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Severe arrhythmias occur predictably on reperfusion after 5 minutes of coronary occlusion in the rat. There is little data available on whether ischemic preconditioning (PC) of hearts can reduce the incidence of such arrhythmias. The effect of PC (three cycles of 2 minutes of coronary occlusion and 5 minutes of reperfusion) on development of arrhythmias after a subsequent 5-minute coronary artery occlusion and reperfusion was studied. Rats (n=16 each group) underwent 5-minute occlusion and reperfusion alone or preceded by PC; arrhythmias were monitored during ischemia and for 10 minutes of reperfusion, and biopsies were taken for creatine phosphate and adenosine triphosphate in ischemic and nonischemic zones of the left ventricle. PC reduced the incidence of ventricular tachycardia (VT) during occlusion (81% control versus 13% PC, p<0.001). On subsequent reperfusion, ventricular fibrillation (VF) developed in zero PC animals versus 13 (81%) of controls (p<0.001), and irreversible VF in zero of PC versus seven (44%) of controls (p=0.007). VT occurred in four (25%) of PC versus all (100%) of controls (p<0.001). PC reduced mean duration of VT plus VF from 320±54 to 5±1 seconds (p<0.001) and delayed arrhythmia onset from 8±2 to 85±35 seconds after reperfusion. There was no difference in creatine phosphate levels in the ischemic zone at the end of reperfusion in PC animals compared with controls without irreversible VF (16.2 ±4.1 versus 15.5 ±3.9 nmol/mg protein, p=NS). There was no relation between creatine phosphate levels and occurrence of VT or VF (14.0±5.6 nmol/mg protein VF versus 16.7±3.3 no VF; 16.4±3.5 VT versus 15.4±4.5 no VT; p=NS). Adenosine triphosphate levels in the ischemic zone were unaffected by PC (15.5±2.1 versus 14.5±1.9 nmol/mg protein, PC versus control). When a coronary occlusion of 5 minutes duration is preceded by PC, the usually severe reperfusion arrhythmias are markedly attenuated. This protective effect of PC is not likely to be related to alterations in high-energy phosphate compounds.