Knight Antony R, Misra Anil, Quirk Kathleen, Benwell Karen, Revell Dean, Kennett Guy, Bickerdike Mike
Department of Molecular Pharmacology, Vernalis Research, Oakdene Court, 613 Reading Road, Winnersh, Wokingham, RG41 5UA, UK.
Naunyn Schmiedebergs Arch Pharmacol. 2004 Aug;370(2):114-23. doi: 10.1007/s00210-004-0951-4. Epub 2004 Jul 30.
In the present study we compared the affinity of various drugs for the high affinity "agonist-preferring" binding site of human recombinant 5-HT(2A), 5-HT(2B) and 5-HT(2C) receptors stably expressed in monoclonal mammalian cell lines. To ensure that the "agonist-preferring" conformation of the receptor was preferentially labelled in competition binding experiments, saturation analysis was conducted using antagonist and agonist radiolabels at each receptor. Antagonist radiolabels ([3H]-ketanserin for 5-HT(2A) receptor and [3H]-mesulergine for 5-HT(2B) and 5-HT(2C) receptor) bound to a larger population of receptors in each preparation than the corresponding agonist radiolabel ([125I]-DOI for 5-HT(2A) receptor binding and [3H]-5-HT for 5-HT(2B) and 5-HT(2C) receptor binding). Competition experiments were subsequently conducted against appropriate concentrations of the agonist radiolabels bound to the "agonist-preferring" subset of receptors in each preparation. These studies confirmed that there are a number of highly selective antagonists available to investigate 5-HT2 receptor subtype function (for example, MDL 100907, RS-127445 and RS-102221 for 5-HT(2A), 5-HT(2B) and 5-HT(2C) receptors respectively). There remains, however, a lack of highly selective agonists. (-)DOI is potent and moderately selective for 5-HT(2A) receptors, BW723C86 has poor selectivity for human 5-HT(2B) receptors, while Org 37684 and VER-3323 display some selectivity for the 5-HT(2C) receptor. We report for the first time in a single study, the selectivity of numerous serotonergic drugs for 5-HT2 receptors from the same species, in mammalian cell lines and using, exclusively, agonist radiolabels. The results indicate the importance of defining the selectivity of pharmacological tools, which may have been over-estimated in the past, and highlights the need to find more selective agonists to investigate 5-HT2 receptor pharmacology.
在本研究中，我们比较了多种药物对稳定表达于单克隆哺乳动物细胞系中的人重组5-HT(2A)、5-HT(2B)和5-HT(2C)受体的高亲和力“激动剂偏好”结合位点的亲和力。为确保在竞争结合实验中优先标记受体的“激动剂偏好”构象，在每个受体上使用拮抗剂和激动剂放射性标记进行饱和分析。与相应的激动剂放射性标记（5-HT(2A)受体结合用[125I]-DOI，5-HT(2B)和5-HT(2C)受体结合用[3H]-5-HT）相比，拮抗剂放射性标记（5-HT(2A)受体用[3H]-酮色林，5-HT(2B)和5-HT(2C)受体用[3H]-美舒麦角）在每个制剂中与更多数量的受体结合。随后针对每个制剂中与“激动剂偏好”受体亚群结合的适当浓度的激动剂放射性标记进行竞争实验。这些研究证实，有许多高选择性拮抗剂可用于研究5-HT2受体亚型功能（例如，MDL 100907、RS-127445和RS-102221分别用于5-HT(2A)、5-HT(2B)和5-HT(2C)受体）。然而，仍然缺乏高选择性激动剂。(-)DOI对5-HT(2A)受体有效且具有中等选择性，BW723C86对人5-HT(2B)受体的选择性较差，而Org 37684和VER-3323对5-HT(2C)受体表现出一定的选择性。我们在一项单一研究中首次报道了众多血清素能药物在哺乳动物细胞系中对来自同一物种的5-HT2受体的选择性，并且仅使用激动剂放射性标记。结果表明定义药理学工具选择性的重要性，其选择性在过去可能被高估了，并强调需要找到更具选择性的激动剂来研究5-HT2受体药理学。
