Two pharmacologically distinct sodium- and chloride-coupled high-affinity γ-aminobutyric acid transporters are present in plasma membrane vesicles and reconstituted preparations from rat brain

Electrogenic sodium- and chloride-dependent γ-aminobutyric acid (GABA) transport in crude synaptosomal membrane vesicles is partly inhibited by saturating levels of either of the substrate analogues cis-3-aminocyclohexanecarboxylic acid (ACHC) or β-alanine. However, both of them together potently and fully inhibit the process. Transport of β-alanine, which exhibits an apparent K_m of about 44 μM, is also electrogenic and sodium and chloride dependent and competitively inhibited by GABA with a K_i of about 3 μM. This value is very similar to the K_m of 2-4 μM found for GABA transport. On the other hand, ACHC does not inhibit β-alanine transport at all. Upon solubilization of the membrane proteins with cholate and fractionation with ammonium sulfate, a fraction is obtained which upon reconstitution into proteoliposomes exhibits 4- to 10-fold-increased GABA transport. This activity is fully inhibited by low concentrations of ACHC and is not sensitive at all to β-alanine. GABA transport in this preparation exhibits an apparent Km of about 2.5 μM and it is competitively inhibited by ACHC (K_i ≈ 7 μM). These data indicate the presence of two GABA transporter subtypes in the membrane vesicles: the A type, sensitive to ACHC, and the B type, sensitive to β-alanine.