Fig. 6.

³H-cAMP accumulation in response to CGP12177 (A, C, and E) and pindolol (B, D, and F) in transiently transfected cells expressing β₁-WT (A and B), β₁-TM4 (C and D), and β₂-WT (E and F). The CGP12177 responses were examined in the absence and presence of 3, 10, and 30 nM cimaterol (A and C); and 0.3, 1, and 3 nM cimaterol (E). Bars represent basal ³H-cAMP accumulation, that in response to 10 µM isoprenaline or that in response to 0.3–30 nM cimaterol alone. Data points are mean ± S.E.M. of triplicate determinations. These single experiments are representative of (A) six, (B) eight, (C) four, (D) three, (E) four, and (F) four separate experiments. This figure examines different evidence for the absence or presence of the secondary conformation. At β₁-WT, low concentrations of CGP12177 (e.g., 3 nM, −8.5) inhibit the response to cimaterol, whereas higher concentrations of CGP12177 cause a stimulatory response (e.g., 100 nM, −7 and above). This creates a “dip” in the curve (CGP12177 + fixed concentration of cimaterol) and is strongly suggestive of interaction at two different conformations. This is absent at β₁-TM4 and β₂-WT, suggesting single-conformation interaction. Similarly, the response to pindolol is biphasic at β₁-WT but monophasic at β₁-TM4 and β₂-WT, again suggesting two conformations at β₁-WT but only one conformation at β₁-TM4 and β₂-WT.