Inhibition by amiloride analogues of Na+-dependent hexose uptake in LLC-PK1/Cl4 cells.

Amiloride and four analogues of amiloride were shown to inhibit Na+-dependent, phlorizin-sensitive hexose uptake by a clone of pig kidney cells, LLC-PK1/Cl4. The analogues tested were: 5-(N-ethyl-N-isopropyl)amiloride (EIPA), 5-(N-methyl-N-isobutyl)amiloride (MIBA), 3',4'-dichlorobenzamil, and phenamil. The transport substrate was the nonmetabolizable glucose analogue alpha-methyl-D-glucoside. Blockade of Na+-K+ transport at the basolateral membranes or removal of divalent cations from the assay medium had little effect on the initial rate of hexose uptake, whereas MIBA remained an effective inhibitor under both conditions. The inhibitions by EIPA of Na+-H+ exchange and hexose-dependent Na+ uptake could be distinguished by appropriate choice of concentrations of the inhibitor. Hexose transport inhibition does not appear to be secondary to other known effects of the amilorides. Inhibition by all analogues is enhanced when they are tested in low (2 mM) Na+ medium, where they show half-maximal inhibition in the range of 100-300 microM. More detailed kinetic analysis of inhibition by EIPA shows it to be competitive with Na+ with a Ki of 73-107 microM. It is concluded that the amilorides are acting directly on the hexose transporter.