Rapid mitochondrial repolarization upon reperfusion after cardiac ischemia

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.