低剂量毒死蜱暴露对新生大鼠黑质多巴胺能神经元发育及行为的影响

Effects of low-dose chlorpyrifos exposure on dopaminergic neurons in the midbrain substantia nigra and neural behavioral development in neonatal rats

目的探讨低剂量毒死蜱(chlorpyrifos,CPF)暴露对新生大鼠中脑黑质多巴胺能(DA)神经元发育和神经行为的影响。方法将出生后11 d Sprague-Dawley大鼠随机分为CPF组、二甲基亚砜(DMSO)组和生理盐水(NS)组。CPF组在大鼠出生后11～14 d经腹部皮下注射低剂量(每日5 mg/kg)CPF,其他两组分别注射DMSO和NS作为对照。在生后15 d、20 d、30 d及60 d四个时间点,观察大鼠体重增长、脑外观、脑系数和脑含水量变化;免疫组织化学方法检测黑质DA神经元酪氨酸羟化酶(TH)的表达;免疫透射电镜观察DA神经元亚细胞结构改变;生后30 d和60 d行旷场实验、握力实验、斜坡实验及Morris水迷宫实验检测大鼠神经行为变化。结果各组大鼠在各观察时间点体重增长、脑外观、脑系数、脑含水量未见异常。CPF组自生后30 d开始与NS和DMSO组相比,不仅黑质TH表达进行性减少,部分DA神经元亚细胞结构发生改变,且逐渐出现活动减少,动作协调障碍及学习记忆能力受损。结论大鼠脑发育期暴露低剂量CPF,可诱导中脑黑质DA神经元迟发性进行性丢失,并影响大鼠远期运动和学习记忆能力。

Objective To explore the effects of low-dose chlorpyrifos（CPF） exposure on dopaminergic（DA） neurons in the midbrain substantia nigra and neural behavioral development in neonatal rats.Methods Postnatal 11 day old Sprague-Dawley rats were randomly assigned into CPF,menstruum dimethysulfoxide（DMSO） and normal saline（NS）groups.The rats in the CPF group were injected with low-dose CPF（5 mg/kg·d） on postnatal days 11-14.The two control groups were injected with DMSO or NS respectively.The rats were sacrificed on postnatal days 15,20,30,and 60.Body weight gain,outward appearance of brain tissue,the coefficient of brain and the water content of brain tissue were measured.Tyrosine hydroxylase（TH） expression in DA neurons in the midbrain substantial nigra was examined by immunohistochemical straining.Immune electron microscopy was used to examine the subcellular structure of DA neurons.Open field test,grip strength test,slope test and Morris water maze test were used to examine the neurobehavioral changes.Results The outward appearance of brain tissue was normal in the three groups.There were no significant differences in the absolute value of body weight gain,the coefficient of brain and the water content of brain tissue among the three groups.CPF exposure decreased the level of TH immunoreactivity（P0.05） in the substantia nigra of CPF group since postnatal day 30 compared with the DMSO and NS groups.The subcellular structures of some DA neurons in the CPF group were impaired.Decreased motor activity and learning and memory impairments were observed in the CPF group compared with those in the DMSO and NS groups（P0.05）since postnatal day 30.Conclusions CPF exposure during the neonatal period can cause long-term motor activity and learning and memory impairments in accompany with DA neurons damage in the midbrain substantia nigra.