Locations of amino acids in brain slices from the rat. Tetrodotoxin-sensitive release of amino acids

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Abstract
1. Amino acids, particularly glutamate, γ-aminobutyrate, aspartate and glycine, were released from rat brain slices on incubation with protoveratrine (especially in a Ca2+-deficient medium) or with ouabain or in the absence of glucose. Release was partially or wholly suppressed by tetrodotoxin. 2. Tetrodotoxin did not affect the release of glutamine under various incubation conditions, nor did protoveratrine accelerate it. 3. Protoveratrine caused an increased rate of formation of glutamine in incubated brain slices. 4. Increased K+ in the incubation medium caused release of γ-aminobutyrate, the process being partly suppressed by tetrodotoxin. 5. Incubation of brain slices in a glucose-free medium led to increased production of aspartate and to diminished tissue contents of glutamates, glutamine and glycine. 6. Use of tetrodotoxin to suppress the release of amino acids from neurons in slices caused by the joint action of protoveratrine and ouabain (the latter being added to diminish reuptake of amino acids), it was shown that the major pools of glutamate, aspartate, glycine, serine and probably γ-aminobutyrate are in the neurons. 7. The major pool of glutamine lies not in the neurons but in the glia. 8. The tricarboxylic cycle inhibitors, fluoroacetate and malonate, exerted different effects on amino acid contents in, and on amino acid release from, brain slices incubated in the presence of protoveratrine. Fluoroacetate (3mm) diminished the content of glutamine, increased that of glutamate and γ-aminobutyrate and did not affect respiration. Malonate (2mm) diminished aspartate and γ-aminobutyrate content, suppressed respiration and did not affect glutamine content. It is suggested that malonate acts mainly on the neurons, and that fluoroacetate acts mainly on the glia, at the concentrations quoted. 9. Glutamine was more effective than glutamate as a precursor of γ-aminobutyrate. 10. It is suggested that glutamate released from neurons is partly taken up by glia and converted there into glutamine. This is returned to the neurons where it is hydrolysed and converted into glutamate and γ-aminobutyrate.