Do agonists promote rapid internalization of beta-adrenergic receptors?

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Abstract
We used elution of radioligands at low pH to quantitate intracellular .beta.-adrenergic receptors on intact S49 lymphoma cells. We validated this method with respect to cell viability, .beta.-adrenergic receptor integrity, and transferrin receptors on these cells. On control cells, about 15% of the radiolabeled .beta.-adrenergic antagonists [3H]dihydroalprenolol and [125I]iodocyanopindolol specifically bound at 37.degree. C could not be eluted at low pH; these binding sites appear to be intracellular receptors that are inaccessible to the surface-restricted antagonist [3H]CGP-12177 [tritiated (.+-.)-4-(3-t-butylamino-2-hydroxypropoxy)benzimidazole-2-one hydrochloride]. Incubation of cells with the agonist isoproterenol at 37.degree. C for 15 min did not change the number of [3H]dihydroalprenolol binding sites but reduced [3H]CGP-12177 binding sites by 50% or more. However, all specifically bound [3H]CGP-12177 and [3H]dihydroalprenolol were eluted by acid. In addition, the number of acid-elution-resistant [125I]iodocyanopindolol binding sites was not increased in cells coincubated with 1 .mu.M isoproterenol and [125I]iodocyanopindolol for 15 min at 37.degree. C, even though those sites show a loss in apparent affinity for isoproterenol of about 2 orders of magnitude, a loss previously attributed to internalization. We conclude that the early phase of agonist-mediated desensitization of .beta.-adrenergic receptors in S49 cells does not coincide with the movement of receptors to intracellular sites; instead, agonist-modified receptors remain in association with the plasma membrane and are accessible to the extracellar environment. These "redistributed" receptors together with "cell-surface" and "intracellular" receptors represent three classes of .beta.-adrenergic receptors that can be selectively identified in intact target cells.

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