Acta Pharmacologica Sinica 2008 June; 29 (6): 661-669; doi: 10.1111/j.1745-7254.2008.00794.x

Aim: To observe the effect of β₂-adrenergic agonist clenbuterol on ischemia/reperfusion (I/R) injury in isolated rat hearts and hydrogen peroxide (H₂O₂)-induced cardiomyocyte apoptosis.

Methods: Isolated rat hearts were subjected to 30 min global ischemia and 60 min reperfusion on a Langendorff apparatus. Cardiac function was evaluated by heart rate, left ventricular end-diastolic pressure (LVEDP), left ventricular systolic pressure, maximal rise rate of left ventricular pressure (+dp/dt_max), and the coronary effluent (CF). Lactate dehydrogenase (LDH) in the coronary effluent, malondialdehyde (MDA), superoxide dismutase (SOD), and Ca²⁺-ATPase activity in the cardiac tissue were measured using commercial kits. The apoptotic cardiomyocyte was detected by terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling (TUNEL) assay. Bax/Bcl-2 mRNA levels and the expression of caspase-3 were detected by RT-PCR and immunoblotting, respectively. Cultured newborn rat cardiomyocytes were pre-incubated with clenbuterol, and oxidative stress injury was induced by H₂O₂. Cell viability and cardiomyocyte apoptosis were evaluated by flow cytometry (FCM).

Results: In the isolated rat hearts after I/R injury, clenbuterol significantly improved diastolic function (LVEDP and CF) and Ca²⁺-ATPase activity. Treatment with clenbuterol increased SOD activity and decreased the MDA level and LDH release compared with the I/R group (P<0.05). Moreover, clenbuterol decreased apoptosis, which was associated with a reduction in TUNEL-positive cells, Bax/Bcl-2 mRNA, and caspase-3 expres-sion. In H₂O₂-induced cardiomyocyte injury, clenbuterol increased cell viability and attenuated cardiomyocyte apoptosis. Pretreatment with ICI118551 (selective β₂-adrenergic antagonist) decreased these effects compared with the clenbuterol-treated group (P<0.05).

Conclusion: Clenbuterol ameliorated ventricular diastolic function by enhaning Ca²⁺-ATPase activity and reduced oxidative stress and cardiac myocyte apoptosis in an experimental rat model of myocardium I/R. It decreased cardiomyocyte apoptosis induced by H₂O₂ in vitro. It plays a key role in the cardiac protection against myocardium I/R injury.

Keywords: clenbuterol; β₂-adrenergic receptor; myocardium ischemia/reperfusion; apoptosis; oxidative stress; Ca²⁺-ATPase