Examined in the present study, allopurinol are xanthine oxidase inhibitors for use in rat hearts in vivo dialysis technology and ventricular myocardial intersitial adenosine production can increase. The microdialysis probe was implanted in the left ventricular myocardium of anesthetized rat hearts and the tissue in the vicinity of the dialysis was perfused with Tyrode's solution containing adenosine 5′-monophosphate (AMP) through the dialysis probe at a rate of 1.0 ml/min to assess the activity of ecto-5′-nucleotidase. Allopurinol (10 μM) significantly increased the level of adenosine in rat heart dialysate (n=6, p<0.05), which was inhibited by chelerythrine, 10 μM, an antagonist of protein kinase C (PKC). Another free radical scavenger, coenzyme Q10 (CoQ10, 100 μM) or ascorbic acid (Vitamin C; 100 μM) also increased adenosine production. In addition, allopurinol enhanced the diacylglycerol (DAG; 50 μM)-induced also increases in adenosine production by 71.5±12.0% (n=6, P<0.05), to a level significantly (P<0.05) greater than the increase caused by DAG alone (33.0±10.6%). In the presence of allopurinol (10 μM), a marked elevation of AMP-primed dialysate adenosine in ischemia/reperfused rat hearts was observed. Free radical generation may suppress adenosine production via activation of PKC. The results suggest that oxidative stress may cause inactivation of nucleotidase, adenosine production in rat heart.