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Metabolism of [U‐ 13 C 5 ]Glutamine in Cultured Astrocytes Studied by NMR Spectroscopy: First Evidence of Astrocytic Pyruvate Recycling
Author(s) -
Sonnewald U.,
Westergaard N.,
Jones P.,
Taylor A.,
Bachelard H. S.,
Schousboe A.
Publication year - 1996
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1996.67062566.x
Subject(s) - glutamine , citric acid cycle , tricarboxylic acid , glutamate receptor , biochemistry , metabolism , alanine , pyruvate carboxylase , astrocyte , glutamic acid , chemistry , amino acid , biology , enzyme , receptor , central nervous system , neuroscience
Metabolism of [U‐ 13 C 5 ]glutamine was studied in primary cultures of cerebral cortical astrocytes in the presence or absence of extracellular glutamate. Perchloric acid extracts of the cells as well as redissolved lyophilized media were subjected to nuclear magnetic resonance and mass spectrometry to identify 13 C‐labeled metabolites. Label from glutamine was found in glutamate and to a lesser extent in lactate and alanine. In the presence of unlabeled glutamate, label was also observed in aspartate. It could be clearly demonstrated that some [U‐ 13 C 5 ]glutamine is metabolized through the tricarboxylic acid cycle, although to a much smaller extent than previously shown for [U‐ 13 C 5 ]glutamate. Lactate formation from tricarboxylic acid cycle intermediates has previously been demonstrated. It has, however, not been demonstrated that pyruvate, formed from glutamate or glutamine, may reenter the tricarboxylic acid cycle after conversion to acetyl‐CoA. The present work demonstrates that this pathway is active, because [4,5‐ 13 C 2 ]glutamate was observed in astrocytes incubated with [U‐ 13 C 5 ]glutamine in the additional presence of unlabeled glutamate. Furthermore, using mass spectrometry, mono‐labeled alanine, glutamate, and glutamine were detected. This isotopomer could be derived via the action of pyruvate carboxylase using 13 CO 2 produced within the mitochondria or from labeled intermediates that had stayed in the tricarboxylic acid cycle for more than one turn.