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) signaling has been demonstrated in naive cell lines and in transgenic mice overexpressing cardiac beta(2)-AR, it is unclear whether the dominant cardiac beta-AR subtype, beta(1)-AR, shares the ability of spontaneous activation. In the 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 with variable receptor densities. A contractile response to a nonselective beta-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 infection 10-1000) led to a dose-dependent expression of beta(1)- or beta(2)-AR with a maximal density of 1207 +/- 173 (36-fold over the wild-type control value) and 821+/-38 fmol/mg protein (69-fold), respectively. Using confocal immunohistochemistry, we directly visualized the cellular distribution of beta(1)-AR and beta(2)-AR and found that both subtypes were distributed on the cell surface membrane and transverse tubules, resulting in a striated pattern. In the absence of ligand, beta(2)-AR expression resulted in graded increases in baseline cAMP and contractility up to 428% and 233% of control, 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, failed 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 overexpression in cardiac myocytes elicits significant physiological responses due to spontaneous receptor activation; however, this property is beta-AR subtype specific because beta(1)-AR does not exhibit agonist-independent spontaneous activation.
Zhou
Y Y
YY
Laboratory of Cardiovascular Science, Gerontology Research Center, National Institute on Aging, National Institutes of Health, Baltimore, Maryland, USA.
Yang
D
D
Zhu
W Z
WZ
Zhang
S J
SJ
Wang
D J
DJ
Rohrer
D K
DK
Devic
E
E
Kobilka
B K
BK
Lakatta
E G
EG
Cheng
H
H
Xiao
R P
RP
D000322:Adrenergic Agonists / Q000494:pharmacology; D000818:Animals; D006352:Heart Ventricles / Q000166:cytology / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009206:Myocardium / Q000378:metabolism*; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
D000322:Adrenergic Agonists; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2