Spontaneous activation of beta 2-but not beta(1)-adrenoceptors expressed in cardiac myocytes from beta(1)beta(2) double knockout mice

Although ligand-free, constitutive beta(2)-adrenergic receptor (AR) signali ng has been demonstrated in naive cell lines and in transgenic mice overexp ressing cardiac beta(2)-AR, it is unclear whether the dominant cardiac beta -AR subtype, beta(1)-AR, shares the ability of spontaneous activation. In t he present study, we expressed human beta(1) -or beta(2)-AR via recombinant adenoviral infection in ventricular myocytes isolated from beta(1)beta(2)- AR double knockout mice, creating pure beta(1)-AR and beta(2)-AR systems wi th variable receptor densities. A contractile response to a nonselective be ta-AR agonist, isoproterenol, was absent in double knockout mouse myocytes but was fully restored after adenoviral beta(1)-AR or adenoviral beta(2)-AR infection. Increasing the titer of adenoviral vectors (multiplicity of inf ection 10-1000) led to a dose-dependent expression of beta(1)- or beta(2)-A R with a maximal density of 1207 +/- 173 (36-fold over the wild-type contro l value) and 821 +/- 38 fmol/ mg protein (69-fold), respectively. Using con focal immunohistochemistry, we directly visualized the cellular distributio n of beta(1)-AR and beta(2)-AR and found that both subtypes were distribute d on the cell surface membrane and transverse tubules, resulting in a stria ted pattern. In the absence of ligand, beta(2)-AR expression resulted in gr aded increases in baseline cAMP and contractility up to 428% and 233% of co ntrol, respectively, at the maximal beta(2)-AR density. These effects were specifically reversed by a beta(2)-AR inverse agonist, ICI 118,551 (10(-7) M). In contrast, overexpression of beta(1)-AR, even at a greater density, f ailed to enhance either basal cAMP or contractility; the alleged beta(1)-AR inverse agonist, CGP 20712A (10(-6) M), had no significant effect on basal contraction in these cells. Thus, we conclude that acute beta(2)-AR overex pression in cardiac myocytes elicits significant physiological responses du e to spontaneous receptor activation; however, this property is beta-AR sub type specific because beta(1)-AR does not exhibit agonist-independent spont aneous activation.