Ca2+ entry through voltage-gated Ca2+ channels and subsequent insulin secretion in pancreatic b-cells is a function of membrane potential. Due to the electrogenic nature of the Na+K+-ATPase, stimulation of the Na+K+-ATPase may generate a hyperpolarizing current, inhibiting Ca2+ entry. Glucocorticoids have been shown previously to inhibit insulin release at least partially by blunting Ca2+ entry. The aim of the present study was to examine whether glucocorticoids increase Na+K+-ATPase activity in insulin secreting cells. Na+K+-ATPase activity was estimated by the mean of outward current generated by the enzyme which was measured using the whole cell configuration of the patch clamp method. The addition of 5 mM KCl to a bath solution lacking potassium initiated an outward current of 0.54 ± 0.02 pA/pF (n=18) in INS-1 cells and of 0.75 ± 0.1 pA/pF (n=11) in isolated mouse islet cells. INS-1 cells were hyperpolarized by - 16.9 ± 2.7 mV, mouse islet cells by -9.3 ± 1.3 mV. Ouabain (1 mM) or omission of ATP from the pipette solution almost completely inhibited the effects of KCl. Pretreatment of INS-1 cells with 100 nM dexamethasone for 5h-24h increased outward current significantly to 0.65 ± 0.04 pA/pF (n=12) an effect which was antagonized by RU486. These data suggest that dexamethasone upregulates Na+K+-ATPase. The concomitant hyperpolarisation may contribute to the inhibition of insulin secretion after glucocorticoid excess.