Coupling of beta2-adrenoceptor to Gi proteins and its physiological relevance in murine cardiac myocytes.
Abstract
-Transgenic mouse models have been developed to manipulate beta-adrenergic receptor
(betaAR) signal transduction. Although several of these models have altered betaAR
subtypes, the specific functional sequelae of betaAR stimulation in murine heart,
particularly those of beta2-adrenergic receptor (beta2AR) stimulation, have not been
characterized. In the present study, we investigated effects of beta2AR stimulation
on contraction, [Ca2+]i transient, and L-type Ca2+ currents (ICa) in single ventricular
myocytes isolated from transgenic mice overexpressing human beta2AR (TG4 mice) and
wild-type (WT) littermates. Baseline contractility of TG4 heart cells was increased
by 3-fold relative to WT controls as a result of the presence of spontaneous beta2AR
activation. In contrast, beta2AR stimulation by zinterol or isoproterenol plus a selective
beta1-adrenergic receptor (beta1AR) antagonist CGP 20712A failed to enhance the contractility
in TG4 myocytes, and more surprisingly, beta2AR stimulation was also ineffective in
increasing contractility in WT myocytes. Pertussis toxin (PTX) treatment fully rescued
the ICa, [Ca2+]i, and contractile responses to beta2AR agonists in both WT and TG4
cells. The PTX-rescued murine cardiac beta2AR response is mediated by cAMP-dependent
mechanisms, because it was totally blocked by the inhibitory cAMP analog Rp-cAMPS.
These results suggest that PTX-sensitive G proteins are responsible for the unresponsiveness
of mouse heart to agonist-induced beta2AR stimulation. This was further corroborated
by an increased incorporation of the photoreactive GTP analog [gamma-32P]GTP azidoanilide
into alpha subunits of Gi2 and Gi3 after beta2AR stimulation by zinterol or isoproterenol
plus the beta1AR blocker CGP 20712A. This effect to activate Gi proteins was abolished
by a selective beta2AR blocker ICI 118,551 or by PTX treatment. Thus, we conclude
that (1) beta2ARs in murine cardiac myocytes couple to concurrent Gs and Gi signaling,
resulting in null inotropic response, unless the Gi signaling is inhibited; (2) as
a special case, the lack of cardiac contractile response to beta2AR agonists in TG4
mice is not due to a saturation of cell contractility or of the cAMP signaling cascade
but rather to an activation of beta2AR-coupled Gi proteins; and (3) spontaneous beta2AR
activation may differ from agonist-stimulated beta2AR signaling.
Type
Journal articleSubject
Adrenergic beta-AgonistsAdrenergic beta-Antagonists
Animals
Calcium
Calcium Channels
Calcium Channels, L-Type
Cells, Cultured
Colforsin
Cyclic AMP
Ethanolamines
GTP-Binding Proteins
Heart
Heart Ventricles
Humans
Imidazoles
Isoproterenol
Male
Mice
Mice, Transgenic
Myocardial Contraction
Myocardium
Norepinephrine
Pertussis Toxin
Propanolamines
Receptors, Adrenergic, beta-2
Thionucleotides
Virulence Factors, Bordetella
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Show full item recordScholars@Duke
Robert J. Lefkowitz
The Chancellor's Distinguished Professor of Medicine
Dr. Lefkowitz’s memoir, A Funny Thing Happened on the Way to Stockholm, recounts his
early career as a cardiologist and his transition to biochemistry, which led to his
Nobel Prize win.
Robert J. Lefkowitz, M.D. is Chancellor’s Distinguished Professor of Medicine and
Professor of Biochemistry and Chemistry at the Duke University Medical Center. He
has bee

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