PBN SPIN-TRAPPING OF FREE-RADICALS IN THE REPERFUSION-INJURED HEART -LIMITATIONS FOR PHARMACOLOGICAL INVESTIGATIONS

Post-ischemic reperfusion causes cardiac dysfunction and radical-induc ed lipid peroxidation (LPO) detectable by ESR spin trapping. This stud y deals with the applicability of the spin trapping technique to pharm acological investigations during myocardial reperfusion injury. The us e of the spin trap phenylbutylnitrone (PBN, 3 mM) in isolated rat hear ts demonstrated the release of alkoxyl radicals (a(N) = 1.39 mT, a(H)( beta) = 0.19 mT) formed particularly within the first 15 min of reperf usion following 30 min of ischemia. The decline of radicals, after 10 min of reperfusion, was accompanied by recovery of function in 80% of the hearts. The radical concentration in the coronary effluent (maximu m after 7.5 min) was reduced by the infusion of 1 mM mercaptopropionyl glycine (MPG, 2.7 +/- 0.5 U/ml, p < 0.001) or 5 mu M vitamin E (11.7 /- 0.8 U/ml, p < 0.001), compared to the (PBN-containing) control (29. 7 +/- 4.3 U/ml). Moreover, functional recovery (left ventricular devel oped pressure, LVDP 91.6 +/- 20% of pre-ischemic level, p < 0.05) was improved by the hydrophilic radical scavenger MPG, compared to the (PB N containing) control (LVDP 50.5 +/- 15.7% of baseline). PBN alone led to higher functional recovery (p < 0.05) and reduced VF (duration of ventricular fibrillation; 7.10 +/- 0.36 min/30 min, p < 0.05), compare d to the untreated (PBN-free) control (LVDP 26.6 +/- 11.8%; VF 19.42 /- 3.64 min/30 min). The Ca antagonist verapamil (0.1 mu M), MPG, and the lipophilic vitamin E showed cardioprotection in the absence of PEN : post-ischemic recovery of LVDP was 25.4 +/- 6.8% (p < 0.05), 39.6 +/ - 12.7% (p < 0.05) and 52.4 +/- 2.6% (p < 0.01), respectively, compare d to the corresponding untreated control (13.3 +/- 6.6%). Whereas vera pamil and vitamin E were able to protect the heart when present alone, they offered no additive effect in the presence of PBN. Therefore, PB N can be used to estimate the radical scavenger properties of an agent in the heart. However, because of the protective properties of PBN it self, the results of simultaneous investigations of the effects of oth er compounds, such as Ca antagonists or lipophilic radical scavengers, on heart function may be limited.