Proline transport by synaptosomal membrane vesicles isolated from rat brain: energetics and inhibition by free fatty acids

Abstract

Synaptosomal membrane vesicles were employed to study the energetics of proline transport and the inhibition of proline transport by unsaturated free fatty acids. Active uptake of proline into synaptosomal membrane vesicles requires extravesicular Na+ and is primarily driven by a Na+ gradient created by diluting K+-loading vesicles into Na+-containing buffers. Uptake of proline under these conditions is enhanced up to 2-fold by a valinomycin-induced diffusion potential (interior negative). Proline transport is reduced in the absence of external Cl- or internal K+. Strong (40-90%) inhibition of proline uptake occurs upon collapse of the Na+ gradient by ionophores such as gramicidin D or activation of the action potential Na+ channel by veratridine or Tityus serrulatus venom. Less (15-25%) inhibition is obtained with the proton ionophore carbonyl cyanide m-chlorophenylhydrazone, which also prevents the stimulation of proline uptake by the valinomycin-induced diffusion potential. Unsaturated free fatty acids inhibit proline uptake. This inhibition is greatest for arachidonic acid and was somewhat less for oleic acid. The saturated fatty acids palmitic and stearic have little or no inhibitory capacity. Endogenous unsaturated free fatty acids may exert similar inhibitory effects on the reuptake systems for neuroactive amino acids and thus modulate their action in the CNS.