FIGURE 8.

G_s-biased pepducins do not interact with the orthosteric binding site of the β₂AR antagonist but ICL3-9 is sensitive to the inverse agonist ICI118,551. A, HEK 293 cells stably expressing FLAG-β₂AR were incubated with 1 nm [¹²⁵I]cyanopindolol for 1 h at room temperature in HBSS with calcium and magnesium and 0.1% BSA in the presence or absence of 3–30 μm pepducin or 100 μm propranolol. 0.05% DMSO was included in non-pepducin-stimulated cells. The data are represented by the mean of three independent experiments ± S.D. B, HEK 293 cells were stimulated with 1 μm isoproterenol or 5 μm pepducin in the presence of 500 μm IBMX for 10 min at 37 °C with or without a 10-min preincubation with 100 μm propranolol. 0.05% DMSO was included in non-pepducin-stimulated cells. The data are represented by the mean of three independent experiments ± S.D. C, HEK 293 cells were preincubated with ICI118,551 for 10 min and stimulated with 100 nm isoproterenol or 5 μm pepducin in the presence of 500 μm IBMX for 10 min in DMEM with 10% FBS. 0.05% DMSO was included in non-pepducin-stimulated cells. The data are represented by the mean of three independent experiments ± S.D.