Renal na+-phosphate cotransport in X-linked hyp mice responds appropriately to NA+ gradient, membrane potential, and pH

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Abstract
To investigate the mechanism for the 50% decrease in Vmax of the high-affinity phosphate transport system in the renal brush-border membrane of X-linked Hyp mice, we compared the effects of external Na+ concentration, membrane potential, pH, phosphonoformic acid (PFA), and arsenate on Na+-Pi cotransport in brush-border membrane vesicles prepared from normal mice and Hyp littermates. The affinity of the Na+-Pi cotransport system for Na+ (apparent Km = 60 ± 7 and 64 ± 2 mM for normal and Hyp mice, respectively) and the Na+-Pi stoichiometry estimated from Hill plots (2.5 ± 0.2 and 2.9 ± 0.6 for normal and Hyp mice, respectively) were similar in brush-border membranes of both strains. Inside-negative membrane potential, generated by anions of different permeabilities, stimulated Na+-Pi cotransport and inside-positive membrane potential generated by valinomycin, and a K+ gradient (outside > inside) inhibited Na+-Pi cotransport to the same extent in brush-border membranes derived from normal mice and Hyp littermates. The pH dependence of Na+-Pi cotransport was similar in brush-border membrane vesicles of normal and Hyp mice. The ratio of Na+-Pi cotransport measured at pH 7.5 relative to that at pH 6.5 was 2.9 ± 0.6 in normal mice and 2.9 ± 0.7 in Hyp mice. PFA was a competitive inhibitor of Na+-Pi cotransport in brush-border membranes of both normal and Hyp mice. However, the apparent Ki for PFA was significantly lower in Hyp mice (0.31 ± 0.01 and 0.19 ± 0.02 mM in normal and Hyp mice, respectively, P < 0.05). In contrast, a similar Ki for arsenate inhibition was observed in brush-border membranes of normal and Hyp mice. These results demonstrate that the decrease in Vmax of the high-affinity Na+-Pi cotransport system in renal brush-border membranes of X-linked Hyp mice cannot be attributed to an impaired response of the transporter to the Na+ gradient driving force, membrane potential, or external pH.
Funding Information
  • Medical Research Council
  • Kidney Foundation of Canada

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