大鼠星形胶质瘤细胞系C6细胞μ受体的表达  

Identification of opioid receptor expression in rat C6 glioma cells

作　　者：曹菲[1] 田玉科[1] 许爱军[1] 高峰[1] 肖兴鹏[1] 陈莎莎 彭伟[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院麻醉科教研室,武汉市430030

出　　处：《中华麻醉学杂志》2008年第6期508-510,共3页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金资助项目（30672027）;高等学校博士学科点专项科研基金资助项目（20060487048）

摘　　要：目的鉴定大鼠星形胶质瘤细胞系C6细胞μ受体的表达。方法星形胶质瘤细胞系C6细胞培养、消化、传代后，接种于50ml培养瓶中，待细胞生长至融合率达80％～90％时，采用RTPCR法测定μ受体mRNA的表达，免疫组化SP法测定μ受体蛋白的表达。将C6细胞接种于无菌的盖玻片上，待细胞生长至融合率达40％～50％时，孵育、去脂后，随机分为5组：A组加入PBS20μl；B组加入μ受体选择性激动剂DAMGO1μmol／L20μl；C组加入吗啡1μmol／L20μl；D组先加入k受体选择性抑制剂nor-Binaltorphimine 1 μmol／L 20μl，10min后再加入吗啡1μmol／L 20μl；E组先加入纳洛酮1μmol/L 20μl，10min后再加入吗啡1μmol/L 20μl。采用FV500型激光扫描共聚焦显微镜测定给予激动剂前、后细胞内游离钙离子浓度（[Ca^2＋]i）。结果C6细胞存在μ受体mRNA，且μ受体主要分布于细胞膜及细胞浆；A组、B组和E组[Ca^2＋]i无变化（P〉0．05），C组和D组[Ca^2＋]i呈一过性升高（P〈0．05）。结论标准条件下培养的大鼠星形胶质细胞系C6细胞膜表面可能存在μ3型受体。Objective To identify the μ opioid receptor expression in the cytomembrane of rat C6 glioma cells. Methods The C6 cells of the rat astrocytic glioma cell line were donated by the cell bank of Tongji Medical college cell engineering platform. After being cultured for 2-3 days the C6 glioma cells were dispersed, underwent passage and were then subcultured in sterile cell culture flasks. When the C6 cells reached 80%-90% confluency the expression of μ opioid receptor mRNA and protein was determined by RT-PCR and immunocyto-chemistry respectively. When the C6 cells reached 40%-50% confluency, they were inoculated onto sterile cover glass and were randomly divided into 5 groups after incubatiorl and degrease： group A was treated with PBS 20 μl; group B with 1 μmol/L DAMGO （opioid receptor selective agonist） 20 μl; group C with 1 μmol/L morphine 20 μl; group D was pretreated with 1 μmol/L nor-binaltorphimine （k opioid receptor selective antagonist） 20 μl and 10 min later 1 μmol/L morphine 20μl was added and group E was pretreated with 1 μmol/L naloxone 20 μl and 10 min later 1 μmol/L morphine 20 μl was added. [ Ca^2＋ ] i in C6 cells was determined with FV 500 laser scanning eonfocal microscope before and after the addition of different drugs. Results The opioid receptor mRNA was identified in C6 glioma cells by RT-PCR. Immuno-histochemistry showed that opioid receptors were mainly distributed in the cytomembrane and cytoplasm. There was no significant change in [ Ca^2＋ ]i after addition of PBS, morphine or naloxone ＋ morphine in group A, B and E. [Ca^2＋ ]i was significantly increased temporarily in group C and D. Conclusion There may be μ3 opioid receptors in the eytomembrane of rat C6 glioma cells cultured under standard condition.

关 键 词：星形细胞瘤 星形胶质细胞 受体 阿片样物质 

分 类 号：R739.4[医药卫生—肿瘤]