Monczor Federico, Fernandez Natalia, Legnazzi Bibiana Lemos, Riveiro Maria Eugenia, Baldi Alberto, Shayo Carina, Davio Carlos
Laboratorio de Radioisótopos, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires. Junin 956 PB, 1113, Capital Federal, Argentina.
Mol Pharmacol. 2003 Aug;64(2):512-20. doi: 10.1124/mol.64.2.512.
Knowing the importance for research and pharmacological uses of proper ligand classification into agonists, inverse agonists, and antagonists, the aim of this work was to study the behavior of tiotidine, a controversial histamine H2 receptor ligand. We found that tiotidine, described previously as an H2 antagonist, actually behaves as an inverse agonist in U-937 cells, diminishing basal cAMP levels. [3H]Tiotidine showed two binding sites, one with high affinity and low capacity and the other with low affinity and high capacity. The former site disappeared in the presence of guanosine 5'-O-(3-thio)triphosphate, indicating that it belongs to a subset of receptors coupled to G-protein, showing the classic binding profile for an agonist. Considering the occupancy models developed up to now, the only one that explains tiotidine dual behavior is the cubic ternary complex (CTC) model. This model allows G-protein to interact with the receptor even in the inactive state. We showed by theoretical simulations based on the CTC model of dose-response and binding experiments that tiotidine biases the system to a G-protein-coupled form of the receptor that is unable to evoke a response. This theoretical approach was supported by experimental results in which an unrelated G-protein-coupled receptor that also signals through Galphas-protein (beta2-adrenoreceptor) was impeded by tiotidine. This interference clearly implies that tiotidine biases the system to Galphas-coupled form of the H2 receptor and turns Galphas-protein less available to interact with beta2-adrenoreceptor. These findings not only show that tiotidine is an H2 inverse agonist in U-937 cells but also provide experimental support for the CTC model.
鉴于将合适的配体正确分类为激动剂、反向激动剂和拮抗剂对于研究及药理用途的重要性，本研究旨在探究替奥替丁（一种存在争议的组胺H2受体配体）的行为。我们发现，先前被描述为H2拮抗剂的替奥替丁，在U - 937细胞中实际上表现为反向激动剂，可降低基础cAMP水平。[3H]替奥替丁显示出两个结合位点，一个具有高亲和力和低容量，另一个具有低亲和力和高容量。前一个位点在鸟苷5'-O-(3-硫代)三磷酸存在时消失，表明它属于与G蛋白偶联的受体亚群，呈现出激动剂的经典结合特征。考虑到目前已建立的占据模型，唯一能解释替奥替丁双重行为的是立方三元复合物（CTC）模型。该模型允许G蛋白即使在受体的非活性状态下也能与之相互作用。我们通过基于剂量反应和结合实验的CTC模型进行的理论模拟表明，替奥替丁使系统偏向于受体的一种与G蛋白偶联的形式，这种形式无法引发反应。这一理论方法得到了实验结果的支持，在实验中，替奥替丁阻碍了另一种同样通过Gαs蛋白信号传导的无关G蛋白偶联受体（β2肾上腺素受体）。这种干扰清楚地表明，替奥替丁使系统偏向于H2受体与Gαs偶联的形式，并使Gαs蛋白与β2肾上腺素受体相互作用的可用性降低。这些发现不仅表明替奥替丁在U - 937细胞中是一种H2反向激动剂，还为CTC模型提供了实验支持。
