Effect of ouabain upon diuretic-sensitive K + transport in cultured cells evidence for separate modes of operation of the transporter

Abstract

(1) Unidirectional K + ( 86Rb) influx and efflux were measured in subconfluent layers of MDCK renal epithelial cells and HeLa carcinoma cells. (2) In both MDCK and HeLa cells, the furosemide-inhibitable and chloride-dependent component of K + influx/efflux was stimulated 2-fold by a 30 min incubation in 1 · 10 −3 M ouabain. (3) Measurements of net K + loss and Na + gain in ouabain-treated cells at 1 h failed to show any diuretic sensitive component, confirming the exchange character of the diuretic-sensitive fluxes. (4) Prolonged incubations for 2.5 h in ouabain revealed a furosemide- and anion-dependent K + (Cl −) outward net flux uncoupled from net Na + movement. Net K + (Cl −) outward flux was half-maximally inhibited by 2 μM furosemide. (5) After 2.5 h ouabain treatment, the anion and cation dependence of the diuretic-sensitive K + influx/efflux were essentially unchanged when compared to untreated controls.