Regulation of the Cellular Localization and Signaling Properties of the α_1B- and α_1D-Adrenoceptors by Agonists and Inverse Agonists

Abstract

The regulation of the cellular distribution and intracellular signaling properties of the α_1B- and α_1D- adrenoceptor (α₁-AR) subtypes was examined in stably transfected Rat 1 fibroblasts. In unstimulated cells, α_1B-AR expression was noted primarily on the cell surface. Treatment with phenylephrine induced internalization of the α_1B-AR and promoted association with arrestin 2. The internalized α_1B-AR colocalized with the transferrin receptor, an endosomal marker. In unstimulated fibroblasts, the α_1D-AR was detected in a perinuclear orientation and was colocalized with arrestin 2 in a compartment also containing the transferrin receptor. After treatment with prazosin, which exhibits inverse agonist properties, the α_1D-AR was redistributed from intracellular sites to the cellular periphery and was no longer associated with the transferrin receptor or arrestin 2. α_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 inositol phosphate formation but had no effect on ERK activity. In α_1B -AR-expressing cells, phenylephrine stimulated both inositol phosphate formation and ERK activity. These data show that: 1) there are differences in the cellular localization of the α₁-AR subtypes; 2) the α_1B-AR exhibits expected G protein-coupled receptor activity regarding cellular localization, agonist-mediated internalization, and coupling to second messengers; and 3) the α_1D-AR is constitutively active and, as a result, is localized to intracellular compartments involved in receptor recycling.