Abstract
The mitochondrial membrane potential (ΔΨ(m)) drives oxidative phosphorylation and alterations contribute to cardiac pathologies, but real-time assessment of ΔΨ(m) has not been possible. Here we describe noninvasive measurements using mitochondrial heme b(L) and b(H) absorbances, which rapidly respond to ΔΨ(m). Multi-wavelength absorbance spectroscopy enabled their continuous monitoring in isolated mitochondria and the perfused heart. Calibration of heme b absorbance in isolated mitochondria revealed that reduced heme b(L) relative to total reduced heme b (fb(L) = b(L)/(b(L) + b(H))) exhibits a sigmoidal relationship with ΔΨ(m). Extrapolating this relationship to the heart enabled estimation of ΔΨ(m) as 166 ± 18 mV (n = 25, mean ± s.d.). We used this approach to assess how ΔΨ(m) changes during ischemia-reperfusion injury, an unknown limiting the understanding of ischemia-reperfusion injury. In perfused hearts, ΔΨ(m) declined during ischemia and rapidly reestablished upon reperfusion, supported by oxidation of the succinate accumulated during ischemia. These findings expand our understanding of ischemia-reperfusion injury.