Redox-induced regulation of the Na/K ATPase was studied in dispersed rat cerebellar granule cells. Intracellular thiol redox state was modulated using glutathione (GSH) conjugating agents and membrane-permeable ethyl ester of GSH (et-GSH) and Na/K ATPase transport and hydrolytic activity monitored as a function of intracellular reduced thiol concentration. Depletion of cytosolic and mitochondrial GSH pools caused an increase in free radical production in mitochondria and rapid ATP deprivation with a subsequent decrease in transport, but not hydrolytic activity of the Na/K ATPase. Selective conjugation of cytosolic GSH did not affect free radical production and Na/K ATPase function. Unexpectedly, over-loading of cerebellar granule cells with GSH triggered global free radical burst originating most probably from GSH auto-oxidation. The latter was not followed by ATP depletion but resulted in suppression of active K⁺ influx and a modest increase in mortality. Suppression of transport activity of the Na/K ATPase was observed in granule cells exposed to both permeable et-GSH and impermeable GSH with inhibitory effects of external and cytosolic GSH being additive. The obtained data indicate that redox state is a potent regulator of the Na/K ATPase function. Shifts from an "optimal redox potential range" to higher or lower levels cause suppression of the Na/K pump activity.