Imbalance between energy supply and demand has been proposed as a significant contributor to progressive heart failure. To gain insight into this hypothesis, we designed experiments to establish a relationship between mitochondrial and contractile activity in intact cardiac myocytes and via mitochondrial respiration in isolated organelle preparations. We are testing a method for real-time, simultaneous monitoring of contractile and mitochondrial activities in isolated cardiac myocytes, using tetramethylrhodamine methyl ester (TMRM) fluorescence measurements of mitochondrial membrane potential (ψ mito ). Porcine left ventricular myocytes were placed in a perfusion chamber in an IonOptix MultiCell contractility system, continuously superfused with PSS + 2 mM CaCl 2 + TMRM and subjected to electrical field stimulation. Contractions appeared matched with increases in TMRM signal. Studies in mouse cardiomyocytes dual-loaded with TMRM and calcein (a fluorescent Ca 2+ indicator) sought to correct for motion artifact by rationing TMRM/calcein signals; treatment with a mitochondrial uncoupler, FCCP, significantly decreased TMRM signal, suggesting that differences in calcein-normalized TMRM fluorescence tracked changes in ψ mito . In separate studies, we examined respiratory activity in isolated cardiac mitochondria from young adult (2–5 months) wildtype (WT) and cMyBP-C knockout (KO) mice, using polarographic methods. Mitochondria from both groups showed no significant differences in O 2 consumption in response to either glutamate/malate or succinate and coupling index was similar between groups. Preliminary experiments in older mice (9–10 months) suggest higher state 4 and 3 respiration and a higher respiratory control ratio in WT vs. KO mice during glutamate/malate-driven respiration. These latter results suggest that lack of cMyBP-C elicits an age-dependent mitochondrial dysfunction, which may promote inefficient matching of bioenergetic supply with contractility. Future experiments will seek to validate a real-time method to quantify ψ mito coincident with contractility and further test our hypothesis of bioenergetic mismatch in isolated cMyBP-C KO cardiac myocytes.
Kalogeris et al. (Sun,) studied this question.