From identification of fluorescent flavoproteins to mitochondrial redox indicators in intact tissues

Abstract

Development of the use of flavin and nicotinamide-adenine nucleotide fluorescence in monitoring the redox state of the free mitochondrial NADH/NAD+ couple in cells, tissues and organs is reviewed. A break-through was the identification of dihydrolipoamide dehydrogenase (FpL) as the major NAD-linked fluorescent flavoprotein of mitochondria. This mitochondrial matrix flavoprotein is in equilibrium with the free NADH/NAD+ pool and its mid-potential is sufficiently near to that of NADH/NAD+ so that its percentage reduction follows that of the latter. Possibilities of monitoring mitochondrial and cytosolic NADH depend on the population density of mitochondria and thus are tissue-dependent. Upon a shift toward reduction, fluorescence intensities of NADH and flavins swing to reciprocal directions, so that the NADH/flavin fluorescence ratio can be used to increase the sensitivity of redox monitoring. This method is attaining widening use in studies on metabolic regulation under normal and pathological conditions.