Salmeterol-induced desensitization, internalization and phosphorylation of the human β₂-adrenoceptor

1 Partial agonists of the β₂-adrenoceptor which activate adenylyl cyclase are widely used as bronchodilators for the relief of bronchoconstriction accompanying many disease conditions, including bronchial asthma. The bronchodilator salmeterol has both a prolonged duration of action in bronchial tissue and the ability to reassert this activity following the temporary blockade of human β₂-adrenoceptors with antagonist. 2 We have compared the activation and desensitization of human β₂-adrenoceptor stimulation of adenylyl cyclase induced by salmeterol, adrenaline and salbutamol in a human lung epithelial line, BEAS-2B, expressing β₂-adrenoceptor levels of 40 - 70 fmol mg ¹, and in human embryonic kidney (HEK) 293 cell lines expressing 2 - 10 pmol mg^-1. The efficacy observed for the stimulation of adenylyl cyclase by salmeterol was only ≅ 10% of that observed for adrenaline in BEAS-2B cells expressing low levels of β₂-adrenoceptor, but similar to adrenaline in HEK 293 cells expressing very high levels of receptors. Salmeterol pretreatment of these cells induced a rapid and stable activation of adenylyl cyclase activity which resisted extensive washing and β₂-adrenoceptor antagonist blockade, consistent with binding to a receptor exosite and or to partitioning into membrane lipid. 3 The desensitization and internalization β₂-adrenoceptors induced by the partial agonists salmeterol and salbutamol were considerably reduced relative to the action of adrenaline. Consistent with these observations, the initial rate of phosphorylation of the receptor induced by salmeterol and salbutamol was much reduced in comparison to adrenaline. 4 Our data suggest that the reduction in the rapid phase of desensitization of β₂-adrenoceptors after treatment with salmeterol or salbutamol is caused by a decrease in the rate of β₂-adrenoceptor kinase (βARK) phosphorylation and internalization. In contrast, the rate of cyclic AMP-dependent protein kinase (PKA)-mediated phosphorylation by these partial agonists appears to be similar to adrenaline.