Evidence for intravesicular beta-adrenergic receptors in membrane fractions from desensitized cells: binding of the hydrophilic ligand CGP-12177 only in the presence of alamethicin.

Two fractions of beta-adrenergic receptors have been separated from desensitized C62B cells by centrifugation on a sucrose gradient. Using two beta-adrenergic receptor (beta AR) antagonists, 125I-iodopindolol and (3H)CGP-12177, the different binding characteristics of these fractions have been investigated. While iodopindolol binds to beta AR both in a light (30% sucrose) and a heavy (45-50% sucrose) peak, CGP-12177 binds only to those in the heavy peak. Incubation of these two gradient fractions with the antibiotic alamethicin selectively increases the binding of CGP-12177 only in the light peak. This result suggests that the desensitization-induced light peak beta AR exist on the inside of vesicles. The hydrophilic ligand CGP-12177 is capable of reaching these receptors only through alamethicin-formed pores.