In contrast to ␤ 1 -adrenoreceptor (␤ 1 -AR) signaling, ␤ 2 -AR stimulation in cardiomyocytes augments L-type Ca 2؉ current in a cAMP-dependent protein kinase (PKA)-dependent manner but fails to phosphorylate phospholamban, indicating that the ␤ 2 -AR-induced cAMP/PKA signaling is highly localized. Here we show that inhibition of G i proteins with pertussis toxin (PTX) permits a full phospholamban phosphorylation and a de novo relaxant effect following ␤ 2 -AR stimulation, converting the localized ␤ 2 -AR signaling to a global signaling mode similar to that of ␤ 1 -AR. Thus, ␤ 2 -AR-mediated G i activation constricts the cAMP signaling to the sarcolemma. PTX treatment did not significantly affect the ␤ 2 -ARstimulated PKA activation. Similar to G i inhibition, a protein phosphatase inhibitor, calyculin A (3 ؋ 10 ؊8 M), selectively enhanced the ␤ 2 -AR but not ␤ 1 -AR-mediated contractile response. Furthermore, PTX and calyculin A treatment had a non-additive potentiating effect on the ␤ 2 -AR-mediated positive inotropic response. These results suggest that the interaction of the ␤ 2 -AR-coupled G i and G s signaling affects the local balance of protein kinase and phosphatase activities. Thus, the additional coupling of ␤ 2 -AR to G i proteins is a key factor causing the compartmentalization of ␤ 2 -AR-induced cAMP 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.)