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1 Second Department of Internal Medicine, Sapporo Medical University, Sapporo, Japan
2 Department of Pharmacology, Sapporo Medical University, Sapporo, Japan; Second Department of Internal Medicine, Sapporo Medical University, Sapporo, Japan
* To whom correspondence should be addressed. E-mail: miura{at}sapmed.ac.jp.
The aim of this study was to examine the hypothesis that acceleration of gap junction (GJ) closure during ischemia contributes to anti-infarct tolerance afforded by preconditioning (PC). First, effects of PC on GJ communication during ischemia were assessed. Isolated buffer-perfused rabbit hearts were subjected to 5-min global ischemia with or without PC with 2 cycles of 5-min
ischemia/5-min reperfusion or a GJ blocker (2 mM heptanol), and then the tissue excised from the ischemic region was incubated in anoxic buffer containing lucifer yellow (LY, 2.5 mg/ml), a tracer of GJ permeability, for 20 min at 37 °C. PC and heptanol significantly reduced the area to which LY was transported in the ischemic myocardium by 39% and by 54%, respectively. In the second series of experiments, three GJ blockers (heptanol, 18 
-glycyrrhetinic acid, 2,3-butanedione monoxime) that were infused after the onset of ischemia reduced infarct size after 30-min ischemia/2-h reperfusion to an extent equivalent to that in the case of PC. In the third series of experiments, Western blotting for connexin-43 (Cx43) showed that PC shortened the time to the onset of ischemia-induced Cx43 dephosphorylation but reduced the extent of Cx43 dephosphorylation during a 30-min period of ischemia. Calphostin C, a PKC inhibitor, abolished preservation of phosphorylated Cx43 but not earlier onset of Cx43 dephosphorylation after ischemia in the preconditioned myocardium. These results suggest that PC-induced reduction of GJ permeability during ischemia, presumably by PKC-mediated Cx43 phosphorylation, contributes to infarct size limitation.
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