Document Detail

Probing the origin of acetyl-CoA and oxaloacetate entering the citric acid cycle from the 13C labeling of citrate released by perfused rat hearts.
MedLine Citation:
PMID:  9334176     Owner:  NLM     Status:  MEDLINE    
We present a strategy for simultaneous assessment of the relative contributions of anaplerotic pyruvate carboxylation, pyruvate decarboxylation, and fatty acid oxidation to citrate formation in the perfused rat heart. This requires perfusing with a mix of 13C-substrates and determining the 13C labeling pattern of a single metabolite, citrate, by gas chromatography-mass spectrometry. The mass isotopomer distributions of the oxaloacetate and acetyl moieties of citrate allow calculation of the flux ratios: (pyruvate carboxylation)/(pyruvate decarboxylation), (pyruvate carboxylation)/(citrate synthesis), (pyruvate decarboxylation)/(citrate synthesis) (pyruvate carboxylation)/(fatty acid oxidation), and (pyruvate decarboxylation)/(fatty acid oxidation). Calculations, based on precursor-product relationship, are independent of pool size. The utility of our method was demonstrated for hearts perfused under normoxia with [U-13C3](lactate + pyruvate) and [1-13C]octanoate under steady-state conditions. Under these conditions, effluent and tissue citrate were similarly enriched in all 13C mass isotopomers. The use of effluent citrate instead of tissue citrate allows probing substrate fluxes through the various reactions non-invasively in the intact heart. The methodology should also be applicable to hearts perfused with other 13C-substrates, such as 1-13C-labeled long chain fatty acid, and under various conditions, provided that assumptions on which equations are developed are valid.
B Comte; G Vincent; B Bouchard; C Des Rosiers
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  272     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  1997 Oct 
Date Detail:
Created Date:  1997-11-17     Completed Date:  1997-11-17     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  26117-24     Citation Subset:  IM    
Department of Nutrition, University of Montréal, Montréal, Québec H3C 3J7, Canada.
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MeSH Terms
Acetyl Coenzyme A / metabolism*
Carbon Isotopes
Citrates / metabolism*
Citric Acid Cycle*
Myocardium / metabolism*
Oxaloacetates / metabolism*
Oxaloacetic Acids
Rats, Sprague-Dawley
Reg. No./Substance:
0/Carbon Isotopes; 0/Citrates; 0/Oxaloacetates; 0/Oxaloacetic Acids; 72-89-9/Acetyl Coenzyme A

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