Volume-sensitive, Cl-dependent K transport in resealed human erythrocyte ghosts

Abstract

Potassium influx and efflux in Cl and NO3 media were measured in resealed ghosts prepared from human red cells. Cl-dependent K influx was three times that in intact cells and, as in intact cells, was partially supported by Br but not by thiocyanate (SCN). In other properties, this flux differed from that in intact cells: substitution of N-methylglucamine for Na did not decrease but rather increased Cl-dependent K influx, the affinity for external K was reduced, with a Km of 21.3 +/- 12.5 mM, and inhibition by furosemide and bumetanide was incomplete. Furosemide at 1 mM inhibited Cl-dependent influx by 26 and 51% at 4 and 20 mM K, respectively. Bumetanide inhibited Cl-dependent K influx by 0 and 55% at concentrations of 10 microM and 1 mM, respectively, in 4 mM K, with no further inhibition at 20 mM K. Neither the magnitude nor the properties of the flux were altered by preparing ghosts in the presence of 1,4-dithiothreitol, indicating that sulfhydryl oxidation was not responsible for the altered flux in ghosts. Treatment with N-ethylmaleimide (NEM) either before or after ghost preparation did not increase Cl-dependent K influx. However, Cl-dependent influx in ghosts could be augmented by increasing ghost volume or ATP content. Resealed human erythrocyte ghosts thus exhibit a volume- and ATP-sensitive, Cl-dependent K flux that differs substantially from the putative Na-K-Cl cotransport in intact cells in that it is independent of Na, is relatively resistant to furosemide and bumetanide, and has a low affinity for K.(ABSTRACT TRUNCATED AT 250 WORDS)