Pancreatic beta-cells of sulfonylurea receptor type 1 knock-out (SUR1^−/−) mice exhibit an oscillating membrane potential (V _m) demonstrating that hyper-polarisation occurs despite the lack of K_ATP channels. We hypothesize that glucose activates the Na⁺/K⁺-ATPase thus increasing a hyper-polarising current. Elevating glucose in SUR1^−/− beta-cells resulted in a transient fall in V _m and [Ca²⁺]_c independent of sarcoplasmic and endoplasmic reticulum Ca²⁺-activated ATPase (SERCA) activation. This was not affected by K⁺ channel blockade but inhibited by ATP depletion and by ouabain. Increasing glucose also reduced [Na⁺]_c, an effect reversed by ouabain. Exogenously applied insulin decreased [Na⁺]_c and hyper-polarised V _m. Inhibiting insulin signalling in SUR1^−/− beta-cells blunted the glucose-induced decrease of [Ca²⁺]_c. Tolbutamide (1 mmol/l) disclosed the SERCA-independent effect of glucose on [Ca²⁺]_c in wild-type beta-cells. The data show that in SUR1^−/− beta-cells, glucose activates the Na⁺/K⁺-ATPase presumably by increasing [ATP]_c. Insulin can also stimulate the pump and potentiate the effect of glucose. Pathways involving the pump may thus serve as potential drug targets in certain metabolic disorders.