Differences in Efficacy and Na+ Sensitivity between α2B and α2D Adrenergic Receptors

Abstract

The ability of Na⁺ ions to modulate coupling of α_2B- and α_2D-adrenergic receptors to G proteins was investigated in isolated membranes from transfected PC12 and NIH 3T3 fibroblast cells. The initial rate of epinephrine-stimulated [³⁵S]GTPγS binding was higher for α_2D-receptors (the rat homolog of the α_2A-receptor) in both cell types, whereas both α_2B- and α_2D-receptor responses were higher in PC12 cell membranes. Pertussis toxin completely blocked agonist-stimulated binding. Graded increases in Na⁺ caused a progressive loss of basal GTP binding, indicative of its ability to reduce the level of the active R* state of the receptor. This inhibitory effect of Na⁺ was more pronounced in PC12/α_2B than PC12/α_2D membranes. Epinephrine-stimulated GTP binding in PC12/α_2B membranes was also more sensitive to Na⁺ inhibition than in PC12/α_2D membranes. In saturation [³⁵S]GTPγS binding studies, the presence of Na⁺ reduced apparent GTP affinity, and its effect was greater in PC12/ α_2B membranes, consistent with a greater reduction in the active R* conformation of the receptor. The higher efficacy of epinephrine at α_2D receptors and their lesser sensitivity to Na⁺ are both indicative of a more stable R* state. Together these results suggest that differences in the modulatory influence of Na⁺ within a family of G_i-coupled receptors may reflect differences in the stability of the active R* state.