Mitochondria play key roles in physiology and in disease. Understanding the mechanisms that define mitochondrial function, the regulation of energy balance in the cell, and the ways in which mitochondrial dysfunction impacts on cell physiology represents a key challenge in modern cellular pathophysiology. This requires a study of mitochondrial function in relation to the physiology of the cell, and so has driven approaches to measure and characterize key aspects of mitochondrial function within living cells. Central to this advance has been the use of fluorescent indicators that report mitochondrial membrane potential, mitochondrial redox state, rates of free radical generation, ATP, and so on. In this chapter we provide a critical appraisal and guide to the measurement of mitochondrial bioenergetics within living cells and discuss both the strengths and potential pitfalls of the most useful probes for mitochondrial membrane potential using confocal microscopy and flow cytometry.
Johnson LV et al (1981) Monitoring of relative mitochondrial membrane potential in living cells by fluorescence microscopy. J Cell Biol 88:526–535PubMedCrossRefGoogle Scholar
Reers M et al (1991) J-aggregate formation of a carbocyanine as a quantitative fluorescent indicator of membrane potential. Biochemistry 30:4480–4486PubMedCrossRefGoogle Scholar
Chinopoulos C et al (1999) Depolarization of in situ mitochondria due to hydrogen peroxide-induced oxidative stress in nerve terminals: inhibition of alpha-ketoglutarate dehydrogenase. J Neurochem 73:220–228PubMedCrossRefGoogle Scholar
Rottenberg H, Wu S (1998) Quantitative assay by flow cytometry of the mitochondrial membrane potential in intact cells. Biochim Biophys Acta 1404:393–404PubMedCrossRefGoogle Scholar
Zamzami N et al (1995) Sequential reduction of mitochondrial transmembrane potential and generation of reactive oxygen species in early programmed cell death. J Exp Med 182:367–377PubMedCrossRefGoogle Scholar