Effects of a range of beta(2) adrenoceptor agonists on changes in intracellular cyclic AMP and on cyclic AMP driven gene expression in cultured humanairway smooth muscle cells

1 The effects of the selective beta(2) adrenoceptor agonists salbutamol, te rbutaline and salmeterol and the non-selective beta adrenoceptor agonist is oprenaline on [H-3]-cyclic AMP formation and cyclic AMP response element (C RE) driven luciferase expression, assessed using the construct p6CRE/luc, w ere studied in primary cultures of human airway smooth muscle (HASM) cells. 2 Optimal transfection conditions for transient expression of pGL3 Control were 4 mu g DNA/well(-1) in a 6 well plate and 1.8 mu l Transfectam/mu g DN A. Expression was maximal at 48-72 h. 3 Salbutamol (maximum response 19%, EC50 0.6 mu M), terbutaline (maximum re sponse 38%, EC50 2.3 mu M) and salmeterol (maximum response 18%, EC50 0.001 2 mu M) were all partial agonists for cyclic AMP formation compared with is oprenaline (EC50 0.08 mu M). However, all of the beta(2) adrenoceptor agoni sts produced increases in CRE-driven luciferase activity, in cultured HASM transfected with the vector p6CRE/luc, which were equivalent or greater (sa lmeterol) than those seen with isoprenaline. 4 Both salbutamol and salmeterol were more potent at increasing luciferase expression than in elevating cyclic AMP levels in these cells. The potency ratios (EC50 ((cyclic AMP))/EC50 ((LUC))) for the agents studied were isopr enaline: 0.2 fold, terbutaline: 3 fold, salbutamol: 24 fold, salmeterol: 38 fold. 5 These data suggest that important quantitative differences exist in the a bility of beta(2) adrenoceptor agonists to increase whole cell cyclic AMP l evels in airway smooth muscle and to drive gene expression via a CRE-driven mechanism.