The mitochondrial permeability transition, release of cytochrome c and cell death. Correlation with the duration of pore openings in situ.
We investigated the relationship between opening of the permeability transition pore (PTP), mitochondrial depolarization, cytochrome c release, and occurrence of cell death in rat hepatoma MH1C1 cells. Treatment with arachidonic acid or induces PTP opening in situ with similar kinetics, as assessed by the calcein loading-Co(2+) quenching technique (Petronilli, V., Miotto, G., Canton, M., Colonna, R., Bernardi, P., and Di Lisa, F. (1999) Biophys. J. 76, 725-734). Yet depolarization, as assessed from the changes of mitochondrial tetramethylrhodamine methyl ester (TMRM) fluorescence, is rapid and extensive with arachidonic acid and slow and partial with. Cyclosporin A-inhibitable release of cytochrome c and cell death correlate with the changes of TMRM fluorescence but not with those of calcein fluorescence. Since pore opening must be accompanied by depolarization, we conclude that short PTP openings are detected only by trapped calcein and may have little impact on cell viability, while changes of TMRM distribution require longer PTP openings, which cause release of cytochrome c and may result in cell death. Modulation of the open time appears to be the key element in determining the outcome of stimuli that converge on the PTP.
Petronilli
V
V
Consiglio Nazionale delle Ricerche Unit for the Study of Biomembranes at the Departments of Biomedical Sciences and Biological Chemistry, University of Padova, Viale Giuseppe Colombo 3, I-35100 Padova, Italy. petro@ci.bio.unipd.it
Penzo
D
D
Scorrano
L
L
Bernardi
P
P
Di Lisa
F
F
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D016718:Arachidonic Acid / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects*; D003574:Cytochrome c Group / Q000378:metabolism*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology*; D051381:Rats; D014407:Tumor Cells, Cultured
D003574:Cytochrome c Group; D016718:Arachidonic Acid; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D000001:Calcimycin