RT Journal Article
SR Electronic
T1 Salmeterol Efficacy and Bias in the Activation and Kinase-Mediated Desensitization of β2-Adrenergic Receptors
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP mol.114.096800
DO 10.1124/mol.114.096800
A1 Luis E Gimenez
A1 Faiza Baameur
A1 Sharat J. Vayttaden
A1 Richard B. Clark
YR 2015
UL http://molpharm.aspetjournals.org/content/early/2015/03/17/mol.114.096800.abstract
AB Salmeterol is a long acting β2-adrenergic receptor (β2AR) agonist widely used as a bronchodilator for the treatment of persistent asthma and COPD in conjunction with steroids. Previous studies demonstrated that salmeterol showed weak efficacy for activation of adenylyl cyclase; however, its efficacy in the complex desensitization of β2AR remains poorly understood. In the present work we provide insights into the roles played by GRK/arrestin and PKA in salmeterol-mediated desensitization through BRET studies of liganded-β2AR binding to arrestin, and kinetic studies of cAMP turnover. First, BRET demonstrated a much-reduced efficacy for salmeterol recruitment of arrestin to the β2AR relative to isoproterenol. The ratio of BRETISO/BRETSALM after 5 min stimulation was 20, decreasing to 5 after 35 min, reflecting a progressive decline in BRETISO, and a stable BRETSALM. Second, to assess salmeterol efficacy for functional desensitization, we examined the kinetics of salmeterol-induced cAMP accumulation (0-30 min) in human airway smooth muscle cells (HASM) in the presence and absence of PDE inhibition. Analysis of shaping of cAMP turnover for both agonists demonstrated significant salmeterol desensitization, although reduced relative to isoproterenol. Using an isoproterenol rescue protocol following either short- (10 min) or long-term (2 and 14 hr) salmeterol pretreatments, we found that salmeterol progressively depressed isoproterenol stimulation, but did not prevent subsequent rescue by isoproterenol and additional isoproterenol-mediated desensitization. Our findings reveal a complex efficacy for functional desensitization demonstrating that while salmeterol shows weak efficacy for adenylyl cyclase activation and GRK/arrestin-mediated desensitization, it acts as a strong agonist in highly amplified PKA-mediated events.