| BACKGROUND AND AIMCellular electrical uncoupling at gap junctions during acute myocardial ischemia is considered to be a sign of irreversible ischemic cell damage. In normal hearts, cellular electrical uncoupling during ischemia contributes to conduction abnormalities and reentrant arrhythmias. Insights into the mechanisms responsible for cellular electrical uncoupling may help in finding new therapeutic strategies to limit the lethal arrhythmias induced by ischemia. Ischemic preconditioning (IPC) activates protective mechanisms that postpone the onset of irreversible myocardial damage during subsequent sustained ischemia. But the precise mechanisms of IPC remain unknown. One of the mediators of IPC protection is the ATP-sensitive potassium channels (KATP)- Previously, sarcolemmal KATP channels were considered to mediate the IPC protection by action potential shortening. But growing evidences have demonstrated that protection of IPC might be mediated via mitochondrial KATP channels(mitoKATp) rather than sarcolemmal KATP channels. Selective mitoKATp channels activation results in antiarrhythmic and cardioprotective effects during ischemia/reperfusion in rabbits and rats. Previous studies showed that IPC can postpone the onset of electrical uncoupling. We studied an isolated rat heart preparation in which the time course ofcellular uncoupling during ischemia was similar to that reported in rabbits and porcine hearts. The purpose of our study was to test the hypothesis that electrical uncoupling induced by myocardial ischemia can be mediated by activation of mitoKATP channels.METHODSRat hearts were perfused on a Langendorff apparatus and subjected to 40min-ischemia followed by 30min-reperfusion (I/R). IPC consists of 2 cycles of 5 min of ischemia followed by 5 min of reperfusion. Hearts were perfused for 3 cycles of 1 min each with Ca2+-free KH buffer followed by 5 min with Ca2+-containing KH buffer, as calcium preconditioning (CPC). The time course of electrical uncoupling induced by acute ischemia was monitored by measuring changes in whole tissue resistance with a four-electrode technique.RESULTSChanges in Rt in no flow ischemia occur in characteristic phases. The mean time of onset of uncoupling in 7 hearts was 13.3+1.0 min in the I/R group. IPC significantly postponed the onset of uncoupling to 22.7+1.3 min of ischemia (P<0.001, vs I/R group). 5-HD, a specific blocker of mitoKATp channels, abolished the effect of IPC (12.6+1.6 min, P > 0.05 vs I/R group). CPC also postponed the onset of uncoupling at 20.5811.28 min of ischemia (P<0.001, vs I/R group), which was blocked by 5-HD, when uncoupling occurred at 13.57+0.75 min (P >0.05, vs I/R group). Diazoxide(DE), a specific opener of mitoKATp channels, mimicked the effect of IPC. When DE was added before sustained ischemia, it delayed the onset of uncoupling induced by ischemia to 18.411.4 min (P<0.05, vs I/R group). The effect of DE on uncoupling was blocked by 5-HD, when the uncoupling started at 12.6+1.0 min (P >0.05, vs I/R group). Blockade of Ca2+ entry by inhibiting the L-type Ca2+ channel with VL reversed the beneficial effect of DE during I/R (13.3+1. 8 min, P>0.05 vs I/R group). N-(2-mercaptopropionyl)glycine, which is a free radical scavenger, also blocked the effect of DE (13.4+2.1 min, P >0.05 vs I/R group).Contractile function was assessed by Left ventricular developed presser ( LVDP) at end-perfusion, and was expressed as percentage of baseline function. Hearts subjected to 40 min of global ischemia had markedly depressed contractile function at end-reperfusion (27+14 %) compared with normal control (68+9%, P<0.001). During reperfusion, hearts in the IPC group showed improved systolic functional recovery, as demonstrated by a higher LVDP compared with that in the I/R group (59+19 %, P <0.05 vs I/R group). This protection was significantly reduced by pretreatment with 5-HD (42+10 %). CPC improved LVDP at end reperfusion (47+9 %, P<0.001 vs I/R group). 5-HD canceled the effect of CPC (20+10 %, P >0.05).
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