Renal ischemia and reperfusion (I/R) injury is the major cause of acute renal failure and may also be involved in the development and progression of some forms of chronic kidney disease. We previously showed that a mineralocorticoid receptor (MR) blockade prevents renal vasoconstriction induced by cyclosporine that leads to acute and chronic renal failure (Feria I, Pichardo I, Juarez P, Ramirez V, Gonzalez MA, Uribe N, Garcia-Torres R, Lopez-Casillas F, Gamba G, Bobadilla NA. Kidney Int 63: 43-52, 2003; Perez-Rojas JM, Derive S, Blanco JA, Cruz C, Martinez de la Maza L, Gamba G, Bobadilla NA. Am J Physiol Renal Physiol 289: F1020-F1030, 2005). Thus we investigated whether spironolactone administration prevents the functional and structural damage induced by renal ischemia-reperfusion (I/R). Five groups were studied: sham-operated animals, rats that underwent 20 min of ischemia and 24 h of reperfusion, and three groups that received spironolactone 1, 2, or 3 days before I/R, respectively. Renal I/R produced significant renal dysfunction and tubular damage. Spironolactone administration completely prevented a decrease in renal blood flow, the development of acute renal failure, and tubular apoptosis. The protection conferred by spironolactone was characterized by decreasing oxidative stress, as evidenced by a reduction in kidney lipoperoxidation, increasing expression of antioxidant enzymes, and restoration of urinary NO(2)/NO(3) excretion. Endothelial nitric oxide synthase expression was upregulated by a mineralocorticoid receptor blockade in I/R groups; in addition, an increase in activating phosphorylation of this enzyme at residue S1177 and a decrease in inactivating phosphorylation at T497 were observed. In conclusion, our study shows that spironolactone administration prevents the renal injury induced by I/R, suggesting that aldosterone plays a central role in this model of renal injury.