-HSD, in the absence of a change in end-distal Na⁺delivery, suggests enhanced Na⁺ reabsorption in the collecting ducts.