TABLE 2

Log EC50 values and % maximum response to isoprenaline for CRE-SPAP production for the agonists cimaterol and CGP12177 at the human β1-WT and chimeric β1/β2-adrenoceptors

Mutations in the β1-WT mean that each TM region in turn is mutated to that of the β2-WT (see Table 1). Log KB values for several antagonists for inhibition of the cimaterol and CGP12177 responses were determined. These data were obtained from stable cell lines, and n in the table refers to the number of separate experiments. The table shows that the most consistent and highly significant difference between the β1-WT receptor and all of the TM-swap mutants is that obtained from β1-TM4 when CGP12177 is the agonist. As CGP12177 agonist responses occur at the secondary low-affinity conformation of the β1-WT receptor, this suggests that the β1-TM4 receptor mutations alter this conformation in some way.

Cimaterol Log EC50% IsoprenalinenLog KB CGP20712AnLog KB BisoprololnLog KB ICI118551nLog KB PropranololnLog KB CarvedilolnLog KB CGP12177n
Cimaterol as agonist: β1 stable cell lines
β1-WT−8.25 ± 0.1495.8 ± 2.98−9.18 ± 0.074−8.48 ± 0.175−6.96 ± 0.045−8.52 ± 0.0812−9.89 ± 0.135−9.55 ± 0.128
β1-TM1−7.91 ± 0.05*96.5 ± 3.311−9.35 ± 0.109−8.65 ± 0.125−6.96 ± 0.059−8.66 ± 0.0716−9.88 ± 0.085−9.77 ± 0.0715
β1-TM2−8.19 ± 0.0398.9 ± 3.015−8.78 ± 0.128−8.15 ± 0.077−7.47 ± 0.03*14−8.52 ± 0.0324−9.53 ± 0.188−9.90 ± 0.0917
β1-TM3−8.82 ± 0.03*102.4 ± 2.59−9.34 ± 0.129−8.43 ± 0.074−7.31 ± 0.04*10−8.53 ± 0.0615−9.92 ± 0.165−9.49 ± 0.0613
β1-TM4−8.15 ± 0.0299.2 ± 1.810−9.50 ± 0.1311−8.81 ± 0.045−8.07 ± 0.06*9−8.99 ± 0.04*15−9.81 ± 0.126−9.63 ± 0.0817
β1-TM5−8.48 ± 0.05103.0 ± 1.79−9.09 ± 0.0411−7.91 ± 0.035−7.01 ± 0.0611−8.71 ± 0.0417−9.96 ± 0.096−9.68 ± 0.0917
β1-TM6−8.59 ± 0.03*103.2 ± 2.210−8.52 ± 0.078−8.10 ± 0.155−7.92 ± 0.05*12−8.90 ± 0.05*18−10.13 ± 0.096−9.58 ± 0.0614
β1-TM7−8.54 ± 0.05106.1 ± 2.510−8.45 ± 0.087−8.19 ± 0.065−7.80 ± 0.05*12−8.95 ± 0.05*18−10.06 ± 0.033−9.39 ± 0.0513
CGP12177 Log EC50% IsoprenalinenLog KB CGP20712AnLog KB BisoprololnLog KB ICI118551nLog KB PropranololnLog KB Carvediloln
CGP12177 as agonist: β1 stable cell lines
β1-WT−8.18 ± 0.0873.8 ± 5.610−7.16 ± 0.0614−5.83 ± 0.144−5.90 ± 0.159−6.18 ± 0.0613−7.25 ± 0.187
β1-TM1−7.83 ± 0.0553.7 ± 3.910−7.41 ± 0.1311−5.77 ± 0.184−5.79 ± 0.093−6.55 ± 0.1213−7.72 ± 0.137
β1-TM2−8.27 ± 0.0953.5 ± 3.914−7.13 ± 0.0813−6.72 ± 0.187−5.87 ± 0.056−6.87 ± 0.15*18−8.52 ± 0.15*,**7
β1-TM3−8.07 ± 0.0370.5 ± 3.69−7.45 ± 0.0512−5.83 ± 0.134−5.83 ± 0.097−6.31 ± 0.0516−7.45 ± 0.096
β1-TM4−9.27 ± 0.06*,**29.2 ± 1.4*,**10−9.49 ± 0.08*,**9−8.83 ± 0.21*,**7−7.93 ± 0.11*,**11−8.85 ± 0.09*,**15−9.73 ± 0.21*,**7
β1-TM5−7.91 ± 0.0360.1 ± 3.410−6.68 ± 0.04*11−5.31 ± 0.135−5.71 ± 0.158−6.23 ± 0.0915−7.53 ± 0.167
β1-TM6−7.83 ± 0.02*63.5 ± 2.811−5.97 ± 0.08*,**12−5.22 ± 0.143−5.79 ± 0.0411−6.05 ± 0.0619−7.24 ± 0.187
β1-TM7−7.08 ± 0.02*,**68.5 ± 3.811−6.86 ± 0.0510−5.67 ± 0.055−6.67 ± 0.03*,**13−6.42 ± 0.0616−7.35 ± 0.126
  • * P < 0.001; one-way ANOVA with post hoc Newman–Keuls comparing values from the mutant receptors with those obtained from the β1-WT. Thus, the log EC50 for CGP12177 at β1-TM4 is different from that obtained from the β1-WT with P < 0.001; **P < 0.001; one-way ANOVA with post hoc Newman–Keuls comparing each value with all other values in this set. Thus, the log EC50 value for CGP12177 at β1-TM4 is different from those obtained for β1-WT, β1-TM1, β1-TM2, β1-TM3, β1-TM5, β1-TM6, and β1-TM7, with P < 0.001 in all cases.