FCCP is cardioprotective at concentrations that cause mitochondrial oxidation without detectable depolarisation.
OBJECTIVE
The role of mitochondria and in particular of mitochondrial uncoupling in the mechanism of cardioprotection is not defined. In the accompanying paper we have shown that pretreatment of isolated rat hearts with a low concentration (100 nM) of FCCP, prior to global ischaemia, is cardioprotective, while 300 nM FCCP exacerbates injury. Here we define the mitochondrial responses to increasing concentrations of FCCP and also to explore the equivalence of the cardioprotective doses of diazoxide.


METHODS
Changes in mitochondrial respiration in response to FCCP and diazoxide were determined in isolated rat ventricular myocytes. In addition, mitochondrial state was monitored using confocal microscopy to record mitochondrial potential (TMRM) and redox state (NADH) during FCCP and diazoxide treatment. Myocytes were also voltage-clamped and whole cell currents recorded in response to 100 nM FCCP.


RESULTS
FCCP (10-1000 nM) caused significant dose-dependent increase in oxygen consumption. Diazoxide (30 microM) failed to cause any measurable change in mitochondrial function. FCCP at 100 nM caused mitochondrial oxidation, but no change in mitochondrial membrane potential or (sarc)K(ATP) channel current, while at 300 nM, FCCP caused significant mitochondrial depolarisation. Diazoxide failed to induce any mitochondrial oxidation or depolarisation.


CONCLUSIONS
Concentrations of FCCP that cause mitochondrial oxidation without depolarisation are cardioprotective. Higher FCCP concentrations dissipate mitochondrial membrane potential and exacerbate injury. This establishes the principle that mild mitochondrial uncoupling activates a protective mechanism. Diazoxide did not cause mitochondrial oxidation or mitochondrial depolarisation, suggesting it induces protection via another mechanism.
Brennan
Jonathan P
JP
Cardiac Physiology (Cardiovascular Division), King's College London, SE1 7EH, UK.
Berry
Roger G
RG
Baghai
Max
M
Duchen
Michael R
MR
Shattock
Michael J
MJ
D000818:Animals; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology*; D002478:Cells, Cultured; D003981:Diazoxide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D018613:Microscopy, Confocal; D008929:Mitochondria, Heart / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D051381:Rats; D017208:Rats, Wistar; D014475:Uncoupling Agents / Q000494:pharmacology*; D014665:Vasodilator Agents / Q000494:pharmacology
D014475:Uncoupling Agents; D014665:Vasodilator Agents; D009243:NAD; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D003981:Diazoxide