TABLE 1

Agonist binding to wild-type and CAM β 2AR and stimulation of cAMP accumulation by wild-type β 2AR

Competition binding assays were performed on HEK 293 membranes stably expressing wild-type or CAM β 2AR, and cAMP accumulation studies were performed in intact HEK 293 cells stably expressing the wild-type β 2AR as described under Materials and Methods. The Ki values were determined according to the method of Cheng and Prusoff using the IC50 values from the competition binding isotherms (Cheng and Prusoff, 1973). The affinity shift is the ratio between the Ki values for WT and CAM β 2AR. Ki/KiPE is the ratio between the Ki values for phenethylamine and each drug. The potencies (EC50 values) of agonists to stimulate cAMP accumulation were determined by nonlinear regression analysis. Values are presented as mean ± S.E. from three to eight independent experiments.

[3H]CGP-12177 Binding
Wild Type CAM cAMP Accumulation
−logKi ± S.E. Ki/KiPE −logKi ± S.E. Ki/KiPE Affinity Shift −logEC50 ± S.E. % MaximalStimulation
Epinephrine 7.16 ± 0.08 407 8.89 ± 0.13 16218 54 7.00 ± 0.11 100
Phenylephrine 5.62 ± 0.08 12 6.85 ± 0.10 148 17 5.65 ± 0.09 40 ± 5
Synephrine 5.00 ± 0.07 2.8 6.25 ± 0.18 37 18 4.98 ± 0.09 24 ± 3
Halostachine 5.08 ± 0.06 3.4 5.90 ± 0.09 17 6.6 4.73 ± 0.14 19 ± 4
Norepinephrine 5.40 ± 0.11 7.1 7.29 ± 0.07 407 78 5.61 ± 0.07 101 ± 10
Dopamine 4.35 ± 0.07 0.6 5.33 ± 0.05 4.5 10 4.50 ± 0.10 46 ± 7
Methyldopamine 5.37 ± 0.04 6.6 7.20 ± 0.09 331 68 5.40 ± 0.11 88 ± 10
Norphenylephrine 4.64 ± 0.04 1.2 5.35 ± 0.08 4.7 5.1 N.D. N.D.
Octopamine 4.81 ± 0.16 1.8 5.09 ± 0.17 2.6 1.9 N.D. N.D.
Hydroxyphenethylamine 4.53 ± 0.05 1.0 4.88 ± 0.04 1.6 2.2 N.D. N.D.
Methylphenethylamine 4.18 ± 0.07 0.4 4.54 ± 0.02 0.7 2.3 N.D. N.D.
Phenethylamine 4.55 ± 0.38 1.0 4.68 ± 0.42 1.0 1.3 N.D. N.D.
  • N.D., not determined.