(A) A representative Western blot (top panel) of GRK2 in cultured adult mouse cardiomyocytes infected with adenovirus expressing GRK2 (Adv-GRK2) or β-Gal (Adv-β-Gal) at m.o.i. 100 for 24 h and quantified data (bottom panel) (n=4; *p<0.01 v.s. β-Gal). (B) Increased GRK2 expression enhances β2AR to Gi signaling in adult mouse cardiomyocytes cultured for 24 h. In cultured cardiomyocytes infected with Adv-GRK2 or Adv-β-Gal (m.o.i. 100), overexpression of GRK2 enhances the receptor to Gi signaling, as evidenced by PTX-induced augmentation of β2AR contractile response. *P<0.001 v.s. other three groups with a two-way repeated measures analysis of variance (ANOVA); (C, D) Disruption of Gi signaling with PTX abolishes GRK2-induced dysfunction of βAR contractile response in GRK2 TG mice. In vivo assessment of left ventricular (LV) function of GRK2 TG mice and littermate control (LC) mice (n=6–7 in each group; *P<0.001 v.s. GRK2 TG mice without PTX with a two-way repeated measures analysis of variance (ANOVA); †p<0.05 GRK2 TG v.s. LC in the absence of PTX with post hoc testing and Bonferroni’s F test). For single cell experiments (panel B), cells were pretreated with PTX (1.5 µg/ml) 3 hours before agonist stimulation. For in vivo studies in panels C&D, one dose of PTX was administered i.p. 72 hours before hemodynamic measurements.