Glutathione-dependent inactivation of sodium-dependent phosphate transport across rat renal brush-border membrane

Abstract

Thiol/disulfide is fundamental in protein function; we previously observed an inhibitory effect of thiol oxidants on the Na-dependent phosphate (Pi) uptake into renal brush border membrane vesicles (BBMV). We examined whether oxidation of glutathione (GSH) is involved in the mechanism. Vesicular thiols were measured by liquid chromatography. BBMV were incubated with reagents before an influx of Pi. Diamide (5 mM) reduced the capacity of the Pi uptake. Subsequent treatment with dithiothreitol (5 mM) blocked the inhibitory effect of diamide. Vesicular GSH was not modified only by the incubation, whereas it was oxidized by the treatment with diamide, and reduced by dithiothreitol. Furthermore, in vivo treatment with cAMP provided GSH-depleted BBMV without any influence on Pi uptake. Diamide did not inhibit the transport of Pi into GSH-depleted vesicles, but it did inhibit the uptake when GSH was introduced into the vesicles. In conclusion, a GSH-dependent mechanism is involved in the inhibitory effect of diamide on sodium-dependent Pi transport across the renal brush-border membrane.