FUNCTIONAL-HETEROGENEITY OF AN ISOLATED MITOCHONDRIAL POPULATION REVEALED BY CYTOFLUOROMETRIC ANALYSIS AT THE SINGLE ORGANELLE LEVEL

Isolated rat Liver mitochondria were incubated under various metabolic conditions to determine their membrane potential (MMP) as measured co ntinuously by a tetraphenylphosphonium (TPP+)-selective electrode, By how cytometry, a parallel analysis of fluorescence emissions observing single mitochondria stained with the lipophilic cation hloro-1,1',3,3 '-tetraethylbenzimidazolcarbocyanine iodide (JC-l) revealed linear cor relation between the median orange fluorescence (FL2) due to J-aggrega te formations and MMP values measured by TPP+. No correlation was dete cted with the green fluorescence (FL1) emission. A significantly highe r correlation appeared between the FL2/FL1 ratio and MMP values. Withi n the same mitochondrial population, cytofluorimetric analysis reveale d the presence of various classes of organelles with different MMP, wh ose distribution was dependent on metabolic condition. The highest fun ctional heterogeneity was found in deenergized mitochondria, while the highest homogeneity was observed during the first phase of the phosph orylative process. Thus, these data suggest that the cytofluorimetric use of JC-l provides direct experimental evidence for the hypothesis o f functional mitochondrial heterogeneity, at least with respect to the ir membrane potential. (C) 1996 Academic Press, Inc.