Ethanol Prevents Oxidant-Induced Mitochondrial Permeability Transition Pore Opening in Cardiac Cells

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The purpose of this study was to determine if ethanol prevents the mitochondrial permeability transition pore (mPTP) opening via glycogen synthase kinase 3β (GSK-3β).


Cardiac H9c2 cells were exposed to ethanol (10–1000 μM) for 20 min. GSK-3β activity was determined by measuring its phosphorylation at Ser9. Mitochondrial membrane potential (ΔΨm) was assessed by imaging (confocal microscopy) H9c2 cells loaded with tetramethylrhodamine ethyl ester (TMRE). To activate GSK-3β, cells were transfected with constitutively active GSK-3β (GSK-3β-S9A-HA) mutant plasmid.


Treatment of cardiac cells with low doses of ethanol (10–500 μM) significantly enhanced GSK-3β phosphorylation, indicating that ethanol can inactivate GSK-3β in H9c2 cells. The effect of ethanol on GSK-3β activity was reversed by the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 and ethanol could enhance Akt phosphorylation, implying that the PI3K/Akt pathway accounts for the action of ethanol. Ethanol prevented oxidant (H2O2)-induced loss ΔΨm, an effect that was reversed by LY294002, indicating that ethanol can modulate the mPTP opening caused by oxidant stress through the PI3K/Akt pathway. Ethanol failed to preserve ΔΨm in cells transfected with the constitutively active GSK-3β (GSK-3β-S9A-HA) mutant, suggesting that ethanol prevents the mPTP opening by inactivating GSK-3β.


These data suggest that ethanol prevents the mPTP opening through inactivation of GSK-3β. The PI3K/Akt signaling pathway is responsible for inactivation of GSK-3β by ethanol.

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