Turnover of β1- and β2-adrenergic receptors after down-regulation or irreversible blockade

Kim Neve, P. B. Molinoff

Research output: Contribution to journalArticle

52 Citations (Scopus)

Abstract

The turnover of β1- and β2-adrenergic receptors was measured after both isoproterenol-induced down-regulation and irreversible blockade of receptors. Changes in the density of receptors were quantified using the radioligands 125I-iodopindolol and 125I-iodocyanopindolol. Treatment of intact L6 myoblasts or C6 glioma cells with N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) inactivated β-adrenergic receptors on membranes prepared from these cells. At a concentration of 100 μM EEDQ, more than 90% of β1- and β2-adrenergic receptors were inactivated within 2 hr of treatment. Recovery of β-adrenergic receptors on intact cells after inactivation by EEDQ required more than 24 hr and was prevented by cycloheximide, an inhibitor of protein synthesis. The kinetics of recovery of the density of receptors were analyzed in terms of a model that allows estimation of the rate constants for receptor appearance in and disappearance from the membrane, assuming that the rate of appearance of receptors is constant and the rate of disappearance of receptors is proportional to the number of receptors. β2-Adrenergic receptors on L6 myoblasts were incorporated into the membrane at a rate of 28 fmol/mg or protein/hr and had a half-life of 12.6 hr. On C6 glioma cells, β1- and β2-adrenergic receptors appeared at rates of 13.3 and 6.6 fmol/mg of protein/hr, respectively, with half-lives of 9.4 and 6.4 hr. Recovery of receptors on C6 cells after isoproterenol-induced down-regulation was inhibited by cycloheximide. The rate of recovery of β1- and β2-adrenergic receptors was reduced after treatemnt with isoproterenol for 8 hr when compared to recovery after treatment with EEDQ. The major effect of treatment with isoproterenol was a persistent decrease in the rate of appearance of β1- and β2-adrenergic receptors (rate of synthesis and insertion into the membrane after treatment with isoproterenol = 4.0 fmol/mg of protein/hr). Since treatment with isoproterenol did not alter the rate of cell division or total protein synthesis, the isoproterenol-induced alteration was probably a specific effect on the rate of synthesis of β-adrenergic receptors.

Original languageEnglish (US)
Pages (from-to)104-111
Number of pages8
JournalMolecular Pharmacology
Volume30
Issue number2
StatePublished - 1986
Externally publishedYes

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Adrenergic Receptors
Down-Regulation
Isoproterenol
Myoblasts
Cycloheximide
Glioma
Membranes
Proteins
Iodocyanopindolol
Protein Synthesis Inhibitors
Cell Division
Half-Life
Cell Membrane

ASJC Scopus subject areas

  • Pharmacology

Cite this

Turnover of β1- and β2-adrenergic receptors after down-regulation or irreversible blockade. / Neve, Kim; Molinoff, P. B.

In: Molecular Pharmacology, Vol. 30, No. 2, 1986, p. 104-111.

Research output: Contribution to journalArticle

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abstract = "The turnover of β1- and β2-adrenergic receptors was measured after both isoproterenol-induced down-regulation and irreversible blockade of receptors. Changes in the density of receptors were quantified using the radioligands 125I-iodopindolol and 125I-iodocyanopindolol. Treatment of intact L6 myoblasts or C6 glioma cells with N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) inactivated β-adrenergic receptors on membranes prepared from these cells. At a concentration of 100 μM EEDQ, more than 90{\%} of β1- and β2-adrenergic receptors were inactivated within 2 hr of treatment. Recovery of β-adrenergic receptors on intact cells after inactivation by EEDQ required more than 24 hr and was prevented by cycloheximide, an inhibitor of protein synthesis. The kinetics of recovery of the density of receptors were analyzed in terms of a model that allows estimation of the rate constants for receptor appearance in and disappearance from the membrane, assuming that the rate of appearance of receptors is constant and the rate of disappearance of receptors is proportional to the number of receptors. β2-Adrenergic receptors on L6 myoblasts were incorporated into the membrane at a rate of 28 fmol/mg or protein/hr and had a half-life of 12.6 hr. On C6 glioma cells, β1- and β2-adrenergic receptors appeared at rates of 13.3 and 6.6 fmol/mg of protein/hr, respectively, with half-lives of 9.4 and 6.4 hr. Recovery of receptors on C6 cells after isoproterenol-induced down-regulation was inhibited by cycloheximide. The rate of recovery of β1- and β2-adrenergic receptors was reduced after treatemnt with isoproterenol for 8 hr when compared to recovery after treatment with EEDQ. The major effect of treatment with isoproterenol was a persistent decrease in the rate of appearance of β1- and β2-adrenergic receptors (rate of synthesis and insertion into the membrane after treatment with isoproterenol = 4.0 fmol/mg of protein/hr). Since treatment with isoproterenol did not alter the rate of cell division or total protein synthesis, the isoproterenol-induced alteration was probably a specific effect on the rate of synthesis of β-adrenergic receptors.",
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