This study determined whether nucleotides that bind to purinergic receptors (P2R) regulate the expression or function of serum- and glucocorticoid-inducible kinase-1 (SGK1) in mouse renal inner medullar collecting duct cells (mIMCD-3). The SGK1 protein was detected by Western blotting. A significant reduction of cytosolic SGK1 expression was observed in the cells pretreated with P2R agonist adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS), and the reduction could be reversed by P2R antagonists. This reduction was also observed in cells that were pretreated with agonists for P2R subtypes. Using ELISA, we observed a reduced SGK1 kinase activity in ATPgammaS-pretreated cells. This effect was reversed by P2R antagonists. Furthermore, an increase of SGK1 kinase activity in aldosterone-pretreated cells was suppressed by ATPgammaS. These studies demonstrate for the first time that SGK1 can be downregulated by nucleotides in renal collecting duct epithelial cells, likely via the activation of P2R, and suggest that activation of renal purinergic signaling regulates a SGK1-dependent pathway that is known to modulate ion transport in the renal collecting duct.