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.
(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 to approximately 13 and to approximately 2 microM for, respec tively, EAAT 1 and EAAT 2) and caused an apparent inhibition of [3H]L-glutamate uptake in "mini-slices" of guinea pig cerebral cortex (IC50 to approximately 12 microM). In slices (350 microM) of guinea pig cerebral cortex, 5 microM 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 microM 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 microM, intracellular L-CCG III could reach > 10 mM and metabolism might be affected directly.
Moussa
Charbel El-Hajj
Cel-H
Department of Anatomy and Histology, The University of Sydney, Australia.
Mitrovic
Ann D
AD
Vandenberg
Robert J
RJ
Provis
Tanya
T
Rae
Caroline
C
Bubb
William A
WA
Balcar
Vladimir J
VJ
D027322:Amino Acid Transport System X-AG / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000600:Amino Acids, Dicarboxylic / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology*; D000818:Animals; D002247:Carbon Isotopes; D002540:Cerebral Cortex / Q000187:drug effects* / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D027341:Excitatory Amino Acid Transporter 1 / Q000378:metabolism; D027342:Excitatory Amino Acid Transporter 2 / Q000378:metabolism; D018698:Glutamic Acid / Q000187:drug effects / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D006168:Guinea Pigs; D066298:In Vitro Techniques; D019344:Lactic Acid / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008968:Molecular Conformation; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D011766:Pyruvate Carboxylase / Q000378:metabolism; D019289:Pyruvic Acid / Q000378:metabolism; D014982:Xenopus laevis; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D027322:Amino Acid Transport System X-AG; D000600:Amino Acids, Dicarboxylic; D002247:Carbon Isotopes; D027341:Excitatory Amino Acid Transporter 1; D027342:Excitatory Amino Acid Transporter 2; D005973:Glutamine; C035261:(alpha-carboxycyclopropyl)glycine; D019344:Lactic Acid; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; D019289:Pyruvic Acid; D011766:Pyruvate Carboxylase