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Characterization of depolarization and repolarization phases of mitochondrial membrane potential fluctuations induced by tetramethylrhodamine methyl ester photoactivation
Author(s) -
Falchi Angela M.,
Isola Raffaella,
Diana Andrea,
Putzolu Martina,
Diaz Giacomo
Publication year - 2005
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2005.04586.x
Subject(s) - oligomycin , mitochondrial permeability transition pore , depolarization , membrane potential , mitochondrion , iodoacetic acid , biophysics , biochemistry , atp synthase , chemistry , nad+ kinase , rotenone , atp–adp translocase , biology , inner mitochondrial membrane , enzyme , atpase , programmed cell death , apoptosis
Depolarization and repolarization phases (D and R phases, respectively) of mitochondrial potential fluctuations induced by photoactivation of the fluorescent probe tetramethylrhodamine methyl ester (TMRM) were analyzed separately and investigated using specific inhibitors and substrates. The frequency of R phases was significantly inhibited by oligomycin and aurovertin (mitochondrial ATP synthase inhibitors), rotenone (mitochondrial complex I inhibitor) and iodoacetic acid (inhibitor of the glycolytic enzyme glyceraldehyde‐3‐phosphate dehydrogenase). Succinic acid (mitochondrial complex II substrate, given in the permeable form of dimethyl ester) abolished the rotenone‐induced inhibition of R phases. Taken together, these findings indicate that the activity of both respiratory chain and ATP synthase were required for the recovery of the mitochondrial potential. The frequency of D phases prevailed over that of R phases in all experimental conditions, resulting in a progressive depolarization of mitochondria accompanied by NAD(P)H oxidation and Ca 2+ influx. D phases were not blocked by cyclosporin A (inhibitor of the permeability transition pore) or o ‐phenyl‐EGTA (a Ca 2+ chelator), suggesting that the permeability transition pore was not involved in mitochondrial potential fluctuations.