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目的 :建立共同稳定表达大鼠 μ阿片受体及人咪唑啉 1受体的哺乳动物细胞表达系统。 方法 :采用脂质体介导的方法 ,将编码大鼠 μ阿片受体和人咪唑啉 1受体的基因导入CHO细胞中 ,以放射配体 受体结合试验分析受体的表达水平 ,用cAMP积累试验分析表达受体的功能。结果 :在共同稳定表达大鼠 μ阿片受体及人咪唑啉 1受体的CHO细胞中 ,表达的 μ阿片受体对3 H 二丙诺啡 (diprenorphine)的Kd 和Bmax值分别为 (0 .2 4± 0 .0 2 )nmol/L和 (1.83±0 .13)pmol/mg蛋白 ;表达的咪唑啉 1受体对3 H 可乐定的Kd 和Bmax值分别为 (3.72± 0 .2 5 )nmol/L和 (4 3.5 9± 6 .83)fmol/10 6细胞。表达的 μ阿片受体抑制福司科林 (forskolin)刺激下cAMP的积累 ,表达的咪唑啉 1受体抑制吗啡慢性处理、纳洛酮催促引起的cAMP超射。结论 :首次建立了共同稳定表达 μ阿片受体和咪唑啉 1受体的细胞模型。
OBJECTIVE: To establish a mammalian cell expression system co-stably expressing rat μ opioid receptor and human imidazoline 1 receptor. METHODS: The gene encoding rat mu opioid receptor and human imidazoline 1 receptor was introduced into CHO cells by liposome-mediated method. The expression of receptor was analyzed by radioligand binding assay. Accumulate experiments analyzing the function of expressing receptors. Results: In CHO cells co-stably expressing rat mu opioid receptor and human imidazoline 1 receptor, the Kd and Bmax values of expressed mu opioid receptor for 3 H diprenorphine were (0. 2 4 ± 0. 2) nmol / L and (1.83 ± 0. 13) pmol / mg protein respectively. The Kd and Bmax values of 3 H-clonidine at the imidazoline 1 receptor were (3.72 ± 0.52 ) nmol / L and (4 3.59 ± 6 .83) fmol / 106 cells. The expressed mu opioid receptor inhibits the accumulation of cAMP stimulated by forskolin, and the expressed imidazoline 1 receptor inhibits the chronic morphine treatment and the cAMP hyperintense induced by naloxone. CONCLUSION: The first co-expression of μ opioid receptor and Imidazoline 1 receptor cell model was established.