Improved estimation of anaplerosis in heart using 13c-nmr.
Cohen, David M., and Richard N. Bergman.
Metabolic Research Unit, Department of Physiology and Biophysics,
University of Southern California School of Medicine, Los Angeles, CA
90033. Laboratory of Cerebral Metabolism, Bulding 36, Room 1A-07,
National Institutes of Health, Bethesda, MD 20892.
APStracts 4:0204E, 1997.
Anaplerotic enzymes (such as pyruvate carboxylase or malic enzyme)
catalyze reactions that fill up the pools of the citric acid cycle
(CAC), thereby increasing the total mass of CAC intermediates.
Relative anaplerosis (y) denotes the ratio of anaplerotic flux to the
flux catalyzed by citrate synthase. We examine conventional methods
(16, 17) of measurement of y using 13C-labeled precursors and
analysis of 13C-glutamate labeling by nuclear magnetic resonance
spectroscopy. Through mathematical analysis and computer simulation,
we show that isotopic enrichment of the pool of pyruvate that is
substrate for anaplerosis will severely decrease the accuracy of
estimates of y using conventional methods (16, 17), no matter how
small the mass of the pool of pyruvate. Suppose that the recycling
parameter R denotes the fraction of molecules of pyruvate which
contain carbons derived from intermediates of the CAC. Each means of
estimation of relative anaplerosis in the peer-reviewed literature
assumes that R=0, although this assumption has not been confirmed by
experiment. We show that conventional formulas (either using
fractional enrichments of carbons or isotopomer analysis) (16, 17)
actually estimate at most y*(1(R) instead of y, during administration
of [2-13C]acetate and unlabeled pyruvate. Using a new formula for
estimation of y, we recalculate values of y from the literature, and
find them approximately 50% too low. We assume that all anaplerosis
is via pyruvate and that the difference in isotopic enrichment
between cytosolic and mitochondrial malate is negligible.
Received 2 April1 997; accepted in final form 16 September 1997.
APS Manuscript Number E151-7.
Article publication pending Am. J. Physiol. (Endocrinol. Metab.).
ISSN 1080-4757 Copyright 1997 The American Physiological Society.
Published in APStracts on 7 October 1997