Exercise training provides cardioprotection by activating and coupling endothelial nitric oxide synthase via a β₃-adrenergic receptor-AMP-activated protein kinase signaling pathway

Exercise training confers sustainable protection against ischemia/reperfusion injury. However, the mechanism by which this process occurs is not fully understood. Previously, it was shown that β₃-adrenergic receptors (β₃-ARs) play a critical role in regulating the activation of endothelial nitric oxide synthase (eNOS) in response to exercise and play a critical role in exercise-mediated cardioprotection. Intriguingly, a deficiency in β₃-ARs led to increased myocardial injury following exercise training. The purpose of the current study was to determine mechanisms by which β₃-ARs are linked to eNOS activation and to determine the mechanism responsible for the exacerbated ischemia/reperfusion injury displayed by β₃-AR deficient (β₃-AR KO) mice after exercise training. Wild-type (n = 37) and β₃-AR KO ( n = 40) mice were subjected to voluntary wheel running for 4 weeks. Western blot analysis revealed that neither protein kinase B nor protein kinase A linked β₃-ARs to eNOS following exercise training. However, analysis revealed a role for AMP-activated protein kinase (AMPK). Specifically, exercise training increased the phosphorylation of AMPK in the hearts of wild-type mice, but failed to do so in the hearts of β₃-AR KO mice. Additional studies revealed that exercise training rendered eNOS less coupled and increased NOS-dependent superoxide levels in β₃-AR KO mice. Finally, supplementing β₃-AR KO mice with the eNOS coupler, tetrahydrobiopterin, during the final week of exercise training reduced myocardial infarction. These findings provide important information that exercise training protects the heart in the setting of myocardial ischemia/reperfusion injury by activating and coupling eNOS via the stimulation of a β₃-AR-AMPK signaling pathway.