| pA2 online © Copyright 2004 The British Pharmacological Society |
071P
University of Bath Summer Meeting July 2004 |
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Site of action of β-adrenoceptor agonists at the human β1-adrenoceptor Jillian G Baker. Institute of Cell Signalling, University of Nottingham, Queen’s Medical Centre, Nottingham NG7 2UH. UK |
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Print abstractThe human β1 receptor has recently been proposed to have two active agonist conformations: the classical agonist isoprenaline activates the “catecholamine site” whereas CGP 12177 has agonist activity best explained by action at a separate “secondary site” (Konkar et al., 2000; Baker et al., 2003a). Agonists binding to the catecholamine site elicit responses very sensitive to antagonism by the selective β1-antagonist CGP 20712A whereas agonist actions occurring via the secondary site are relatively resistant to CGP 20712A antagonism. The aim of this study was to determine through which site the agonist actions of a range of classical β-agonists occurs.
CHO-K1 cells stably expressing both the human β1-adrenoceptor (at 1100fmol/mg protein) and a SPAP reporter gene under the transcriptional control of six cAMP response elements were used. The rate of SPAP secretion over 1 hour, 5-6 hours after the addition of agonist, was measured as previously described (Baker et al., 2003a).
Isoprenaline stimulated an increase in CRE-SPAP secretion (log EC50 = -8.48 ± 0.06; 2.3 ± 0.3 fold over basal; n=6) that was antagonised by CGP20712A to yield a log KD value of -9.25 ± 0.14 (n=6). The responses to the endogenous agonists adrenaline and noradrenaline were similarly antagonised. However, most other β-agonists stimulated responses that were more sensitive to CGP 20712A blockade.
agonist |
Log EC50 |
% isoprenaline |
Log KD CGP 20712A |
n |
cimaterol |
-8.81 ± 0.12 |
97.6 ± 5.6 |
-9.86 ± 0.10 |
8 |
formoterol |
-8.87 ± 0.09 |
91.6 ± 2.3 |
-9.78 ± 0.08 |
9 |
terbutaline |
-6.18 ± 0.09 |
84.5 ± 5.2 |
-9.76 ± 0.05 |
7 |
BRL 37344 |
-6.84 ± 0.14 |
79.8 ± 6.8 |
-9.70 ± 0.16 |
7 |
salbutamol |
-6.22 ± 0.17 |
86.5 ± 6.2 |
-9.64 ± 0.13 |
7 |
tulobuterol |
-6.72 ± 0.20 |
78.3 ± 3.0 |
-9.62 ± 0.10 |
7 |
clenbuterol |
-7.62 ± 0.14 |
75.4 ± 5.8 |
-9.56 ± 0.16 |
9 |
fenoterol |
-7.83 ± 0.15 |
88.1 ± 3.3 |
-9.56 ± 0.11 |
8 |
isoprenaline |
-8.48 ± 0.06 |
100 |
-9.25 ± 0.14 |
6 |
noradrenaline |
-7.67 ± 0.13 |
93.7 ± 5.7 |
-9.17 ± 0.12 |
8 |
adrenaline |
-7.46 ± 0.08 |
94.8 ± 4.2 |
-9.16 ± 0.07 |
7 |
dobutamine |
-7.10 ± 0.11 |
93.4 ± 2.4 |
-9.09 ± 0.11 |
8 |
CGP 12177 |
-8.63 ± 0.13 |
75.4 ± 3.4 |
-7.73 ± 0.11 |
6 |
Table. Log EC 50 values, % maximum of each agonist response relative to the isoprenaline max in each experiment, and log KD values for CGP 20712A for each agonist expressed as mean ± S.E.M. for n separate experiments.
This study shows that the responses elicited by many classical β-adrenoceptor agonists were very sensitive to CGP 20712A antagonism suggesting that these responses were occurring via the catecholamine site. However, as most responses were more sensitive to antagonism than isoprenaline, it remains to be established whether this is related to ligand efficacy and receptor phosphorylation in a manner to that previously reported at the human β2-adrenoceptor (Baker et al., 2003b).
Baker J.G. et al., (2003a). Mol. Pharmacol.63, 1312-1321
Baker J.G. et al., (2003b). Mol. Pharmacol. 64, 679-688
Konkar A.A. et al., (2000) J. Pharmacol. Exp. Ther.294, 923-932