Role of Na+ in alpha-aminoisobutyric acid uptake by membrane vesicles from mouse fibroblasts transformed by simian virus 40

Abstract

The uptake of alpha-amino[(3)H]isobutyric acid (AIB) was studied in membrane vesicles from mouse fibroblasts transformed by simian virus 40 to examine the features of the Na(+)-stimulated and Na(+)-dependent AIB transport process. The simultaneous addition of NaCl and AIB to these vesicles produced a transient accumulation, or "overshoot," of amino acid 3-4 times the equilibrium value. Both the initial rate of uptake and the rate of fall of intravesicular AIB after maximal accumulation were sensitive to the temperature of incubation. The overshoot of AIB uptake was enhanced with Na(+) salts of highly permeant lipophilic anions, such as SCN(-) and NO(3) (-), and was decreased by the addition of SO(4) (2-), a relatively impermeant ion. Gramicidin D, which enhances the membrane conductance of Na(+) electrogenically, decreased the overshoot, while a potassium diffusion potential, induced by valinomycin (in K(+)-preloaded membrane vesicles), produced a Na(+)-dependent overshoot of AIB uptake. When vesicles were preincubated with both Na(+) and AIB, followed by the generation of an interior negative membrane potential (by the addition of SCN(-)), an overshoot of AIB uptake resulted. However, this did not occur in the absence of Na(+). It is concluded that, apart from its role in the generation of a transmembrane electrochemical potential, Na(+) is essential for the overshoot of AIB uptake.