Inward Na + currents associated with the cloned intestinal Na +/glucose cotransporter expressed in Xenopus oocytes have been studied using the twomicroelectrode voltage-clamp method. The steady-state current/voltage relations showed voltage-dependent (Vm from +20 to --75 mV) and relatively voltage-independent (Vm from -75 to -150 mV) regions. The apparent Imax for Na + and glucose increased with negative membrane potentials, and the apparent Ko.5 for glucose (Ko~)~) depended on Vm and [Na]o. Increasing [Na]o from 7 to 110 mmot/1 had the same effect in decreasing KoG.~ from 0.44 to 0.03 mmol/1 as increasing the Vm from --40 to --150 inV. The I/V curves under saturating conditions (20 retool/1 external sugars and 110 retool/1 [Na]o) were identical for D-glucose, D-galactose, e-methyl Dglucopyranoside and 3-O-methyl D-glucoside. The specificity of the cotransporter for sugars was: D-glucose, Dgalactose, e-methyl D-glucopyranoside > 3-O-methyl Dglucoside ~> I>xylose > D-allose ,> D-mannose. Ki for phlorizin (~ 10 gmol/1) was independent of Vm at saturating [Na]o. We conclude that a variety of sugars are transported by the cloned Na+/glucose cotransporter at the same maximal rate and that membrane potential affects both the maximal current and the apparent Ko.5 of the cotransporter for Na + and glucose.