CLONING AND PHARMACOLOGICAL CHARACTERIZATION OF A RAT KAPPA-OPIOID RECEPTOR
CLONING AND PHARMACOLOGICAL CHARACTERIZATION OF A RAT KAPPA-OPIOID RECEPTOR
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DOI：
10.1073/pnas.90.21.9954
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发表时间：
1993-11-01
影响因子：
11.1
通讯作者：
AKIL, H
中科院分区：
文献类型：
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作者：
MENG, F;XIE, GX;AKIL, H
通过对两个 PCR 片段进行低严格筛选，从大鼠纹状体文库中分离出全长 cDNA，其中一个片段跨越小鼠 δ 阿片受体的跨膜结构域 3-6，另一个未鉴定但与来自大鼠大脑的小鼠 δ 受体同源。该新型cDNA具有长的开放阅读框，编码380个残基的蛋白质，与小鼠δ受体具有59%的同一性，并且拓扑结构与七螺旋鸟嘌呤核苷酸结合蛋白偶联受体一致。转染该克隆编码区的 COS-1 细胞显示出与 kappa 阿片受体选择性配体（例如强啡肽 A 和 U-50,488）以及非选择性阿片配体（例如布马佐辛、乙基酮环佐辛和纳洛酮）的高亲和力结合。完全不结合（或以低亲和力结合）的是强啡肽A-(2-13)、纳洛酮和左啡烷的对映体[即(+)-纳洛酮和右啡烷]以及选择性μ和δ阿片受体配体。 kappa 受体激动剂激活表达的受体导致 cAMP 的抑制。最后，原位杂交揭示了大鼠大脑中 mRNA 的分布，与 kappa 选择性配体标记的结合位点的分布非常吻合。这些观察结果表明我们已经克隆了编码 kappa1 亚型的大鼠 kappa 受体的 cDNA。
A full-length cDNA was isolated from a rat striatal library by using low-stringency screening with two PCR fragments, one spanning transmembrane domains 3-6 of the mouse delta opioid receptor and the other unidentified but homologous to the mouse delta receptor from rat brain. The novel cDNA had a long open reading frame encoding a protein of 380 residues with 59% identity to the mouse delta receptor and topography consistent with a seven-helix guanine nucleotide-binding protein-coupled receptor. COS-1 cells transfected with the coding region of this clone showed high-affinity binding to kappa opioid receptor-selective ligands such as dynorphin A and U-50,488 and also nonselective opioid ligands such as bremazocine, ethylketocyclazocine, and naloxone. Not bound at all (or bound with low affinity) were dynorphin A-(2-13), enantiomers of naloxone and levophanol [i.e., (+)-naloxone and dextrorphan], and selective mu and delta opioid receptor ligands. Activation of the expressed receptor by kappa receptor agonists led to inhibition of cAMP. Finally, in situ hybridization revealed a mRNA distribution in rat brain that corresponded well to the distribution of binding sites labeled with kappa-selective ligands. These observations indicate that we have cloned a cDNA encoding a rat kappa receptor of the kappa1 subtype.