Mitochondrial Nadh in The Langendorff Rat Heart Decreases in Response to Increases in Work

Abstract

The mechanism that tunes the rate of respiration to a changing ATP demand during alterations in work of the intact myocardium has not been elucidated (3, 4, 5, 12). The phosphate potential (ATP/ADP ratio) and the substrate oxidation potential (e.g. redox potential difference of the NADH/NAD⁺ couple and 0₂/H₂0 couple) have been proposed as potential regulatory factors of oxidative phosphorylation. Although studies of isolated mitochondria have demonstrated that the rate of respiration can be controlled by changes in the concentration of ADP and P¡ (7), in intact myocardium no significant change in the phosphorylation potential has been found (3,4). Whereas elevation of the NADH concentration can increase the maximum rate of oxidative phosphorylation and stimulate ATP synthesis in isolated mitochondria (18), conflicting data on the NADH redox state during increase of work in intact myocardium have been found. In isolated rabbit heart no change was found in the NAD(P)H redox state during changes in afterload and heart rate (12). An increase of the mitochondrial NADH/NAD⁺ ratio was found in response to increased heart rate in the rat heart (16). In contrast, a decrease of the intramitochondrial NADH/NAD⁺ ratio was reported for isolated papillary rabbit muscle in response to twitches (9). Methodological differences could explain these contradictory results. Measurement of the intracellular autofluorescence of NADH (365 nm excitation, 470 nm emission), allows qualitative evaluation of the mitochondrial NADH/NAD⁺ redox state since NADH fluoresces when excited with ultraviolet light, and NAD⁺ does not (6). The NADH fluorescence measurement can be disturbed by ischemia, oximetric effect, endogenous substrate depletion, movement and photo-bleaching (1,2, 11, 12, 14, 15, 16).