Glutamate, aspartate, and gamma-aminobutyrate transport by membrane vesicles prepared from rat brain.

To prepare membrane vesicles, nerve terminal preparations (synaptosomes) isolated from rat cerebral cortex were first subjected to hypotonic lysis. After collecting the membranes contained in this fraction by centrifugation, membrane vesicles were then reconstituted during incubation in a potassium salt solution at 37°C. The transport of glutamate, aspartate, or y-aminobutyrie acid (GABA) was measured by transferring vesicles to 10 vol of 0.1 M NaCl solution containing the radioactive substrate. Transport was temperature dependent and exhibited saturation kinetics with an apparent K, of 2.5 brM. The rates and extent of L-glutamate and L-aspartate uptake were equivalent and were greater than those for GABA. Valinomycin increased the rate of uptake of each of these substances suggesting a role for an electrogenic component in transport. Consonant with this notion, external K+ and Rb+ decreased uptake of all three compounds. External thiocyanate also increases the rate of glutamate, aspartate, and GABA transport. Uptake of these neuroactive amino acids was absolutely dependent on external Na+; no other monovalent cation tested substitutes for it. Gramicidin D and nigericin inhibit glutamate transport by abolishing both the Na+ and Kf gradients. Monensin inhibits uptake by selectively dissipating the Na+ gradient. For both glutamate and GABA transport, the Na+ and K+ gradients are synergistic and not additive.