Abstract

We recently reported that α_2A-adrenoceptor (AR) desensitization and down-regulation occurs after 24-h treatment with epinephrine (EPI) (0.3 μM) in BE(2)-C cells that express both α₂- and β₂-ARs. The same concentration of norepinephrine (NE) has no effect. The effect of EPI is prevented by β₂-AR blockade and is associated with an increase in G protein-coupled receptor kinase 3 (GRK3) expression. Because differences in agonist-induced down-regulation of the α_2A-versus α_2B-ARs have been reported, the present study examines the effects of simultaneous activation of α_2B- and β₂-ARs on α_2B-AR number and signaling. We studied NG108 cells that naturally express α_2B-ARs, and BN17 cells, NG108 cells transfected to express the human β₂-AR. In NG108 cells, α_2B-AR desensitization and down-regulation require treatment with 20 μM EPI or NE; GRK expression was not changed. In BN17 cells expressing β₂-ARs, the threshold EPI concentration for α_2B-AR desensitization and down-regulation was reduced to 0.3 μM; 10 μM NE was required for the same effect. Furthermore, 24-h EPI or NE treatments that produced desensitization also resulted in a selective 2-fold up-regulation of GRK3; GRK2 was unchanged. The β-AR antagonist alprenolol (1 μM) and GRK3 antisense (but not sense) DNA blocked 0.3 μM EPI- and 10 μM NE-induced desensitization and down-regulation of the α_2B-AR as well as GRK3 up-regulation. In conclusion, simultaneous activation of α_2B- and β₂-ARs results in a 67-fold decrease in the threshold concentration of EPI required for α_2B-AR down-regulation. This lower threshold for down-regulation is associated with α_2B- and β₂-AR dependent up-regulation of GRK3 expression.