Lane C M, Elde R, Loh H H, Lee N M
Department of Pharmacology, University of Minnesota, Minneapolis 55455.
Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11234-8. doi: 10.1073/pnas.89.23.11234.
An opioid-binding protein has recently been purified from bovine brain and cloned, and its cDNA sequence has been obtained. Indirect evidence suggests that this protein has a role in opioid-receptor function. However, because direct testing of its function by expression of its cDNA has not yet been possible and because its structure bears no resemblance to G protein-coupled receptors, the role of this protein in opioid-receptor activity is still in question. An antibody raised to a portion of the predicted amino acid sequence of opioid-binding cell-adhesion molecule (OBCAM) specifically labeled the surface of NG108-15 cells, as visualized by immunofluorescence with confocal microscopy. Furthermore, chronic treatment of these cells with opioid agonist, which down-regulates opioid receptors, reduced OBCAM immunoreactivity (ir). Down-regulation of both opioid receptors and OBCAM-ir was greatest after chronic treatment of NG108-15 cells with delta-opioid agonists, as well as with nonselective agonists such as etorphine, whereas other agonists including [D-Ala2-N-MePhe4-Gly-ol]enkephalin, morphine, levorphanol, dynorphin A-(1-13), and U-50,488H were less effective or ineffective. Chronic treatment of NG108-15 cells with muscarinic agonists had no effect on OBCAM-ir. Furthermore, NG108-15 cells transfected with an antisense construct to OBCAM have a reduced density of opioid-binding sites as well as reduced OBCAM-ir. Taken together, these results strongly suggest that OBCAM has a role in opioid-receptor function in NG108-15 cells.
一种阿片样物质结合蛋白最近已从牛脑中纯化并克隆出来，其cDNA序列也已获得。间接证据表明该蛋白在阿片样物质受体功能中起作用。然而，由于通过其cDNA表达直接测试其功能尚不可能，且其结构与G蛋白偶联受体毫无相似之处，该蛋白在阿片样物质受体活性中的作用仍存在疑问。针对阿片样物质结合细胞粘附分子（OBCAM）预测氨基酸序列的一部分产生的抗体，通过共聚焦显微镜免疫荧光观察，特异性标记了NG108 - 15细胞的表面。此外，用阿片样物质激动剂对这些细胞进行慢性处理，可下调阿片样物质受体，降低了OBCAM免疫反应性（ir）。在用δ-阿片样物质激动剂以及非选择性激动剂如埃托啡对NG108 - 15细胞进行慢性处理后，阿片样物质受体和OBCAM - ir的下调最为明显，而其他激动剂，包括[D - Ala2 - N - MePhe4 - Gly - ol]脑啡肽、吗啡、左啡诺、强啡肽A -（1 - 13）和U - 50,488H的效果较差或无效。用毒蕈碱激动剂对NG108 - 15细胞进行慢性处理对OBCAM - ir没有影响。此外，用针对OBCAM的反义构建体转染的NG108 - 15细胞，阿片样物质结合位点的密度降低，OBCAM - ir也降低。综上所述，这些结果强烈表明OBCAM在NG108 - 15细胞的阿片样物质受体功能中起作用。
