Use of stable isotopes and gas chromatography-mass spectrometry in the study of different pools of neurotransmitter amino acids in brain slices.
A method was developed for simultaneous determination of endogenous and newly synthesized neurotransmitter amino acids (4-aminobutyric acid, glutamate and aspartate) and glutamine in brain in vitro. Brain slices were incubated in artificial cerebrospinal fluid in the presence of 13C-labeled precursors (glucose, pyruvate or acetate). After the incubation, the slices were homogenized in cold 80% ethanol and the supernatants were evaporated to dryness. The resultant residues were derivatized with N-methyl-N-(tert.-butyldimethylsilyl)trifluoroacetamide and analyzed by capillary gas chromatography-mass spectrometry in the electron-impact mode. N(O)-tert.-Butyldimethylsilyl derivatives of the naturally occurring amino acids, their 13C-enriched counterparts and deuterated internal standards were detected as their [M-57]+ fragments using selected-ion monitoring. The method was shown applicable to studying compartmentation of neurotransmitter amino acids.
Kapetanovic
I M
IM
Preclinical Pharmacology Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892.
Yonekawa
W D
WD
Kupferberg
H J
HJ
D000596:Amino Acids / Q000032:analysis*; D000818:Animals; D002247:Carbon Isotopes; D002623:Chemistry Techniques, Analytical; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006624:Hippocampus / Q000032:analysis*; D008297:Male; D051379:Mice; D018377:Neurotransmitter Agents / Q000032:analysis*; D005680:gamma-Aminobutyric Acid / Q000032:analysis
D000596:Amino Acids; D002247:Carbon Isotopes; D018377:Neurotransmitter Agents; D005680:gamma-Aminobutyric Acid