These results suggest that PI3K regulates the opening of the mitochondrial permeability transition pore in rat hearts reperfused with low pressure or postconditioning.

In conclusion, we showed that prolonged reperfusion beyond 60 min was not useful for function assessment and did not change infarct size measurement, on Langendorff rat model of ischemia-reperfusion.

We hypothesized that low-pressure reperfusion may limit myocardial necrosis and attenuate postischemic contractile dysfunction by inhibiting mitochondrial permeability transition pore (mPTP) opening. Male Wistar rat hearts (n = 36) were perfused according to the Langendorff technique, exposed to 40 min of ischemia, and assigned to one of the following groups: 1) reperfusion with normal pressure (NP = 100 cmH(2)O) or 2) reperfusion with low pressure (LP = 70 cmH(2)O). Creatine kinase release and tetraphenyltetrazolium chloride staining were used to evaluate infarct size. Modifications of cardiac function were assessed by changes in coronary flow, heart rate (HR), left ventricular developed pressure (LVDP), the first derivate of the pressure curve (dP/dt), and the rate-pressure product (RPP = LVDP x HR). Mitochondria were isolated from the reperfused myocardium, and the Ca(2+)-induced mPTP opening was measured using a potentiometric approach. Lipid peroxidation was assessed by measuring malondialdehyde production. Infarct size was significantly reduced in the LP group, averaging 17 +/- 3 vs. 33 +/- 3% of the left ventricular weight in NP hearts. At the end of reperfusion, functional recovery was significantly improved in LP hearts, with RPP averaging 10,392 +/- 876 vs. 3,969 +/- 534 mmHg/min in NP hearts (P < 0.001). The Ca(2+) load required to induce mPTP opening averaged 232 +/- 10 and 128 +/- 16 microM in LP and NP hearts, respectively (P < 0.001). Myocardial malondialdehyde was significantly lower in LP than in NP hearts (P < 0.05). These results suggest that the protection afforded by low-pressure reperfusion involves an inhibition of the opening of the mPTP, possibly via reduction of reactive oxygen species production.

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Introduction: Estrogen effect can be mediated by three receptors: Classical estrogen receptors: alpha (ERα) and beta (ERβ), and recently identified G protein‐coupled estrogen receptor1 (GPER1). Hypothesis: We investigated the role of ERα, ERβ and GPER1 in mediating rapid estrogen‐induced cardioprotection in male mice hearts subjected to ischemia/reperfusion using wild type (WT) and gene specific knockout animals. Methods: Isolated hearts from wild type (WT: C57BL/6NCrL), ERα‐/‐, ERβ‐/‐ and GPER1‐/‐ were perfused using Langendorff apparatus with Krebs Henseleit buffer (control) or with the addition of estrogen (40 nM). Hearts were subjected to 18 min global ischemia followed by 60 min reperfusion. Cardiac function was recorded during the entire experiment and myocardial infarct size measured by TTC staining at the end of the reperfusion. Mitochondria calcium retention capacity (CRC) required to induce the mitochondrial permeability transition pore (mPTP) opening was assessed after 10 min reperfusion. Protein levels were measured by Western Blot in whole heart lysates after 5 min treatment just before ischemia, and after 10 min reperfusion. LY294002, U0126 and Chelerythrin‐Cl were used as inhibitor of PI‐3K/Akt, MAPK/ERK and PKC translocation, respectively. Results: In WT, ERα‐/‐ and ERβ‐/‐, estrogen treatment significantly improved cardiac functional recovery, reduced infarct size and improved mitochondrial CRC. However, estrogen effects were completely absent in GPER1‐/‐. Estrogen treatment during 5 min before ischemia induced up‐regulation of Akt, GSK‐3β, and ERK1/2 phosphorylation in WT mouse as compared with control but not in GPER1‐/‐. However, after 10 min reperfusion estrogen effect was still oserved on GSK‐3β, but not on Akt and ERK1/2,. Chelerythrin‐Cl prevented estrogen‐induced cardioprotection effect and U126 abolished estrogen effect on mitochondrial CRC while LY294002 could not prevent estrogen effect on GSK‐3β observed in WT. P<0.05 and n=3‐6. Conclusion: Rapid activation of GPER1 induces cardioprotection effect against ischemia/reperfusion injury. Estrogen effects through GPER1 are associated with phosphorylation of Akt, GSK‐3β and ERK1/2, translocation of PKC, and inhibition of the mPTP opening. Grant Funding Source: American Heart Association

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