Na+-dependent hexose transport in vesicles from cultured renal epithelial cell line

Abstract

Apical membrane vesicles were prepared from cultured epithelia formed by LLC-PK1 cells using a calcium precipitation technique. alpha-Methylglucoside uptake into this vesicle preparation was markedly stimulated by sodium and inhibited by phlorizin. In addition, a transient "overshoot" of intravesicular alpha-methylglucoside concentration above its equilibrium value was observed under initial sodium gradient conditions. The specificity of this sodium-dependent hexose transporter closely resembled that found in the mammalian kidney brush border membrane, e.g., alpha-methylglucoside, D-glucose, and D-galactose apparently share the transporter while 2-deoxy-D-glucose, mannose, and fructose do not. Kinetic analysis of the sodium-dependent component of alpha-methylglucoside flux into LLC-PK1 apical membrane vesicles indicates the existence of single transporter with Km congruent to 2 mM and Vmax congruent to 3 nmol.min-1.mg protein-1. Measurement of alpha-methylglucoside uptake as a function of sodium concentration is consistent with a sodium:sugar stoichiometry of approximately 2:1.l There is a good correlation over time between the development of the concentrating capacity of the intact epithelium for alpha-methylglucoside and the transport properties of the vesicle preparation.