Role of PI3Kα and sarcolemmal ATP-sensitive potassium channels in epoxyeicosatrienoic acid mediated cardioprotection

Aims: Epoxyeicosatrienoic acids (EETs) are cytochrome P450 epoxygenase metabolites of arachidonic acid that have known cardioprotective properties. While the mechanism(s) remains unknown, evidence suggests that phosphoinositide 3-kinase (PI3K) and sarcolemmal ATP-sensitive potassium channels (pmK(ATP)) are important. However the role of specific PI3K isoforms and corresponding intracellular mechanisms remains unknown. Methods and results: To study this, mice hearts were perfused in Langendorff mode for 40 min of baseline and subjected to 20 or 30 min of global no-flow ischemia followed by 40 min of reperfusion. C57BL6 mice perfused with 11,12-EET (1 mu M) had improved postischemic recovery, whereas co-perfusion with PI3K alpha inhibitor, PI-103 (0.1 mu M), abolished the EET-mediated effect. In contrast, blocking of PI3K beta or PI3K gamma isoforms failed to inhibit EET-mediated cardioprotection. In addition to the improved post-ischemic recovery, increased levels of p-Akt, decreased calcineurin activity and decreased translocation of proapoptotic protein BAD to mitochondria were noted in EET-treated hearts. Perfusion of 11,12-EET to Kir6.2 deficient mice (pmK(ATP)) failed to improve postischemic recovery, decrease calcineurin activity and translocation of proapoptotic protein BAD, however increased levels of p-Akt were still observed. Patch-clamp experiments demonstrated that 11,12-EET could not activate pmK(ATP) currents in myocytes pre-treated with PI-103. Mechanistic studies in H9c2 cells demonstrate that 11,12-EET limits anoxia-reoxygenation triggered Ca2+ accumulation and maintains mitochondrial Delta psi m compared to controls. Both PI-103 and glibenclamide (10 mu M, pmK(ATP) inhibitor) abolished EET cytoprotection. Conclusion: Together our data suggest that EET-mediated cardioprotection involves activation of PI3K alpha, upstream of pmK(ATP), which prevents Ca2+ overload and maintains mitochondrial function. (c) 2012 Elsevier Ltd. All rights reserved.