Abstract
Although ligand-free, constitutive β2-adrenergic receptor (AR) signaling has been demonstrated in naive cell lines and in transgenic mice overexpressing cardiac β2-AR, it is unclear whether the dominant cardiac β-AR subtype, β1-AR, shares the ability of spontaneous activation. In the present study, we expressed human β1- or β2-AR via recombinant adenoviral infection in ventricular myocytes isolated from β1β2-AR double knockout mice, creating pure β1-AR and β2-AR systems with variable receptor densities. A contractile response to a nonselective β-AR agonist, isoproterenol, was absent in double knockout mouse myocytes but was fully restored after adenoviral β1-AR or adenoviral β2-AR infection. Increasing the titer of adenoviral vectors (multiplicity of infection 10-1000) led to a dose-dependent expression of β1- or β2-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 β1-AR and β2-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, β2-AR expression resulted in graded increases in baseline cAMP and contractility up to 428% and 233% of control, respectively, at the maximal β2-AR density. These effects were specifically reversed by a β2-AR inverse agonist, ICI 118,551 (10-7 M). In contrast, overexpression of β1-AR, even at a greater density, failed to enhance either basal cAMP or contractility; the alleged β1-AR inverse agonist, CGP 20712A (10-6 M), had no significant effect on basal contraction in these cells. Thus, we conclude that acute β2-AR overexpression in cardiac myocytes elicits significant physiological responses due to spontaneous receptor activation; however, this property is β-AR subtype specific because β1-AR does not exhibit agonist-independent spontaneous activation.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 887-894 |
| Number of pages | 8 |
| Journal | Molecular Pharmacology |
| Volume | 58 |
| Issue number | 5 |
| State | Published - 2000 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Medicine(all)
- Molecular Medicine
- Pharmacology
Cite this
Spontaneous activation of β2- but not β1-adrenoceptors expressed in cardiac myocytes from β1β2 double knockout mice. / Zhou, Ying Ying; Yang, Dongmei; Zhu, Wei Zhong; Zhang, Sheng Jun; Wang, Ding Ji; Rohrer, Dan K.; Devic, Eric; Kobilka, Brian K.; Lakatta, Edward; Cheng, Heping; Xiao, Rui Ping.
In: Molecular Pharmacology, Vol. 58, No. 5, 2000, p. 887-894.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Spontaneous activation of β2- but not β1-adrenoceptors expressed in cardiac myocytes from β1β2 double knockout mice
AU - Zhou, Ying Ying
AU - Yang, Dongmei
AU - Zhu, Wei Zhong
AU - Zhang, Sheng Jun
AU - Wang, Ding Ji
AU - Rohrer, Dan K.
AU - Devic, Eric
AU - Kobilka, Brian K.
AU - Lakatta, Edward
AU - Cheng, Heping
AU - Xiao, Rui Ping
PY - 2000
Y1 - 2000
N2 - Although ligand-free, constitutive β2-adrenergic receptor (AR) signaling has been demonstrated in naive cell lines and in transgenic mice overexpressing cardiac β2-AR, it is unclear whether the dominant cardiac β-AR subtype, β1-AR, shares the ability of spontaneous activation. In the present study, we expressed human β1- or β2-AR via recombinant adenoviral infection in ventricular myocytes isolated from β1β2-AR double knockout mice, creating pure β1-AR and β2-AR systems with variable receptor densities. A contractile response to a nonselective β-AR agonist, isoproterenol, was absent in double knockout mouse myocytes but was fully restored after adenoviral β1-AR or adenoviral β2-AR infection. Increasing the titer of adenoviral vectors (multiplicity of infection 10-1000) led to a dose-dependent expression of β1- or β2-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 β1-AR and β2-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, β2-AR expression resulted in graded increases in baseline cAMP and contractility up to 428% and 233% of control, respectively, at the maximal β2-AR density. These effects were specifically reversed by a β2-AR inverse agonist, ICI 118,551 (10-7 M). In contrast, overexpression of β1-AR, even at a greater density, failed to enhance either basal cAMP or contractility; the alleged β1-AR inverse agonist, CGP 20712A (10-6 M), had no significant effect on basal contraction in these cells. Thus, we conclude that acute β2-AR overexpression in cardiac myocytes elicits significant physiological responses due to spontaneous receptor activation; however, this property is β-AR subtype specific because β1-AR does not exhibit agonist-independent spontaneous activation.
AB - Although ligand-free, constitutive β2-adrenergic receptor (AR) signaling has been demonstrated in naive cell lines and in transgenic mice overexpressing cardiac β2-AR, it is unclear whether the dominant cardiac β-AR subtype, β1-AR, shares the ability of spontaneous activation. In the present study, we expressed human β1- or β2-AR via recombinant adenoviral infection in ventricular myocytes isolated from β1β2-AR double knockout mice, creating pure β1-AR and β2-AR systems with variable receptor densities. A contractile response to a nonselective β-AR agonist, isoproterenol, was absent in double knockout mouse myocytes but was fully restored after adenoviral β1-AR or adenoviral β2-AR infection. Increasing the titer of adenoviral vectors (multiplicity of infection 10-1000) led to a dose-dependent expression of β1- or β2-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 β1-AR and β2-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, β2-AR expression resulted in graded increases in baseline cAMP and contractility up to 428% and 233% of control, respectively, at the maximal β2-AR density. These effects were specifically reversed by a β2-AR inverse agonist, ICI 118,551 (10-7 M). In contrast, overexpression of β1-AR, even at a greater density, failed to enhance either basal cAMP or contractility; the alleged β1-AR inverse agonist, CGP 20712A (10-6 M), had no significant effect on basal contraction in these cells. Thus, we conclude that acute β2-AR overexpression in cardiac myocytes elicits significant physiological responses due to spontaneous receptor activation; however, this property is β-AR subtype specific because β1-AR does not exhibit agonist-independent spontaneous activation.
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M3 - Article
VL - 58
SP - 887
EP - 894
JO - Molecular Pharmacology
JF - Molecular Pharmacology
SN - 0026-895X
IS - 5
ER -