Glial cell Na,K-ATPase is suggested to participate in extracellular K+ concentration ([K+]o) control by being activated when [K+]o rises in the brain. The extent of that activation directly depends on the Na/K pump affinity to [K+]o, intracellular Na+ ([Na+]i) and, indirectly on pump cycle regulation by membrane potential (Vm). In the present investigation, these Na/K pump properties were studied with the whole-cell patch-clamp technique in cultured mouse oligodendrocytes (N20.1 cell line). N20.1 cells possess ouabain-sensitive Na/K pump current (Ip) with a maximal density of 0.5-0.6 pA/pF (estimated for conditions of Na/K pump stimulation by saturating [Na+]i, [ATP]i, [K+]o and at positive Vm). This current was half-inhibited at 83 +/- 31 microM ouabain, and half-activated by [Na+]i of 9.6 +/- 1.1 mM, by [K+]o of 2.0 +/- 0.1 mM and by membrane potential at about -65 mV. High levels of nervous activity may increase [K+]o from 3 to 12 mM which would only increase Na/K pump current by 40% due to the direct effect of [K+]o. However, elevated [K+]o would also depolarize the glial cell membrane which would indirectly activate Ip and together with the direct effect of [K+]o would increase Ip as much as 2-2.5-fold. These data suggest that glial cell Na/K pump regulation by Vm may be an important factor in determining the participation of the Na/K pump in [K+]o homeostasis in the nervous system.