Determination of malate‐aspartate shuttle activity in intact hearts using 13C NMR spectroscopy and modeling

Abstract

The malate‐aspartate (M‐A) shuttle, which transports cytosolic NADH into mitochondria, provides an important mechanism to regulate metabolic activity in these two compartments. However, direct measurement of M‐A shuttle flux has not been possible. In this study, we developed a novel approach that combines dynamic 13C NMR spectroscopy with biology detailed model of cardiac metabolism to directly quantify M‐A shuttle flux from experimental data. Dynamic 13C NMR spectra were acquired from isolated rat hearts perfused with either short chain (2.5mM [2‐13C]acetate) or long chain (0.32mM [U‐13C]palmitate) fatty acids (FA). A 3rd group of hearts were perfused with 2.5mM [2‐13C]acetate plus unlabeled glucose (5.0mM) and lactate (1.0mM) to induce high cytosolic redox state (HRX). TCA cycle flux (VTCA) and M‐A shuttle activity (VM‐A) were determined by least‐square fitting of a novel multi‐domain kinetic model of cardiac metabolism to dynamic 13C NMR spectra. Our results showed that, regardless of chain length, both VTCA and VM‐A were unchanged in FA only groups. At HRX, VTCA was unaltered compared to the other two groups, however, VM‐A was increased by 40% (P<0.05). Therefore, our proposed method provides the opportunity to directly quantify metabolic communication between subcellular compartments via M‐A shuttle under various pathophysiological conditions. This work was supported by R01HL073315 & R01HL086935.