柴胡口服液改善成年BTBR自闭症小鼠海马神经发生及短期记忆缺陷

Chaihu Oral Liquid improves hippocampal neurogenesis and short-term memory defects in adult BTBR autism mice

目的研究柴胡口服液(ChaiHu oral liquid,CHOL)对BTBR自闭症模型小鼠(BTBR T^(+)Itpr3^(tf)/J)海马齿回神经发生及短期记忆缺陷的影响。方法7~8周龄成年雄性健康C57BL/6J(C57)[体质量(24.72±1.35)g]与BTBR小鼠[体质量(27.7±1.42)g]随机分为4组(n=4):C57对照组、C57药物处理组、BTBR对照组、BTBR药物处理组。C57和BTBR药物处理组小鼠予以柴胡口服液(10μL/g)灌胃,1次/d,共7 d;对照组给予等体积的生理盐水。记录小鼠每日体质量变化,最后一次给药后,于次日开始进行理毛、埋珠、新物体识别和开场实验,共计4 d。最后收取脑组织样本,采用免疫荧光染色检测海马齿回内NeuN、DCX、SOX2/GFAP的表达。结果行为学结果显示:与C57对照组比较,BTBR对照组小鼠在新物体识别实验中表现出对旧物体更明显的偏好(P<0.01),而BTBR药物处理后新物体识别指数得到提升(P<0.05);与C57对照组比较,BTBR对照组小鼠理毛、埋珠实验检测的重复刻板行为、开场实验检测的运动距离及体质量明显增加(P<0.05),而BTBR药物处理组小鼠重复刻板行为、运动距离和体质量较之BTBR对照组无明显改变;与C57对照组比较,C57药物处理组小鼠新物体识别指数、重复刻板行为、运动能力、体质量变化差异无统计学意义。免疫荧光染色结果显示:与C57对照组比较,BTBR对照组小鼠海马齿回内NeuN、DCX标记的成熟神经元和新生神经元的数量,以及SOX/GFAP标记的神经祖细胞的数量明显减少(P<0.01),而BTBR药物处理组相比BTBR对照组明显增多(P<0.05)。C57对照组与C57药物处理组小鼠海马齿回内NeuN、DCX及SOX2/GFAP的数量差异无统计学意义。结论柴胡口服液治疗促进成年BTBR小鼠海马齿回神经祖细胞、新生神经元及成熟神经元的产生,能够改善小鼠短期学习记忆功能障碍。

Objective To investigate the effects of Chaihu Oral Liquid(CHOL)on the neurogenesis in hippocampal dentate gyrus(DG)and short-term memory defects in BTBR T^(+)Itpr3^(tf)/J(BTBR)autism model mice.Methods Healthy adult male C57 BL/6 J(C57)mice(7~8 weeks old,24.72±1.35 g)and BTBR mice(7~8 weeks old,27.7±1.42 g)were randomly divided into groups as follows(n=4 in each group):C57 control,C57 drug treatment,BTBR control,and BTBR drug treatment.Mice in the treatment groups were given CHOL solution gavage(10μL/g)once a day for 7 consecutive days,while those of the control group received equal volume of normal saline.Daily changes of body weight were recorded.After the last administration,hair grooming,bead embedding,new object recognition and open field experiments were conducted on the next day and lasted for 4 d.Finally,the samples of mice brain tissue were harvested,and immunofluorescence staining was performed to detect the NeuN,DCX,SOX2/GFAP labeled nerve cells in hippocampal DG.Results Behavioral results suggested that mice in the BTBR control group showed a more obvious preference for old objects in the new object recognition experiment(P<0.01),as compared with the C57 control group,while the new object recognition index was significantly improved in the BTBR drug treatment group(P<0.05).Moreover,the BTBR control group presented more repetitive behavior and longer movement distance than the C57 control during the hair grooming,bead embedding and open field experiments,as well as higher body weight(P<0.05),whereas the BTBR treatment group had no significant changes in comparison with the BTBR control regarding these aspects.There were no notable differences between the C57 control and C57 treatment groups in terms of new object recognition index,repetitive behavior,motor ability and body weight.Immunofluorescence staining results indicated that the numbers of NeuN labeled mature neurons and DCX labeled newborn neurons in hippocampal DG were greatly decreased in the BTBR control group,so as the number of SOX/GFAP labeled neural progenitor cells(P<0.01),while the number of neurocytes was remarkably increased in the BTBR drug treatment group(P<0.05).There were no significant differences in the numbers of NeuN,DCX and SOX2/GFAP labeled neurocytes between the C57 control and the C57 treatment groups.Conclusion CHOL promotes the generation of hippocampal DG nerve progenitor cells,new and mature neurons in adult BTBR mice,and improves the short-term learning and memory dysfunction.