E xogenous gene delivery to alter the function of the heart is a potential novel therapeutic strategy for treatment of cardiovascular diseases such as heart failure (HF). Before gene therapy approaches to alter cardiac function can be realized, efficient and reproducible in vivo gene techniques must be established to efficiently transfer transgenes globally to the myocardium. We have been testing the hypothesis that genetic manipulation of the myocardial β-adrenergic receptor (β-AR) system, which is impaired in HF, can enhance cardiac function. We have delivered adenoviral transgenes, including the human β₂-AR (Adeno-β₂AR), to the myocardium of rabbits using an intracoronary approach. Catheter-mediated Adeno-β₂AR delivery produced diffuse multichamber myocardial expression, peaking 1 week after gene transfer. A total of 5 × 10¹¹ viral particles of Adeno-β₂AR reproducibly produced 5- to 10-fold β-AR overexpression in the heart, which, at 7 and 21 days after delivery, resulted in increased in vivo hemodynamic function compared with control rabbits that received an empty adenovirus. Several physiological parameters, including dP/dt_max as a measure of contractility, were significantly enhanced basally and showed increased responsiveness to the β-agonist isoproterenol. Our results demonstrate that global myocardial in vivo gene delivery is possible and that genetic manipulation of β-AR density can result in enhanced cardiac performance. Thus, replacement of lost receptors seen in HF may represent novel inotropic therapy.