PBN spin trapping of free radicals in the reperfusion-injured heart. Limitations for pharmacological investigations.
Post-ischemic reperfusion causes cardiac dysfunction and radical-induced lipid peroxidation (LPO) detectable by ESR spin trapping. This study deals with the applicability of the spin trapping technique to pharmacological investigations during myocardial reperfusion injury. The use of the spin trap phenylbutylnitrone (PBN, 3 mM) in isolated rat hearts demonstrated the release of alkoxyl radicals (aN = 1.39 mT, aHbeta = 0.19 mT) formed particularly within the first 15 min of reperfusion 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 (maximum after 7.5 min) was reduced by the infusion of 1 mM mercaptopropionylglycine (MPG, 2.7+/-0.5 U/ml, p < 0.001) or 5 microM 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 developed pressure, LVDP 91.6 +/-20% of pre-ischemic level, p < 0.05) was improved by the hydrophilic radical scavenger MPG, compared to the (PBN-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), compared 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 microM), MPG, and the lipophilic vitamin E showed cardioprotection in the absence of PBN: 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, compared to the corresponding untreated control (13.3+/-6.6%). Whereas verapamil and vitamin E were able to protect the heart when present alone, they offered no additive effect in the presence of PBN. Therefore, PBN can be used to estimate the radical scavenger properties of an agent in the heart. However, because of the protective properties of PBN itself, the results of simultaneous investigations of the effects of other compounds, such as Ca antagonists or lipophilic radical scavengers, on heart function may be limited.
Institute for Heart Research, Slovak Acadamy of Sciences, Bratislava, Slovak Republic.
N
N
Vrbjar
S
S
Zöllner
R F
RF
Haseloff
M
M
Pissarek
I E
IE
Blasig
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001145:Arrhythmias, Cardiac / Q000209:etiology / Q000503:physiopathology / Q000517:prevention & control; D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy; D016166:Free Radical Scavengers / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives / Q000494:pharmacology; D006321:Heart / Q000187:drug effects / Q000503:physiopathology; D066298:In Vitro Techniques; D007700:Kinetics; D015227:Lipid Peroxidation; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000378:metabolism* / Q000503:physiopathology / Q000517:prevention & control; D009589:Nitrogen Oxides; D051381:Rats; D017207:Rats, Sprague-Dawley; D013113:Spin Labels; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D016277:Ventricular Function, Left / Q000187:drug effects; D014700:Verapamil / Q000494:pharmacology; D014810:Vitamin E / Q000494:pharmacology
D000975:Antioxidants; D003497:Cyclic N-Oxides; D016166:Free Radical Scavengers; D005609:Free Radicals; C110658:N-(2-mercaptoproprionyl)-glycine; D009589:Nitrogen Oxides; D013113:Spin Labels; D013438:Sulfhydryl Compounds; D014810:Vitamin E; C029217:phenyl-N-tert-butylnitrone; D014700:Verapamil; D005998:Glycine