A 13C NMR double-labeling method to quantitate local myocardial O2 consumption using frozen tissue samples

Abstract

Measurement of local myocardial O2 consumption (V˙o 2) has been problematic but is needed to investigate the heterogeneity of aerobic metabolism. The goal of the present investigation was to develop a method to measure local V˙o 2 using small frozen myocardial samples, suitable for determiningV˙o 2 profiles. In 26 isolated rabbit hearts, 1.5 mmol/l [2-13C]acetate was infused for 4 min, followed by 1.5 min of [1,2-13C]acetate. The left ventricular (LV) free wall was then quickly frozen. High-resolution 13C-NMR spectra were measured from extracts taken from 2- to 3-mm thick transmural layer samples. The multiplet intensities of glutamate were analyzed with a computer model allowing simultaneous estimation of the absolute flux through the tricarboxylic acid cycle and the fractional contribution of acetate to acetyl CoA formation from which localV˙o 2 was calculated. The 13C-derivedV˙o 2 in the LV free wall was linearly related to “gold standard”V˙o 2 from coronary venous O2 electrode measurements in the same region ( r = 0.932, n = 22, P < 0.0001, slope 1.05) for control and lowered metabolic rates. The ratio of subendocardial to subepicardial V˙o 2 was 1.52 ± 0.19 (SE, significantly >1, P< 0.025). Local myocardialV˙o 2 can now be quantitated with this new 13C method to determine profiles of aerobic energy metabolism.