Cl(-)-dependent ATP-driven H+ transport in rabbit renal cortical endosomes

Abstract

An endosomal fraction isolated from rabbit renal cortex by a novel, fast, and simple procedure was enriched in ATP-dependent H+ pumping that was oligomycin insensitive but was inhibited by dicyclohexylcarbodiimide (DCCD), N-ethylmaleimide (NEM), Zn2+, Hg2+, diethylstilbestrol, mersalyl, and 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole. No substantial Na+-H+ exchange was detected. Electrogenicity of the pump was demonstrated using [14C]-SCN-. In addition, these membranes featured ATP-dependent Cl- flux. The ATP-driven H+ pumping had an absolute requirement for Cl-: an inside-negative membrane potential was not a substitute for Cl-. The protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone inhibited ATP-driven Cl- uptake but no inhibition was observed with nigericin. Finally, both ATP-driven H+ pumping and ATP-dependent Cl- flux were inhibited by Cl(-)-channel inhibitors. Part, or all, of the absolute dependence on Cl- may derive from a Cl- channel, the function of which is intimately related to H+ pumping by the ATPase. Flux through this Cl- channel may be regulated by one or more factors, including ATP, membrane potential, and pH.

Physiological implications of the regulation of vacuolar H+-ATPase by chloride ions

“…The role of chloride channels in H + -ATPase function is not explained only by the establishment or dissipation of a potential difference across cellular and intracellular membranes, but some evidence has supported a direct, specific role of Cl -in stimulating H + -ATPase activity (16). Kaunitz et al (17) have shown that acidification of microsomes from the rat renal medulla depends on chloride channels.…”

“…In the kidney, the role of Cl -channels in regulating H + -ATPase activity has been studied in endosomal fractions (16,27,29), brush border membrane vesicles (3) as well as in apical membranes of renal tubules (10,(30)(31)(32).…”

Angiotensin II stimulates vesicular H + -ATPase in rat proximal tubular cells

“…In view of evidence that the activity of H ϩ -ATPase is affected by Cl Ϫ (2,11,12), experiments were carried out in which either Cl Ϫ was removed from the perfusion fluid (Fig. 2B) or AII was added in the presence of the potent anion channel blocker NPPB (see Fig.…”

“…Two mechanisms have been suggested to account for the Cl Ϫ dependence of H ϩ -ATPase activity. First, Cl Ϫ ions are necessary as cotransported anions to maintain electric neutrality during H ϩ ion transport (12,20,(22)(23)(24). The observation that the deletion of Cl Ϫ and exposure to Cl Ϫ channel blockers abolishes AII-stimulated pH recovery after deletion of Na ϩ supports this view.…”

Cl- and regulation of pH by MDCK-C11 cells

“…It has been shown that the presence of ATP activates not only H + -ATPase, but also the Cl -channels connected to it (10); and even after dissipating the PD across endosome membranes with valinomycin, the ATPase still depends on the presence of Cl - (3).…”