Kinetic analysis of agonist-induced down-regulation of the beta 2-adrenergic receptor in BEAS-2B cells reveals high-and low-affinity components

We examined the interrelationships of internalization and downregulation of the beta(2)-adrenergic receptor in response to treatment of the BEAS-2B hu man epithelial cell line with both a series of agonists at high occupancy a nd with various concentrations of fenoterol that gave occupancies from 0.93 to 0.001. We found that the extent of internalization measured after a 30- min treatment increased as a function of coupling efficiency, with ephedrin e, dobutamine, albuterol, fenoterol, and epinephrine giving 0, 7, 17, 48, a nd 55% internalization, respectively. With the exception of dobutamine, the rates of down-regulation (k(deg)) also showed a dependence on agonist coup ling efficiency, giving (in terms of fraction of receptors lost/h) 0.082 wi th ephedrine, 0.250 with dobutamine, 0.148 with albuterol, 0.194 with fenot erol, and 0.212 with epinephrine. Comparison of down-regulation to internal ization showed that weak agonists caused down-regulation in the absence of significant internalization. The extent of internalization caused by fenote rol over a 1000-fold range of occupancy was proportional to agonist occupan cy. However, although no internalization was observed with the low concentr ations (0.2 and 2 nM fenoterol), these concentrations did cause significant down-regulation. Thus, as with partial agonists, it was clear that down-re gulation occurred in the absence of measurable internalization. The kinetic s of agonist-induced down-regulation are consistent with a scheme in which down-regulation proceeds by two pathways; a high-affinity, low-capacity com ponent (EC50 = 0.5 nM) clearly dissociated from internalization and a low-a ffinity, high-capacity component (EC50 = 160 nM) closely correlated with in ternalization.