Continuous detection of superoxide in situ during ischemia and reperfusion in the rabbit heart

In order to clarify the time course of superoxide generation in situ during ischemia and reperfusion in the rabbit heart, we used a method of enhanced chemiluminescence (CL) with 2-methyl-6-[p-methoxyphenyl]-3, 7-dihydroimidazo [1, 2-alpha]pyrazin-3-one (MCLA) as a specific probe for detecting superoxide radicals. The surface of the rabbit heart was exposed to a photomultiplier tube in a light-proof box. We introduced a reversible snare occluder into the box to continuously observe the light emission. An ischemia-reperfusion group (I/R, n = 7) was subjected to 30 mins of coronary occlusion, followed by 90 mins of reperfusion. We performed the same procedure (except for coronary occlusion) in the sham-operated group (n = 4). Another group of rabbits (n = 4) subjected to I/R received superoxide dismutase (SOD: 20 mg/kg, i.v.) during reperfusion to observe the CL response. In the I/R-group, the increase in CL began at 13 +/- 2 (mean +/- SEM) mins and peaked at 52 +/- 12 mins of reperfusion. CL in the I/R-group gradually increased from 818 +/- 350 counts/10 secs in the preischemic period to 1077 +/- 401 counts/10 secs during reperfusion (p < 0.01). In contrast, there was no increase in CL in the sham-operated group. The administration of SOD briefly attenuated CL by 24.1 +/- 6.8% for a period of 24.3 +/- 6.8 mins. The superoxide generation in situ in the ischemic rabbit heart appears to increase gradually and persists for a period following reperfusion.