BACKGROUND: Brief episodes of coronary blood flow interruption, ischemic preconditioning (IP), following a prolonged ischemia induces myocardial tolerance to ischemia and improves myocardial function during reperfusion by undefined mechanism. Recently, it has been suggested that the signal transduction pathway of the cardiomyocyte itself may involve in this protection. The aims of the present study were : (1) to examine the effect of adenosine in early phase of IP, (2) to define the relationship between the adenosine and protein kinase C(PKC) METHOD AND RESULTS: Heart isolated from New Zealand White rabbit (1.2 - 1.5kg body weight, n=78) were perfused with Tyrode solution by non-recirculating Langendorff technique. After stabilization of baseline hemodynamics, the hearts were subjected to receiving 45min global ischemia (I) and 120min reperfusion (R) with or without IP. IP was induced by a single dose of 5min I and 10min R. A part of the IP hearts, calphostin C (200nmol/L), a PKC inhibitor, was administered 5min before IP and sustained during IP regimen. Left ventricular function and coronary flow were monitored. Infarct size was determined by staining with 1% triphenyltetrazolium chloride solution and computerized planimetry. Adenosine concentration in the coronary flow was determined by HPLC. Myocardial cytosolic and membrane PKC activities were measured by (32)P-r-ATP incorporation into PKC specific peptide. Expression of PKC-e and PKC-o was determined by SDS-PAGE and Western blot. IP enhanced improvement of functional recovery (p<0.05, in the left ventricular developed and end-diastolic pressure ; p<0.01, in the coronary flow) during 120min R after 45min I. Preconditioned hearts showed reduction in the infarct size compared with the non-preconditioned hearts (p<0.05) ; however, IP-induced protection was lost by calphostin C. Adenosine release from the cardiomyocytes abruptly increased to 10-20 folds baseline just after IP manipulation and decreased rapidly on reperfusion. Cytosolic PKC activity significantly decreased in the preconidtioned hearts which received 45min I(p<0.05) and 45min I and 120min R(p<0.01), while the membrane fraction increased in the former(p<0.05) and the latter(p<0.01) groups. There was no significant difference in the PKC-o activity among all experimental groups in cytosolic and membrane fraction, however, the membrane PKC-e isoenzyme activity was increased in the preconditioned hearts which received 45min I. CONCLUSION: These results indicate that (1) a single dose of brief ischemia has an infarctlimiting effect and can improve post-ischemic contractile dysfunction after 45min subsequent sustained I ; and (2) increase of adenosine release in the earlier period of IP regimen and translocation of PKC from the cytosol to myocyte membrane may be important processes signal transduction for protection. These results suggest that cardioprotective mechanism responsible for IP in isolated rabbit heart may be initiated by adenosine and PKC.