Agonist-induced β-adrenergic receptor internalization on intact human mononuclear leukocytes: Effect of temperature of mononuclear leukocyte separation

The hydrophilic ligand ³H-CGP 12177 was used to measure β-adrenergic receptors on intact human mononuclear leukocytes (MNLs). A single homogeneous class of receptor sites was found, with K_D value of 0.71 ± 0.04 nmol/L and B_max, of 3.0 ± 0.4 fmol/10⁶ cells (mean ± SEM; n = 12). The receptor affinity (K_D) and density (B_max) were similar when measured on MNLs, purified lymphocytes, and a T-lymphocyte enriched population from the same individual. Preincubation of intact MNLs with 1 μmol/L isoproterenol at 37 ° C for 20 minutes reduced the number of surface receptors, measured by ³H-CGP 12177 binding at 4 ° C for 20 hours, by ~70% (receptor internalization) without affecting K_D. This effect was reversible, and surface receptors completely reappeared when binding was investigated at 37 ° C for 40 minutes. Receptor internalization was similar when either isolated MNLs or whole blood was incubated with isoproterenol. Agonistinduced receptor internalization was stable during MNL isolation from whole blood at 4 ° C but was partially or completely lost from MNLs prepared at 20 ° C.