Opioid efficacy in a C6 glioma cell line stably expressing the human kappa opioid receptor.
Opioid efficacy in a C6 glioma cell line stably expressing the human kappa opioid receptor.
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发表时间：
1999-02
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通讯作者：
A. Remmers;M. Clark;A. Mansour;H. Akil;J. Woods;F. Medzihradsky
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作者：
A. Remmers;M. Clark;A. Mansour;H. Akil;J. Woods;F. Medzihradsky
一个C6神经胶质瘤细胞系稳定转染人κ阿片受体（kappaOR）被用来表征受体结合和G蛋白激活通过kappaOR由一系列全面的阿片配体。[3 H] 5 α，7 α，8 β（-）-N-甲基-N-（7-Cl-吡咯烷基）-1-氧杂螺（4，5）癸-8-基）苯乙酰胺（U69593）的配体结合亲和力与猴脑细胞膜中观察到的相似，在钠和GDP存在下低10倍。肽和非肽激动剂都最大程度地刺激[35 S] GTP γ S结合。[35 S] GTP γ S结合的刺激通过用百日咳毒素预处理细胞来阻断。对于作为μ阿片受体拮抗剂的几种配体，观察到通过kappaOR对[35 S] GTP γ S结合的部分刺激，表明了药物作用的另一种机制。替氟多和左洛啡烷的异构体刺激[35 S] GTP γ S结合的能力表明，左洛啡烷（一种苯并吗喃衍生物）的手性碳比苯并二氮卓类衍生物替氟多中的手性碳具有更大程度的立体选择性。此外，（-）替氟酮（替氟酮的较弱异构体，也是一种γ-氨基丁酸A受体激动剂）刺激[35 S] GTP γ S结合。相反，d-喷他佐辛、（+）SKF 10047、（+）环唑辛和d-乙基酮环唑辛没有显示激动剂活性。kappaOR选择性拮抗剂norbinaltorphimine竞争性抑制乙基酮环唑辛、环唑辛和纳洛啡的活性异构体对[35 S] GTP γ S结合的刺激，抑制程度相同，表明所有三种配体均通过kappaOR引发效应。结果表明，这些细胞表达kappaOR的同质群体，并且它们的[35 S] GTP γ S结合特性使它们成为评估kappaOR功效的极好手段。
A C6 glioma cell line stably transfected with the human kappa opioid receptor (kappaOR) was used to characterize receptor binding and G protein activation via the kappaOR by a comprehensive series of opioid ligands. The ligand-binding affinity for [3H]5alpha,7alpha, 8beta(-)-N-methyl-N-(7-Cl-pyrrolidinyl)-1-oxaspiro(4, 5)dec-8-yl)benzene acetamide (U69593) was similar to that observed in monkey brain membranes and was 10-fold lower in the presence of sodium and GDP. Both peptide and nonpeptide agonists maximally stimulated [35S]GTPgammaS binding. The stimulation of [35S]GTPgammaS binding was blocked by pretreatment of cells with pertussis toxin. Partial stimulation of [35S]GTPgammaS binding via the kappaOR was observed for several ligands that are antagonists at the mu opioid receptor, suggesting an additional mechanism of drug action. The ability of isomers of tifluadom and levallorphan to stimulate [35S]GTPgammaS binding indicates that the chiral carbon of levallorphan, a benzomorphan derivative, imparts a greater degree of stereoselectivity than does the chiral carbon in the benzodiazepine derivative tifluadom. In addition, (-)tifluadom, the less potent isomer of tifluadom, which is also a gamma-aminobutyric acidA receptor agonist, stimulated [35S]GTPgammaS binding. In contrast, d-pentazocine, (+)SKF10047, (+)cyclazocine, and d-ethylketocyclazocine displayed no agonist activity. kappaOR-selective antagonist norbinaltorphimine competitively inhibited the stimulation of [35S]GTPgammaS binding by the active isomers of ethylketocyclazocine, cyclazocine, and nalorphine to the same degree, indicating that all three ligands are eliciting an effect via the kappaOR. The results suggest that these cells express a homogeneous population of kappaOR, and that their [35S]GTPgammaS-binding properties make them an excellent means to assess kappaOR efficacy.