Effects of L-Glutamate Transport Inhibition by a Conformationally Restricted Glutamate Analogue (2S,1'S,2'R)-2-(Carboxycyclopropyl)Glycine (L-CCG III) on Metabolism in Brain Tissue In Vitro Analysed by NMR Spectroscopy |
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Authors: | Moussa Charbel El-Hajj Mitrovic Ann D Vandenberg Robert J Provis Tanya Rae Caroline Bubb William A Balcar Vladimir J |
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Institution: | (1) Department of Anatomy and Histology, The University of Sydney, Australia;(2) Department of Pharmacology, The University of Sydney, Australia;(3) Department of Biochemistry, The University of Sydney, Australia;(4) Institute for Biomedical Research, The University of Sydney, Australia;(5) Department of Molecular Pharmacology, Faculty of Pharmaceutical Sciences, Kanazawa University, Ishikawa, Japan |
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Abstract: | (2S,1'S,2'R)-2-(Carboxycyclopropyl)glycine (L-CCG III) was a substrate of Na+-dependent glutamate transporters (GluT) in Xenopus laevis oocytes (IC50 13 and 2 M for, respectively, EAAT 1 and EAAT 2) and caused an apparent inhibition of 3H]L-glutamate uptake in mini-slices of guinea pig cerebral cortex (IC50 12 M). In slices (350 M) of guinea pig cerebral cortex, 5 M L-CCG III increased both the flux of label through pyruvate carboxylase and the fractional enrichment of glutamate, GABA, glutamine and lactate, but had no effect on total metabolite pool sizes. At 50 M L-CCG III decreased incorporation of 13C from 3-13C]-pyruvate into glutamate C4, glutamine C4, lactate C3 and alanine C3. The total metabolite pool sizes were also decreased with no change in the fractional enrichment. Furthermore, L-CCG III was accumulated in the tissue, probably via GluT. At lower concentration, L-CCG III would compete with L-glutamate for GluT and the changes probably reflect a compensation for the missing L-glutamate. At 50 M, intracellular L-CCG III could reach > 10 mM and metabolism might be affected directly. |
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Keywords: | NMR spectroscopy guinea pig brain GLAST GLT EAAT 1 EAAT 2 toxic amino acids |
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