Abstract
Morphine has been shown to protect the myocardium against ischemia-reperfusion injury through inhibition of glycogen synthase kinase-3β (GSK-3β). Given that GSK-3β is known to modulate the mitochondrial permeability transition pore (mPTP), we investigated the role of mPTP in the cardioprotective effect of morphine and the GSK-3β inhibitor SB216763 [SB; 3-(2,4-dichlorophenyl)-4(1-methyl-1H-indol-3-yl)-1H-pyrrole-2,5-dione] during ischemia-reperfusion. Both morphine (0.3 mg/kg) and SB (0.6 mg/kg) reduced infarct size in a model of regional myocardial ischemia-reperfusion in rats (13 ± 1 and 14 ± 3% of the area at risk versus 33 ± 4% in controls; p < 0.05). Morphine and SB protected the ischemic myocardium against Ca2+-induced mPTP opening as demonstrated by the increased capacity of mitochondria to retain Ca2+ when they were isolated from the ischemic zone 10 min after the onset of reperfusion (59 ± 8 and 66 ± 3 versus 29.5 ± 6 nmol Ca2+/mg · protein, respectively; p < 0.05). This was associated with a restoration of mitochondrial oxidative phosphorylation parameters. In isolated adult rat cardiomyocytes subjected to anoxia-reoxygenation, morphine (2 μM), SB (3 μM), and the direct mPTP inhibitor cyclosporine A (3 μM) delayed mPTP opening as assessed by the calcein loading Co2+-quenching technique. This was accompanied by an increase in cell survival as measured by nuclear staining with propidium iodide. These in vitro effects of morphine on inhibition of mPTP opening during anoxia-reoxygenation were suppressed by the phosphatidylinositol 3-kinase (PI3-kinase) inhibitor wortmannin (0.1 μM). These data indicate that the infarct-limiting effect of morphine and SB is linked by a cause-effect relationship, which leads to an increased mitochondrial resistance and inhibition of mPTP opening through the PI3-kinase pathway and subsequent inactivation of GSK-3β.
Footnotes
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This work was supported by Grant 2004004775 from the Fondation de France.
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F.N.O. was supported by the Ministère de la Recherche et de la Technologie.
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C.P.-M. was supported by the Fondation Lefoulon Delalande-Institut de France.
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R.A. was supported by a doctoral grant from Région Ile-de-France.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.108.138008.
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ABBREVIATIONS: mPTP, mitochondrial permeability transition pore; AAR, area at risk; calcein-AM, calcein acetoxymethyl ester; FCCP, carbonyl cyanide 4-trifluoromethoxyphenylhydrazone; ROS, reactive oxygen species; GSK-3β, glycogen synthase kinase-3β; PI3-kinase, phosphatidyl-inositol 3-kinase; I/R, ischemia-reperfusion; SB216763 (SB), 3-(2,4-dichlorophenyl)-4(1-methyl-1H-indol-3-yl)-1H-pyrrole-2,5-dione; TBAR, thiobarbituric acid-reactive substance; TmPTP, time for mitochondrial permeability transition pore opening.
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↵1 Both authors contributed equally to this work.
- Received February 13, 2008.
- Accepted April 22, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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