Heterodimerization With 5-HT2BR Is Indispensable for β2AR-Mediated Cardioprotection.

Abstract

The β2-adrenoceptor (β2-AR), a prototypical GPCR (G protein-coupled receptor), couples to both Gs and Gi proteins. Stimulation of the β2-AR is beneficial to humans and animals with heart failure presumably because it activates the downstream Gi-PI3K-Akt cell survival pathway. Cardiac β2-AR signaling can be regulated by crosstalk or heterodimerization with other GPCRs, but the physiological and pathophysiological significance of this type of regulation has not been sufficiently demonstrated. Here, we aim to investigate the potential cardioprotective effect of β2-adrenergic stimulation with a subtype-selective agonist, (R,R')-4-methoxy-1-naphthylfenoterol (MNF), and to decipher the underlying mechanism with a particular emphasis on the role of heterodimerization of β2-ARs with another GPCR, 5-hydroxytryptamine receptors 2B (5-HT2BRs). Using pharmacological, genetic and biophysical protein-protein interaction approaches, we studied the cardioprotective effect of the β2-agonist, MNF, and explored the underlying mechanism in both in vivo in mice and cultured rodent cardiomyocytes insulted with doxorubicin, hydrogen peroxide (H2O2) or ischemia/reperfusion. In doxorubicin (Dox)-treated mice, MNF reduced mortality and body weight loss, while improving cardiac function and cardiomyocyte viability. MNF also alleviated myocardial ischemia/reperfusion injury. In cultured rodent cardiomyocytes, MNF inhibited DNA damage and cell death caused by Dox, H2O2 or hypoxia/reoxygenation. Mechanistically, we found that MNF or another β2-agonist zinterol markedly promoted heterodimerization of β2-ARs with 5-HT2BRs. Upregulation of the heterodimerized 5-HT2BRs and β2-ARs enhanced β2-AR-stimulated Gi-Akt signaling and cardioprotection while knockdown or pharmacological inhibition of the 5-HT2BR attenuated β2-AR-stimulated Gi signaling and cardioprotection. These data demonstrate that the β2-AR-stimulated cardioprotective Gi signaling depends on the heterodimerization of β2-ARs and 5-HT2BRs.