Cannabinoid receptor agonist efficacy for stimulating [35S]GTPγS binding to rat cerebellar membranes correlates with agonist-induced decreases in GDP affinity
Cannabinoid receptor agonist efficacy for stimulating [35S]GTPγS binding to rat cerebellar membranes correlates with agonist-induced decreases in GDP affinity
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DOI：
10.1074/jbc.273.27.16865
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发表时间：
1998-07-03
影响因子：
4.8
通讯作者：
Childers, SR
中科院分区：
文献类型：
--
作者：
Breivogel, CS;Selley, DE;Childers, SR
本文研究了大麻素刺激大鼠小脑膜[S-35]鸟苷-5‘-O-(3-硫代三磷酸)([S-35]鸟苷-5’-O-(3-硫代三磷酸)-([S-35]鸟苷-O-(3-硫代三磷酸))结合的作用及其与国内生产总值的关系。动力学分析表明，[35S]GTP与S的结合达到稳态水平，激动剂Win 55212-2与Gdp的结合率与浓度成正比。[S-35]GTPγS以两种速率(t(1/2)=7和170min)发生解离，Win 55212-2增加了表现出较快速率的位点的比例。在没有国内生产总值的情况下，[S-35]GTP-γS与高亲和力和低亲和力的膜结合，而Win 55212-2则没有作用。与30亩Gdp、[S-35]GTP与S结合中低亲和力部位，与Win 55212-2诱导高亲和力[S-35]GTP与S结合而不影响低亲和力部位，Gdp竞争高亲和力[S-35]GTP与S结合，在没有Win 55212-2的情况下与高、中亲和力结合，在Win 55212-2存在的情况下具有高、低亲和力。大麻素配体表现出不同的能力，在GDP存在的情况下最大限度地刺激[S-35]GTP-S结合。不同配基间的药效差异随着GDP浓度的增加而增大。GDP竞争曲线显示，激动剂诱导的低亲和力GDPKi值与激动剂的E-max值成正比，表明激动剂的效果取决于GDP从G蛋白中置换。
The relationship between GDP and cannabinoid-stimulated [S-35]guanosine-5'-O-(3-thiotriphosphate) ([S-35]GTP gamma S) binding was investigated in rat cerebellar membranes. Kinetic analyses showed that [35S]GTP gamma S binding reached steady-state levels and that the association rate was increased by the agonist WIN 55212-2 proportional to the concentration of GDP. Dissociation of [S-35]GTP gamma S occurred with two rates (t(1/2) = 7 and 170 min), and WIN 55212-2 increased the proportion of sites exhibiting the faster rate. Without GDP, [S-35]GTP gamma S bound to membranes with high and low affinity, and WIN 55212-2 had no effect. With 30 mu M GDP, [S-35]GTP gamma S bound to low and intermediate affinity sites, and WIN 55212-2 induced high affinity [S-35]GTP gamma S binding without affecting low affinity sites, GDP competed for high affinity [S-35]GTP gamma S binding with high and intermediate affinity in the absence of WIN 55212-2 and with high and low affinity in the presence of WIN 55212-2. Cannabinoid ligands displayed differential abilities to maximally stimulate [S-35]GTP gamma S binding in the presence of GDP. Efficacy differences among ligands increased with increasing GDP concentrations. GDP competition curves revealed that agonists induced low affinity GDP Ki values that were proportional to agonist E-max values, indicating that agonist efficacy is determined by displacement of GDP from G-proteins.