天麻素注射液通过调节背侧纹状体miR-30a表达改善大鼠甲基苯丙胺成瘾的实验研究

Effect of Gastrodin Injection on Improving Methamphetamine Addiction in Rats by Regulating the Expression of miR-30a in Dorsal Striatum

目的:研究天麻素注射液通过调节背侧纹状体miR-30a表达改善大鼠甲基苯丙胺成瘾的作用。方法:SD大鼠分为正常对照组、甲基苯丙胺组、天麻素10 mg/kg、20 mg/kg组、天麻素20 mg/kg+miR-30a抑制慢病毒组,制备甲基苯丙胺自身给药模型并给予不同剂量天麻素注射液腹腔注射、miR-30a抑制慢病毒背侧纹状体注射,评价复吸行为及背侧纹状体中miR-30a及BDNF蛋白的表达;培养SH-SY5Y神经元细胞,用甲基苯丙胺、天麻素处理或转染NC模拟物、miR-30a模拟物,检测miR-30a及BDNF的表达、验证miR-30a与BDNF 3’UTR的靶向结合。结果:与假手术对照组比较,甲基苯丙胺组大鼠的触鼻次数明显增多,背侧纹状体中miR-30a的表达明显下调、BDNF蛋白的表达明显上调(P <0. 05);与甲基苯丙胺组比较,天麻素10 mg/kg、20 mg/kg组大鼠的触鼻次数明显减少,背侧纹状体中miR-30a的表达明显上调、BDNF的蛋白表达明显下调(P <0. 05);天麻素20 mg/kg+miR-30a抑制慢病毒组大鼠的触鼻次数明显多于天麻素20 mg/kg组(P <0. 05),背侧纹状体中miR-30a的表达明显低于正常对照组(P <0. 05)、BDNF的蛋白表达明显多于天麻素20 mg/kg组(P <0. 05)。与空白对照组比较,甲基苯丙胺组SH-SY5Y细胞中miR-30a的表达明显下调、BDNF的蛋白表达明显上调(P <0. 05),与甲基苯丙胺组比较,天麻素1 mmol/L组SH-SY5Y细胞中miR-30a的蛋白表达明显上调、BDNF的表达明显下调(P <0. 05);miR-30a模拟物组SH-SY5Y细胞中miR-30a的表达明显高于NC模拟物组(P <0. 05),BDNF的表达及野生型BDNF荧光素酶报告基因的荧光活性明显低于NC模拟物组(P <0. 05)。结论:天麻素注射液能上调背侧纹状体miR-30a表达、改善大鼠甲基苯丙胺成瘾,上调miR-30a并靶向抑制BDNF表达可能是天麻素发挥上述作用的分子机制。

Objective: To study the effect of gastrodin injection on methamphetamine addiction in rats by regulating the expression of miR-30 a in dorsal striatum. Methods: SD rats were divided into the control group,methamphetamine group,gastrodin groups( 10 mg/kg,20 mg/kg)and 20 mg/kg gastrodin + miR-30 a inhibiting lentivirus group. The self-administered model of methamphetamine was established,gastrodin was intraperitoneally injected while miR-30 a inhibiting lentivirus was injected into dorsal striatum. The relapse behavior and the protein expressions of miR-30 a and BDNF in dorsal striatum were evaluated. SH-SY5 Y neurons were cultured and treated with methamphetamine and gastrodin or transferred with NC mimic and miR-30 a mimic. Then the expressions of miR-30 a and BDNF were determined and the targeted binding of miR-30 a to BDNF 3’UTR was verified. Results: Compared with the control group,the number of nasal touches was significantly increased in methamphetamine group,the expression of miR-30 a in dorsal striatum was significantly decreased,the expression of BDNF was significantly increased. Compared with methamphetamine group,the number of nasal touches in 10 mg/kg and 20 mg/kg gastrodin groups were significantly decreased,the expression of miR-30 a in dorsal striatum was significantly increased,the expression of BDNF was significantly decreased. Compared with 20 mg/kg gastrodin group,the number of nasal touches was significantly increased in 20 mg/kg gastrodin+ miR-30 a inhibiting lentivirus group,the expression of miR-30 a in dorsal striatum was significantly decreased,the expression of BDNF was significantly increased. Compared with the control group,the expression of miR-30 a in SH-SY5 Y cells in methamphetamine group was significantly decreased,the expression of BDNF was significantly increased. Compared with methamphetamine group,the expression of miR-30 a in SH-SY5 Y cells in gastrodin group was significantly increased,the expression of BDNF was significantly decreased. The expression of miR-30 a in SH-SY5 Y cells in miR-30 a mimic group was significantly higher than that in NC group,the expression of BDNF and the fluorescence activity of wild-type BDNF luciferase reporter gene were significantly lower than those in the NC mimic group. Conclusion: Gastrodin injection can increase the expression of miR-30 a in dorsal striatum and improve methamphetamine addiction in rats,its molecular mechanism may be increasing miR-30 a and targeted inhibiting BDNF expression.