Opioid Receptor Mediates The Cardioprotective Effect Of Ischemic Postconditioning

BACKGROUNDThe ischemic preconditioning phenomenon was introduced in 1986 by Murry, who found that several brief episodes of ischemia-reperfusion prior to a prolonged ischemic insult significantly decreased the myocardial injury. Intensive research has shown that a preconditioning response may be elicited by a variety of chemical and physical stimuli. In 2003, Vinten-Johansen and his colleagues reported that brief episodes of myocardial ischemia and reperfusion employed during reperfusion after a prolonged ischemic insult have been demonstrated to play the cardioprotective role. This phenomenon was termed "ischemic postconditioning". The mechanism of postconditioning is not yet clearly known. Activation of the opioid receptors by opioid peptides has also been investigated as a potent exogenous trigger of preconditioning in myocardial tissue. Adult rat ventricular cardiac myocytes have been demonstrated the presence of k- and 5-opioid receptors, but not μ-opioid receptors. It has been also demonstrated that the mechanism of cardioprotection by opioid involved Gi/o, protein kinase C, tyrosine kinase,mitogen-activated protein kinase and ATP sensitive potassium channel. It is know that there is a calciumactivated potassium channel (KCa) in the mitochondrion of cardiomyocyte, which can play the trigger of cardioprotection in ischemic preconditioning as well as the activation of opioid receptor. So we hypothesized that opioid receptor and KCa channel participate in the cardioprotection of ischemic postconditioning. The objective of this study was to investigate whether KCa and opioid receptors contribute to cardioprotective effect elicited by ischemic postconditioning.OBJECTIVES1. To observe the cardioprotection of ischemic postconditioning, and determine the possible synergistic cardioprotective effect of ischemic postconditioning combined with ischemic preconditioning.2. To investigate the effect of opioid receptors in the cardioprotection elicited by ischemic postconditioning and the downstream mechanism.3. To verify the role of calcium activated potassium channel and mitochondrial permeability transition pore in the cardioprotection by ischemic postconditioning.METHODS1. Langendorff isolated perfused rat heart was used and left ventricle hemodynamics were evaluated. The hearts were subjected to 60 or 30 min of global ischemia followed by 120 min of reperfusion.2. Ischemic postconditioning was achieved by 6 cycles of 10 s reperfusion/10 s global ischemia starting at the beginning of the reperfusion.3. Ischemic preconditioning was achieved by 2 cycles of 5min global ischemia/5min reperfusion starting before the ischemia.4. Lactate dehydrogenase (LDH) in the effluent from the isolated perfused heart was spectrophotometrically assayed.5. Myocardial viability was evaluated by measurement of formazan, a product of3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide, which is proportional to myocardial viability.6. Opening of mitochondrial permeability transition pore was measured spectrophotometrically.7. Isolated ventricular myocytes was prepared by an enzyme method.8. Cardiomyocytes was subjected to 60min hypoxia and 60min reoxygenation.9. Hypoxia postconditioning was achieved by 3 cycles of 5 min reoxygenation/5 min hypoxia starting at the beginning of reoxygenation.RESULTS1. Synergistic cardioprotective effect of ischemic postconditioning and preconditioning on heart subjected to severe ischemia/reperfusion injury(1) Compared with hearts of ischemia and reperfusion (I/R), ischemic preconditioning or postconditioning increased ±dP/dtmax, reduced LDH release and increased formazan content.(2) Compared with I/R hearts, combination of ischemic preconditioning and postconditioning increased left ventricular developed pressure (LVDP), rate-pressure product (RPP), maximal rise/fall rate of left ventricular pressure (±dP/dtmax) and formazan content, and reduced LDH release. The cardioprotection of combined conditioning was stronger than preconditioning or postconditioning alone.2. Activation of opioid receptors contribute to the cardioprotection ofpostconditioning(1) In isolated rat heart, pretreatment with a selective 5-opioid antagonist naltrindoleat Ixl0"7mol/L for 15 min attenuated cardioprotection of ischemic posconditioning.In isolated ventricular myocytes naltrindole at SxlO^mol/L also reduced the increasedcell viability of ischemic postconditioning.(2) In isolated rat heart, pretreatment with a selective K-opioid antagonist nor-BNI at3><10"8 mol/L for 15 min attenuated cardioprotection of ischemic posconditioning. In isolated ventricular myocytes nor-BNI at 5xlO'6mol/L also reduced the increased cell viability of ischemic postconditioning.3. Role of calcium activated potassium channel and mitochondrial permeability transition pore in cardioprotection induced by ischemic postconditioning(1) Pretreatment with paxilline (1 umol/L), an inhibitor of KCa, attenuated the cardioprotective effect of ischemic postconditioning.(2) CaCh (200 umol/L) reduced absorbency at 520 nm of cardiac mitochondrial suspension isolated from I/R heart, while the reduction in mitochondria from Postconditioned heart was attenuated.CONCLUSIONS1. In the Langendorff perfused rat heart model subjected to severe ischemia/reperfusion, ischemic postconditioning has a similar effect to that of ischemic preconditioning in cardioprotection. Combination of both conditioning was stronger than preconditioning or postconditioning alone.2. In the present conditions, the cardioprotection elicited by ischemic postconditioning was probably mediated by activation of opioid receptors and the calcium activated potassium channel, and the inhibition of opening of mitochondrial permeability transition pore.