TCA cycle kinetics in the rat heart by analysis of C-13 isotopomers using indirect H-1[C-13] detection

This study was designed to test the hypothesis that indirect H-1 [C-13] det ection of tricarboxylic acid (TCA) cycle intermediates using heteronuclear multiple quantum correlation-total correlation spectroscopy (HMQC-TOCSY) nu clear magnetic resonance (NMR) spectroscopy provides additional C-13 isotop omer information that better describes the kinetic exchanges that occur bet ween intracellular compartments than direct C-13 NMR detection. NMR data we re collected on extracts of rat hearts perfused at various times with combi nations of [2-C-13]acetate, propionate, the transaminase inhibitor aminooxy acetate, and 13C multiplet areas derived from spectra of tissue glutamate w ere fit to a standard kinetic model of the TCA cycle. Although the two NMR methods detect different populations of 13C isotopomers, similar values wer e found for TCA cycle and exchange fluxes by analyzing the two data sets. P erfusion of hearts with unlabeled propionate in addition to [2-C-13]acetate resulted in an increase in the pool size of all four-carbon TCA cycle inte rmediates. This allowed the addition of isotopomer data from aspartate and malate in addition to the more abundant glutamate. This study illustrates t hat metabolic inhibitors can provide new insights into metabolic transport processes in intact tissues.