阿特拉津对青春期大鼠黑质区多巴胺能神经元的损伤作用

Induction of dopaminergic neuron degeneration by atrazine in substantia nigra in adolescent rats

目的:探讨阿特拉津对青春期大鼠黑质区多巴胺能神经元的损伤作用。方法:将4周龄的Wistar雄性大鼠40只随机分为4组,分别为对照组(1%甲基纤维素)、阿特拉津低(50 mg/kg)、中(100 mg/kg)和高(200 mg/kg)剂量组,每组10只,连续灌胃45 d后处死。收集中脑黑质部分,采用免疫组织化学法(SP法)检测神经元细胞中酪氨酸羟化酶(TH)的变化,并采用实时定量PCR法(qPCR)和Western blot检测多巴胺能神经元损伤相关基因mRNA和蛋白的表达情况。结果:免疫组化结果显示,200 mg/kg ATR组中TH阳性细胞数较对照组显著减少,差异具有统计学意义(P<0.01)。qPCR结果显示,与对照组相比,100和200 mg/kg ATR染毒组TH mRNA和Bcl-2 mRNA的表达量显著降低;200 mg/kg ATR组p53 mRNA和Caspase-9 mRNA的表达量显著增加,差异具有统计学意义(P<0.05)。Western blot结果显示,与对照组相比,100和200 mg/kg ATR染毒组TH和Bcl-2的蛋白表达量显著降低;100和200 mg/kg ATR组p53蛋白表达量显著增加;50、100和200 m g/kg A TR组Caspase-9蛋白表达量均显著增加,差异均具有统计学意义(P均<0.05)。结论:阿特拉津可以影响大鼠黑质区多巴胺能神经元损伤相关基因mRNA和蛋白的表达。

OBJECTIVE：To study effects of atrazine （ATR） on neuronal degeneration in adolescent rats. METHODS：4 weeks old Wistar male rats were randomly subdivided into 4 groups： control and ATR test groups. All rats were treated with ATR： 50,100 and 200 mg/kg body weight per day （five times per week） for 45 days by oral gavage. They were sacrificed at 12 h after the last ATR injection. The entire midbrain was collected and stored frozen at -80℃. The presence of tyrosine hydroxylase（TH） was detected by immunohistochemistry SP in dopaminergic neurons. TH expression in mRMA and protein was measured using real-time PCR and Western blot.RESULTS：Immunohistochemical results showed that,compared with the control group,the number of TH positive cells in the 200 mg/kg ATR group was significantly decreased （P〈0.01）. In addition,qPCR results show that expression of TH mRNA and Bcl-2 mRNA was significantly decreased in the 100 and 200 mg/kg ATR groups,and expression of p53 mRNA and Caspase-9 mRNA was significantly increased in the 200 mg/kg ATR group,the difference was statistically significant （P〈0.05）. Western blot results show that expression of TH and Bcl-2 protein was significantly decreased in the 100 and 200 mg/kg ATR groups compared with the control group. Expression of p53 protein in the ATR group was significantly increased at 100 and 200 mg/kg,and Caspase-9 protein in he 200 mg/kg ATR group was significantly increased,the difference was statistically significant （P〈0.05）.CONCLUSION：Atrazine caused damage to the dopaminergic neurons in the substantia nigra of rats and activated the apoptotic pathway which led to the dopaminergic neuronal death.