Hopkinson H E, Latif M L, Hill S J
Institute of Cell Signalling, Queen's Medical Centre, Nottingham NG7 2UH. School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham NG7 2UH.
Br J Pharmacol. 2000 Sep;131(1):124-30. doi: 10.1038/sj.bjp.0703535.
Constitutive activity of the beta(2)-adrenoceptor, which is sensitive to inhibition by an inverse agonist such as ICI 118551, has been readily demonstrated in recombinant systems expressing constitutively-active mutant receptors or over-expressing the wild-type beta(2)-adrenoceptor. Here we demonstrate the presence of constitutive beta(2)-adrenoceptor activity in BC3H1 cells which endogenously express this receptor. In BC3H1 cells, only ICI 118551 behaved as an inverse agonist at beta(2)-adrenoceptors, while propranolol, ICI 118551, atenolol and, to a lesser extent, alprenolol exhibited inverse agonism in CHO-beta(2)4 cells transfected with cDNA for the human beta(2)-adrenoceptor (310 fmol. mg protein(-1)). The level of expression of beta2-adrenoceptors in BC3H1 cells was not high (78 fmol.mg protein-1) and the efficiency of receptor - effector coupling in this cell line was much lower than in the recombinant CHO-beta(2)4 cells (as judged by the partial agonist nature of both salbutamol and clenbuterol). ICI 118551 (log K(D)-9.73+/-0.07) and propranolol (log K(D)-9.25+/-0.12) both behaved as conventional competitive antagonists of isoprenaline-stimulated cyclic AMP accumulation in high expressing CHO-beta(2)4 cells. In contrast, ICI 118551 appeared to act as a non-competitive antagonist in BC3H1 cells and in low expressing CHO-beta(2)6 cells (50 fmol.mg protein(-1)). This non-competitive effect of ICI 118551 in BC3H1 cells was also observed when either salbutamol was used as agonist, or the incubation period with isoprenaline was extended to 30 min. The possibility that these effects of ICI 118551 are due to an interaction with different affinity states (R, R* and R') of the receptor is discussed.
β₂ - 肾上腺素能受体的组成性活性对如ICI 118551等反向激动剂的抑制敏感,在表达组成性活性突变受体或过表达野生型β₂ - 肾上腺素能受体的重组系统中已很容易得到证实。在此我们证明了内源性表达该受体的BC3H1细胞中存在组成性β₂ - 肾上腺素能受体活性。在BC3H1细胞中,只有ICI 118551在β₂ - 肾上腺素能受体上表现为反向激动剂,而普萘洛尔、ICI 118551、阿替洛尔以及程度稍轻的烯丙洛尔在转染了人β₂ - 肾上腺素能受体cDNA(310 fmol·mg蛋白⁻¹)的CHO - β₂4细胞中表现出反向激动作用。BC3H1细胞中β₂ - 肾上腺素能受体的表达水平不高(78 fmol·mg蛋白⁻¹),并且该细胞系中受体 - 效应器偶联效率远低于重组CHO - β₂4细胞(通过沙丁胺醇和克伦特罗的部分激动剂性质判断)。ICI 118551(log K(D) - 9.73 ± 0.07)和普萘洛尔(log K(D) - 9.25 ± 0.12)在高表达CHO - β₂4细胞中均表现为异丙肾上腺素刺激的环磷酸腺苷积累的传统竞争性拮抗剂。相反,ICI 118551在BC3H1细胞和低表达CHO - β₂6细胞(50 fmol·mg蛋白⁻¹)中似乎表现为非竞争性拮抗剂。当使用沙丁胺醇作为激动剂或将与异丙肾上腺素的孵育时间延长至30分钟时,也观察到ICI 118551在BC3H1细胞中的这种非竞争性作用。讨论了ICI 118551的这些作用是否由于与受体的不同亲和力状态(R、R*和R')相互作用所致。