Messier T L, Dorman C M, Braüner-Osborne H, Eubanks D, Brann M R
Receptor Technologies Inc., Winooski, VT 05404, USA.
Pharmacol Toxicol. 1995 May;76(5):308-11. doi: 10.1111/j.1600-0773.1995.tb00152.x.
Many receptors stimulate proliferation of NIH 3T3 cells in a ligand dependent fashion. Based on this observation, we developed a high throughput assay of cloned receptor pharmacology. In this assay, receptors are transiently co-expressed with the marker enzyme beta-galactosidase. Receptors that induce cellular proliferation select and amplify the cells that also express the marker, thus the ability of ligands to alter receptor activity are reported as changes in enzyme activity. In the present study, we used this assay to evaluate the ability of agonist ligands to stimulate four cloned receptors. The agonists phenylephrine, carbachol, substance P and nerve growth factor selectively stimulated cells transfected with the alpha-1b adrenergic, m4 muscarinic, NK1 neurokinin and trkA neurotrophin receptors, respectively. These data demonstrate that a high throughput colorimetric assay performed in 96 well plates can be used to evaluate the pharmacology of ligands for cloned receptors belonging to a wide range of functional and pharmacological classes.
许多受体以配体依赖的方式刺激NIH 3T3细胞增殖。基于这一观察结果，我们开发了一种用于克隆受体药理学的高通量检测方法。在该检测中，受体与标记酶β-半乳糖苷酶瞬时共表达。诱导细胞增殖的受体选择并扩增同时表达该标记的细胞，因此配体改变受体活性的能力通过酶活性的变化来报告。在本研究中，我们使用该检测方法来评估激动剂配体刺激四种克隆受体的能力。激动剂去氧肾上腺素、卡巴胆碱、P物质和神经生长因子分别选择性地刺激用α-1b肾上腺素能、m4毒蕈碱、NK1神经激肽和trkA神经营养因子受体转染的细胞。这些数据表明，在96孔板中进行的高通量比色检测可用于评估属于广泛功能和药理类别的克隆受体的配体药理学。
