Transient mitochondrial permeability transition pore opening mediates preconditioning-induced protection.
BACKGROUND
Transient (low-conductance) opening of the mitochondrial permeability transition pore (mPTP) may limit mitochondrial calcium load and mediate mitochondrial reactive oxygen species (ROS) signaling. We hypothesize that transient mPTP opening and ROS mediate the protection associated with myocardial preconditioning and mitochondrial uncoupling.


RESULTS
Isolated perfused rat hearts were subjected to 35 minutes of ischemia/120 minutes of reperfusion, and the infarct-risk-volume ratio was determined by tetrazolium staining. Inhibiting mPTP opening during the preconditioning phase with cyclosporine-A (CsA, 0.2 micromol/L) or sanglifehrin-A (SfA, 1.0 micromol/L) abolished the protection associated with ischemic preconditioning (IPC) (20.2+/-3.6% versus 45.9+/-2.5% with CsA, 49.0+/-7.1% with SfA; P<0.001); and pharmacological preconditioning with diazoxide (Dzx, 30 micromol/L) (22.1+/-2.7% versus 46.3+/-3.0% with CsA, 48.4+/-5.5% with SfA; P<0.001), CCPA (the adenosine A1-receptor agonist, 200 nmol/L) (24.9+/-4.5% versus 54.4+/-6.6% with CsA, 42.6+/-9.0% with SfA; P<0.001), or 2,4-dinitrophenol (DNP, the mitochondrial uncoupler, 50 micromol/L) (15.7+/-2.7% versus 40.8+/-5.5% with CsA, 34.3+/-3.1% with SfA; P<0.001), suggesting that mPTP opening during the preconditioning phase is required to mediate protection in these settings. Inhibiting ROS during the preconditioning protocols with N-mercaptopropionylglycine (MPG, 1 mmol/L) also abolished the protection associated with IPC (20.2+/-3.6% versus 47.1+/-3.8% with MPG; P<0.001), diazoxide (22.1+/-2.7% versus 56.3+/-3.8% with MPG; P<0.001), and DNP (15.7+/-2.7% versus 50.7+/-6.6% with MPG; P<0.001) but not CCPA (24.9+/-4.5% versus 26.5+/-8.4% with MPG; P=NS). Further experiments in adult rat myocytes demonstrated that diazoxide induced CsA-sensitive, low-conductance transient mPTP opening (represented by a 28+/-3% reduction in mitochondrial calcein fluorescence compared with control; P<0.01).


CONCLUSIONS
We report that the protection associated with IPC, diazoxide, and mitochondrial uncoupling requires transient mPTP opening and ROS.
Hausenloy
Derek
D
The Hatter Institute and Centre for Cardiology, University College London, UK.
Wynne
Abigail
A
Duchen
Michael
M
Yellon
Derek
D
D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D058646:Adrenergic alpha-1 Receptor Agonists; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D002316:Cardiotonic Agents / Q000494:pharmacology; D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology; D003981:Diazoxide / Q000494:pharmacology; D004553:Electric Conductivity; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D017136:Ion Transport / Q000187:drug effects / Q000502:physiology; D019157:Ischemic Preconditioning, Myocardial; D007783:Lactones / Q000494:pharmacology; D046911:Macromolecular Substances; D008297:Male; D008929:Mitochondria, Heart / Q000502:physiology*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017202:Myocardial Ischemia / Q000503:physiopathology*; D015428:Myocardial Reperfusion Injury / Q000503:physiopathology / Q000517:prevention & control*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology; D010085:Oxidative Phosphorylation / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013141:Spiro Compounds / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology
D058646:Adrenergic alpha-1 Receptor Agonists; D000316:Adrenergic alpha-Agonists; D002316:Cardiotonic Agents; D007473:Ion Channels; D007783:Lactones; D046911:Macromolecular Substances; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species; D013141:Spiro Compounds; D014475:Uncoupling Agents; D014665:Vasodilator Agents; C121329:sanglifehrin A; C058394:2-chloro-N(6)cyclopentyladenosine; D016572:Cyclosporine; D000241:Adenosine; D003981:Diazoxide; D019297:2,4-Dinitrophenol