Thiol-Dependent passive K/Cl transport in sheep red cells: IV. Furosemide inhibition as a function of external Rb+, Na+, and Cl−

Abstract

The effect of the loop diuretic furosemide (4-chloro-N-furfuryl-5-sulfamoyl-anthranilic acid) on the thiol-dependent, ouabain-insensitive K(Rb)/Cl transport in low K+ sheep red cells was studied at various concentrations of extracellular Rb+, Na+ and Cl-. In Rb+-free NaCl media, 2 X 10(-3) M furosemide inhibited only one-half of thiol-dependent K+ efflux. In the presence of 23 mM RbCl, however, the concentration of furosemide to produce 50% K+ efflux inhibition (IC50) was 5 X 10(-5) M. In Rb+ containing NaCl media, the inhibitory effect of 10(-3) M furosemide was equal to that caused by NO-3 replacement of Cl- in the medium. The apparent synergistic action of furosemide and external Rb+ on K+ efflux was also seen in the ouabain-insensitive Rb+ influx. A preliminary kinetic analysis suggests that furosemide binding alters both maximal K+(Rb+) transport and apparent external Rb+ affinity. In the presence of external Rb+, Na+ (as compared to choline) exerted a small but significant augmentation of the furosemide inhibition of K+(Rb+) fluxes. There was no effect of Cl- on the IC50 value of furosemide. As there is no evidence for coupled Na+K+ cotransport in low K+ sheep red cells, furosemide may modify thiol-dependent K+(Rb+)/Cl flux or Rb+ (and to a slight degree Na+) modulate the effect of furosemide.

Molecular Physiology and Pathophysiology of Electroneutral Cation-Chloride Cotransporters

“…It was shown that transfection of HEK-293 cells with rbKCC1 cDNA produced a significant increase of an 86 Rb ϩ efflux mechanism that was activated by twofold when cells were exposed to hypotonicity. rbKCC1 activity assessed as Cl Ϫ -dependent 86 Rb ϩ uptake was stimulated by NEM and inhibited by the presence of 2 mM furosemide, indicating that rbKCC1 indeed encoded a K ϩ -Cl Ϫ cotransporter with similar characteristics to those previously reported in erythrocytes (232,233). Although ion transport kinetics for Cl Ϫ and Rb ϩ dependency of 86 Rb ϩ uptake were assessed, K m values were reported only as Ͼ25 mM for Rb ϩ and as Ͼ50 mM for extracellular Cl Ϫ .…”

Plasmodium falciparum likely encodes the principal anion channel on infected human erythrocytes

“…First, although a number of transporters in other systems are inhibited by furosemide, its affinity for other transporters varies over a wide range, with published inhibitory concentration (IC 50 ) values from nano-to millimolar. [20][21][22] Second, the data in Figure 1D were adequately fitted by the Michaelis-Menten equation (solid line), implicating a 1:1 stoichiometry of furosemide binding to PSAC and further supporting the overwhelming predominance of a single ion channel type.…”

Molecular Characterization of a Putative K-Cl Cotransporter in Rat Brain

“…These cotransporters directly couple the movement of cations and Cl Ϫ ions in an electrically neutral fashion, and therefore, net ion movement is dependent solely on the chemical gradients of the transported ions. Two such cationchloride cotransporters (CCC) 1 include the Na-K-Cl and K-Cl cotransporters. Both of these transporters are sensitive to the sulfamoylbenzoic acid "loop" diuretics furosemide and bumetanide; however, the K-Cl cotransporter has a much lower affinity for these drugs than the Na-K-Cl cotransporter (1).…”

“…Two such cationchloride cotransporters (CCC) 1 include the Na-K-Cl and K-Cl cotransporters. Both of these transporters are sensitive to the sulfamoylbenzoic acid "loop" diuretics furosemide and bumetanide; however, the K-Cl cotransporter has a much lower affinity for these drugs than the Na-K-Cl cotransporter (1). The K-Cl cotransporter was first described as a swellingactivated K ϩ efflux pathway in avian red blood cells (2).…”