Fish acclimated to ion-deficient water exhibit proliferation of branchial chloride cells. The objective of the present study was to investigate the role of cortisol in this response using the corticosteroid receptor antagonists RU486 and spironolactone. RU486 is a potent antagonist of the glucocorticoid actions of cortisol, whereas spironolactone exhibits high-affinity binding to mineralocorticoid receptors, with a resulting blockade of mineralocorticoid properties in mammals. Untreated rainbow trout, as well as rainbow trout given a single intraperitoneal implant of coconut oil alone, coconut oil containing RU486 (0.5 mg g(-1)) or coconut oil containing spironolactone (0.1 mg g(-1)), were exposed to either dechlorinated city-of-Ottawa tapwater or artificial softwater for 7 days. Neither corticosteroid antagonist nor acclimation condition affected circulating plasma cortisol levels, plasma ion concentrations or gill Na(+)-K(+)-ATPase activity. Kidney Na(+)-K(+)-ATPase activity was significantly higher in softwater-acclimated fish than in fish held in dechlorinated tapwater. In addition, whereas RU486 treatment was found to be without effect on gill morphometrics, treatment with spironolactone inhibited the proliferation of chloride cells normally associated with acclimation to ion-deficient water. The results of the present study provide further evidence for the mineralocorticoid actions of cortisol in freshwater fish, specifically in eliciting chloride cell proliferation. Furthermore, these results support the hypothesis that distinct glucocorticoid and mineralocorticoid receptor populations are present in teleost fish, despite the apparent absence of the classic mineralocorticoid hormone, aldosterone.