Regulation of the cellular localization and signaling properties of the alpha(1B)- and alpha(1D)-adrenoceptors by agonists and inverse agonists

The regulation of the cellular distribution and intracellular signaling pro perties of the alpha(1B) - and alpha(1D) - adrenoceptor (alpha(1)-AR) subty pes was examined in stably transfected Rat 1 fibroblasts. In unstimulated c ells, alpha(1B)-AR expression was noted primarily on the cell surface. Trea tment with phenylephrine induced internalization of the alpha(1B)-AR and pr omoted association with arrestin 2. The internalized alpha(1B)-AR colocaliz ed with the transferrin receptor, an endosomal marker. In unstimulated fibr oblasts, the alpha(1D)-AR was detected in a perinuclear orientation and was colocalized with arrestin 2 in a compartment also containing the transferr in receptor. After treatment with prazosin, which exhibits inverse agonist properties, the alpha(1D)-AR was redistributed from intracellular sites to the cellular periphery and was no longer associated with the transferrin re ceptor or arrestin 2. alpha(1D)-AR-expressing cells exhibited a high degree of basal activity for both inositol phosphate formation and extracellular signal regulated kinase (ERK), which was reduced by treatment with prazosin . In these cells, phenylephrine induced a dose-dependent increase in inosit ol phosphate formation but had no effect on ERK activity. In alpha(1B)-AR-e xpressing cells, phenylephrine stimulated both inositol phosphate formation and ERK activity. These data show that: 1) there are differences in the ce llular localization of the alpha(1)-AR subtypes; 2) the alpha(1B)-AR exhibi ts expected G protein-coupled receptor activity regarding cellular localiza tion, agonist-mediated internalization, and coupling to second messengers; and 3) the alpha(1D)-AR is constitutively active and, as a result, is local ized to intracellular compartments involved in receptor recycling.