-Transgenic mouse models have been developed to manipulate beta-adrenergic receptor (betaAR) signal transduction. Although several of these models have altered betaAR subtypes, the specific functional sequelae of betaAR stimulation in murine heart, particularly those of beta2-adrenergic receptor (beta2AR) stimulation, have not been characterized. In the present study, we investigated effects of beta2AR stimulation on contraction, [Ca2+]i transient, and L-type Ca2+ currents (ICa) in single ventricular myocytes isolated from transgenic mice overexpressing human beta2AR (TG4 mice) and wild-type (WT) littermates. Baseline contractility of TG4 heart cells was increased by 3-fold relative to WT controls as a result of the presence of spontaneous beta2AR activation. In contrast, beta2AR stimulation by zinterol or isoproterenol plus a selective beta1-adrenergic receptor (beta1AR) antagonist CGP 20712A failed to enhance the contractility in TG4 myocytes, and more surprisingly, beta2AR stimulation was also ineffective in increasing contractility in WT myocytes. Pertussis toxin (PTX) treatment fully rescued the ICa, [Ca2+]i, and contractile responses to beta2AR agonists in both WT and TG4 cells. The PTX-rescued murine cardiac beta2AR response is mediated by cAMP-dependent mechanisms, because it was totally blocked by the inhibitory cAMP analog Rp-cAMPS. These results suggest that PTX-sensitive G proteins are responsible for the unresponsiveness of mouse heart to agonist-induced beta2AR stimulation. This was further corroborated by an increased incorporation of the photoreactive GTP analog [gamma-32P]GTP azidoanilide into alpha subunits of Gi2 and Gi3 after beta2AR stimulation by zinterol or isoproterenol plus the beta1AR blocker CGP 20712A. This effect to activate Gi proteins was abolished by a selective beta2AR blocker ICI 118,551 or by PTX treatment. Thus, we conclude that (1) beta2ARs in murine cardiac myocytes couple to concurrent Gs and Gi signaling, resulting in null inotropic response, unless the Gi signaling is inhibited; (2) as a special case, the lack of cardiac contractile response to beta2AR agonists in TG4 mice is not due to a saturation of cell contractility or of the cAMP signaling cascade but rather to an activation of beta2AR-coupled Gi proteins; and (3) spontaneous beta2AR activation may differ from agonist-stimulated beta2AR signaling.

Background-Myocardial contractile response to ␤ 1 -and ␤ 2 -adrenergic receptor (AR) stimulation is severely impaired in chronic heart failure, in which G i signaling and the ratio of ␤ 2 /␤ 1 are often increased. Because ␤ 2 -AR but not ␤ 1 -AR couples to G s and G i with the G i coupling negating the G s -mediated contractile response, we determined whether the heart failure-associated augmentation of G i signaling contributes differentially to the defects of these ␤-AR subtypes and, if so, whether inhibition of G i or selective activation of ␤ 2 -AR/G s by ligands restores ␤ 2 -AR contractile response in the failing heart. Methods and Results-Cardiomyocytes were isolated from 18-to 24-month-old failing spontaneously hypertensive (SHR) or age-matched Wistar-Kyoto (WKY) rat hearts. In SHR cardiomyocytes, either ␤-AR subtype-mediated inotropic effect was markedly diminished, whereas G i proteins and the ␤ 2 /␤ 1 ratio were increased. Disruption of G i signaling by pertussis toxin (PTX) enabled ␤ 2 -but not ␤ 1 -AR to induce a full positive inotropic response in SHR myocytes. Furthermore, screening of a panel of ␤ 2 -AR ligands revealed that the contractile response mediated by most ␤ 2 -AR agonists, including zinterol, salbutamol, and procaterol, was potentiated by PTX, indicating concurrent G s and G i activation. In contrast, fenoterol, another ␤ 2 -AR agonist, induced a full positive inotropic effect in SHR myocytes even in the absence of PTX. Conclusions-We conclude that enhanced G i signaling is selectively involved in the dysfunction of ␤ 2 -but not ␤ 1 -AR in failing SHR hearts and that disruption of G i signaling by PTX or selective activation of ␤ 2 -AR/G s signaling by fenoterol restores the blunted ␤ 2 -AR contractile response in the failing heart. (Circulation. 2003;108:1633-1639.)