第十九届全国神经精神药理学学术会议论文摘要

2021年10月15-17日，广东广州

专题1：神经退行性疾病

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NMDA受体对中枢神经系统的影响

孙丽丛，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：N-甲基-D-门冬氨酸（NMDA）受体是中枢神经系统重要的兴奋性神经递质，主要分布于大脑和脊髓等。研究发现，NMDA参与许多神经系统疾病的过程如脑卒中、癫痫、抑郁症和强迫症等。在正常机体的中枢神经系统内，NMDA受体被激活后，Na+，K+和Ca2+的通透性增加，可产生一系列生化改变，影响突触效率，进而促进学习和记忆能力的形成，并促进海马神经干细胞增殖。但NMDA受体对中枢神经系统也存在不利影响，表现为以下2个方面。①兴奋性增高：有研究表明，在急性脑缺血中，NMDA受体亚基NR2A和NR2B的mRNA及蛋白表达均明显上调，导致细胞内钙超载，引起细胞毒性作用，甚至可导致细胞凋亡；当NMDA受体被过度激活时，过多的神经元突触后膜发生同步性去极化，持续性放电，引发癫痫；当炎性疼痛产生时，NMDA受体被激活，NMDA受体磷酸化水平升高，使神经元的兴奋作用放大，最终导致神经元敏化。②兴奋性降低：NMDA受体可调控γ-氨基丁酸（GABA）能中间神经元的功能，其活性异常及不同亚基缺失可导致GABA能中间神经元功能异常，从而引发抑郁症和强迫症等。本文通过对NMDA受体在中枢神经系统中的影响进行综述，以期为NMDA受体异常导致疾病的治疗提供参考。

关键词：N-甲基-D-门冬氨酸受体；中枢神经系统；兴奋性神经递质；γ-氨基丁酸；神经干细胞；学习能力；记忆能力

基金项目：河北省自然科学基金（H2020208032）

通讯作者：张丹参，E-mail：zhangds2011@126.com

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姜黄素对阿尔茨海默病治疗作用及其机制

孙 婷，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：姜黄素是从姜科或天南星科一些植物的根茎中提取的一种二酮类化合物，是中药姜黄发挥药理作用的主要活性成分。近年来国内外研究发现，姜黄素的药理活性广泛，具有抗氧化、抗炎、神经保护、降血脂、抗肿瘤等作用。阿尔茨海默病（AD）即老年痴呆症，是一种常见的中枢神经退行性疾病，其临床特征主要表现为学习记忆能力减退和认知功能障碍，其发病机制复杂，神经递质乙酰胆碱缺失、氧化应激、炎症反应及细胞凋亡等均可导致AD。姜黄素治疗AD有显著的疗效，本文主要从其作用机制方面进行综述。①抗氧化应激：姜黄素可以通过提高SOD和GSH-Px活性、降低MDA含量，抑制自由基生成并加强其清除，减轻氧化应激；②抗炎症反应：姜黄素可以降低1L-1β和TNF-α炎症因子的表达，减少炎症介质的产生，发挥神经保护作用；③抑制细胞凋亡：姜黄素通过激活NF-κB信号通路，调控细胞凋亡相关基因Bax和Bcl-2表达，降低促凋亡因子Bax表达，增强抑制凋亡因子Bcl-2表达而阻止细胞凋亡；还可通过抑制糖原合成酶激酶3β活性抑制细胞凋亡；④调节胆碱能系统损伤：乙酰胆碱（ACh）是与学习记忆相关的一种重要的神经递质，姜黄素通过降低乙酰胆碱酯酶活性，增加胆碱乙酰转移酶活性，增加ACh含量而改善ACh的代谢平衡。综上所述，姜黄素可通过抗氧化应激、抗炎症反应、抑制细胞凋亡及调节胆碱能系统损伤等治疗AD。本文对姜黄素在治疗AD作用及机制进行综述，以期为其临床应用及研究提供参考。

关键词：阿尔茨海默病；姜黄素；作用机制

基金项目：河北省自然科学基金（H2020208032）

通讯作者：张丹参，E-mail：zhangds2011@126.com

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氨基酸类神经递质对神经系统疾病的影响及可能机制

杨静涵，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：小分子氨基酸如天冬氨酸（Asp）、谷氨酸（Glu）、甘氨酸（Gly）和γ-氨基丁酸（GABA）是人体内重要的氨基酸类神经递质，在人体内属低水平游离氨基酸，参与维系脑部的血液循环，调控人脑的功能，神经系统疾病如阿尔茨海默病（AD）与脑内氨基酸关系密切。氨基酸之间比例失衡会导致多种神经系统疾病的发生。有研究表明，AD和脑缺血等中枢神经系统疾病模型动物中的兴奋性氨基酸类神经递质Glu和Asp有不同程度的升高；抑制性氨基酸类神经递质GABA和Gly则发生了不同程度的降低，而神经系统疾病的治疗药物可以抑制这种明显的改变，提示一个治疗脑部疾病的新方向。氨基酸类神经递质与神经元信息传递、营养发育和学习记忆等过程有着紧密的联系，Glu/GABA比值在一定范围内的升高可提高学习记忆能力，比值过高则产生抑制作用，蛇床子素可降低AD大鼠脑内Glu和Glu/GABA比值，这可能是其改善认知的机制。神经系统疾病是一类病因复杂、容易复发且难以被及时诊断的疾病，多种神经系统疾病仅能被缓解而无法治愈。因此，深入了解氨基酸类神经递质在此类疾病中的变化，有助于揭示该类疾病发生发展进程及药物作用机制，为该类疾病的药物研发提供新靶点。

关键词：氨基酸；神经递质；神经系统疾病；学习记忆

基金项目：河北省自然科学基金（H2020208032）

通讯作者：张丹参，E-mail：zhangds2011@126.com

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小胶质细胞在治疗神经退行性疾病中的作用

张智超，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：小胶质细胞（MG）来源于卵黄囊，在血脑屏障形成之前进入脑组织，成为大脑中的免疫细胞。MG参与一系列神经退行性疾病（ND）的发生，神经系统紊乱可导致炎症和MG的激活，不同信号刺激MG可呈现不同的活化表型与功能。MG在ND发生中起双重作用，静息态MG被激活，可释放促炎因子，而活化的MG还可释放抗炎因子，对神经元起修复和保护作用。脑卒中和脑缺氧等急性ND中，MG释放炎症介质对神经细胞有益，可减轻继发性损伤；阿尔茨海默病和帕金森病等慢性ND，慢性炎症使得MG被长时程激活，持续释放一系列炎症因子，导致氧化应激，造成神经组织损伤。本文从MG的角度对治疗ND的策略进行综述。①调节MG代谢：MG可通过脑内微环境的代谢变化，改变转录表型和功能，当受到炎症刺激时，MG代谢方式会从氧化磷酸化减少转为糖酵解增加。对糖酵解代谢进行靶向性治疗有望改善神经炎症反应。②抑制MG分泌促炎因子：MG类似巨噬细胞有吞噬、清除作用，可清除掉凋亡神经细胞，疾病早期MG可自行清除，晚期神经毒性作用抑制MG的吞噬作用，分泌的促炎细胞因子会加速疾病的发展。抑制MG分泌促炎因子，激发MG吞噬功能可缓解ND。

关键词：小胶质细胞；神经退行性疾病；治疗策略

基金项目：河北省自然科学基金（H2020208032）

通讯作者：张丹参，E-mail：zhangds2011@126.com

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Imperatorin alleviates Aβ-induced spatial learning memory impairment and neuroinflam⁃mation in model mice of Alzheimer disease

WAN Hang-juan,LUO Li,LIU Xin,HE Wei

(Department of Pharmacology,Gannan Medical University,Key Laboratory of Cerebrovascular Pharmacology of Jiangxi Province,Key Laboratory of Prevention and Treatment of Cardiovascular and Cerebrovascular Diseases of Ministry of Edu⁃cation,Ganzhou 341000,China）

Abstract:OBJECTIVETo investigate the effects of imperatorin on the spatial learning memory impairment and neuroinflammation in model mice of Alzheimer disease(AD)induced by intracerebroventricular injection of Aβ1-42.METHODSMouse model of AD was established by injection of Aβ1-42into the lateral ventricles.Im⁃peratorin(2.5 and 5.0 mg·kg-1,daily)was inject⁃ed by intraperitoneally 1 h after intracerebroven⁃tricular injection for 13 d.The effect of imperato⁃rin on the spatial learning and memory impair⁃ment was assessed by eight arm maze tests.The levels of cytokines TNF-α,IL-1β,IL-6,IL-18 and chemokines MCP-1 in mouse cortex and hip⁃pocampus were detected by ELISA.The protein expression of NF-κB P65,TLR4,MyD88,p-P38,p-ERK,and p-JNK were detected by Western blotting.RESULTSAs compared with the AD model group,imperatorin treatment significantly attenuated Aβ1-42-induced spatial learning and memory impairment assessed by eight arm maze tests.In addition,imperatorin significantly reduced the levels of cytokines TNF-α,IL-1β,IL-6,IL-18 and chemokines MCP-1 in the cerebral cortex and hippocampus.Meanwhile,Western blotting results showed that imperatorin treat⁃ment significantly down-regulated the protein expression of NF-κB P65,TLR4,MyD88,p-P38,p-ERK,and p-JNK.CONCLUSIONImperatorin has neuroprotective effects in the Aβ1-42induced AD model mice and its mechanism may be partially associated with the inhibition of inflam⁃matory response in the cortex and hippocampus.

Key words:imperatorin;Alzheimer disease;Aβ1-42;learning and memory impairment;inflam⁃matory response

Corresponding author:HE Wei,E-mail：hewei86 087@163.com

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欧前胡素对Aβ1-42致阿尔茨海默病模型小鼠神经细胞凋亡Smac/DIABLO信号通路的影响

罗 丽，万航娟，何 蔚

（赣南医学院，江西省脑血管药理重点实验室，心脑血管疾病防治教育部重点实验室，江西 赣州 341000）

摘要：目的研究欧前胡素（Imp）对Aβ1-42致阿尔茨海默病（AD）模型小鼠Smac/DIABLO介导神经细胞凋亡线粒体途径的影响。方法雄性小鼠随机分为正常对照组、AD模型组、AD模型+Imp 2.5 mg·kg-1组和AD模型+Imp 5.0 mg·kg-1组，每组10只。通过脑室内注射Aβ1-42制备小鼠AD模型，Imp（2.5和5.0 mg·kg-1）在手术后当天开始ip给药，每天1次，连续给药13 d。给药后第14天，分离小鼠大脑皮质组织，通过Western印迹法检测皮质组织中Smac/DIABLO、X染色体连锁凋亡抑制蛋白（XIAP）、胱天蛋白酶原3、活化的胱天蛋白酶3、胱天蛋白酶原9、活化的胱天蛋白酶9、聚腺苷二磷酸核糖多聚酶（PARP）和活化的PARP的蛋白表达。通过胱天蛋白酶3和胱天蛋白酶9的活性检测试剂盒检测胱天蛋白酶3和胱天蛋白酶9蛋白活性。结果与正常对照组比较，模型组Smac/DIABLO、活化的胱天蛋白酶3、活化的胱天蛋白酶9和活化的PARP蛋白表达及胱天蛋白酶3和胱天蛋白酶9蛋白活性均显著升高（P＜0.05），XIAP、胱天蛋白酶原3、胱天蛋白酶原9和PARP蛋白表达均显著下降（P＜0.05）；与AD模型组相比，Imp（2.5 和 5.0 mg·kg-1）药物治疗组Smac/DIABLO和活化的胱天蛋白酶3蛋白表达及胱天蛋白酶3和胱天蛋白酶9蛋白活性均显著下降（P＜0.05），活化的胱天蛋白酶9和活化的PARP蛋白表达无明显变化，XIAP和胱天蛋白酶原3的蛋白表达均明显升高（P＜0.05），胱天蛋白酶原9和PARP无明显变化。结论Imp可能通过抑制Smac/DIABLO介导的线粒体途径降低Aβ1-42致AD模型小鼠神经细胞凋亡，从而发挥对AD的神经保护作用。关键词：欧前胡素；阿尔茨海默病；细胞凋亡；线粒体途径

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Effect of rhynchophylline on behavior of zebrafish with Alzheimer disease

LIU Kai-fei,WU Shi-min,LI Xun-yi,HUANG Yao,GUAN Guo-kai,HUANG Xue-hong,CHEN Yi-fei

（Guilin Medical University,Guilin 541000,China）

Abstract:OBJECTIVETo investigate the effect of rhynchophylline on behavior of zebrafish with Alzheimer disease induced by AlCl3.METH⁃ODSTake a video of the zebrafish before train⁃ing,which is convenient for judging whether the zebrafish has been trained successfully.Then,the zebrafish were trained for 7 d.The 60 zebraf⁃ish that were successfully trained were randomly divided into6 groups:normal group,model group,positive group,low-dose rhynchophylline group,middle-dose group,and high-dose group.The normal group was video-recorded,while the model group,positive group,low-dose group,middle-dose group,and high-dose group were given AlCl3for modeling.After that,the model group was videotaped,and the other groups were given drug intervention.Both the positive group and the rhynchophylline administration group were administered for 6 d,and finally all the administration groups were videotaped.After all the video is finished,the behavioral analysis software smart 3.0 was used for behavioral anal⁃ysis,and conclusions are drawn by analyzing the data.RESULTSThe data of the 6 groups of zebrafish staying in the red short arm area were used for comparative analysis:there was a signif⁃icant difference between the normal group and the model group(P＜0.05),the model group and the positive group,and the middle-dose group of rhynchophylline There are significant differences in the high-dose group(P＜0.05).Comparative analysis with the data of the percentage of zebraf⁃ish in the red short arm area of the 6 groups:the normal group and the model group are signifi⁃cantly different(P＜0.05),There were significant differences between the model group and posi⁃tive group,the dose of the rhynchophylline administration group high-dose group and the middle-dose group(P＜0.05).Comparing the data of the swimming distance of the 6 groups of zebrafish in the red short arm area:the normal group and the model group are not significantly different(P＞0.05),while the model group is only significantly different from the high-dose group(P＜0.05).However,the comparison of the percentage of zebrafish swimming distance in the red short arm area of the 6 groups of data:the normal group and the model group are signifi⁃cantly different(P＜0.05),the model group and the positive group,and the rhynchophylline administration group There were significant differ⁃ences between the dose group and the high-dose group(P＜0.05).CONCLUSIONRhynchophyl⁃line can improve the behavior of zebrafish with Alzheimer disease.

Key words：rhynchophylline;Alzheimer disease;zebrafish;behavioral analysis

Foundation item:National Natural Science Foundation of China(8216140711);Natural Sci⁃ence Foundation of Guangxi Zhuang Autono⁃mous Region(2017GXNSFAA198255);Natural Science Foundation of Guangxi Zhuang Autono⁃mous Region(2018GXNSFBA138028);Guangxi Key Laboratory of Brain and Cognitive Neurosci⁃ence Open Project Funding(GKLBCN-2018010 5-03);2019 College Student Innovation and En⁃trepreneurship Project Training Plan(20191060 1038);and the Fourth Training Plan for Thou⁃sands of Young and Mid-aged Mainstay Teach⁃ers in Guangxi Colleges and Universities.

Corresponding author:CHEN Yi-fei,E-mail:chenyifei_pharm@glmc.edu.cn

T1-8

雌激素介导线粒体途径的神经保护作用

戴月英，赵雨薇，甄艳杰，沈丽霞

（河北北方学院药学系，河北 张家口 075000）

摘要：阿尔茨海默病（AD）是多因素引起的神经退行性疾病，以认知障碍、执行力障碍为主要的临床表现，发病机制有Aβ级联和Tau蛋白异常学说。诸多研究表明，Aβ沉积和Tau过度磷酸化之间存在密切的联系。除此之外，由于线粒体被视为细胞的能量体，因而AD线粒体机制也成为AD发病机制研究领域。Aβ可直接与线粒体膜结合，从而改变线粒体动力学和功能，导致能量代谢异常，最终引起细胞凋亡等一系列链式反应。研究表明，线粒体功能障碍会加剧Aβ沉积和Tau蛋白异常磷酸化，而这2种病理变化反过来又能促进线粒体损伤，通过线粒体依赖性凋亡通路来诱导细胞凋亡。流行病学调查显示，绝经后妇女较同龄男性AD发病率高1.5～3.0倍，认为与女性绝经后雌激素降低相关。研究表明，雌激素有神经元保护作用。雌激素通过线粒体途径发挥神经保护作用及其机制的研究，为AD防治提供一定的思路。

关键词：阿尔茨海默病；雌激素；线粒体

基金项目：河北省自然科学基金（H2019405057）；河北省高等学校科学技术研究项目（ZD2020136）

通讯作者：沈丽霞，E-mail：shenlixiacn@163.com

T1-9

槲皮素神经保护作用研究进展

赵雨薇，戴月英，甄艳杰，沈丽霞

（河北北方学院药学系，河北 张家口 075000）

摘要：阿尔茨海默病（AD）主要病理特征有β-淀粉样蛋白沉积形成的斑块、胞内Tau蛋白异常磷酸化导致的神经纤维缠结、神经元突触缺失与凋亡。雌激素替代治疗在防治和延迟AD发生的同时，也增加了致癌的风险，限制了其在临床上的应用。槲皮素作为一种植物雌激素，与雌激素结构特征相似，可发挥雌激素样神经保护作用，且无致畸、致癌和致死作用，因此受到了广泛关注。研究表明，槲皮素能调控核因子E2相关因子2易位，维持氧化还原平衡和线粒体功能的稳定；介导MAPK信号通路，抑制对氧磷酶2的表达，抵抗氧化应激损伤；抑制β-分泌酶（BACE1）基因表达，促进非淀粉样蛋白途径，减少Aβ积聚；激活PI3K/AKT/GSK-3β信号通路，抑制Tau蛋白异常磷酸化；降低一氧化氮和一氧化氮合酶蛋白水平，减少核转录因子κB的核转位，发挥抗炎特性；还可与雌激素受体结合，对神经元凋亡起到保护作用。槲皮素的神经保护作用机制复杂，涉及多条信号通路相互作用。

关键词：阿尔茨海默病；槲皮素；植物雌激素；神经保护

基金项目：河北省自然科学基金（H2019405057）；河北省高等学校科学技术研究项目（ZD2020136）

通讯作者：沈丽霞，E-mail：shenlixiacn@163.com

T1-10

姜黄素类似物促神经分化活性及其作用机制

房红志1，吴 敏1，阳泽界1，石 浩1，许文博1，张 双1，李莎莉1，唐根云1，2

（湖南医药学院1.基础医学院，2.脑与神经内分泌疾病湖南省高等学校重点实验室，湖南 怀化 418000）

摘要：目的对姜黄素类似物进行促神经分化活性筛选，发现活性最佳的化合物后进行分子作用机制研究。方法利用Neuro-2a细胞进行药物毒性和促神经分化活性筛选，用MTT法进行细胞毒性实验；药物诱导Neuro-2a分化后，采用神经突起专一性抗体β-tubulin Ⅲ进行免疫荧光染色，拍照后用Image J软件对细胞分化率和神经突的长度进行测量，进行统计分析。用Western印迹法对活性最佳化合物激活的分子信号通路关键蛋白进行检测，并用药理学抑制剂进行验证。结果通过对12个姜黄素类似物的促神经分化活性筛选，其中双去氧基姜黄素具有较好的促神经分化活性，在很低的浓度就能够促进Neuro-2a细胞分化，1，2和4 μmol·L-1均能够促进细胞分化率和神经突起长度显著增加，并且成浓度依赖性。随后采用4 μmol·L-1进行分子作用机制研究，发现双去氧基姜黄素通过激活ERK/MEK和PI3K/AKT通路诱导Neuro-2a细胞分化，并且用抑制剂U0126和LY200942得到验证。结论双去氧基姜黄素具有较好的促神经分化活性，对治疗神经退行性疾病具有潜在的研究价值。

关键词：姜黄素；神经分化；抑制剂

基金项目：国家自然科学基金（81703821）；湖南省教育厅项目（2018B530）；湖南医药学院自然科学基金（2015ZKB01）；国家级大学生创新创业训练计划（S202012214012）；国家级大学生创新创业训练计划（S202012214012）

T1-11

Glutathione peroxidase 4 deficit-mediated fer⁃roptosis contributes to dopaminergic neuron degeneration under synucleinopathy

SUN Jie1,2,3,4＊, LU Dan-hua1,2,3,4＊, LI Kun1,2,3,4,LIN Xiao-min1,2,3,4, PAN Ming-hai1,2,3,4, GONG Hai-biao1,2,3,4,SUN Wan-yang1,2,3,WANG Meng1,2,3,4,LIANG Lei1,2,3,4,Kurihara Hiroshi1,2,3,LI Yi-fang1,2,3,DUAN Wen-jun1,2,3,4,ZHANG Li5,HE Rong-rong1,2,3,4

[1.Guangdong Engineering Research Center of Chinese Medicine & Disease Susceptibility,2.International Cooperative Laboratory of Tradi⁃tional Chinese Medicine Modernization and Innovative Drug Development of Chinese Ministry of Education (MOE),College of Pharmacy,3.Guangdong Province Key Laboratory of Phar⁃macodynamic Constituents of TCM and New Drugs Research,College of Pharmacy,4.School of Traditional Chinese Medicine,5.Key Laboratory of Central CNS Regeneration(Ministry of Educa⁃tion),Guangdong-Hong Kong-Macau Institute of CNS Regeneration,Jinan University,Guangzhou 510632,China]

Abstract:OBJECTIVEIntracellular aggre⁃gation of α-synuclein(SNCA)is one of the core pathological features of neurodegenerative disor⁃ders including Parkinson disease,whilst the detailed mechanism for consequently neuron loss has not been fully illustrated.Since the altered phospholipid homeostasis has been suggested to play a role in synucleinopathy,this study aims to depict the fully-featured status of phospholip⁃ids and the targets reposing α-synuclein-related neurotoxicity.METHODSSNCAA53T transgenic mice were utilized as the model of Parkinson disease.Behavioral tests including pole test,rotarod test and gait analysis were conducted to assess the motor features of Parkinsonism.Tyro⁃sine hydroxylase were determined by immunohis⁃tochemistry.Glutathione,dopamine,3,4-dihy⁃droxyphenylacetic acid and homovanillic acid were determined by HPLC-ECD analysis.Assess⁃ment of lipid peroxidation included MDA assay and Liperfluo staining.Phospholipid-omics was analyzed based on LC-MS/MS.Investigation on mechanism was relied on Western blotting and qPCR assay.The injection of AAV into midbrain was achieved by ultra-micro injection pump to obtain the target genotype.RESULTSSNCAA53T transgenic mice displayed progres⁃sively deteriorated motor coordination functions and the mechanisms were related with lipid per⁃oxidation and ferroptosis,which might help to explain the parkinsonism.These hydroperoxides were observed on plasm membrane and were further characterized by LC-MS/MS-based phos⁃pholipid-omics analysis.α-synucleinA53T trans⁃genic mice displayed distinct patterns of phos⁃pholipid peroxidation in midbrain regions com⁃pared to wild type littermates.Among different subtypes of oxidized phospholipids,oxidative phosphatidylcholine(PC-ox)was more promi⁃nently elevated.Phospholipid peroxidation is believed as a biomarker of ferroptosis,which is largely a specialized death program caused by insufficiency of glutathione peroxidase-4(GPX4),the only known enzyme that can reduce lipid hydroperoxides within biological membranes.The deficiency of Gpx4 was demonstrated to be responsible for α -synuclein-induced lipid peroxi⁃dation,and the cell lines and mouse models underwent genetic Gpx4 downregulation showed exacerbated dopaminergic neuron loss and par⁃kinsonism.On the other hand,the potentiation of Gpx4 expression remarkably inhibited dopami⁃nergic ferroptotic death and behavioral deficits in a mouse model with synucleinopathy.CONCLU⁃SIONA cellular pathway that Gpx4 deficit-medi⁃ated phospholipid peroxidation and behavioral consequence participated in synucleinopathy,suggesting a potential strategy targeting Gpx4 to alleviate PD symptoms.

Key words:Parkinson disease;ferroptosis;glutathione peroxidase-4; phospholipid-omics;α-synuclein

Foundation item:National Key Research and Development Program of China(2017YFC1700 404);Natural Science Foundation of China(81873209);Natural Science Foundation of China(U1801284);Natural Science Foundation of China(81903821);Natural Science Foundation of China(81973718);the Local Innovative and Research Teams Project of Guangdong Pearl River Talents Program(2017BT01Y036);GDUPS(2019),Fun⁃damental Research Funds for the Central Univer⁃sities(21620448);Natural Science Foundation of Guangdong Province(2019A 1515010909);Nat⁃ural Science Foundation of Guangdong Prov⁃ince (2021A1515011297);Science and Tech⁃nology Program of Guangzhou(201903010062);Science and Technology Program of Guang⁃zhou(907158833068),and the Innovation Team Project of Guangdong Provincial Department of Education(2020KCXT D003)

Corresponding author:HE Rong-rong,E-mail:rongronghe@jnu.edu.cn

＊Co-first author.

T1-12

Therapeutic efficacy of novel memantine nitrate MN-08 in animal models of Alzheimer disease

WU Liang-miao,ZHOU Xin-hua,CAO Yi-wan,Shing Hung MAK,ZHA Ling,LI Ning,SU Zhi-yang,HAN Yi-fan,WANG Yu-qiang,Maggie Pui Man Hoi,SUN Ye-wei,ZHANG Gao-xiao,YANG Xi-fei,ZHANG Zai-jun

(Institute of New Drug Research,Jinan University College of Pharmacy,Guangzhou 510632,China)

Abstract:OBJECTIVEAlzheimer disease(AD)is a leading cause of dementia in elderly individuals and therapeutic options for AD are very limited.Over-activation of N-methyl-D-aspar⁃tate(NMDA)receptors,amyloid β (Aβ)aggrega⁃tion,a decrease in cerebral blood flow(CBF),and downstream pathological events play impor⁃tant roles in the disease progression of AD.This study seeks to explore the efficacy and mecha⁃nism of action of MN-08,a novel memantine ni⁃trate,in established animal models of AD.METH⁃ODSMN-08′s effectiveness as a preventative and therapeutic agent was tested in 2-to 8-month-old APP/PS1 transgenic mice and 9-to 12-month-old 3×Tg-AD mice,respectively.The neuroprotective mechanism of MN-08 was tested in the glutamate cell model.The pharmacokinet⁃ics and safety of MN-08 in vivo were determined in normal rats and beagle dogs.For the behavioral test,Western blotting analysis,pathology,ELISA test and in vitro cell tests,investigators were blinded to the experimental grouping and drug treatment.RESULTSMN-08,a novel meman⁃tine nitrate,was found to inhibit Aβ accumulation,prevent neuronal and dendritic spine loss,and consequently attenuate cognitive deficits in 2-month-old APP/PS1 transgenic mice(for a 6-month preventative course)and in the 8-monthold triple-transgenic(3×Tg-AD)mice(for a 4-month therapeutic course).In vitro,MN-08 could bind to and antagonize NMDA receptors,inhibit the calcium influx,and reverse the dysregula⁃tions of ERK and PI3K/Akt/GSK3β pathway,sub⁃sequently preventing glutamate-induced neuro⁃nal loss.In addition,MN-08 had favorable phar⁃macokinetics,blood-brain barrier penetration,and safety profiles in rats and beagle dogs.CON⁃CLUSIONThe novel memantine nitrate MN-08 may be a useful therapeutic agent for AD.

Key words:Alzheimer disease;MN-08;meman⁃tine nitrate

Corresponding author:ZHANG Zai-jun,E-mail:zaijunzhang@jnu.edu.cn

T1-13

TBN improves motor function and prolongs survival in SOD1G93Aand TDP-43M337Vmouse model of amyotrophic lateral sclerosis

HUANG Chun-hui,LI Jun,ZHANG Gui-liang,LIN Ying-qi,LI Cai-juan,ZHENG Xiao,SONG Xi-cheng,HAN Bo-feng,GUO Bao-jian,TU Zhu-chi,ZHANG Jun,SUN Ye-wei,WANG Yu-qiang,YAN Sen,ZHANG Zai-jun

(Institute of New Drug Research,Jinan University College of Pharmacy,Guangzhou 510632,China)

Abstract:OBJECTIVEAmyotrophic lateral sclerosis(ALS)is a fatal neurodegenerative dis⁃ease characterized by progressive loss of upper and lower motor neurons that results in skeletal muscle atrophy,weakness and paralysis.Oxida⁃tive stress plays a key role in the pathogenesis of ALS,including familial forms of the disease arising from mutation of the gene coding for superoxide dismutase 1(SOD1).Moreover,although the pathogenesis of ALS is unclear,the abnormal accumulation of TAR DNA-binding pro⁃tein of 43 ku(TDP-43)is a pathological feature that exists in almost all patients.Thus far,there is no drug that can cure ALS/FTLD.Tetramethyl⁃pyrazine nitrone(TBN)is a derivative of tetra⁃methylapyrazine,derived from the traditional Chinese medicine Ligusticum chuanxiong,which has been widely proven to have therapeutic effects on models of various neurodegenerative diseases.TBN is currently under clinical investi⁃gation for several indications including a phase Ⅱtrial of ALS.Here,we explored the therapeutic effect of TBN in the SOD1G93A and TDP-43M337VALS mouse model.METHODSIn the SOD1G93A transgenic mouse model,TBN was administered to mice by intraperitoneal or intragastric injection after the onset of motor deficits.At the same time,we unilaterally and bilaterally injected the TDP-43M337Vvirus into the striatum of the WT mouse,and gave the TBN treatment after the mice developed a phenotype.After administering these two models for a period of time,we con⁃ducted behavioral tests,including rotarod test,balance beam test,climbing pole test,etc,to evaluate the efficacy of TBN on SOD1G93A and TDP-43M337Vmodels.Furthermore,we explored the possible mechanism of action of TBN in the treatment of ALS through Western blotting and immunohistochemistry/immunofluorescence staining analysis.RESULTSIn the SOD1G93A transgenic mouse model,TBN slowed the pro⁃gression of motor neuron disease as evidenced by improved motor performance,reduced spinal motor neuron loss and the associated glial response,and decreased skeletal muscle fiber denervation and fibrosis.TBN treatment activated mitochondrial antioxidant activity through the PGC-1α/Nrf2/HO-1 pathway and decreased the expression of human SOD1.What′s more,in the TDP-43M337Vmice model,the results showed that in mice with unilateral injection of TDP-43M337V,TBN improved motor deficits and cognitive im⁃pairment in the early stages of disease progres⁃sion.In mice with bilateral injection of TDP-43M337Vinto the striatum,TBN not only improved motor function but also prolonged survival rate.Moreover,we show that its therapeutic effect may be through activation of the Akt/mTOR/GSK-3β and AMPK/PGC-1α/Nrf2 signaling pathways.In summary,TBN is a promising agent for the treat⁃ment of ALS/FTLD.CONCLUSIONTBN has shown good efficacy in both SOD1 and TDP-43 ALS-related models,and it may act by activating the AMPK/PGC-1α/Nrf2 signaling pathway,which shows some light for the development of ALS therapeutic drugs.

Key words:amyotrophic lateral sclerosis;tetra⁃methylpyrazine nitrone;superoxide dismutase 1

Corresponding author:ZHANG Zai-jun,E-mail:zaijunzhang@jnu.edu.cn

T1-14

PDE4抑制剂罗氟普兰通过促进自噬抑制小胶质细胞活化作用机制

刘 璐，汪海涛，徐江平

（南方医科大学药学院，广东 广州 510515）

摘要：目的神经炎症参与神经退行性疾病的发展进程，而小胶质细胞活化是神经炎症的特征之一。罗氟普兰（ROF）是一种新型致呕吐潜能低的磷酸二酯酶4（PDE4）抑制剂。我们之前发现，ROF可能通过促进自噬从而抑制活化的小胶质细胞中促炎因子的产生，但其机制尚不明确。本研究旨在探究ROF促进自噬抑制小胶质细胞活化作用及机制。方法使用Aβ25-35诱导BV-2细胞活化，采用ELISA试剂盒检测白细胞介素6（IL-6）、IL-1β和肿瘤坏死因子α（TNF-α）含量；建立神经炎症细胞及炎症小体活化细胞模型，模型条件下用自噬激活剂雷帕霉素诱导自噬，用自噬特异性抑制剂3-MA等抑制自噬及转染PDE4质粒，实现敲低PDE4基因；使用AMPK抑制剂和ULK1抑制剂评价是否阻断药物作用，上述情况下检测自噬相关信号通路中关键蛋白及激酶的变化。在C57小鼠侧脑室定位注射Aβ25-35构建小鼠炎症模型，通过新物体识别等行为学实验方法观察模型小鼠学习记忆能力的损伤及PDE4抑制剂能否改善学习记忆的能力。取脑组织，采用免疫荧光双标法检测海马和皮质中小胶质细胞自噬相关蛋白的表达。用ELISA和Western印迹法分别检测小胶质细胞炎症因子的水平及M1和M2表型标志物的水平。结果①Aβ25-35处理BV-2小胶质细胞显著增加细胞的活化水平，ROF可以有效减轻Aβ25-35诱导的小胶质细胞活化，并降低Iba1的表达水平；同时，给予ROF或者敲低PDE4B能显著降低模型条件下炎症因子的水平。②Aβ25-35处理BV-2小胶质细胞后自噬水平下降，给予ROF，显著上升；在给予ULK1抑制剂后，ROF的促自噬作用则被阻断。③小鼠侧脑室注射凝聚态的Aβ25-35，明显影响小鼠的学习记忆能力；ig给予ROF改善其学习记忆能力，海马和皮质中小胶质细胞活化水平也明显下降，促炎因子降低，其自噬蛋白水平升高；ig给予ROF和ULK1抑制剂组逆转这些现象。结论PDE4抑制剂ROF可能通过AMPK/ULK1信号转导通路促进自噬，从而抑制Aβ25-35诱导BV-2小胶质细胞炎症因子，减轻海马和皮质中小胶质活化，也可能依赖ULK1蛋白，从而产生神经保护作用。

关键词：磷酸二酯酶4；罗氟普兰；神经炎症；小胶质细胞

基金项目：国家自然科学基金（81773698）；国家自然科学基金（81974501）

通讯作者：徐江平，E-mail：jpx@smu.edu.cn

T1-15

Urolithin A protects dopaminergic neurons in experimental models of Parkinson disease by promoting mitochondrial biogenesis through SIRT1/PGC-1α signaling pathway

LIU Jia,QIU jing-ru,WANG Bao-zhu,SUN De-qing,YU Shu-yan,LOU Hai-yan

(Department of Pharmacology,School of Basic Medical Sciences,Cheeloo College of Medicine,Shandong University,Jinan 250012,China)

Abstract:OBJECTIVEMitochondrial dys⁃function contributes to the pathogenesis of neuro⁃degenerative diseases such as Parkinson dis⁃ease(PD).Therapeutic strategies targeting mito⁃chondrial dysfunction hold considerable promise for the treatment of PD.Urolithin A(UA)is a gut metabolite produced from ellagic acid-containing foods such as pomegranates,berries,and wal⁃nuts.Recent reports have highlighted the protec⁃tive role of UA in several neurological disorders including Alzheimer disease and ischemic stroke.However,the potential role of UA in PD has not been characterized.In this study,the role of UA in 6-OHDA-induced neurotoxicity in cell cultures and mouse model of PD was investi⁃gated.METHODSIn vitro,PC12 cells were exposed to 6-OHDA in the presence or absence of UA.For in vivo study,C57BL/6 mice were ste⁃reotactic injected with 6-OHDA to induce experi⁃mental PD model.UA(10 mg·kg-1)was intraperi⁃toneal injected for 7 d before surgery.RESULTSUA protected against 6-OHDA cytotoxicity and apoptosis in PC12 cells.Prior administration of UA to 6-OHDA lesioned mice ameliorated both motor deficits and nigral-straital dopaminergic neurotoxicity.Moreover,UA attenuated 6-OHDA-induced mitochondrial dysfunction in PC12 cells accompanied by enhanced mitochondrial biogen⁃esis.Mechanically,the neuroprotective effects of UA were mediated by SIRT1-PGC-1α signalingmediated mitochondrial biogenesis.CONCLU⁃SIONThese data provide new insights into the novel role of UA in promoting mitochondria bio⁃genesis and suggest that UA may have potential therapeutic applications for PD.

Key words:urolithin A;Parkinson disease;mito⁃chondrial biogenesis

T1-16

Effect of betaine on sustained hypertension induced learning and memory injury in SHR rats

CHEN Jiang-yan,HAN Wen-dong,GAO Shan

(Shandong First Medical University,Jinan 271000,China)

Abstract:OBJECTIVETo observe the effects of betaine on learning memory in SHR rats through behavioral experiments,and then to investigate the mechanisms of betaine to improve the cognitive impairment caused by hyperten⁃sion.METHODS4-month-old male SHR rats were randomly divided into 4 groups,including betaine 10,30 and 100 mg·kg-1groups and SHR group,and administered by gavage,10 mL·kg-1once a day for 4 weeks.The control group was WKY rats,and the same amount of saline was given by gavage.The learning and memory behaviors of the rats were analyzed and evaluated through behavioral testing.RESULTSThe results of Morris water maze showed that persistent hypertension decreased the time in the target quadrant and the number of platform crossing of the rats,and the intervention with different con⁃centrations of betaine increased the time in the target quadrant and the number of platform crossing of SHR in a dose-dependent manner.Then,persistent hypertension increased the escape latency of rats,and betaine inhibited this change.The results of new object recognition showed that the recognition index of SHR group decreased,and the recognition index of SHR rats increased with different concentrations of betaine intervention in a dose-dependent manner.The Y-maze results showed that persistent hypertension decreased the alternation in rats,and the alternation in SHR rats increased after the intervention with different concentrations of betaine,and in a dose-dependent manner.CON⁃CLUSIONPersistent hypertension can cause learning memory impairment in rats.Betaine intervention has an ameliorative effect on the reduction of learning memory ability in rats caused by hypertension.

Key words:betaine;hypertension;learning and memory

Foundation item:National Natural Science Foundation of China(81400182);and Natural Science Foundation of Shandong Province(BS2014YY045)

T1-17

Baroreflex deficiency aggravates learning and memory disorders in rats

ZHU De-rong＊,FENG Zhao-yang＊,JI Wei,QI Hui-bin,KONG De-ping,ZHANG Fang-fang,YU Hai-yang,GAO Yong-feng,TAN Rui,ZHAO Xiao-min

(Institute of Pharmacology,Shandong First Medi⁃cal University & Shandong Academy of Medical Sciences,Tai′an 271016,China)

Abstract:OBJECTIVEMicroglia M1/M2 po⁃larization play pro-inflammatory and anti-inflam⁃matory roles,respectively,which is involved in memory decline.There is a close relationship between impaired baroreflex function and memory impairment.The present study was designed to investigate whether arterial baroreflex deficiency induced by sinoaortic denervation(SAD)affected inflammation through modulation of M1/M2 polar⁃ization leading to the aggravation of learning and memory disorders in rats.METHODSAdult male SD rats were divided into four groups:the sham control,the SAD,the sham+scopolamine,the SAD+scopolamine.In another experiment,there were also four groups:the sham control,the SAD,the SAD+scopolamine and the SAD+scopolamine+ketanserin.All rats were examined for various behaviors using Morris water maze test,new object recognition test,and light dark shuttle test and Y maze test 4 weeks after sham or SAD surgery.CD16,CD206,IL-10,IL-6,IL-1β and TNF-α from hippocampus using Western blotting,immunofluorescence and turbidimetry.RESULTSCompared with the sham+scopol⁃amine,the SAD+scopolamine rats showed the reduced crossing times in Morris water maze test,the longer residence time in dark box during light dark shuttle test,and the decreased alterna⁃tion ratio in Y maze test.The level of CD206,IL-10,T-AOC and GSH was decreased,whereas CD16,IL-6,TNF-α,MDA was increased in the hippocampus of SAD+scopolamine rats.Addi⁃tionally,all the above changes were improved in the SAD+scopolamine+ketanserin rats when compared with the SAD+scopolamine.CONCLU⁃SIONArterial baroreflex dysfunction aggravates learning and memory disorders in rats,which may be related to the polarization of microglia.

Key words:arterial baroreflex;learning and memory;microglia;polarization;ketanserin

Foundation item:Natural Science Foundation of Shandong Province(ZR2017MH048)

Corresponding author:ZHAO Xiao-min,E-mail:xmzhao@sdfmu.edu.cn;TAN Rui,E-mail:rtan@sdfmu.edu.cn.

＊Co-first author.

T1-18

甜菜碱通过RAS影响大鼠高血压蓄积致学习记忆障碍

韩文东，陈姜艳，刘丽媛，王 浩，高 山

〔山东第一医科大学（山东省医学科学院），山东泰安 271016〕

摘要：目的高血压蓄积会导致体内氧自由基增多和炎症反应加剧，从而诱发许多心脑疾病的发生，其中包括学习记忆障碍。本研究探讨甜菜碱（Bet）是否通过抗氧化对高血压导致学习记忆障碍大鼠产生保护作用。方法雄性SD大鼠，160～180 g，制备2K1C高血压模型。造模成功后，随机分为6组：对照组、模型组、Bet 10 和 30 mg·kg-1、抑制剂组（A779+Bet 30 mg·kg-1）和A779组，给药4周。采用水迷宫、新物体识别和Y迷宫检测大鼠学习记忆能力；用Western印迹法检测海马中APP，Aβ，PS-1，Catalase和HO-1蛋白表达；ELISA检测血浆中Ang1-7含量。结果①行为学实验结果表明，穿越平台次数、目标象限停留时间、总探索时间、认知指数、总进臂次数和交替比模型组显著低于对照组，给予Bet可抑制此作用，并呈剂量依赖性。②Western印迹法结果显示，与对照组相比，模型组大鼠海马中APP，Aβ和PS-1含量均增加，盐酸Bet可抑制此变化，并呈剂量依赖性；模型组海马中Catalase和HO-1含量均低于对照组，盐酸Bet可抑制此变化并呈剂量依赖性。③ELISA结果显示，模型组血浆中Ang1-7含量均低于对照组，盐酸Bet可抑制此变化，并呈剂量依赖性；A799可逆转上述盐酸Bet的作用。结论盐酸Bet对高血压蓄积致学习记忆障碍的保护可能是通过作用于Ang1-7/Mas轴，增强抗氧化作用，从而改善大鼠学习记忆障碍。

关键词：甜菜碱；高血压蓄积；学习记忆障碍；氧化应激

基金项目：国家自然科学基金青年科学基金（81400182）；山东省博士基金（BS2014YY045）

通讯作者：高 山，E-mail：gshan84117@163.com

T1-19

组胺及其受体在阿尔茨海默病发病中的生物学作用

刘丽媛，张 猛，王 浩

（山东第一医科大学药理学研究所，山东 泰安 271016）

摘要：阿尔茨海默病（AD）为中枢退行性疾病。组胺为内源性生物胺，与其受体构成中枢组胺能系统，在脑稳态的维持和高级脑功能中发挥重要作用。近年发现，组胺及其受体在神经调控及AD发生发展中发挥关键作用，可能是治疗AD的潜在靶点。本文从组胺受体的脑内分布及功能、组胺在AD发生发展中的作用及机制对组胺及其受体在AD发病中的生物学作用进行综述。组胺通过激动其受体发挥作用。组胺受体包括H1～H4亚型，4种亚型在脑内均有分布，发挥多种生物学功能，其中目前研究最广泛的是组胺H3受体（H3R），其分布于下丘脑、大脑皮质、海马、杏仁核、纹状体和基底神经节等多个部位。H3R通过对组胺合成的反馈抑制来调控组胺的释放。同时，其也可作为异源受体，参与细胞信号转导、调控其他神经递质的释放。H3R可能通过胆碱能通路、Aβ损伤、Tau蛋白通路和海马神经发生等多种机制参与AD的发生。总之，组胺与其受体组成的组胺能系统广泛分布于中枢神经系统，与AD发生密切相关，其中组胺H3R拮抗剂研究广泛，可通过多种机制改善AD，可成为AD的治疗的潜在靶点。

关键词：阿尔茨海默病；组胺；胆碱能通路；Aβ损伤；Tau蛋白

基金项目：国家自然科学基金（81441111）；国家自然科学基金（81601229）

T1-20

Inhibition of glycolysis mitigate microglialactivation mediated neuroinflammation in vitro and in vivo

CHENG Jun-jie1,2,SUN Ren-juan2,ZHEN Xue-chu2,ZHENG Long-Tai2

(1.Department of Pharmacy,Shenzhen Children′s Hospital,Shenzhen 518038,China;2.Jiangsu Key Laboratory of Neuropsychiatric Diseases and College of Pharmaceutical Sciences,Soochow University,Suzhou 215123,China)

Abstract:OBJECTIVEMicroglial activationmediated neuroinflammation plays an important pathological basis in the progression of many neurodegenerative diseases.Activated microglia cells show a metabolic shift from oxidative phos⁃phorylation to aerobic glycolysis.However,the molecular mechanism underlying the role of glycolysis in microglial activation and progres⁃sion of neuroinflammatory diseases have not yet been fully understood.METHODSThe antiinflammatory effects and its underlying mecha⁃nisms of glycolytic inhibition in vitro were exam⁃ined in lipopolysaccharide(LPS)activated BV-2 microglial cells or primary microglial cells by enzyme-linked immunosorbent assay (ELISA),quantitative reverse transcriptase polymerase chain reaction (RT-PCR),Western blotting,immunoprecipitation,Flow cytometry and nuclear factor kappa B (NF-κ B)luciferase reporter assays.In vivo,the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)-or LPS-induced Par⁃kinson disease(PD)models were constructed to explored the anti-inflammatory and neuropro⁃tective effects of glycolytic inhibitor.RESULTSInhibition of glycolysis by specific inhibitors[2-DG and 3-bromopyruvic acid(3-BPA)],knockdown of glucose transporter type 1(Glut-1)or hexoki⁃nase(HK) Ⅱ abolished LPS-induced expres⁃sion of proinflammatory genes in microglia cells.Mechanistic studies demonstrated that glyco⁃lytic inhibitors significantly inhibited LPS-induced mTOR phosphorylation,IKKβ phosphorylation,IκB phosphorylation,IκB degradation,nuclear translocation of P65 and NF-κ B luciferase activity.Furthermore,LPS-induced P65 acetyla⁃tion on lysine 310,which is mediated by NAD-dependent protein deacetylase sirtuin-1 and is critical for NF-kB activation,were inhibited by glycolytic inhibitors.A coculture study revealed that 2-DG reduced the cytotoxicity of activated microglia toward MES23.5 dopaminergic neuron cells with no direct protective effect.In vivo,2-DG significantly ameliorated MPTP or LPS induced DA neuron loss and glial cell activation.CONCLUSIONGlycolysis is actively involved in microglial activation.Inhibition of glycolysis can ameliorate microglial activation-related neuroinflammatory diseases.

Key words:microglia cells;neuroinflammatory diseases;glycolytic inhibition

Corresponding author:ZHENG Long-tai,E-mail:zhenglongtai@suda.edu.cn;ZHEN Xue-chu,E-mail:zhenxuechu@suda.edu.cn

T1-21

山茱萸有效成分对老年快速老化小鼠的药理作用及其机制

李雅莉，马登磊，张 兰，李 林

（首都医科大学宣武医院药学部，神经变性病教育部重点实验室，北京市神经药物工程研究中心，北京 100053）

摘要：目的阿尔茨海默病（AD）是一种与年龄相关，以神经元退行性病变导致个体行为异常为主要特征的神经系统疾病。快速老化小鼠（SAM）是具有快速老化特征的小鼠品系，SAMP8小鼠表现不可逆的快速老化，表现与老年人和AD患者类似的老化症状。山茱萸环烯醚萜苷（CIG）是本室从中药山茱萸中提取的有效部位。方法给予12月龄SAMP8小鼠ig CIG治疗至14月龄后，观察SAMP8小鼠的行为学改变和老化状态等变化及相应的作用机制。结果①CIG能降低14月龄SAMP8小鼠的老化指数评分（根据小鼠反应性、皮毛等程度进行评分，指数越高表明小鼠的老化程度越严重），增高存活率，改善转棒实验中运动协调能力。②CIG能降低SAMP8小鼠纹状体区tau蛋白在Thr205和Ser396位点的过度磷酸化。③CIG能降低APP相关代谢蛋白的表达。④CIG增加突触相关蛋白表达。⑤GIG降低坏死性凋亡相关蛋白表达。结论CIG具有延缓衰老的作用，有利于预防和治疗AD，并且对于晚期AD患者可能起到改善生活质量的作用。

关键词：山茱萸；快速老化小鼠；tau蛋白

T1-22

sPINK1抑制泛素依赖蛋白酶体的活性损伤神经元

陈 聪，易华伟，沈 晨，高童谣，唐 淳，张纬萍

（浙江大学医学院，浙江 杭州 310043）

摘要：目的PINK1有线粒体膜定位的fPINK1和胞浆分布的sPINK1，2种PINK1在老年人、阿尔茨海默病患者脑内的神经元均显著增加，它们都能够磷酸化泛素Ser65位点。fPINK1通过磷酸化泛素诱导线粒体自噬，发挥神经保护作用，而sPINK1的作用尚不清楚。本文揭示sPINK1在神经元中的作用及机制。方法通过噬神经元的腺相关病毒（AAV）在小鼠海马CA1神经元，分别过表达GFP、sPINK1和sPINK1及S65A突变的泛素（Ub/S65A，不能被磷酸化的泛素）和Ub/S65E（模拟磷酸化的泛素），通过分析小鼠神经行为、形态和生化等变化，揭示sPINK1的作用，并在蛋白质水平分析sPINK1的作用机制。结果过表达sPINK1可损伤小鼠海马依赖的学习记忆功能，导致神经元损伤，包括线粒体肿胀、神经突起发生退行性变化和突触减少，以及神经元内的蛋白质聚集增加、星形胶质细胞和小胶质细胞反应性激活。sPINK1的作用能够被Ub/S65A抑制，而Ub/S65E能够模拟sPINK1的作用。在蛋白质水平，泛素化的GFP能够被纯化的蛋白酶体降解；而泛素被sPINK1磷酸化后，GFP被蛋白酶体降解的速率显著降低。结论sPINK1可通过磷酸化泛素，抑制泛素依赖的蛋白质降解，损伤神经元。

关键词：PINK1；蛋白酶体；神经元

T1-23

Roles of NAMPT and NAD decline in patho⁃genesis of Parkinson disease in mice

CHEN Cong,SHEN Chen,GAO Tong-yao,WANG Tong,LU Yun-bi,ZHANG Wei-ping

（Zhejiang University School of Medicine,Hangzhou 310043,China）

Abstract:OBJECTIVENicotinamide phos⁃phoribosyltransferase (NAMPT) is the key enzyme for the synthesis of nicotinamide ade⁃nine dinucleotide (NAD) in the salvaging pathway.NAD is an essential co-enzyme of multiple enzymes involved in cell metabolism and important enzymes such as sirtuins.The level of both NAMPT and NAD decreases upon aging,which may cause neuronal degeneration.However,the roles of NAMPT and NAD in Par⁃kinson disease(PD)remain unknown.This study was to explore the roles of NAMPT and NAD decline in the pathogenesis of PD in mice.METOHDSFloxed nampt gene C57BL/6J mice,including Namptwt/wt,Namptflox/wtand Namptflox/flox,were used.The rAAV-hSyn-Cre-WPRE pA or rAAV-TH-Cre-WPRE pA adeno-associated virus(AAV)was used to conditioning knockout nampt gene in neurons or dopaminergic neurons,respectively.At 2,4,6,and 8 weeks after AAV injection,the motor deficits of mice were evaluat⁃ed.Immunofluorescence and immunohistochem⁃istry were used to evaluate the neuronal injury.Transmission electron microscope was used to evaluate the axonal degeneration.RESULTSThe knockout of nampt gene induced dopaminer⁃gic neuron loss in substantia nigra at 4 weeks but not 2 weeks after AAV injection.At 8 weeks after AAV injection,the Namptflox/floxmice showed a significantly decrease in motor activity in an open-field than Namptwt/wtand Namptflox/wtmice.And some Namptflox/floxmice showed spin behav⁃ior at 6-8 weeks after AAV injection.The dopa⁃minergic neurons in substantia nigra and ventral tegmental area are more susceptible to the knockout of nampt gene than the non-dopaminer⁃gic neurons.Transmission electron microscope examine showed degenerative changes of the myelin in striatum at 4 weeks after AAV injection for the Namptflox/floxmice.The orally supplementary of NAD precursor,nicotinamide ribose,improved the motor activity and decreased neuronal injury for the nampt gene knockout mice.CONCLU⁃SIONDecline of NAMPT and NAD in dopaminer⁃gic neurons is a risk for developing PD,and NAD precursors might be a new strategy for treatment of PD.

Key words:nicotinamide phosphoribosyltrans⁃ferase;nicotinamide adenine dinucleotide;myelin;dopaminergic neurons

T1-24

Conditioning knockout of nampt gene in mouse hippocampus neuron induces neuro⁃degeneration and cognitive deficiency

SHEN Chen,GAO Tong-yao,WANG Tong,CHEN Cong,LU Yun-bi,ZHANG Wei-ping

（Zhejiang University School of Medicine,Hangzhou 310043,China）

Abstract:OBJECTIVENicotinamide phos⁃phoribosyltransferase(NAMPT)is the key en⁃zyme in the NAD(nicotinamide adenine dinucleo⁃tide)salvaging synthesis pathway.The level of NAMPT and NAD decreases during ageing,which causes neurodegenerative diseases.How⁃ever,the study of neuron-and region-specific ge⁃netic disturbance of NAMPT on cognition impair⁃ment is still lacking.This study was to explore the consequences and mechanisms of hippo⁃campus CA1 neuron-specific knockout of NAMPT on mouse cognitive functions.METH⁃ODSFloxed three-month old Nampt(Namptflox/flox)mice were used,and injected rAAV-hSyn-Cre-APRE-pA into the hippocampus CA1 region to specifically knockout the nampt gene.The learn⁃ing and memory of mice was determined at one-month after the intracerebral injection of AAV.We used immunofluorescence and trans⁃mission electron microscopy to detect neuronal injury.Western blotting and ELISA were used to measure the change of protein level and small molecule level.RESULTSOne month after the knockout of nampt gene,the level of NAD signifi⁃cantly decreased in mouse hippocampus.The hippocampus dependent cognition of mice was also significantly decreased when compared to the wild type mice.However,the locomotor ac⁃tivity and anxiety behavior remained un⁃changed.Though there was no neuronal loss,and the neuronal cell bodies remained morpholog⁃ically intact.The level of Tau and MAP2 signifi⁃cantly decreased and degenerative change was found by using transmission electron micro⁃scope,which indicating the injury of both den⁃drites and axons.Meanwhile,the neuronal injury increased the neuroinflammation indicated by the activation of astrocyte and microglia.CON⁃CLUSIONThe decline of NAMPT and NAD causes neurodegeneration and cognition impair⁃ment of mice.

Key words:nicotinamide phosphoribosyltrans⁃ferase;nicotinamide adenine dinucleotide;cogni⁃tive functions

T1-25

鲜天麻对慢性束缚应激诱导小鼠学习记忆损伤的改善作用

黄 红，姜 宁，张亦文，姚彩虹，刘新民

（中国医学科学院北京协和医学院药用植物研究所，北京 100193）

摘要：目的现代科技发展使得人类生活模式、生存空间发生了重大转变，也对人类神经精神活动产生重大影响。这种内外源性应激会导致学习记忆的功能性损伤（亚健康状态），严重降低人们的生活质量。我国传统中药对于应激所致认知功能性损伤的亚健康状态的防护具有独特优势。鲜天麻（FG）是一种药食同源中药，现代药理学研究表明，它具有良好的改善学习记忆作用。本研究采用慢性束缚应激（CRS）诱导小鼠学习记忆障碍模型，探讨FG对CRS所致学习记忆障碍的改善作用。方法除对照组外，其余各组ICR雄鼠采用本实验室自制的束缚器连续造模35 d，每天10 h（22∶00～8∶00），FG汁（每天ig 0.5和1.0 g·kg-1），阳性药组给予多奈哌齐（Don，每天ig 1.6 mg·kg-1），给药3周。新物体识别实验（NORT）和物体位置识别实验（OLRT），分别用于评价小鼠的辨别能力和空间学习记忆能力，避暗实验评价小鼠的被动回避能力。实验数据采用SPSS软件21.0（ANOVA，±s）统计分析，P＜0.05认为差异具有统计学意义。结果CRS能显著性降低OLRT和NORT的辨别指数（P＜0.05），缩短避暗实验动物进入暗室的潜伏期（P＜0.05）。FG和Don治疗组可逆转上述变化（P＜0.05）。结论FG可改善CRS所致的学习记忆障碍。FG是可供长期服用的治疗应激相关记忆障碍的药食同源中药，本研究结果为研发天麻改善记忆减退类产品提供了依据。

关键词：天麻；学习记忆能力；应激

通讯作者：刘新民，E-mail：liuxinmin@hotmail.com

T1-26

左旋丁苯酞对肌萎缩性侧索硬化症细胞模型的保护作用和机制

兰嘉琦，彭雨晨，彭 英

（中国医学科学院药物研究所，北京 100050）

摘要：目的肌萎缩性侧索硬化（ALS）是一种复杂的多因素慢性神经元疾病，TAR DNA结合蛋白43（TDP-43）在细胞中形成的包涵体是其特征性病理标志，并与其另一个病理特征氧化应激密切相关。本研究探讨了神经保护剂左旋丁苯酞（L-NBP）对突变TDP-43所致的ALS相关损伤的可能保护作用和机制。方法建立了稳定转染TDP-43 M337V突变的NSC-34细胞模型以考察L-NBP对细胞损伤的改善作用，对氧化应激的影响，对线粒体的可能作用，对TDP-43 M337V聚集的改善作用，对DNA损伤的保护作用以及对相应蛋白的调控作用，并探讨其可能的作用机制。结果在转染TDP-43 M337V的小鼠运动神经元细胞模型上，L-NBP浓度依赖地提升细胞的存活率，并显著降低TDP-43 M337V造成的氧化应激，可能与其激活Nrf2/HO-1轴的作用有关。L-NBP对于TDP-43 M337V在胞浆内的聚集也有显著的改善作用，但对泛素-蛋白酶体通路和自噬-溶酶体通路均无明显影响。L-NBP显著升高稳转细胞的线粒体膜电位，并对线粒体片段化有一定改善作用，可能与其降低线粒体分裂蛋白FIS1的表达，并促进融合相关蛋白MFN2的表达有关。结论L-NBP可能通过激活Nrf2/HO-1抗氧化信号通路抑制细胞内氧化应激和相关损伤，并降低TDP-43 M337V在胞浆内的聚集，改善线粒体片段化，提高膜电位，进而发挥其抗ALS的作用，提示L-NBP可能成为治疗ALS的新策略。

关键词：左旋丁苯酞；氧化应激；线粒体膜电位

T1-27

N2L,a novel lipoic acid-niacin dimer,attenu⁃ates ferroptosis and decreases lipid peroxida⁃tion in HT22 cells

PENG Wei-jia1,ZHU Ze-yu1,YANG Yang1,HOU Jia-wei2,LU Jun-feng1,CHEN Chen1,LIU Fang3,PI Rong-biao2

(1.School of Pharmaceutical Sciences,2.School of Medicine,Sun Yat-sen University,Guangzhou 510006,China;3.School of Pharmaceutical Sci⁃ences,Guangzhou University of Chinese Medi⁃cine,Guangzhou 510006,China)

Abstract:OBJECTIVEN2L is a novel lipoic acid-niacin dimer regulating lipid metabolism with multifunction,including antioxidant effect.We investigated the protective effect of N2L and the underlying mechanisms under the ferroptosis inducer RAS-selective lethality 3(RSL3)treat⁃ment in HT22 cells.METHODSHT22 cells were pretreated with N2L and then were treated with RSL3 to establish a ferroptosis cell model.MTT assay was used to detect the cell survival rate.Free radical probe(dihydroethidium,DHE)and ferrous probe FerroOrange were used to detect the contents of free radicals and ferrous ions in cells.The ultrastructure of mitochondria of treat⁃ed cells was observed by transmission electron microscope.The expression of ferroptosis-relat⁃ed proteins acyl-CoA synthetase long-chain fami⁃ly member 4(ACSL4),glutathione peroxidase 4(GPX4), cyclooxygenase-2 (COX-2), ferritin Heavy Chain 1(FTH1),nuclear factor E2-related factor 2/heme oxygenase-1,and phosphoryla⁃tion levels of the c-Jun N-terminal kinase(JNK)/extracellular regulated protein kinases (ERK)pathway were detected by Western blotting.RE⁃SULTSRSL3 decreased the cell viability and induced excessive accumulation of(reactive oxy⁃gen species)ROS in HT22 cells.N2L pretreat⁃ment effectively protected HT22 cells against lipid peroxidation.What′s more,N2L recovered GPX4 protein expression and blocked the increase of COX-2 and ACSL4 expressions.Moreover,N2L also significantly prevented FTH1 from downregulation and maintained iron homeo⁃stasis.Finally,N2L pretreatment could decrease JNK/ERK activation induced by RSL3.CON⁃CLUSIONN2L is an excellent ferroptosis inhibi⁃tor,and its anti-ferroptosis mechanism may be related to the reduction of lipid peroxidation and the regulation of iron homeostasis.

Key words:N2L;ferroptosis;lipid peroxidation;c-Jun N-terminal kinase;extracellular regulated protein kinase

Corresponding author:PI Rong-biao,E-mail:pirb@mail.sysu.edu.cn

T1-28

大黄对溃疡性结肠炎治疗作用研究进展

郭子霞1，张丹参1，2，李 炜1

（1.河北北方学院药学系，河北省神经药理学重点实验室，河北 张家口 075000；2.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：溃疡性结肠炎（UC）是一种慢性非特异性肠道疾病。该病病程漫长，易反复发作，严重影响患者身心健康及生活品质。UC具体病因不明，与机体遗传、免疫反应、肠道菌群、环境、精神心理状态等多种因素有关。临床症状有腹痛腹泻、结直肠因溃疡出血出现黏液脓血便。此外患者血液高凝、血小板易聚集造成微循环障碍。中药大黄具有通经解瘀、清热解毒之功效。大黄的多种药理活性对应UC的多种临床症状及病理特点。大黄中鞣质具有收敛止泻作用，当小剂量使用时，鞣质的收敛作用盖过蒽醌类物质的泻下作用，可减轻患者腹泻症状。大黄全成分活血止血，临床研究已证明其可有效减轻上消化道出血症状，其中的没食子酸等成分可促进血小板在创伤部位聚集，促进凝血进而止血。大黄的活血止血功效可缓解UC患者肠道溃疡出血、血便症状。大黄黄酮类成分可通过升高血浆渗透压增加血容量来改善微循环、大黄酚-8-葡萄糖苷可抑制血小板聚集从而抑制血栓形成。此外，大黄蒽醌类物质如芦荟大黄素、大黄酚、大黄酸等可减少炎症因子的释放，缓解UC免疫功能异常引起的炎症反应。综上，大黄可多方面、多角度地对UC起到缓解甚至治疗的作用。

关键词：大黄；溃疡性结肠炎；治疗作用

基金项目：河北省高等学校科学技术研究项目（ZD2020117）

通讯作者：张丹参，E-mail：zhangds2011@126.com

T1-29

Effect of Huanglian Jiedu decoction on cogni⁃tive dysfunction of zebrafish with Alzheimer disease

HUANG Yao,LIU Kai-fei,GUAN Guo-kai,CHEN Yi-fei

(Guilin Medical University,Guilin 541199,China)

Abstract:OBJECTIVETo investigate the effects of Huanglian Jiedu decoction on cognitive dysfunction of zebrafish with Alzheimer disease.METHODS126 g of coptis chinensis,84 g of phellorescent phellorescent chinensis,84 g of scutellaria chinensis and 126 g of gardenia chinensis were immersed in 10,8 and 8 times of water for 30 min,and then extracted at 100℃for 2,1.5 and 1.5 h,respectively.The three extracts were coarse filtered and concentrated into 308 g·L-1and stored in refrigerator for later use.200 zebrafish were selected for behavioral train⁃ing in T-shaped tank for seven days.After that,the behavioral record analysis software Smart 3.0 was used to conduct behavioral analysis.Qualified zebrafish were selected as the blank group.Except for the blank group,zebrafish in the other 5 groups were exposed to AlCl3100 μg·L-1aquaculture water for modeling,and exposed for 6 d.The behavioral record analysis software Smart 3.0 was used to conduct behavioral analy⁃sis,and Select the successfully modeled zebraf⁃ish.After that,huperzine A(4 μg·L-1)and Huan⁃glian Jiedu decoction of low,medium and high doses(154,308 and 616 mg·L-1)were adminis⁃tered respectively,and exposed for six days.Then,the behavior analysis was conducted again through the behavioral record analysis soft⁃ware Smart 3.0 to select qualified zebrafish.After the training,the brain and gut of zebrafish in each group were collected,and the expression changes of N-cadherin,p-P38/p38MAPK and p-Tau in the brain were detected by Western blot⁃ting and qPCR to show the effect of Huanglian Jiedu decoction on cognitive dysfunction of Zebrafish with Alzheimer disease.RESULTSWestern blotting and qPCR results showed that the contents of N-cadherin increased(P＜0.05),and the contents of p-P38/p38MAPK and p-Tau decreased(P＜0.05)compared with the model group,indicating that Huanglian Jiadu decoction had an effect on the cognitive dysfunction of zebrafish with Alzheimer disease.CONCLU⁃SIONHuanglian Jiedu decoction can alleviate cognitive dysfunction of zebrafish model with Alzheimer disease to a certain extent.

Key words:Huanglian Jiedu decoction;Alzheimer disease;zebrafish;cognitive dysfunction

Foundation item:National Natural Science Foundation of china (8216140711);Natural Science Foundation of Guangxi Province(2017GXNSFAA198255);Natural Science Foun⁃dation of Guangxi Province(2018GXNSFBA13 8028);Open Project Program of Guangxi Key Laboratory of Brain and Cognitive Neuroscience,Guilin Medical University(GKLBCN-20180105-03);and 2019 College Student Innovation and Entre⁃preneurship Project Training Program(20191060 1038)

Corresponding author:CHEN Yi-fei,E-mail:chenyifei_pharm@glmc.edu.cn

T1-30

Cornel iridoid glycoside inhibits PP2Ac demethylation by regulationg PME-1

YANG Cui-cui,KUAI Xue-xian,LI Ya-li,ZHANG Li,LI Lin,ZHANG Lan

(Department of Pharmacology,Xuanwu Hospital of Capital Medical University,Key Laboratory for Neu⁃rodegenerative Diseases of Ministry of Education,Beijing 100053,China)

Abstract:OBJECTIVEPP2Ac demethyl⁃ation is regulated by LCMT(a specific leucine carboxyl methyltransferase catalyzing methyla⁃tion of PP2A)and PME(a specific methylester⁃ase catalyzing demethylation of PP2A.This study was to investigate the mechanism of Cor⁃nel iridoid glycoside(CIG)on PP2A catalytic sub⁃unit C(PP2Ac).METHODSRecombined lentivi⁃rus vector was used to deliver PME-1 genetic materials into N2a cells or transfected LCMT-1 siRNA into N2a cells to block the expression of LCMT-1.Twenty-four hours later,cells were rinsed twice with cold PBS(pH 7.4)and CIG at different concentrations(50,100 and 200 g·L-1,respectively)were added for 24 h.Western blotting was used to PP2Ac,demethylaion/methylation PP2Ac,LCMT-1 and PME-1.The ac⁃tivity of PP2A was detected by a biochemical as⁃say.RESULTS①Lentivirus transferred PME-1 was expressed at high level in the N2a cells after transduction.Correspondingly，the demethylation of PP2Ac was increasing and PP2A activity was decreasing after transduction.Treatment with CIG for 24 h reversed the increase of PME-1 and demethylation of PP2Ac without influencing LCMT-1 expression.PP2A activity was also sig⁃nificantly enhanced in CIG treatment group,compared with the cells after PME-1 transduc⁃tion.②LCMT-1 siRNA significantly decreased LCMT-1 expression.CIG did not affect LCMT-1expression.however,demethylation of PP2Ac is increased in siRNA-transfected cells and CIG could reversed the high demethylation of PP2Ac and PP2A activity.CONLUSIONCIG increases methylation of PP2A subunit C by inhibiting PME-1.

Key words:cornel iridoid glycoside;Alzheimer disease;PP2A catalytic subunit C demethylation

Corresponding author:Lin Li,E-mail:linli97@hotmail.com;Lan Zhang,E-mail:lanizhg@hotmail.com

T1-31

Novel compound FLZ alleviates rotenoneinduced Parkinson disease model by sup⁃pressing TLR4/MyD88/NF-κB pathway through microbiota-gut-brain axis

ZHAO Zhe,LI Fang-yuan,NING Jing-wen,BAO Xiu-qi,ZHANG Dan

(State Key Laboratory of Bioactive Substrate and Function of Natural Medicine,Institute of Materia Medica,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing100050,China)

Abstract:OBJECTIVESParkinson disease(PD)is the second most common neurodegener⁃ative disease,but none of the current treatments for PD could halt the progress of the disease due to the limited understanding of the pathogenesis.Increasing evidence proves that the close com⁃munication between the brain and the gastroin⁃testinal system is influenced by gut microbiota in PD pathogenesis,known as microbiota-gut-brain axis.However,the explicit mechanisms of micro⁃biota dysbiosis in PD development have not been well elucidated yet.FLZ,a novel squamosamide derivative,has been proved to be effective in many PD models and is undergoing the phase Ⅰ clinical trial to treat PD in China.The aims of our study are to assess the neuroprotective effects of FLZ treatment on PD and to further explore the underlying microbiota-related mechanisms of PD by using FLZ as a tool.METHODSChronic administration of rotenone(30 mg·kg-1per day)was utilized to induce a mouse model to mimic the pathological process of PD.Behavioral tests and gastrointestinal function tests were conduct⁃ed to evaluate the PD symptoms.Gut microbiota alterations were analyzed by 16s rRNA sequenc⁃ing.The intestinal permeability and blood-brain barrier structures were assessed by various methods.The pro-inflammatory cytokines and LPS levels in the colon,serum,and brain were detected by ELISA.Furthermore,the levels of in⁃flammation and TLR4/MyD88/NF-κB pathway in the substantia nigra(SN)and colon were mea⁃sured.RESULTSBehavioral tests and gastroin⁃testinal function tests found that rotenone-in⁃duced mice showed gastrointestinal dysfunctions(week 3)prior to the motor deficits(week 4).However,FLZ treatment significantly alleviated these PD symptoms.16S rRNA sequencing illus⁃trated that PD-related microbiota alterations in⁃duced by rotenone were reversed by FLZ treatment at various taxa levels.Especially,we identified an increased genus Akkermansia in the Rotenone group(P=0.0006),which could be reversed by FLZ administration(P=0.0070).By reducing microbiota dysbiosis,qPCR results showed that FLZ treatment suppressed intestinal inflammation of rotenone-challenged mice.After⁃wards,transmission electron microscopy(TEM),in vivo FITC permeability assay,bacterial translocation assay,and Western blotting togeth⁃er suggested that FLZ treatment attenuated the intestinal barrier destruction induced by rote⁃none.Subsequently, ELISA results showed that FLZ administration inhibited the leakage of pro-inflammatory cytokines(TNF-α,IL-1β,and IL-6)and LPS into the serum,suggesting the atten⁃uation of systemic inflammation.Then,several experiments including TEM analysis found that FLZ treatment restored blood-brain barrier struc⁃ture.Consequently,the immunofluorescence staining demonstrated that neuroinflammation(increased Iba-1+and GFAP+cells)and dopami⁃nergic neuronal death(reduced TH+cells)in the SN caused by rotenone were remarkably attenu⁃ated.Further mechanistic research proved that the anti-inflammatory effects of FLZ administra⁃tion were mediated through the TLR4/MyD88/NF-κB pathway both in the SN and colon.CONCLU⁃SIONFLZ treatment ameliorates microbiota dys⁃biosis to protect the PD model via inhibiting TLR4 pathway,which contributes to one of the underlying mechanisms beneath its neuroprotec⁃tive effects.Our research also supports the importance of microbiota-gut-brain axis in PD pathogenesis,suggesting its potential role as a novel therapeutic target for PD treatment.

Key words:FLZ;rotenone mouse model;micro⁃biota-gut-brain axis;akkermansia

Corresponding author:ZHAO Zhe,E-mail:zhaozhe@imm.ac.cn

T1-32

斯皮诺素通过减轻炎症改善阿尔茨海默病

杜会枝

（山西大学分子科学研究所，山西 太原 030006）

摘要：目的阿尔茨海默病（AD）是以记忆障碍和学习认知损伤为特征的慢性神经退行性疾病。目前为止，AD的确切发病机制并未完全阐明，主要有β-淀粉样蛋白（Aβ）沉积学说、氧化应激学说、神经炎症学说和细胞凋亡学说等。近年来大量研究发现，神经炎症在AD症的发生发展中起着重要作用，脑内慢性炎症反应是AD的病理特征之一，Aβ沉积激活神经胶质细胞释放促炎因子，如白细胞介素6（IL-6）、肿瘤坏死因子α（TNF-α）和一氧化氮（NO）等，进而损伤周围神经元，引起突触丢失和细胞凋亡，最终导致学习记忆功能受损。减轻炎症被认为是改善甚至是治疗AD症的重要途经之一。斯皮诺素（spinosin），属于黄酮类化合物，是酸枣仁（Ziziphi Spinosae Semen）中主要活性成分之一。本文先在化学角度研究了斯皮诺素对Aβ聚集的影响，又在小鼠海马脑片上探讨了其改善AD的机制。方法和结果 37℃孵育96 h产生的Aβ25-35聚集体与新鲜配制的Aβ25-35单体相比，粒径显著增大；斯皮诺素（终浓度为30 μmol·L-1）与Aβ单体一起孵育后，粒径明显降低，表明斯皮诺素缓解Aβ25-35的聚集。Aβ25-35（5 μmol·L-1）组炎症因子IL-1β和PGE-2含量显著升高；在脑片上预给药斯皮诺素0.5 h（终浓度为 30 μmol·L-1），该 2种炎症因子的含量降低至接近对照组。COX-2蛋白可以降解2-花生四烯酸甘油从而产生PGE-2。本研究检测了各组COX-2蛋白表达。结果显示，斯皮诺素抑制Aβ25-35引起的COX-2蛋白的过表达。随后研究了斯皮诺素对促凋亡蛋白Bax的作用。Aβ25-35处理海马脑片后，上调Bax蛋白表达，增加细胞凋亡；用斯皮诺素预保护后，Bax蛋白表达显著下降，抑制Aβ25-35引起的细胞凋亡。长时程增强（LTP）是反应学习记忆的主要指标之一。采用膜片钳技术，记录兴奋性突触后场电位，研究斯皮诺素对Aβ25-35引起的学习记忆障的改善作用。用斯皮诺素预保护后，LTP比例从Aβ25-35组的108%回升至与对照组的147%，对照组为142%。表明斯皮诺素对Aβ25-35诱导的学习记忆障碍有改善作用。结论斯皮诺素通过抑制COX-2蛋白过表达，减轻炎症，减少细胞凋亡，改善AD症状。

关键词：斯皮诺素；Aβ25-35；神经炎症；阿尔茨海默病

基金项目：山西省自然科学基金（201801D121290）；山西省回国留学人员科研项目（2017-021）；山西医科大学细胞生理学教育部重点实验室开放基金（KLMEC/SXMU-201911）

通讯作者：杜会枝，E-mail：duhuizhi@sxu.edu.cn

T1-33

Erzhi pills ameliorates cognitive dysfunction and alter proteomic profiles of hippocampus in ovariectomized Alzheimer disease model rats induced by D-galactose and Aβ1-40injection

XIE Yong-yan1,2＊,HOU Min3＊,ZHOU Mao-fu1,YAN Bo4,LIU Chao6,YAO Li-hua7,SUN Men-sheng1,HE Kun1, FANG Cong1, CHEN Yao-hui5,HUANG Li-ping1

(1.School of Pharmacy,2.College of traditional Chinese medicine,Jiangxi University of Traditional Chinese Medicine,Nanchang 330004,China;3.the Second People′s Hospital of Jingdezhen,Jingdezhen 333099,China;4.Shandong Qidu Pharmaceutical Co.,Ltd.,Zibo 255400,China;5.Jiangxi Provincial People′s Hospital Affiliated to Nanchang University,Nanchang 330006,China;6.China Pharmaceutical University,Nanjing 211198,China;7.School of Life Science,Jiangxi Science & Technology Normal University,Nanchang330096,China)

Abstract:OBJECTIVEErzhi pills is a clas⁃sic prescription of Chinese medicine originated from Fu Shou Jing Fang in Ming dynasty,with the effects of nourishing kidney-Yin and hemosta⁃sis,black hair,strengthening muscles and bones.As a classical prescription for nourishing kidney-Yin,Erzhi pills has been used to treat senile dementia in China for many years.Herein,our study aimed to investigate the protective effects of Erzhi pills in rat models of Alzheimer disease(AD)induced by ovariectomy as well as D-galactose and Aβ1-40injection and to explore its potential mechanism.METHODSThe model of AD rats was established by ovariectomy com⁃bined with D-galactose and Aβ1-40injection.Ovariectomized rats were randomly divided into four groups:model group,estradiol valerate(0.80 mg·kg-1)group,Erzhipills high(1.50 mg·kg-1)and low(0.75 mg·kg-1)doses group.In addition,rats of sham operation were selected as the sham operated group.Except for the sham oper⁃ated group,rats were injected intraperitoneally with D-galactose(100 mg·kg-1per day,for 49 d)on the 8thday,then they were given intracerebro⁃ventricular injection of A β1-40(10 μ g per rat,1 g·L-1)on the 36thday,while the corresponding drugs were given by gastrointestinal administra⁃tion on the 22ndday.In our study,Morris water maze test was used to evaluate the learning and memory abilities,while ELISA kit was used to an⁃alyze serum estrogen level.The morphology of hippocampal neuron cells was observed by HE staining,and Nissl staining was utilized to ob⁃serve the Nissl body in cytoplasm.Then,the ex⁃pression of ERβ positive cells and hippocampal Aβ1-40and p-Tau404proteins was determined by immunohistochemistry.In order to further explore the molecular mechanism of Erzhi pills preven⁃tion and treatment of AD,proteomics was used to find potential targets and related pathways,Western blotting was used to verify the expres⁃sion of candidate differential protein.According to the results of proteomics in our experiment,Western blotting was used to detect the protein expression of PI3K,Akt,Bcl-2,Bcl-xl,Bad,14-3-3 and GSK3β.RESULTSThe escape latency was significantly shortened,the number of crossing platform was increased,the neuron arrange⁃ment was more orderly and with less nuclear pycnosis in rats of Erzhi pills groups compared to the model group.In rats treated with Erzhi pills,the number of neurons,Nissl bodies,the estro⁃gen levels and ERβ positive cells were increased,while the number of Aβ1-40and p-Tau404positive cells was significantly decreased.Proteomics found that there were more than one hundred differentially expressed proteins of rats treated with Erzhi pills,which were involved in 48 signal⁃ing pathways.Among these proteins,five of them were involved in the PI3K/Akt signal path⁃way.As the down-stream protein of PI3K/Akt sig⁃naling pathway,the content of 14-3-3 protein was significantly increased.Western blotting analysis showed that the expression of p-GSK3β/GSK3β and Bad was decreased,while that of p-Akt/Akt,p-PI3K/PI3K,14-3-3,Bcl-xl and Bcl-2 was up-regulated in rats from the Erzhi pills groups compared with the model group.CON⁃CLUSIONErzhi pills can improve estrogen levels,alter proteomics expression of the hippocampus and activate PI3K/Akt pathway in AD rats,reduce Aβ aggregation,inhibit the hyperphos⁃phorylation of Tau protein,maintain the morphol⁃ogy of hippocampal neurons and decrease the apoptosis of hippocampal neurons,thereby improving the learning and memory abilities of ovariectomized AD rats induced by D-galactose and Aβ1-40injection.This study may provide an experimental basis for the clinical treatment of Erzhi pills.

Key words:Erzhi pills;Alzheimer disease;hip⁃pocampal neurons;proteomics;estrogen;aging;memory;PI3K/Akt signaling pathway

Corresponding author:HUANG Li-ping,E-mail:jxnchlp@163.com;LIU Bo,E-mail:bozhrliu@163.com

＊Co-first author.

T1-34

黄精多糖对阿尔茨海默病模型斑马鱼p38MAPK/N-cadherin的影响

陈毅飞1，刘凯菲1，吴世敏1，李讯怡1，黄 瑶1，黄典凤1，蒋才武2

（1.桂林医学院药学院，广西 桂林 541199；2.广西中医药大学药学院，广西 南宁 530200）

摘要：目的研究黄精多糖对阿尔茨海默病模型斑马鱼的影响。方法将斑马鱼置于T迷宫中训练7 d，将训练成功的40条斑马鱼分为4组：空白组、模型组、阳性组的和黄精多糖组。模型组、阳性组和黄精多糖组在100 μg·L-1的AlCl3中造模6 d；阳性组暴露在4 μg·L-1的石杉碱甲药液，黄精多糖组暴露在6 g·L-1黄精多糖药液，均给药6 d，模型组不予以给药，空白组不做任何处理，并用录像的方法对斑马鱼各阶段进行记录，对各组在EC区的时间进行分析。给药结束后取出脑组织，采用Western印迹法测定N-cadherin，P38和p-P38蛋白因子的表达。结果在行为学上，分析在EC区所待时间，空白组、阳性组和黄精多糖组分别与模型组相比均有统计学差异（P＜0.05）。在蛋白水平上，与模型组比较，阳性组和黄精多糖组P38和p-P38蛋白下调，而N-cadherin蛋白上调（P＜0.05）。结论黄精多糖通过上调N-cadherin蛋白水平，阻碍P38磷酸化，改善阿尔茨海默病模型斑马鱼的学习记忆能力。

关键词：阿尔茨海默病；斑马鱼；黄精多糖；N-cadherin；P38蛋白

基金项目：国家自然科学基金（8216140711）；广西壮族自治区自然科学基金（2017GXNSFAA198255），广西壮族自治区自然科学基金（2018GXNSFBA13 8028）；广西脑与认知神经科学重点实验室开放课题（GKLBCN-20180105-03）；2019大学生创新创业项目训练计划（201910601038）；第四期广西高等学校千名中青年骨干教师培育计划

通讯作者：蒋才武，E-mail：cwjiang@126.com

T1-35

Effect of rhynchophylline on p38MAPK /N-cadherin in zebrafish with Alzheimer disease

LIU Kai-fei,WU Shi-min,LI Xun-yi,HUANG Yao,GUAN Guo-kai,CHEN Xiao-long,CHEN Yi-fei

(Guilin Medical University,Guilin 541000,China)

Abstract:OBJECTIVETo investigate the effect of rhynchophylline on zebrafish with Alzheimer disease induced by AlCl3.METHODSZebrafish were trained in T maze for7 d.60 zebrafish were selected and divided into 6 groups:blank group,model group,positive group,low-dose group,medium-dose group and high-dose group,with 10 in each group.Feeding zebrafish adaptively for 2 d,starting training on the third day,and recording its movement track;Except the blank group,the other five groups were modeled and their motion trajectories were recorded.After modeling,the drug group was giv⁃en rhynchophylline,while the positive group was given huperzine A,and the movement track was recorded.The recorded movement track was analyzed by behavioral record analysis software smart 3.0.The expression levels of P38,p-P38 protein and N-cadherin protein in zebrafish brain tissue were detected by Western blotting.RESULTSThe data of time spent in the green short arm area of six groups of zebrafish were compared and analyzed:there was significant dif⁃ference between blank group and model group(P＜0.05),and there was significant difference between model group and positive group,low dose group,middle dose group and high dose group(P＜0.05).Comparative analysis of the time percentage of zebrafish in the green arm area of six groups:there was significant differ⁃ence between the blank group and the model group(P＜0.05),and there was significant differ⁃ence between the model group and the positive group,the rhynchophylline administration group,low dose group,middle dose group and high dose group(P＜0.05).The protein was detected by Western blotting.Compared with the model group,the expression level of N-cadherin protein was up-regulated(P＜0.05).Compared with the model group,the expression levels of p-P38 pro⁃tein in blank group,positive group and rhyncho⁃phylline administration group were down-regulat⁃ed(P＜0.05).CONCLUSIONUncaria rhyncho⁃phylline can down-regulate the ratio of p-P38/P38 and up-regulate N-cadherin,thus improving the learning and memory ability of zebrafish with Alzheimer disease.

Key words:rhynchophylline;Alzheimer disease;p38MAPK;N-cadherin

Foundation item:National Natural Science Foundation of China(8216140711);Natural Sci⁃ence Foundation of Guangxi Zhuang Autono⁃mous Region(2017GXNSFAA198255);Natural Science Foundation of Guangxi Zhuang Autono⁃mous Region(2018GXNSFBA138028);Guangxi Key Laboratory of Brain and Cognitive Neurosci⁃ence Open Project Funding (GKLBCN-20180105-03);2019 College Student Innovation and Entrepreneurship Project Training Plan(201910601038);and the Fourth Training Plan for Thousands of Young and Mid-aged Mainstay Teachers in Guangxi Colleges and Universities

Corresponding author:CHEN Yi-fei,E-mail:chenyifei_pharm@glmc.edu.cn

专题2：药物成瘾与精神疾病

T2-1

热休克蛋白70功能抑制剂亚甲蓝缓解吗啡依赖小鼠纳洛酮催促戒断症状

宫 琦1，魏守鹏1，李语玲2，梁 慧1，梁建辉1，3

（北京大学1.基础医学院药理学系，3.药学院分子与细胞药理学系，北京 100083；2.同济大学医学院附属上海市东方医院药学部，上海 200120）

摘要：目的探讨热休克蛋白70（Hsp70）抑制剂亚甲蓝对吗啡依赖小鼠纳洛酮催促戒断症状的药理作用。方法采用自发活动视频分析系统（Dig Behv-LG）测定亚甲蓝2.5～10 mg·kg-1对雄性昆明小鼠自主活动的影响。给雄性昆明小鼠连续2 d sc给予吗啡，第1天吗啡剂量为每次30 mg·kg-1，第2天吗啡剂量为每次60 mg·kg-1，每天2次，建立吗啡依赖小鼠模型。每次给予吗啡前5 min，ip给予不同剂量的亚甲蓝（0，2.5，5和10 mg·kg-1）进行干预试验。第3天，sc给予吗啡60 mg·kg-1后3 h，ip给予纳洛酮8 mg·kg-1，并观察和记录30 min内小鼠的跳跃次数以及注射纳洛酮前后30 min小鼠体重的变化。结果亚甲蓝在2.5～10 mg·kg-1剂量范围内，对小鼠自主活动无明显影响。吗啡依赖小鼠纳洛酮催促戒断试验显示，亚甲蓝可以剂量依赖性地抑制小鼠的催促戒断跳跃，并明显抑制纳洛酮催促戒断小鼠的体重丢失。结论作为Hsp70抑制剂，亚甲蓝对吗啡依赖小鼠纳洛酮催促戒断症状具有明确的抑制作用。

关键词：吗啡；纳洛酮；催促戒断；热休克蛋白70；亚甲蓝

基金项目：国家自然科学基金（81773705）

通讯作者：梁建辉，E-mail：liangjh@bjmu.edu.cn

T2-2

Effects of functional and transcriptional Hsp70 inhibitors on development and expres⁃sion of conditioned place preference induced by morphine in rats

GONG Qi1,WEI Shou-peng1,LI Yu-ling2,LIANG Hui1,LIANG Jian-hui1,3

(1.Department of Pharmacology,School of Basic Medical Science,3.Department of Molecular and Cellular Pharmacology,School of Pharmaceutical Sciences,Peking University,Beijing 100083,China;2.Department of Pharmacy,Shanghai East Hospi⁃tal,School of Medicine,Tongji University,Shanghai 200120,China)

Abstract:OBJECTIVETo investigate the effects of heat shock protein 70(Hsp70)in the development as well as expression of morphine induced conditioned place preference(CPP)in rats using Hsp70 inhibitors.METHODSThe unbi⁃ased procedures of CPP lasted for 7 d and included three phases:preconditioning(D1-D3),conditioning(D4-D6,6 sessions)and test(D7).Here,morphine 5 mg·kg-1injected in a subcuta⁃neous(sc)manner can induce significant place preference.Inhibitors of Hsp70 were injected into the right lateral ventricle during either the condi⁃tioning phase or the test phase separately.Fur⁃thermore,the expression of Hsp70 in certain areas of the mesocorticolimbic system was also studied following the intervention of N-formyl-3,4-methylenedioxybenzylidine-γ -butyrolactam(KNK437),a transcriptional inhibitor of Hsp70.RESULTSPifithrin-μ(PES),a selective functional inhibitor acting on the substrate binding domain(SBD)of Hsp70,dose-dependently suppressed both the acquisition and expression of morphineinduced CPP.Similar function was observed after the intracerebroventricular injection (icv) of KNK437.The other functional inhibitor methy⁃lene blue,targeting the nucleotide-binding area,showed a significant tendency of inhibitory phar⁃macological effect on the expressional and devel⁃opment phases of morphine-induced CPP.Following the interventions of KNK437,we found that the level of Hsp70 was significantly decreased in the NAcs both in the acquisition and expres⁃sion of morphine induced CPP.CONCLUSIONHsp70 in NAcs plays a critical role in mediating the psychological dependence induced by morphine.

Key words:morphine;reward;conditioned place preference;heat shock protein 70;methylene blue

Foundation item:National Natural Science Foundation of China(81773705)

Corresponding author:LIANG Jian-hui,E-mail:liangjh@bjmu.edu.cn

T2-3

Modeling and simulation for individualized therapy of amisulpride in Chinese patients with schizophrenia:focus on inter interindi⁃vidual variability,therapeutic reference range and laboratory alert level

HUANG Shan-qing＊,LI Lu＊,WANG Zhan-zhang,XIAO Tao,LI Xiao-lin,LIU Shu-jing,ZHANG Ming,LU Hao-yang,WEN Yu-guan,SHANG De-wei

(The Affiliated Brain Hospital of Guangzhou Medical University,Guangzhou 510370,China)

Abstract:OBJECTIVETo explain the high inter-individual variability and the frequency of exceeding the therapeutic reference range and the laboratory alert level of amisulpride,a popula⁃tion pharmacokinetic model in Chinese patients with schizophrenia was built based on therapeu⁃tic drug monitoring data to guide individualized therapy.METHODSPlasma concentration data(330 measurements from 121 patients)were ana⁃lyzed using a nonlinear mixed-effects model⁃ing approach with first-order conditional estima⁃tion with interaction(FOCE I).The concentra⁃tions of amisulpride were detected by HPLC-MS/MS.Age,weight,sex,combination medication history and renal function status were evaluated as main covariates.The model was internally val⁃idated using goodness-of-fit,bootstrap and nor⁃malized prediction distribution error.Recom⁃mended dosage regimens for patients with key covariates were estimated on the basis of Monte Carlo simulations and the established model.RESULTSA one-compartment model with firstorder absorption and elimination was found to adequately characterize amisulpride concentra⁃tion in Chinese patients with schizophrenia.The population estimates of the apparent volume of distribution(V/F)and apparent clearance(CL/F)were 12.7 L and 1.12 L·h-1,respectively.Age sig⁃nificantly affected the clearance of amisulpride and the final model was as follow:CL/F=1.04×(AGE/32)-0.624(L·h-1).To avoid exceeding the lab⁃oratory alert level(640 μg·L-1),the model-based simulation results showed that the recommended dose of amisulpride was no more than 600 mg per day for patients aged 60 years,800 mg per day for those aged 40 years and 1200 mg per day for those aged 20 years,respectively.CON⁃CLUSIONDosage optimization of amisulpride can be carried out according to age to reduce the risk of adverse reactions.The model can be used as a suitable tool for designing individual⁃ized therapy for Chinese patients with schizo⁃phrenia.

Key words:amisulpride;population pharmacoki⁃netics;therapeutic drug monitoring;modeling and simulation;individualized therapy

＊Co-first author.

T2-4

盐酸羟哌吡酮（YL-0919）的快速抗抑郁作用及其电生理机制

尹勇玉1，张黎明1，李云峰1，2

（军事科学院军事医学研究院1.毒物药物研究所，2.军事认知与脑科学研究所，北京 100850）

摘要：目的探究盐酸羟哌吡酮（YL-0919）快速抗抑郁的行为学效应及其电生理机制。方法采用慢性应激模型、开窗测验、强迫游泳测验、行为类别分析等评价YL-0919的抗抑郁活性，采用在体多通道电生理记录技术、在体诱导神经场电位技术探究YL-0919快速抗抑郁作用的电生理机制。结果①连续8周的慢性应激可使食蟹猴表现出显著的抑郁样行为，包括蜷缩行为、自我挠体行为等显著增加，环境探索、自主运动等显著下降；开窗潜伏期显著增加，开窗时间显著下降；慢性给予YL-0919（1.2 mg·kg-1，ig）9 d即可显著逆转由慢性应激所导致的系列抑郁样行为，具体表现为显著降低蜷缩行为、自我挠体行为等消极行为，显著增加环境探索、自主运动等积极行为，显著增加食蟹猴在开窗测验上的探索兴趣；氟西汀（FLX）却需要慢性ig给予17 d才会表现出抗抑郁行为学效应，提示YL-0919在该模型上具有较快速的抗抑郁活性。②在体多通道电生理研究表明单次ip给予氯胺酮（Ket，10 mg·kg-1）在给药1.5和24 h后均可显著减少大鼠在强迫游泳测验上的不动时间，表现出显著的抗抑郁行为学效应，同时可以显著增加锥体神经元的放电活动并显著抑制中间神经元的放电活动，对mPFC中的E∶I平衡产生快速调节作用；而在相同实验条件下，FLX却无上述作用。进一步研究发现慢性ig给予FLX（10 mg·kg-1）21 d，末次给药24 h后表现出显著的抗抑郁行为学效应，同时对mPFC中E∶I平衡产生显著性的调节作用。③ YL-0919（2.5 mg·kg-1，ip）可显著增加mPFC中锥体神经元的放电活动并抑制中间神经元的放电活动，可对mPFC中E∶I平衡产生显著性的调节作用，而5-HT1A受体拮抗剂WAY-100635（0.3 mg·kg-1，ip）可显著阻断YL-0919 对mPFC中E∶I功能性平衡的影响，而5-HT6受体拮抗剂SB271046（10 mg·kg-1，ip）不能阻断 YL-0919对mPFC中E∶I平衡的影响，提示5-HT1A受体在YL-0919对mPFC中E∶I平衡的影响中发挥着重要的调节作用。④在体诱导神经场电位记录结果表明，ig给予YL-0919（2.5 mg·kg-1）7 d即可显著增加大鼠海马齿状回群峰电位幅值，WAY-100635（0.3 mg·kg-1，ip）可显著阻断YL-0919对海马神经可塑性的影响，SB271046（10 mg·kg-1，ip）不能阻断YL-0919对海马神经可塑性的影响，提示5-HT1A受体在YL-0919对海马神经可塑性的影响中发挥着重要的调节作用。结论YL-0919在食蟹猴慢性应激模型上具有快速的抗抑郁行为学效应；mPFC中E∶I功能性平衡可能是抗抑郁药起效的重要限速环节，YL-0919通过5-HT1A受体快速调节mPFC中E∶I平衡并增强海马神经可塑性，这可能是其抗抑郁效应快速起效的关键机制之一。

关键词：YL-0919；抑郁症；E∶I平衡；慢性不可预知应激；快速起效

通讯作者：张黎明，E-mail：zhanglm0308@163.com；李云峰，E-mail：lyf619@aliyun.com

T2-5

基于胶质细胞18 ku转位蛋白的快速起效抗抑郁新药研究

张黎明1，姚俊琪1，姚如梦1，刘 畅1，袁 瑾1，李云峰1，2

（军事科学院军事医学研究院1.毒物药物研究所，2.军事认知与脑科学研究所，北京 100850）

摘要：目的脑内18 ku转位蛋白（TSPO）主要位于胶质细胞上，是介导神经类固醇（如别孕烯醇酮）合成的关键蛋白。本研究拟在动物行为、细胞和分子水平探讨TSPO在精神-认知障碍疾病治疗中的靶标价值，并研究自主研发的TSPO激活剂YL-IPA08的抗精神-认知障碍的药理学作用及其机制。方法①采用TSPO敲除小鼠进行YL-IPA08给药后的强迫游泳实验、悬尾实验和高架十字迷宫实验检测抗抑郁和抗焦虑行为效应。②采用PCPA和5，7-DHT等药理学技术干扰大脑5-HT系统后，探索5-HT系统在YL-IPA08抗抑郁和抗焦虑中的作用。③采用大鼠慢性不可预知应激模型、小鼠慢性束缚应激模型和小鼠皮质酮饮水模型，以糖水偏好实验检测糖水偏嗜度这一核心指标评价YLIPA08抗抑郁起效时程。采用Western印迹以及高尔基染色进行相应的神经生物学机制研究。④用大鼠在体多通道电生理记录研究YL-IPA08对内侧前额皮层（mPFC）部位谷氨酸能锥体神经元和γ-氨基丁酸（GABA）能中间神经元放电频率的影响。结果①TSPO敲除可取消YL-IPA08的抗抑郁、抗焦虑效应，进一步确证了YL-IPA08的靶标特异性。②5-HT系统被干扰之后可以阻断YL-IPA08的行为学效应，进一步微透析实验研究发现，YL-IPA08可以快速增加大鼠海马细胞间隙5-HT水平，而TSPO拮抗剂PK11195可以完全阻断YL-IPA08诱导的海马5-HT升高，上述结果提示5-HT系统的完整性可能参与了YL-IPA08的抗抑郁、抗焦虑效应。③YL-IPA08给药1周可显著发挥抗抑郁效应，而氟西汀则需3周，提示YL-IPA08具有快速起效的优势。YL-IPA08的快速抗抑郁效应可能与快速增加mPFC神经元树突复杂性有关。④YL-IPA08单次给药使大鼠mPFC中谷氨酸能锥体神经元放电频率增加，GABA能中间神经元放电频率降低，提示YL-IPA08抗抑郁抗焦虑效应可能与兴奋锥体神经元，抑制中间神经元有关，即YL-IPA08可快速调节mPFC的E∶I功能性再平衡，这可能是其快速起效的重要启动机制。结论①TSPO选择性配体YL-IPA08在多种抑郁动物模型上具有快速起效的抗抑郁作用（1周）；② YL-IPA08快速调节mPFC中谷氨酸锥体神经元和GABA中间神经元的E∶I功能性再平衡，进而快速增强锥体神经元树突复杂性，这可能是其快速抗抑郁的重要机制。本研究为前期课题组提出的“单胺（5-HT）-谷氨酸/GABA长神经环路”候选假说提供了有力证据，为快速起效抗抑郁新药的发现及靶标机制研究提供了有益的借鉴。

关键词：18 ku转位蛋白；抑郁症；焦虑症；神经可塑性；YL-IPA08

通讯作者：李云峰，E-mail：lyf619@aliyun.com

T2-6

大麻二酚通过Sigma1R/Akt/GSK3 β/CREB信号通路减轻甲基苯丙胺诱导的大鼠复吸和PC12细胞神经毒性

刘 柳1，李 娟2，王 婵1，许 悦1，杨根梦1，林纾丞1，张树威1，澹 忆1，张慧洁1，王浩伟1，曾晓锋1，刘建兴1

（昆明医科大学1.法医学院，2.基础医学院，云南 昆明 650500）

摘要：目的甲基苯丙胺（METH），俗称“冰毒”，是一种精神依赖性极强的新型合成毒品，在全球范围内被广泛滥用。防治METH滥用主要通过减少吸食者对METH渴求和防止METH复吸发生2条途径。大麻二酚（CBD）因其在治疗药物依赖和防止药物复吸方面具有潜在药理学价值而受到广泛关注。在我们的前期研究中，通过建立METH诱导的大鼠条件位置偏爱（CPP）模型来研究CBD的药理学作用。方法和结果 研究证实，ip给予CBD 20，40和80 mg·kg-1可显著减少METH（2 mg·kg-1）诱导的大鼠CPP，且该作用可能涉及CBD对Sigma1R/Akt/GSK3β/CREB信号通路的调节。在此基础上，本研究继续采用METH诱导的大鼠CPP模型来探讨CBD在METH戒断和复吸中的作用。研究发现，对大鼠ip给予METH 2 mg·kg-1可以诱导CPP产生，待大鼠经历5 d的METH戒断后，再次注射METH 0.5 mg·kg-1以恢复METH诱导的CPP；而在戒断期内连续5 d注射CBD 20，40和80 mg·kg-1不仅能促进大鼠CPP灭绝，还能有效防止METH诱导的大鼠CPP恢复，并且Sigma1R和Akt/GSK3β/CREB信号通路的表达水平变化参与该过程。为了进一步验证上述调控机制，本研究建立了体外PC12细胞模型，在METH暴露前，分别采用CBD、Sigma1R拮抗剂BD1047、Sigma1R基因敲低的慢病毒预处理细胞，随后使用Western印迹法检测Sigma1R，Akt，GSK3β，CREB等相关蛋白的表达。结果表明，CBD可减轻METH诱导的PC12细胞神经毒性并且通过拮抗Sigma1R的功能或表达抑制METH对Akt/GSK3β/CREB信号通路的激活作用。结论CBD可能是一种防止METH复吸和神经毒性的药物，而Sigma1R可能是CBD药理学作用的重要靶点，但其具体的干预作用和机制还需要更多的研究证明。

关键词：大麻二酚；甲基苯丙胺；复吸；神经毒性；Sigma1R

基金项目：国家自然科学基金地区基金项目（8196070173）；云南省科技厅-昆明医科大学联合专项（202001AY070001015）

通讯作者：曾晚锋，E-mail：zxf2004033@163.com；刘建兴，E-mail：444319658@qq.com

T2-7

甲基苯丙胺和HIV-Tat蛋白通过Nrf2/NQ01/HO-1通路协同诱导小胶质细胞氧化应激和自噬

杨根梦，沈宝玉，王 婵，许 悦，林纾丞，洪仕君，李利华，曾晓锋

（昆明医科大学法医学院，云南 昆明 650500）

摘要：目的探讨Nrf2通路在甲基苯丙胺（METH）和HIV-Tat蛋白协同诱导小胶质细胞自噬中的作用。方法以滥用METH和滥用METH同时感染HIV死亡者，C57BL/6J小鼠，BV2细胞和原代小胶质细胞为研究对象。分别通过激活Nrf2和敲除或沉默Nrf2后，观察人体脑组织，小鼠纹状体组织和小胶质细胞内LC3-Ⅱ，Beclin1，ATG5，ATG7，T-Nrf2，N-Nrf2，HO-1和NQ01的变化和小胶质细胞内氧化应激水平。结果人体脑组织：METH组和METH+AIDS组纹状体内小胶质细胞被激活，同时Nrf2和LC3-Ⅱ表达水平明显升高，且METH+AIDS组表达水平比METH组高。体内实验：METH和HIV-Tat蛋白协同激活纹状体内小胶质细胞并诱导自噬水平明显升高。激活Nrf2后，两者协同诱导自噬水平降低。敲除Nrf2后，两者协同诱导自噬水平升高。与WT小鼠相比，Nrf2-KO后，自噬表达水平更高。体外实验：METH和HIV-Tat蛋白协同诱导小胶质细胞自噬和氧化应激水平明显升高，且具有METH和HIV-Tat蛋白浓度依赖性。激活Nrf2后，两者协同诱导自噬和氧化应激水平降低。沉默Nrf2后，两者协同诱导自噬和氧化应激水平明显升高。结论滥用METH同时感染HIV对激活小胶质细胞和Nrf2以及诱导自噬具有协同作用。METH和HIVTat蛋白可协同诱导小胶质细胞自噬和氧化应激，Nrf2通路可通过调节氧化应激来调控两者协同诱导的自噬作用。该研究可为毒品滥用的HIV感染者寻找有效的药物干预靶点提供理论依据和新思路。

关键词：甲基苯丙胺；氧化应激；自噬

通讯作者：曾晓锋，E-mail：zxf2004033@163.com

T2-8

Can Na+/K+ATPase be a novel target to treat anxiety?A hint from its regulatory effect on neuroinflammation

HUANG Song-qiang1, ZHU Meng-yuan2,LI Shan-shan1,NIE Xiao-wei3,BIAN Jin-song1,2

[1.Department of Pharmacology,School of Medi⁃cine,Southern University of Science and Technology,Shenzhen 518055,China;2.Department of Phar⁃macology,Yong Loo Lin School of Medicine,National University of Singapore,117600,Singapore;3.Shenzhen Institute of Respiratory Diseases,the First Affiliated Hospital(Shenzhen People′s Hospi⁃tal),Southern University of Science and Technolo⁃gy,Shenzhen 518020,China]

Abstract:OBJECTIVENa+/K+-ATPase(NKA)is large membrane protein expressed uni⁃versally which is indispensable for the mainte⁃nance of ionic gradient as well as neuronal excit⁃ability.The role of NKA in inflammatory regula⁃tion is still unclear.Inflammatory responses are initiated upon the activation of inflammasomes.In order to investigate the crosslink between NKA and inflammasome,NKAα1 knockout(KO)N2a cells were generated using CRISPR/Cas9 system.METHODS AND RESULTSqPCR results showed that NLRP1 and NLRP3 were upregulated in response to NKA α 1 loss while both NLRC4 and AIM2 remained unaffected.Meanwhile,consistent with the change in NLRP1 and NLRP3,both the mRNA level of ASC and IL-1β were significantly increased in NKAα1 KO cells.These data indicated that NKAα1 interfer⁃ence might influence the level of NLRP1 and NLRP3 inflammasomes in neuronal cells.Further evidence indicating the potential link between NKA and inflammasome pathway were provided using cytokine array assay where all the differen⁃tiated protein detected were closely linked to NLRP1 and NLRP3.To confirm this effect,we also observed the transcriptional levels of inflam⁃masome proteins in the brain cortex from both NKAα1+/+and NKAα1+/-mice.In line with the observation gained in NKAα1 KO cells,the mRNA level of NLRP1,NLRP3 and IL-1β were significantly upregulated in NKAα1+/-mice brain.Interestingly,in the primary cultured astrocytes,treatment with LPS/ATP significantly reduced the mRNA and protein levels of NKAα1 expression.These data imply that a negative regulation loop between NKAα1 and inflammation may exist in the central nervous system.Since neuroinflam⁃matory mechanism is currently considered the most potential of interventions to target anxiety,we therefore perform behavioural experiments to investigate the role of NKAα1 in anxiety.Chronic restraint stress (CRS)for10 d significantly reduced the time and frequency of entering the open arm and prolonged the retention time in the closed arm in the elevated plus-maze test.In the open field test,CRS also reduced both duration and frequency of entering into the central region.Although NKA α 1 loss itself did not alter the behaviour performance in the normal condition,it exacerbated CRS-induced above behaviour abnormalities.CONCLUSIONNKAα1 is regulat⁃ed upon inflammatory challenger and may be a novel target to treat anxiety.

Key words:Na+/K+ATPase;anxiety;inflamma⁃some;neuroinflammation

Corresponding author:BIAN Jin-song,E-mail:bianjs@sustech.edu.cn

T2-9

尼古丁对大脑奖赏系统的调控作用及脑源性神经营养因子相关机制

常 芮，黄泽怡，陈运凡，谭日东，廖海琳，谭思杰

（南华大学衡阳医学院，湖南 衡阳 421001）

摘要：目的烟草依赖是严重的公共卫生问题，尼古丁是烟草中的主要精神活性物质。本研究运用颅内自我刺激技术（ICSS）和条件性位置偏爱（CPP）模型，研究尼古丁对大脑奖赏系统功能的影响，并进一步探讨脑源性神经营养因子（BDNF）相关信号通路在尼古丁成瘾中的作用。方法通过立体定位手术在大鼠腹侧被盖区-伏隔核通路上方植入刺激微电极，sc给予不同剂量的尼古丁，检测ICSS的电流阈值以衡量大脑奖赏系统的功能。结果尼古丁≥0.1 mg·kg-1能显著降低ICSS的阈值；此外，与雄性大鼠相比，雌性大鼠的阈值下降更显著，表明尼古丁的奖赏效应存在性别差异。在尼古丁戒断时，ICSS电流阈值显著升高，表明尼古丁戒断能降低大脑奖赏系统的功能。在CPP实验中发现，尼古丁0.5 mg·kg-1组小鼠的CPP得分显著高于生理盐水组，表明尼古丁0.5 mg·kg-1能诱导小鼠形成CPP，而BDNF高亲和力受体TrkB的阻断剂可以降低小鼠CPP得分。单次尼古丁0.5 mg·kg-1注射对小鼠伏隔核和前额叶皮质BDNF表达无影响，但可显著降低海马BDNF的表达；连续7 d给小鼠注射尼古丁0.5 mg·kg-1，停药2 h后，海马BDNFTrkB信号通路表达显著上调，下游AKT表达显著升高，且Hip CA3和DG内GluA1蛋白表达增加。结论急性尼古丁能激活大脑奖赏系统，而尼古丁戒断时大脑奖赏系统功能下调；尼古丁对奖赏系统的调控作用可能与BDNF-TrkB信号通路有关。

关键词：尼古丁；脑源性神经营养因子；奖赏系统

T2-10

Effect of early psychedelic use on social func⁃tion in adulthood

YU Zhi-peng,SHEN Hao-wei

(Department of Pharmacology,Medical School of Ningbo University,Ningbo 315211,China)

Abstract:Psychedelic is one of psychoac⁃tive drug substances which leads to abnormali⁃ties in thinking,mood and sensory perception via the subtype of 5-HT receptor(5-HT2A receptor).Psychedelic is known to cause schizophrenialike symptoms or worsen schizophrenia.Mean⁃while it causes abnormalities in neural synchroni⁃zation and oscillations.In recent years,with the prevalence of various new psychedelic drugs and the rising population of young addicts,the clinical effects of psychedelic have been highly concerned.Adolescence is an important period for the formation of neuroplasticity,and it is an important stage in the functional development and maturation of the central nervous system.However,it is currently unclear about the endur⁃ing effects of juvenile exposure to psychedelic on social behavior,and neural network function.OBJECTIVEThe chronic exposure model of psy⁃chedelic(2CC-NBOMe,2CC)in rats was used to reveal the enduring effects on social behavior and neural network function in adulthood,which is intended to provide important reference infor⁃mation about the potential enduring effects and clinical applicability of psychedelic.METHODSRepeated exposure of normal saline in adoles⁃cence group(0.1 mL·kg-1,for 14 d),repeated exposure of 2CC group(0.1 mg·kg-1,for 14 d).The duration of adolescent administration was 35-48 d after birth.Adulthood means 9 weeks after birth.Behavioral and somatic neuroelectro⁃physiological experiments were performed 9-12 weeks after birth in the adolescent drug exposure group.RESULTSPsychedelic-exposed animals during adolescence showed a significant decline in social rank and an increase in social willingness.Coherence of theta waves(6-10 Hz)from the hippocampal CA1 to the orbital frontal cortex(Orb)was significantly reduced in the psy⁃chedelic-exposed animals compared to the con⁃trol animals.During the social task,the coher⁃ence of theta waves between CA1 and Orb was significantly reduced in the control animals,but not in the psychedelic-exposed animals.CON⁃CLUSIONThe altered social function caused by chronic psychedelic exposure during adoles⁃cence is associated with abnormal CA1-Orb theta wave coherence.

Key words：psychedelic;orbital frontal cortex;social rank;social willingness

Corresponding author:SHEN Hao-wei,E-mail:shenhaowei@nbu.edu.cn

T2-11

Parvalbumin interneurons in anterior cingu⁃late cortex regulate cognitive behaviors in methylazoxymethanol acetate model of schizophrenia

ZHANG Xiao-qin1,2, XU Le1, YU Zhi-peng1,SHEN Hao-wei1,2

(1.Department of Pharmacology,School of Medicine,Ningbo University,Ningbo 315211,China;2.Key Laboratory of Addiction Research of Zhejiang Province,Ningbo 315010,China)

Abstract:OBJECTIVECognitive dysfunc⁃tion is a core disturbance of schizophrenia,appear to emerge from impaired neural activity.The anterior cingulate cortex(ACC)is an integra⁃tion hub for higher-order thalamic inputs impor⁃tant for complex cognitive tasks such as learning and memory processes,attention and social interaction.Parvalbumin(PV)interneurons could filter information at pyramidal neurons of ACC,and the abnormal PV interneurons have been observed in both humans and animal models of schizophrenia.However,the mechanisms of PV interneurons in ACC regulating cognition in schizophrenia is poorly understood.METHODSThe pregnant mice were injected with methyl⁃azoxymethanol acetate(MAM)on gestational day(GD)16 for the neurodevelopmental MAM model of schizophrenia in our study.We investi⁃gated the cognitive behaviors by a serious of tests such as pre-pulse inhibition,Y maze,novel object and novel location recognition and the intrinsic excitability of PV interneurons and inhibi⁃tory synaptic transmission onto pyramidal cells localized in layer 5 of ACC by whole-cell record⁃ings.Further,the PV interneurons were regulat⁃ed by designer receptor exclusively activated by a designer drug(DREADD)system and the D-serine,a co-agonist of N-methyl-D-aspartate(NMDA)receptors.RESULTS①MAM mice showed the cognitive deficits and hypo-excitability of PV interneurons in ACC.②Restoration of PV interneuron activity in ACC improved cognitive function in MAM mice.③ Inhibition of PV interneu⁃ron activity in ACC was sufficient to cause cogni⁃tive dysfunction in control mice.④ NMDA recep⁃tors of PV interneurons in ACC were impaired in MAM mice.⑤ Deficits of NMDA receptor sig⁃naling specifically in PV interneurons and of cog⁃nitive behaviors in MAM mice were rescued by D-serine.CONCLUSIONPV interneurons in ACC are closely related to cognitive function in the MAM model of schizophrenia and D-serine maybe a potential therapy for schizophrenia.

Key words:methylazoxymethanol acetate;cognitive function;parvalbumin;interneurons;D-serine

Corresponding author:ZHANG Xiao-qin,E-mail:zhangxiaoqin1@nbu.edu.cn

T2-12

Regulation by phosphodiesterase of anxietyand depressive-like behavior induced by alcohol withdrawal in mice

LIU Xiao-qian1＊,WANG Dong1＊,ZHENG Wen-qing1,ZHANG Fang-fang1,ZHANG Han-ting1,2

(1.Institute of Pharmacology,Shandong First Medi⁃cal University & Shandong Academy of Sciences,Tai’an 271016,China;2.Department of Pharma⁃cology,Qingdao University School of Pharmacy,Qingdao 266073,China)

Abstract:OBJECTIVEAlcohol dependence is not only the most common public health prob⁃lem worldwide,but also the cause of many alco⁃hol-related diseases,such as anxiety and depres⁃sion.However,the mechanism of these comor⁃bidities is not clear.Recently,the role of phos⁃phodiesterases(PDEs)in anxiety-and depres⁃sion-like behavior induced by alcohol withdrawal has received increasing attentions.PDEs are a superfamily of enzymes that catalyze the hydroly⁃sis of intracellular cyclic AMP(cAMP)and(or)cyclic GMP (cGMP).There are 11 families(PDE1-PDE11),each of which has 1 to 4 sub⁃types encoded by different genes.In the present study,we identified the key PDEs involved in anxiety-and depression-like behavior induced by alcohol withdrawal.METHODSAfter withdrawal from short-term or chronic intermittent ethanol vapor exposure(STEVE and CIEVE,respectively),adult male C57BL/6J(B6)mice were tested for anxiety-and depression-like behavior using the sucrose preference,open field,elevated plusmaze,tail suspension,forced swimming,hole board and light-dark tests.In addition,the expression of PDE subtypes(PDE2,PDE4,and PDE7)in the hippocampus,amygdala,and stria⁃tum,which are considered as anxiety-and(or)depression-related brain regions,was detected by Western blotting.RESULTS①Differences in anxiety-and depression-like behaviors between STEVE and CIEVE mice.Compared to STEVE mice,CIEVE mice showed greater anxiety-and depression-like behavior in the tests described above.②The expression of PDE2,PDE4 and PDE7 in the anxiety-and depression-related brain regions of CIEVE mice.Compared to the STEVE mice,CIEVE mice showed significant higher expression of PDE4A and PDE7A in the hippo⁃campus(P＜0.05)and PDE4D and PDE7A in the amygdala(P＜0.05).However,the expres⁃sion of PDE2A,PDE4B,PDE4D and PDE7B in the hippocampus,PDE2A,PDE4A,PDE4B and PDE7B in the amygdala,PDE2A,PDE4A,PDE4B,PDE4D,PDE7A and PDE7B in the striatum was not significantly changed.CONCLUSIONThe model of CIEVE showed significant anxiety-and depression-like behavior, with concurrent increases in the expression of PDE4A and PDE7A in the hippocampus and PDE4D and PDE7A in the amygdala.These results suggest that PDE4A,PDE4D,and DE7A may play an important role in the regulation of anxiety-and depression-like behavior induced by alcohol withdrawal.

Key words:phosphodiesterases;alcohol with⁃drawal;behavior

Foundation item:National Natural Science Foundation of China(81773717);National Natural Science Foundation of China(81801510);and Academic Promotion Program of Shandong First Medical University(2019QL011)

Corresponding author:ZHANG Fang-fang,E-mail:15650512848@163.com; ZHANG Han-ting,E-mail:htzhang@qdu.edu.cn

＊Co-first author.

T2-13

Comparison of phosphodiesterase 7 expres⁃sion in brain striatum of C57BL/6J and DBA/2J mice and association with ethanol drinking behavior

ZHANG Fang-fang1,DU Xian1,ZHOU Yan-meng1,ZHENG Wen-qing1,LIU Xiao-qian1,ZHANG Han-ting1,2

(1.Institute of Pharmacology,Shandong First Medi⁃cal University & Shandong Academy of Sciences,Tai’an 271016,China;2.Department of Pharma⁃cology,Qingdao University School of Pharmacy,Qingdao 266073,China)

Abstract:OBJECTIVECyclic adenosine monophosphate(cAMP)-protein kinase A(PKA)signaling has been shown to regulate alcohol consumption.The phosphodiesterase 7(PDE7)enzyme is one of the PDE families responsible for controlling intracellular levels of cAMP.However,the role of PDE7 in alcohol consump⁃tion remains unknown.C57BL/6J(B6)mice innately consume larger amounts of alcohol while DBA/2J(DBA)mice do the opposite,ie,they drink little alcohol.In the present study,we evaluated whether PDE7 plays a role in regulat⁃ing alcohol intake using adult B6 and DBA mice.METHODSAdult male B6 and DBA mice were tested for ethanol(7% and 10%,V/V)intake and preference using the two-bottle choice task.In addition,a separate set of B6 and DBA mice was examined for PDE7 expression in the striatum,a brain region critical for ethanol drinking,using Western blotting.Further,PDE7 subtype expres⁃sion in the striatum of B6 mice in response to ethanol drinking was examined.Finally,the effect of the PDE7 inhibitor BRL-50481 on etha⁃nol consumption was examined in B6 mice.RESULTS①Comparison of ethanol drinking behavior between B6 and DBA mice.Compared to DBA mice,B6 mice had significantly higher ethanol intake and preference,without altering sucrose intake and preference or quinine intake and preference.② Comparison of the expres⁃sion of PDE7 subtypes in the brain striatum of B6 and DBA mice.Compared to DBA mice,naive B6 mice showed significant lower expres⁃sion of PDE7A(P＜0.01),but not PDE7B in the striatum.③PDE7A in the striatum of naive and ethanol-drinking B6 mice.After ethanol drinking for 10 d,B6 mice showed significant increases in expression of PDE7A in the striatum(P＜0.01)relative to naive controls,but no changes in PDE7B.④Effect of BRL-50481 on ethanol drinking behavior in B6 mice.BRL-50481(0.3-3 mf·kg-1)reduced ethanol intake(P＜0.01 for 0.3 and 1 mg·kg-1;P＜0.05 for 3 mg·kg-1)and preference(P＜0.05 for all doses)without altering the total fluid intake in B6 mice.CON⁃CLUSIONPDE7A expression is relatively high in the striatum of alcohol preferring mice such as B6 and can be further increased following etha⁃nol drinking.PDE7A is an important player in the regulation of alcohol consumption.Drugs inhibiting PDE7A can be novel treatments for alcoholism.

Key words:phosphodiesterase;ethanol drinking;behavior

Foundation item:National Natural Science Foundation of China(81773717);National Natural Science Foundation of China (81801510);Academic Promotion Program of Shandong First Medical University(2019QL011);and Science and Technology Development Plan Project in Tai′an City(2017NS0 237).

Corresponding author:ZHANG Fang-fang,E-mail:15650512848@163.com; ZHANG Han-ting,E-mail:htzhang@qdu.edu.cn

T2-14

海马神经发生对甲苯丙胺成瘾小鼠成瘾记忆消退及再唤起的作用

车晓航，白亦君，蔡佳伶，吴春福，杨静玉

（沈阳药科大学生命科学与生物制药学院，辽宁 沈阳 110016）

摘要：目的最近研究表明海马神经发生（HN）可能对记忆产生双向调节作用，即促进HN不但能够促进记忆的形成，而且同时可能诱导记忆的遗忘。目前，包括吗啡、可卡因等药物所导致的成瘾记忆被证明可能与HN有关，但HN在甲苯丙胺（METH）相关成瘾记忆中的作用还有待研究。方法通过METH诱导的条件性位置偏爱模型模拟小鼠对METH的成瘾记忆，然后进一步通过给予7，8-二羟基黄酮（DHF）与替莫唑胺（TMZ）分别在METH成瘾记忆获得前及获得后促进或抑制HN，同时结合神经发生标志物双皮质素（DCX）及5-溴脱氧尿嘧啶核苷（BrdU）考察HN在METH成瘾记忆中的作用。结果条件性位置偏爱实验结果表明，METH能够显著诱导小鼠的成瘾记忆；BrdU和DCX的免疫组化及免疫荧光结果表明DHF及TMZ分别能够促进或抑制小鼠HN。在METH诱导小鼠成瘾记忆获得后，通过DHF促进HN能够显著缩短其成瘾记忆的消退时间并抑制成瘾记忆的再唤起程度，而通过TMZ抑制HN则起到相反的结果；而在METH诱导成瘾记忆获得前，无论给予DHF或TMZ，均能延长小鼠成瘾记忆的消退时间，并增加其再唤起程度。结论在METH成瘾记忆获得后，促进HN能够显著诱导METH成瘾记忆的遗忘，提示基于干预HN的方式可能为METH成瘾行为的治疗提供新的手段与策略。

关键词：甲苯丙胺；成瘾记忆；神经发生

通讯作者：杨静玉，E-mail：yangjingyu2006@gmail.com；吴春福，E-mail：wucf@syphu.edu.cn

T2-15

栀子厚朴汤抗抑郁作用及机制

邢 航，张 阔

（沈阳药科大学生药教研室，辽宁 沈阳 110016）

摘要：目的抑郁症是由各种原因引起以抑郁为主要特征的心境或情感性精神障碍，具有患病率高、复发率高、致残率高等特点。栀子厚朴汤出自于《伤寒论》，由栀子、枳实和厚朴3味药组成。主治伤寒下后，心烦腹满，卧起不安。现代研究表明，栀子厚朴汤具有抗抑郁的作用。本文研究栀子厚朴汤对慢性不可预见温和应激所致抑郁大鼠的保护作用及相关机制。方法采用成年雄性SD大鼠，通过慢性不可预见温和刺激建立大鼠抑郁模型，连续ig给予栀子厚朴汤（3.66，7.32和14.64 g·kg-1）21 d，结合大体打分、糖水偏爱、强迫游泳和旷场等多种行为学方法，明确栀子厚朴汤对慢性不可预见温和应激抑郁大鼠的保护作用；进一步利用酶联免疫和免疫荧光等技术，研究栀子厚朴汤对慢性不可预见温和应激抑郁大鼠的保护机制。结果行为学结果表明，与慢性不可预见温和刺激模型组相比，栀子厚朴汤给药组大鼠外貌评分显著降低，糖水偏爱显著增加，游泳不动时间显著缩短，在旷场中心区域和非边缘区域停留时间显著增加，在边缘区域停留时间显著减少。神经内分泌结果表明，栀子厚朴汤给药组大鼠血清皮质酮抑制率显著增加，血清皮质酮水平显著降低。免疫组织化学结果表明栀子厚朴汤给药组大鼠海马中脑源性神经营养因子（BDNF）表达显著增加，海马齿状回DCX和BrdU/NeuN阳性细胞数显著增加。结论栀子厚朴汤连续灌胃21 d对慢性不可预见温和应激所致抑郁大鼠具有保护作用，其机制可能与增加海马BDNF和增强海马神经发生有关。

关键词：栀子厚朴汤；抑郁；脑源性神经营养因子；神经发生

通讯作者：张 阔，E-mail：zhangkuosyphu@163.com

T2-16

Roles of 5-HT2receptors in effects of DOM,ketamine and methamphetamine on prepulse inhibition in Sprague Dawley rats

JIANG Kai-li1,LI Kai-xi1,2,LIU Xiao-yan1,SU Rui-bin1

(1.State Key Laboratory of Toxicology and Medical Countermeasures,Beijing Key Laboratory of Neu⁃ropsychopharmacology,Beijing Institute of Phar⁃macology and Toxicology,Beijing 100850,China;2.Key Laboratory of Xinjiang Phytomedicine Resource and Utilization,Ministry of Education,Department of Pharmacology,Shihezi University,Shihezi 832003,China)

Abstract:OBJECTIVEPrepulse inhibition(PPI)of the acoustic startle response provides a measure of sensorimotor gating system mecha⁃nisms,which is known to be impaired in schizo⁃phrenia patients.We assessed the effects of the 5-HT2A/2Creceptor agonist(±)2,5-dimethoxy-4-methylamphetamine(DOM),the NMDA receptor antagonist ketamine,the dopamine receptor ago⁃nist methamphetamine(Meth)on PPI and the startle magnitude in SD rats.METHODS AND RESULTSSystemic administration of the three compounds all dose-dependently reduced PPI.However,as far as startle magnitude,only DOM at the doses of 3 mg·kg-1reduced that,while both ketamine and Meth did not change the startle magnitudes.Furthermore,to determine whether 5-HT2Areceptor mediate this effect,the non-spe⁃cific 5-HT2receptor antagonist cyproheptadine,specific 5-HT2Areceptor antagonist ketanserin and specific 5-HT2Creceptor antagonist SB242084 were tested.Cyproheptadine,ketan⁃serin and SB242084 did not alter startle ampli⁃tude by themselves in SD rats and only ketanserin slightly increased PPI at higher dose(3 mg·kg-1).PPI impairment induced by DOM was restored by pretreatment of cyproheptadine(1 mg·kg-1)and ketanserin(1 mg·kg-1),while not by pretreat⁃ment of SB242084(1 mg·kg-1).Damage of PPI induced by ketamine and Meth was not reversed by cyproheptadine(1 and 5 mg·kg-1).CONCLU⁃SIONThe receptor mechanisms underlying the disruption of PPI caused by DOM,ketamine and Meth were different from each other,at least 5-HT2Areceptor was not the junction receptor for which the three chemicals acted.

Key words:prepulse inhibition;5-HT2receptor;startle magnitude;psychoactive substances

T2-17

ABIN-1 targets β-arrestin-2 to attenuate opioids tolerance

ZHANG Yi-xin,ZHOU Pei-lan,LU Feng-feng,SU Rui-bin,GONG Ze-hui

(State Key Laboratory of Toxicology and Medical Countermeasures,Beijing Key Laboratory of Neu⁃ropsychopharmacology,Beijing Institute of Phar⁃macology and Toxicology,Beijing 100850,China)

Abstract:OBJECTIVEChronic morphine exposure reduced analgesic efficiency leads to morphine tolerance which was mediated by βarrestin signaling of MOR,but the further mecha⁃nism remains unclear.METHODS AND RESULTSMorphine tolerance and dependence was attenuated by overexpression of ABIN-1 in mice brain.ABIN-1 in hippocampus or vascular nucleus participated in morphine tolerance and physical dependence.ABIN-1 through AHD2 re⁃gion interacted with MOR to regulate its function.As the result,the peptide of AHD2 could also de⁃lay morphine tolerance as ABIN-1.Two lines of evidence may explain the function of ABIN-1 on morphine tolerance.First,ABIN-1 inhibited the phosphorylation and internalization of MOR after acute or chronic agonists treatment through inter⁃action with MOR.Second,The formation of ABIN-1-β-arrestin-2 complexes promoted the translocation of β-arrestin-2 to plasma mem⁃brane and accelerated the ubiquitination of β-ar⁃restin-2 to degrade it.However,the function kjj⁃mice brain.CONCLUSIONABIN-1 may target βarrestin-2 to regulate MOR function and provides a potential strategy for enhancing morphine anal⁃gesia without increasing analgesic tolerance.

Key words:MOR;ABIN-1;β-arrestin-2;tolerance

T2-18

赖右苯丙胺对前额叶皮质执行功能的影响

陈健民，王志媛，吴世轩，吴 宁，李 锦

（军事科学院军事医学研究院毒物药物研究所，北京 100850）

摘要：目的研究赖右苯丙胺（Lis）与传统认知增强剂右旋苯丙胺（Amp）对前额叶皮质执行功能（工作记忆、持续性主动注意、抑制控制和辨别能力）的药效学差异，为把Lis作为安全高效且低成瘾的认知增强候选化合物提供研究依据。方法建立反映空间工作记忆的Y迷宫连续自发交替任务，反映持续性主动注意及抑制控制的五项选择连续反应时间任务，反映空间信息辨别的Y迷宫延迟自发交替任务以及视觉信息辨别的视觉辨别学习任务，在此基础上研究Lis与Amp的药效学差异。结果在Y迷宫连续自发交替任务中，Lis 0.2～1.5 mg·kg-1和Amp均不影响大鼠自发交替比，Lis 4.5 mg·kg-1显著提高大鼠自发交替比率（P<0.05），而Amp 4.5 mg·kg-1显著降低大鼠自发交替比率（P<0.01），进一步增加剂量至13.5 mg·kg-1，Amp显著抑制大鼠自发交替比（P＜0.01），而赖右苯丙胺对此无显著影响。在5项选择连续反应时间任务中，在长时程高注意任务情况下，Amp 0.05～0.5 mg·kg-1及 Lis 0.1～0.3 mg·kg-1均无提高正确率的作用，而Lis 1.0 mg·kg-1则显著提高大鼠反应正确率（P＜0.05），缩短正确反应潜伏期（P＜0.05）。在高冲动状态下，Amp 0.5 mg·kg-1显著增加大鼠冲动行为（P＜0.01），而Lis 1.0 mg·kg-1并无明显影响。在空间信息辨别的Y迷宫延迟自发交替任务，Amp 0.2～4.5 mg·kg-1均无增加大鼠新臂进臂次数和停留时间百分比，而Lis 4.5 mg·kg-1显著提高新臂进臂次数（P＜0.05）和停留时间百分比（P＜0.01），增加剂量至13.5 mg·kg-1后，Amp显著降低该指标（P＜0.01）而Lis对此无明显影响。在视觉辨别学习任务中，Lis 1.0 mg·kg-1急性给药显著提高大鼠的辨别正确率（P＜0.05），而Amp 0.05～0.5 mg·kg-1无明显影响。结论相比于Amp，Lis能提高空间工作记忆，在长时程高强度注意力任务中能提高持续性主动注意，急性给药能促进视觉信息的辨别学习能力，提示Lis能提高前额叶皮质的执行功能，且安全性较好。本研究为把赖右苯丙胺能作为安全高效且低成瘾的认知增强候选化合物提高研究基础和研究依据。

关键词：赖右苯丙胺；右旋苯丙胺；前额叶皮质；执行功能

通讯作者：吴 宁，E-mail：wuning7671@126.com；李 锦，E-mail：jinli9802@163.com

专题3：脑卒中与脑损伤

T3-1

星形胶质细胞对脑缺血再灌注损伤的保护作用

耿宇涵1，张丹参1,2，李 炜1

（1.河北北方学院药学系，河北省神经药理学重点实验室，河北 张家口 075000；2.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：在缺血性脑损伤中，神经元和星形胶质细胞会迅速耗尽能量和氧气，这个过程最终导致神经元失去存活的能力，发生不可逆转的组织损伤和神经损伤。星形胶质细胞在中枢神经系统中有着十分重要的作用，如调控神经系统的发育、脑的物质代谢和血流。在正常情况下，正常健康的星形胶质细胞会保护神经元，避免氧化应激和兴奋性毒性。星形胶质细胞拥有十分丰富的缝隙连接，且有研究表明，在传导细胞的凋亡信号和细胞间信息交流时，缝隙连接有很重要的作用。连接蛋白是缝隙连接的重要组成部分。在大脑中，缝隙连接作为细胞间通信（GJIC）可能允许能量代谢物和危险分子的传递。星形胶质细胞GJIC和神经元共培养可增强神经元细胞抵抗氧化应激的能力；阻塞星形胶质细胞间隙连接，增加共培养时神经元对谷氨酸细胞毒性的敏感性。

关键词：星形胶质细胞；脑缺血；保护作用

基金项目：河北省重点研发计划项目生物医药专项（20372509D）

通讯作者：张丹参，E-mail：zhangds2011@126.com

T3-2

星形胶质细胞拯救缺血性脑卒中

苏晓梅1，张丹参1,2

（1.河北医科大学基础医学院药理学教研室，河北 石家庄 050017；2.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：星形胶质细胞（AS）是中枢神经系统内含量最丰富的神经胶质细胞，约占脑总体积的50%，其数量约为神经元的10倍。AS与神经元之间存在广泛而复杂的信息传递，以直接或间接作用的方式与神经元发生联系。在生理情况下，AS对神经元发挥营养支持的作用，可参与神经元的糖、脂肪和体液代谢过程。在缺血性脑卒中发生时，由于AS线粒体内可以储存大量糖原，使得其对低氧环境具有弹性和适应性，AS可以通过自身生物能量和线粒体动力学的变化为损伤神经元提供能量支持。不仅如此，在缺血性脑卒中发生时，AS可以通过隧道纳米管或细胞外囊泡与周围的受损神经元进行线粒体交换，释放功能性线粒体。同时，神经元也可以释放受损线粒体并将其转移至AS内，进行内吞和降解，实现线粒体的循环利用，使得线粒体在健康细胞和受损细胞之间发生串扰。此外，AS的终足与中枢神经系统中的微动脉和毛细血管接触，成为神经元网络和血管网络之间的桥梁，通过影响血管直径来调节脑血流。综上，AS可以作为治疗缺血性脑卒中的重要靶点之一，改善缺血性脑卒中的临床治疗效果。

关键词：星形胶质细胞；缺血性脑卒中；生物能量；线粒体转移；脑血流

基金项目：河北省重点研发计划项目生物医药专项（20372509D）

通讯作者：张丹参，E-mail：zhangds2011@126.com

T3-3

姜黄素对脑缺血再灌注损伤的神经保护作用及其机制

谢佳佳，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：姜黄素是从姜黄根茎中提取出来的一种脂溶性多酚类化合物，是姜黄发挥药理作用的主要活性成分。研究表明，姜黄素可减轻多种器官的缺血再灌注损伤，容易透过血脑屏障，可用于治疗中枢神经系统疾病。脑缺血再灌注损伤是在发生缺血性脑卒中后的短时间内，坏死脑组织周围的缺血神经元细胞在恢复血供时，发生加重缺血级联反应，造成脑组织更为严重的再灌注损伤。国内外研究表明，姜黄素可通过抑制脑缺血再灌注损伤的多种病理机制而发挥显著的神经保护作用，本文主要从其作用机制方面进行总结。①激活SOD，加速体内超氧阴离子和羟自由基等ROS的清除。姜黄素作为一种强抗氧化剂，主要通过激活SOD，GSHPx，CAT和GST等抗氧化酶，从而稳定大脑的抗氧化酶系统，加速体内自由基的清除；②降低IL-6和TNF-α的组织浓度，下调NF-κB的表达，抑制炎症反应。姜黄素可以通过介导TLR-4/P38/MAPK途径，降低IL-6，TNF-α和NOS等炎症因子的水平，减少炎症反应，从而改善神经功能；③抑制凋亡。姜黄素可通过激活丝裂原活化蛋白激酶信号通路，降低胱天蛋白酶3活性，抑制凋亡蛋白表达，最终抑制脑缺血再灌注损伤诱导的细胞凋亡；④抑制内质网应激。姜黄素可通过GADD153和胱天蛋白酶12途径抑制内质网应激，CADD153作为一种信号分子，通过影响细胞内的Ca2+代谢和下调Bcl-2等多种途径抑制内质网应激，促进神经功能恢复。综上所述，姜黄素能通过多种分子机制抑制脑缺血再灌注损伤，发挥神经保护作用，具有补充现有缺血性脑卒中治疗方法的潜力。

关键词：姜黄素；脑缺血再灌注损伤；神经保护

基金项目：河北省重点研发计划项目生物医药专项（20372509D）

通讯作者：张丹参，E-mail：zhangds2011@126.com

T3-4

PI3K/Akt信号通路对脑缺血再灌注损伤的作用机制

袁鑫茹，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：PI3K/Akt信号通路是经典的抗凋亡和促存活的信号转导通路之一，在脑卒中、肿瘤、糖尿病和抑郁等疾病的发生中具有重要作用。脑缺血是目前世界上继心脏病和癌症之后的第三大致死性疾病，其最常用的治疗手段是再灌注，但可能引起缺血细胞损伤加重，即脑缺血再灌注损伤（CIRI），其机制非常复杂。大量研究表明，通过激活PI3K/Akt信号通路，可减轻CIRI。因此，PI3K/Akt信号通路在CIRI中发挥重要作用，其机制主要有以下4个方面：①抑制细胞凋亡。CIRI后多数受损细胞最终会死亡或凋亡。据报道，PI3K/Akt信号通路可通过调节mTOR和Bcl-2表达，进而控制凋亡和促凋亡蛋白的合成。此外，PI3K/Akt信号通路还可减少线粒体介导的细胞凋亡；②促进脑血管新生。血管内皮生长因子（VEGF）能够促进神经元和新生血管的生成，激活VEGF介导的PI3K/Akt信号通路可促进脑血管的新生；③促进神经生长因子释放。大脑周围神经损伤后神经营养因子释放减少，神经生长因子与酪氨酸激酶结合后激活PI3K/Akt信号通路，抑制神经元的损伤，从而促使神经营养因子释放增加；④抑制自噬。缺血损伤后可导致自噬过度活化，细胞损伤死亡，PI3K/Akt信号通路的激活可抑制过度活化的自噬水平，减轻细胞损伤。本文总结PI3K/Akt信号通路在CIRI中的作用机制，为治疗CIRI提供参考依据。

关键词：PI3K/Akt信号通路；脑缺血再灌注损伤；作用机制

基金项目：河北省重点研发计划项目生物医药专项（20372509D）

通讯作者：张丹参，E-mail：zhangds2011@126.com

T3-5

TIMP1通过与CD63/整合素β1复合物相互作用并调节下游FAK/RhoA信号保护血脑屏障

唐婧姝，康钰莹，黄龙舰，吴 镭，彭 英

（中国医学科学院&北京协和医学院药物研究所，北京 100050）

摘要：目的创伤性脑外伤（TBI）是青壮年致残和死亡的首位原因。TBI后血脑屏障（BBB）明显受损，而改善BBB功能障碍、维持其完整性可能是TBI治疗的潜在策略。TIMP1参与调节血管重塑以及稳定血管通透性，但分子机制尚不明确。本研究采用整体和离体TBI模型，探讨TIMP1对TBI导致BBB功能失调的改善作用及具体调控机制，以期为TBI的临床治疗和药物研发提供更多策略。方法首先，建立TBI模型小鼠，给予rTIMP1，采用平衡木和转棒实验评估神经功能；采用伊文思兰实验评估BBB通透性。其次，利用缺氧和外源加入IL-1β，在人脑微血管内皮细胞中建立离体TBI模型，分别敲减、过表达TIMP1或外源给予rTIMP1，应用荧光素酶渗漏实验和TEER实验检测细胞屏障功能，利用生物素标记法提取膜蛋白和IF实验检测连接复合体的表达和定位。在机制探讨部分，比较rTIMP1和Ala-rTIMP1（无MMP抑制活性的rTIMP1）对内皮屏障功能的影响。最后采用Co-IP检测TIMP1与CD63/整合素β1相互作用，利用FAK抑制剂明确TIMP1调控BBB的关键通路，采用Rhotekin pulldown检测RhoA活化水平，以及应用IF检测F-actin聚集程度。结果rTIMP1可以改善TBI小鼠神经功能损伤，减少脑组织伊文思蓝渗漏和连接复合体表达缺失。敲减TIMP1增加荧光素酶渗漏、降低TEER、破坏连接复合体的表达和定位，而给予rTIMP1或过表达TIMP1均能逆转上述损伤，表明TIMP1是调控内皮屏障完整性的关键分子。rTIMP1和Ala-rTIMP1均能改善TBI小鼠神经功能损伤和内皮细胞屏障功能损伤，二者作用无显著性差异，表明TIMP1对BBB调控作用可以不依赖于其MMP抑制活性。TIMP1与CD63/整合素β1结合并激活下游FAK信号转导，FAK抑制剂可逆转TIMP1对TBI小鼠血脑屏障的保护作用以及连接复合体的调控作用，表明FAK激活是TIMP1发挥作用的关键节点。此外，TIMP1抑制RhoA活化和F-actin解聚，参与维持细胞骨架稳定性。结论TIMP1与血管内皮细胞表面CD63/整合素β1结合，以受体介导的细胞因子活性调节RhoA活化、抑制细胞骨架重构从而调控连接复合体的表达和分布，减轻BBB损伤进而在TBI中发挥保护作用。本研究提示针对TIMP1及其下游通路的干预可能是TBI治疗的潜在策略。

关键词：TIMP1；创伤性脑外伤；血脑屏障；连接复合体

T3-6

Intranasal ginsenoside Rb1 protects pentyl⁃enetetrazole-induced epileptic mice

LI Juan,LIU Yu-shu,WANG Xi,LIU Ying,MA Qing,TANG Min-ke

(School of Chinese Materia Medica,Beijing Univer⁃sity of Chinese Medicine,Beijing 102488,China)

Abstract:OBJECTIVETo evaluate whether ginsenoside Rb1 has antiepileptic effects on pen⁃tylenetetrazole(PTZ)-induced epileptic mice via intranasal therapeutic administration.METHODSRb1 monoclonal antibody was used to observe the distribution of Rb1 20 mg·kg-1in mouse brain tissues under different administration routes and to explore the feasibility of intranasal Rb1.PTZ was injected intraperitoneally into healthy ICR mice every 48 hours to construct a tonic-clonic epileptic model.Then Rb1 20 or 40 mg·kg-1or valproate 300 mg·kg-1or saline was administered intranasally for 30 d,and PTZ was continued every five days to imitate occa⁃sional convulsions in the clinic.Racine scale(RCS)and wireless electroencephalogram(EEG)monitoring were used to assess the presence and severity of seizure.Immunofluorescence(IF)was performed after drug treatment to evalu⁃ate the effect of Rb1 on brain neuron,microglia and astrocyte in epileptic mice.RESULTSRb1 had specific binding with anti-Rb1 in the brain under different administration routes,and intrana⁃sal Rb1 was able to enter the brain and play a therapeutic role(P＜0.01).PTZ-injured mice pre⁃sented body mass loss,higher seizure stage and shorter seizure latency.At the same time,epilep⁃tic waves,mainly spikes,were detected by wire⁃less EEG.Compared with PTZ group,intranasal Rb1 increased mice weight(P＜0.01)and seizure latency(P＜0.05),reduced seizure stage(P＜0.01)and EEG spikes.In addition,Rb1 significantly reduced neuron loss(P＜0.01)indicated by NeuN staining and decreased the number of acti⁃vated microglia (P＜0.01)indicated byIba-1 staining in the cortex and CA1 area of hippocam⁃pus.Moreover,Rb1 reduced the decrease of GLT-1 and GS expression(P＜0.05)induced by PTZ.CONCLUTIONIntranasal Rb1 has anti-epi⁃leptic effects on PTZ mice.Moreover,Intranasal Rb1 affects the functions of neurons,astrocytes and microglia through regulating the expression of GLT and GS in astrocytes,which may be related to its anti-epileptic effect.

Key words:ginsenoside Rb1;antiepileptic effects; epileptic mice; pentylenetetrazole;wireless electroencephalogram

T3-7

Possible role of translocator protein 18 ku on sepsis associated encephalopathy by mediat⁃ing neuroinflammation

LIU Hai-ping, JIN Gui-lin, HUANG Ya-xin,YUE Rong-cai,YU Chang-xi

(Department of Pharmacology,School of Pharmacy,Fujian Medical University,Fuzhou 350122,China)

Abstract:OBJECTIVETo clarify the role of translocator protein 18 ku(TSPO)on cecum liga⁃tion and puncture(CLP)induced sepsis associat⁃ed encephalopathy(SAE)mice,which consis⁃tently demonstrated astrocyte activation and neu⁃roinflammation.Background SAE,a brain dys⁃function,caused by systemic infection without clinical or laboratory evidence of direct infection.Most patients have symptoms such as long-term cognitive dysfunction.As the pathogenesis of SAE is very complex,neuroinflammation for SAE is one of the causes of the disease.TSPO as a marker of neuroinflammation that has the poten⁃tialto regulate neuroinflammation and SAE.METHODSThe animal model of SAE was in⁃duced by CLP.TSPO ligands and TSPO knock⁃out mice were used for behavioral and molecular biology research.Survival rate of mice within 120 h on CLP mice was observed.The changes of cog⁃nitive function in mice were observed by Morris water maze and open field test.The changes of proinflammatory factors(IL-1β,TNF-α,IL-6)in hippocampus were observed by ELISA;Astro⁃cyte activation,marked by GFAP,in hippocam⁃pal was analyzed by tissue immunofluorescence and Western blotting.RESULTSPretreatment with the TSPO ligands,XBD173 or PK11195,sig⁃nificantly improved the survival rate of CLP mice.The results of Morris water maze showed that TSPO ligands significantly increased the number of crossing the platform and the target quadrant time on CLP mice,suggesting that TSPO ligands may improve the learning and memory ability of CLP mice.Subsequent experiments revealed that TSPO ligands can reduce the inflammatory factors(IL-1β,TNF-α,IL-6)and astrocyte activa⁃tion in hippocampus of CLP mice.Similar results were also confirmed in TSPO knockout CLP mice,suggesting intervention of TSPO can reduce neuroinflammatory response and play a protec⁃tive role on SAE mice.CONCLUSIONTSPO may play a critical role on SAE mice.Targeting TSPO by pharmacological means may improve the survival rate and cognitive function on CLP mice,which may through inhibiting astrocyte acti⁃vation and neuroinflammation in hippocampal.

Key words:translocator protein 18 ku;astro⁃cyte;sepsis associated encephalopathy;cogni⁃tive dysfunction;hippocampal

Foundation item:Joint Funds for the Innovation of Science and Technology of Fujian Province(2019Y9009);and Natural Science Foundation of Fujian Province(2020J01618)

Corresponding author:YU Chang-xi,E-mail:changxiyu@mail.fjmu.edu.cn;JIN Gui-lin,E-mail:jinguilin0611@163.com

T3-8

Neuroprotective effect of cerebroprotein hydrolysate on cerebral ischemia-reperfusion injury mice

SHI Cai-yun1,AN Zi-xuan1,LI Wei1,2

(1.Department of pharmacy, Hebei North University,Zhangjiakou 075000,China;2.Hebei Provincial Key Laboratory of neuropharmacology,Zhangjiakou 075000,China)

Abstract:OBJECTIVETo investigate the neuroprotective effect of cerebroprotein hydroly⁃sate (CH)on cerebral ischemia-reperfusion injury in mice.METHODSA total of 60 male SPF Kunming mice were randomly divided,reforming longa method into sham group(sham),model group(tMCAO,reforming longa method),CH 0.2 and 0.5 g·kg-1groups and positive drug control group(edaravone 0.008 g·kg-1).Neurological deficit score were performed 24 h after opera⁃tion.Mice with scores ranged between 1 and 3 were included in subsequent experiments.Each group had 8 mice.CH edaravone and normal sa⁃line were ip injected for 5 d.The tMCAO group and the sham group were administered the same amount of normal saline as administration groups.TTC staining was used to measure the volume of cerebral infarction;ELISA was per⁃formed to detect the levels of interleukin-6(IL-6),interleukin-1β (IL-1β),brain-derived neurotrophic factor(BDNF)and interferon-γ (IFN-γ)in serum and penumbra.RESULTSTTC staining results showed that there was no infarction in sham group.Compared with tMCAO group,the infarct volume in each administration group was signifi⁃cantly decreased (P＜0.01).ELISA results showed that IL-6,IL-1β and IFN-γ in serum and penumbra were of significant difference between tMCAO group and sham group(P＜0.01),and BDNF was significantly decreased (P＜0.01).Compared with tMCAO group,IL-6,IL-1β and IFN-γ in serum and ischemic penumbra were sig⁃nificantly decreased in all administration groups(P＜0.01),while the content of BDNF was in⁃creased in CH 0.2 g·kg-1group and edaravone 0.008 g·kg-1group(P＜0.05),and other groups were significantly increased(P＜0.01).CONCLU⁃SIONCH could reduce the cerebral infarction vol⁃ume and improve the nerve injury caused by cerebral ischemia-reperfusion.The mechanism was related to inhibit the expression of IL-6,IL-1β and IFN-γ and increase the expression of BDNF possibly.

Key words:cerebral ischemia-reperfusion;injury;cerebroprotein hydrolysate

Fund program:Natural science foundation of Hebei Province(H2020405298)

Corresponding author:LI Wei,E-mail:leewei318@163.com

T3-9

大麻二酚抗高热惊厥的作用机制

于永洲，张 炜

（河北医科大学中西医结合研究所，河北 石家庄 050013）

摘要：目的明确大麻二酚是否对高热惊厥小鼠具有保护作用，及其可能的分子机制和可能的作用靶点。方法应用HE染色观察大麻二酚对高热惊厥小鼠海马神经元的形态学改变；膜片钳观察记录大麻二酚对海马神经元的兴奋性的影响。结果①行为学实验结果表明，大麻二酚能够显著延长高热惊厥小鼠的潜伏期，且具有剂量依赖性。②HE染色结果显示，大麻二酚对高热惊厥诱发的小鼠海马CA1区神经元损伤具有保护作用。③急性海马脑片膜片钳实验结果显示，大麻二酚能够显著降低小鼠海马CA1区锥体细胞的兴奋性，且呈剂量依赖性。结论大麻二酚具有抑制小鼠高热惊厥的作用，其机制可能为通过抑制AMPA受体激活，使得AMPA受体内流进入细胞的阳离子减少，继而降低海马神经元兴奋性，从而发挥对高热惊厥状态下的海马神经元的保护作用。

关键词：大麻二酚；高热惊厥；作用机制；作用靶点

通讯作者：张 炜，E-mail：zhangwei_wz59@163.com

T3-10

山药多糖对脑缺血再灌注损伤的保护作用机制

李朋月1,2，景永帅3，吴兰芳1,2，郑玉光1,2

（1.河北中医学院药学院，河北 石家庄 050200；2.河北省中药炮制技术创新中心，河北 石家庄 050091；3.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：山药是常用药食两用资源，含有蛋白质、多糖、薯蓣皂苷、尿囊素及微量元素等成分。多糖作为山药的主要活性成分之一，具有抗肿瘤、降血脂、降血糖、调节免疫、抗炎、抗氧化和神经保护等作用。脑缺血再灌注损伤可导致一系列严重的脑疾病，导致死亡或不同程度的残疾，研究表明，山药多糖可能通过多种机制发挥对脑缺血再灌注损伤的保护作用。①改善脑组织抗氧化能力。氧化应激是脑缺血再灌注损伤的主要原因之一，在氧化应激条件下，超氧化物歧化酶（SOD）和谷胱甘肽（GSH）等抗氧化酶和氧化应激标志物丙二醛（MDA）在神经细胞对活性氧（ROS）的抵抗中起关键作用——诱导细胞死亡。相关研究结果表明，山药多糖可使细胞抗氧化剂SOD和GSH含量显著增高、MDA含量显著降低。此外，山药多糖也可以通过调节CaMMKβ的表达进而影响Nrf2/HO-1信号通路的激活，减少ROS的积累。②抑制神经元细胞凋亡。当缺血再灌注诱导脑组织产生并积累ROS时，这些自由基会激活线粒体、死亡受体和内质网应激，进而诱导神经元凋亡。研究发现，山药多糖能显著提高BV2细胞的活性，调节BV2细胞诱导的神经细胞凋亡；另外可调节脑缺血再灌注诱导的小胶质细胞凋亡，降低脑梗死面积。③抑制炎性细胞因子过度表达。氧化应激损伤产生的ROS会诱导外周免疫系统和中枢神经系统的炎症反应，从而产生大量的促炎因子如IL-6，IL-1β和TNF-α，促炎因子通过引起血管收缩、微血管阻塞、分泌细胞毒素、破坏血脑屏障的完整性和促进ROS的产生，加剧氧化应激损伤，最终导致神经细胞凋亡。山药多糖能够显著降低脑缺血再灌注损伤模型的IL-6，IL-1β和TNF-α的水平，减少炎症因子对神经元的损伤。本文综述了山药多糖在神经保护方面的作用及机制，以期为山药及其多糖成分的开发利用提供依据。

关键词：山药；多糖；脑损伤再灌注；作用机制

基金项目：河北省中医药管理局中医药科研计划项目（2021104）

通讯作者：郑玉光，E-mail：zyg314@163.com

T3-11

SMU-X通过Mfn2/JNK通路发挥抗炎作用对抗实验性自身免疫性脑脊髓炎疾病进程发展

李梦凡，汪海涛，徐江平

（南方医科大学药学院，广东 广州 510515）

摘要：目的SMU-X是治疗多发性硬化的潜在药物，但SMU-X的作用靶点较多，其治疗多发性硬化的具体机制还未明确。线粒体融合蛋白2（Mfn2）是一种线粒体外膜GTP酶，调节线粒体融合，影响内质网与线粒体的相互作用，并且被认为密切参与炎症小体的形成。本研究旨在确定SMU-X对实验性自身免疫性脑脊髓炎（EAE）小鼠的治疗作用是否与Mfn2有关。方法在BV2小胶质细胞系上构建脂多糖（LPS）刺激模型。采用Western印迹法观察炎症相关蛋白（iNOS和COX-2）变化，ELISA法检测炎症因子（IL-6和TNF-α）水平以及线粒体氧化应激的变化。Western印迹法观察Mfn2蛋白水平、炎症通路蛋白（p38 MAPK和JNK）磷酸化水平的变化。用C57小鼠构建EAE模型，进行临床评分，通过Western印迹法检测髓鞘碱性蛋白（MBP）的表达，采用HE染色和Western印迹法观察脊髓组织炎症细胞浸润及小胶质细胞激活相关蛋白的表达。结果①LPS处理后，BV2细胞激活，炎症因子IL-6和TNF-α及炎症相关蛋白iNOS和COX2显著上调，而给予SMU-X可逆转LPS诱导的BV2细胞激活以及炎症因子和蛋白上调。②LPS处理BV2细胞后，线粒体膜电位降低，超氧阴离子活性氧的生成明显上调，线粒体氧化应激水平上升，而给予SMU-X后可以改善线粒体氧化应激水平。③LPS处理BV2细胞后，Mfn2表达下调，P38和JNK磷酸化水平明显增加，给予SMU-X后可以逆转Mfn2的下调及P38和JNK磷酸化水平的增高。④敲低Mfn2可对抗SMU-X对LPS诱导的BV2细胞的抗炎保护作用。⑤敲低Mfn2可对抗SMU-X对BV2线粒体氧化应激水平的降低作用。⑥敲低Mfn2可对抗SMU-X对LPS诱导的BV2细胞的JNK通路的抑制作用。⑦SMU-X可降低EAE小鼠的临床评分及脊髓白质区炎症细胞浸润，增加脊髓组织结构蛋白MBP的表达。⑧EAE小鼠发病后，小胶质细胞激活标志物Iba1和炎症相关蛋白COX-2的表达明显上调，而给予SMU-X后可以改善这一现象。⑨EAE小鼠发病后，小鼠脊髓白质区Mfn2的表达明显降低，CD11b的表达明显升高，给予SMU-X后可以明显逆转这些变化。结论SMU-X可通过Mfn2/JNK通路发挥抗炎作用，从而对抗EAE小鼠疾病进程的发展。

关键词：多发性硬化；SMU-X；炎症；线粒体融合蛋白2

基金项目：国家自然科学基金（81773698）；国家自然科学基金（81974501）

通讯作者：徐江平，E-mail：jpx@smu.edu.cn

T3-12

抑制磷酸二酯酶4通过调控内质网应激对抗缺血性脑损伤

许炳钿，汪海涛，徐江平

（南方医科大学药学院，广东 广州 510515）

摘要：目的抑制磷酸二酯酶4（PDE4）可减少缺血性脑损伤导致的神经元死亡，然而机制还不清楚。本研究旨在确定抑制PDE4对缺血性脑损伤保护作用的机制。方法在HT-22细胞及原代皮质神经元上构建OGD模型，Western印迹法观察相关蛋白水平的变化，Co-IP实验观察肌醇依赖酶1α（IRE1α）以及肿瘤坏死因子受体相关因子2（TRAF2）的相互作用。在SD大鼠上构建MCAO导致的局灶性脑缺血动物模型，采用TTC染色检测脑梗死体积，Fluoro-Jade C染色实验观察缺血半暗带区神经元凋亡，滚轴实验和胶带黏附实验观察大鼠的运动功能障碍。结果抑制PDE4可减少SD大鼠脑梗死的体积，减少缺血半暗带区神经元凋亡以及改善SD大鼠的运动功能障碍。抑制PDE4或敲低PDE4B可逆转OGD诱导的HT-22细胞凋亡。抑制PDE4可降低SD大鼠缺血半暗带区内质网应激相关蛋白的表达。过表达PDE4B可上调内质网应激，而敲低PDE4B或抑制PDE4可对抗OGD诱导的HT-22细胞内质网应激。抑制PDE4可促进OGD诱导的核因子红系2相关因子2（NFE2L2，Nrf-2）核转位，并降低氧化应激水平。抑制PDE4可增加蛋白激酶B和糖原合酶激酶 3β（GSK3β）的磷酸化水平，而过表达 GSK3β组成性激活质粒GSK3β（S9A）可消除抑制PDE4对Nrf-2核转位、氧化应激和内质网应激的影响。Nrf-2抑制剂ML385、敲低Nrf-2以及下游靶蛋白HO-1均可阻断抑制PDE4的保护作用。抑制PDE4可减少IRE1α与TRAF2的相互作用，并减弱c-Jun N端激酶的磷酸化水平。抑制PDE4在原代皮质神经元中产生了与上述一致的保护作用。结论抑制PDE4可以通过激活Nrf-2保护内质网应激诱导的损伤，从而恢复内质网稳态并防止神经元死亡。

关键词：缺血性脑损伤；磷酸二酯酶4；内质网应激

基金项目：国家自然科学基金（81773698）；国家自然科学基金（81974501）

通讯作者：汪海涛，E-mail：wht821@smu.edu.cn；徐江平，E-mail：jpx@smu.edu.cn

T3-13

神经血管耦合功能障碍与脑损伤药物靶标研究

韩峰

（南京医科大学药学院，江苏 南京 211166）

摘要：目的大脑神经元活动依赖于脑血流持续供应的氧气和葡萄糖等营养物质，其中神经-血管偶联保障与局部大脑活动变化进行时空高度匹配的局部脑血流供应，从而为大脑信息传递和处理保证了充分的能量供应。众多脑损伤相关疾病源于大脑的神经-血管偶联障碍。因此，阐明神经-血管偶联的调控机制对发现精准药物靶标有重要意义。方法利用药理学/基因调控、电生理记录、组学等技术，探究血管源性的分泌蛋白Sema3G缺失介导的神经血管耦合功能障碍在神经系统疾病中的作用。结果①脑血管内皮源性Sema3G基因敲除导致认知功能下降，Sema3G/Nrp2/PlexinA4参与神经元树突棘的成熟及LTP。②Sema3G基因敲除导致血管丛的密度下降，血管发育延迟。靶向Sema3G/Nrp2/PlexinA4的基因调控，可以促进健康血管网络的形成，并促进病变血管的退化，同时改善认知功能。结论脑血管源性Sema3G信号分子介导的跨细胞类型通讯模式参与神经血管耦合障碍和脑损伤病理过程。因此，脑血管源性Sema3G及其下游信号通路为精确调控神经血管耦合提供新思路和药物新靶点。

T3-14

URB597 exerts neuroprotective effect against transient cerebral ischemic injury via restor⁃ing autophagy flux and inhibiting neuronal necroptosis in mice

YUAN Xiao-qian1,2,YE Wen-xuan1,2,LUO Dou-dou1,2,ZHOU Yu1,2,3

(1.Department of Basic Medical Science,School of Medicine,3.State Key Laboratory of Cellular Stress Biology, Xiamen University, Xiamen 361102,China;2.Key Laboratory of Chiral Drugs,Xiamen 361102,China)

Abstract:OBJECTIVEURB597(KDS-4103)is a potent and selective inhibitor of the enzyme fatty acid amide hydrolase(FAAH)and can ele⁃vate the level of oleoylethanolamide(OEA),a naturally occurring endocannabinoid in the brain.However,the effect of URB597 on cerebral isch⁃emic injury in mice remains unclear.METHODSFocal cerebral ischemia was induced by middle cerebral artery occlusion for 30 min followed by reperfusion for 24 h in mice.To observe the dosedependent effect,URB597(0.04-5.00 mg·kg-1,ip)was administered at the same time of reperfu⁃sion.To determine the time-dependent effect,URB597(1.00 mg·kg-1,ip)was administered as a single dose at 0,1,3 or 5 h after reperfusion.Twenty-four hours after brain ischemia,Beder⁃son scoring test and grip strength test were used to evaluate the neurological function;brain in⁃farct volume was assayed by 2,3,5-triphenyltetra⁃zolium chloride (TTC)staining or diffusionweighted magnetic resonance imaging (MRI).Laser speckle imaging(LSI)technique was used to assay the regional cerebral blood flow(rCBF);NeuN immunofluorescence staining was used to observe the neuron survival in the penumbra.To further investigate the underlying mechanism,au⁃tophagy flux related proteins(LC3-Ⅱ,P62 and LAMP2)and necroptosis related proteins(pRIPK3 and pMLKL)were detected by Western blotting and immunofluorescence staining.RESULTSTwenty-four hours after brain ischemia,URB597 dose-dependently improved neurological func⁃tion and reduced brain infarct volume.The most effective dose was 1.00 mg·kg-1;the therapeutic time window was within1 h after ischemic stroke.The protective effect is further confirmed by the results that post-ischemic treatment with URB597(1.00 mg·kg-1)significantly increased neurons survival,promoted autophagy flux and reduced cell necroptosis in cortical penumbra after cerebral I/R.CONCLUSIONURB597 doseand time-dependently exerts a neuroprotective effect against acute cerebral I/R injury.This neu⁃roprotective effect of URB597 may be associated with its restoration of autophagy flux and inhibi⁃tion of neuronal necroptosis in the cortical penumbra.

Key words:URB597;ischemic stroke;ischemic penumbra;neuronal necroptosis;autophagy flux;neuroprotective effect

Foundation item:National Natural Science Foundation of China(81603093);and the Open Research Fund of State Key Laboratory of Cellu⁃lar Stress Biology,Xiamen University(SKLC⁃SB2019KF016)

Corresponding author:ZHOU Yu,E-mail:zhouyu@xmu.edu.cn

T3-15

PPARα activation inhibits astrocyte over acti⁃vation by restoring autophagy flux after tran⁃sient brain ischemia

LUO Dou-dou1,2,YE Wen-xuan1,2,YUAN Xiao-qian1,2,ZHANG Ya-li1,2,JIN Xin1,2,ZHOU Yu1,2,3

(1.Department of Basic Medical Science,School of Medicine,3.State Key Laboratory of Cellular Stress Biology, Xiamen University, Xiamen 361102,China;2.Key Laboratory of Chiral Drugs,Xiamen,361102,China)

Abstract:OBJECTIVEAstrocytes activa⁃tion and glial scar formation are the important causes that hinder the recovery of motor function after cerebral ischemia.However,its precise mechanism has not been clarified.Peroxisome proliferator-activated receptor α (PPAR α )is a ligand-activated nuclear transcriptional factor.This study aims to further clarify the role of PPARα in astrocyte activation after cerebral isch⁃emia and explore the underlying mechanism.METHODSAstrocyte activation in vivo model was induced by transient middle cerebral artery occlusion(tMCAO)in mice and in vitro model was induced by oxygen-glucose deprivation/reox⁃ygenation(OGD/R)in primary culture of mouse astrocyte.The effects of PPARα on astrocyte ac⁃tivation and autophagy flux were observed in the condition of PPARα dysfunction(PPARα null mice)or PPARα activation by oleoylethanol⁃amide(OEA).RESULTSPPARα mainly ex⁃pressed in activated astrocytes during the chron⁃ic phase of brain ischemia and PPARα dysfunc⁃tion promoted astrocytes activation after brain ischemia in vivo and in vitro.After cerebral isch⁃emia,the expressions of LC3-Ⅱ/Ⅰ and P62 both increased in the brain tissue near the infarct core.Autophagic vesicles accumulation was ob⁃served by electron microscopy in astrocytes,and mRFP-GFP-LC3 adenovirus infection assay indi⁃cated the block of autophagy flux.PPARα dys⁃function aggravated autophagy flux block,while PPARα activation preserved the lysosome func⁃tion and restored autophagy flux in astrocytes after OGD/R.Autophagy flux blocker bafilomycin A1 and chloroquine antagonized the effect of OEA on inhibiting astrocyte activation.CONCLU⁃SIONPPARα activation inhibites the over-activa⁃tion of astrocytes by restoring the autophagy flux after cerebral ischemia.

Key words:astrocyte activation;autophagy;au⁃tophagy flux;ischemic stroke;peroxisome prolif⁃erator-activated receptor α;lysosome

Foundation item:National Natural Science Foundation of China(81603093);and the Open Research Fund of State Key Laboratory of Cellu⁃lar Stress Biology,Xiamen University(SKLC⁃SB2019KF016)

corresponding author:ZHOU Yu,E-mail:zhouyu@xmu.edu.cn

T3-16

星形胶质细胞和小胶质细胞对中枢神经系统的作用

孔德平，亓会斌，冯兆阳，谭 瑞，赵晓民

（山东第一医科大学药理研究所，山东 泰安 271016）

摘要：星形胶质细胞与小胶质细胞的作用以及相互作用是一直处于神经胶质细胞研究的前沿且推动着中枢神经系统疾病的全面认识。星形胶质细胞是神经系统中数量最多的细胞，当其正常功能丧失时会引发一系列中枢神经系统疾病，如阿尔茨海默症、肝性脑病等。星形胶质细胞被激活后形成反应性星形胶质细胞A1或A2表型，对中枢神经系统发挥着神经毒性和神经保护性的作用。小胶质细胞是中枢神经系统微环境变化的传感器，它与星形胶质细胞相比，较为敏感。研究表明小胶质细胞与中枢神经系统损伤和炎症密切相关。另外，小胶质细胞也可以被激活形成M1或M2表型从而发挥促炎和抗炎的作用。星形胶质细胞与小胶质细胞不仅可以单独作用于中枢神经系统，还可以相互作用调节中枢神经系统。小胶质细胞决定了反应性星形胶质细胞在中枢神经系统的功能。相反，星形胶质细胞通过其分泌的分子调节改变小胶质细胞的表型和功能以及吞噬作用。本综述讨论了目前星形胶质细胞与小胶质细胞以及两者相互作用对于中枢神经系统的作用，为中枢神经系统疾病提供参考依据。

关键词：星形胶质细胞；小胶质细胞；神经系统

通讯作者：赵晓民，E-mail：zhaoxiaominty@163.com

T3-17

Baroreflex impairment exacerbates LPS-induced inflammation in rat cerebral cortex and hippocampus

JI Wei＊,QI Hui-bin＊,ZHU De-rong,FENG Zhao-yang,KONG De-ping,ZHANG Fang-fang,YU Hai-yang,GAO Yong-feng,TAN Rui,ZHAO Xiao-min

(Institute of Pharmacology,Shandong First Medi⁃cal University & Shandong Academy of Medical Sciences,Tai′an 271016,China)

Abstract:OBJECTIVEPlatelets play a major role in mediating inflammatory response.The present work was designed to investigate whether arterial baroreflex impairment induced by sinoaortic denervation(SAD)affect platelet activation,leading to the exacerbation of cerebral cortex and hippocampus inflammation in rats.METHODSAdult male SD rats were divided into four groups:the sham control,the sinoaortic denervation(SAD),the sham+LPS,the SAD+LPS.In another experiment,there were also four groups:the sham control,the SAD,the SAD+LPS and the SAD+LPS+asprin.Four weeks after sham or SAD surgery,all rats were examined for the level of CD41,CD45,IL-1β and PF-4 in the cerebral cortex and hippocampus using immunofluorescence and ELISA.Blood platelet and leukocyte count,platelet microaggre⁃gation,expression of CD154 and CD62P on platelet surface and platelet-leukocyte aggregate level was detected by flow cytometry.RESULTSCompared with sham+LPS group,the in SAD+LPS group rats exhibited the high level of CD41,CD45,IL-1β and PF-4 in the cerebral cortex and hippocampus.Leukocyte count,platelet microag⁃gregation,expression of CD154 and CD62P on platelet surface and platelet-leukocyte aggregate level was increased,while blood platelet count was decreased in the SAD+LPS.Moreover,all the above changes were improved in the SAD+LPS+asprin group when compared with the SAD+LPS group.CONCLUSIONArterial baroreflex dysfunction exacerbates inflammation in the rat cerebral cortex and hippocampus,which is likely mediated by platelet.

Key words:arterial baroreflex;inflammation;platelet;aspirin

Foundation item:Natural Science Foundation of Shandong Province(ZR2017MH048)

Corresponding author:ZHAO Xiao-min,E-mail:xmzhao@sdfmu.edu.cn;TAN Rui,E-mail:rtan@sdfmu.edu.cn

＊Co-first author.

T3-18

拟人参皂苷F-11通过激活calcineurin介导的TFEB核转位缓解脑缺血后神经元自噬-溶酶体通路的功能障碍

刘月阳，付晓晓，杨静玉，吴春福

（沈阳药科大学生命科学与生物制药学院，辽宁 沈阳 110016）

摘要：目的转录因子-EB（transcription factor EB，TFEB）是一种调控自噬和溶酶体生物合成的重要转录因子。课题组前期研究已证明，TFEB可作为脑缺血治疗的新靶标。拟人参皂苷F-11（PF11）具有显著的脑缺血保护作用，其机制可能与其调节ALP功能相关。为此，本研究深入探究PF11调节脑缺血后ALP功能的作用是否与其调节TFEB的核转位相关及其调节机制。方法采用大鼠永久性脑缺血（pMCAO）及原代皮质神经元氧糖剥夺（OGD）模型，结合慢病毒敲低技术等观察PF11对脑缺血后TFEB核转位的调节。结果PF11（6和12 mg·kg-1，iv）显著增加pMCAO后24 h TFEB的核蛋白水平，并提高Tfeb及其下游靶基因mRNA水平。与在体结果一致，PF11（30 和 100 μmol·L-1）可显著增加OGD后12 h原代神经元TFEB总蛋白表达及核蛋白表达，显著促进TFEB入核，改善溶酶体功能，减少自噬小体及底物的不正常积累。此外，慢病毒敲低原代神经元中的TFEB能够逆转PF11对OGD后12 h ALP功能障碍的缓解作用。进一步研究表明，calcineurin抑制剂环孢菌素（10 μmol·L-1）可显著逆转PF11对神经元OGD后TFEB核转位的促进作用及其对ALP功能的调节。结论PF11可能通过促进calcineurin介导的TFEB核转位来缓解脑缺血后ALP功能障碍，进一步明确了PF11抗脑缺血的自噬相关机制。

关键词：转录因子-EB；脑缺血；氧糖剥夺；溶酶体；自噬

T3-19

氰化钠加重缺氧诱发脑损伤机制

石华香，周梦玮，李鹏飞，周 琥，王丽韫

（军事科学院军事医学研究院毒物药物研究所，抗毒药物与毒理学国家重点实验室，北京 100850）

摘要：目的探究急性氰化钠（NaCN）中毒合并缺氧对小鼠脑损伤的作用。方法观察生理盐水对照组、NaCN 0.26，0.38 和 0.51 mg·kg-1（ip）中毒组小鼠缺氧存活时间。小鼠随机分为对照组、NaCN（3.8 mg·kg-1）处理组、NaCN 复合缺氧30 min（CN-30）组和NaCN复合缺氧60 min（CN-60）组，采用激光散斑成像仪检测小鼠脑血流变化；组织含水量测定脑组织水肿影响；酶标法检测海马组织氧化应激反应指标T-SOD活力和MDA含量变化；免疫荧光法检测皮质和海马组织小胶质细胞炎性标志物Iba1和CD68信号表达变化；Western印迹法测定Iba1和CD68及凋亡信号分子Bcl-2和Bax蛋白表达变化。结果与对照组比较，NaCN中毒显著延长缺氧小鼠存活时间（P＜0.01），0.38 mg·kg-1延长（74.7±17.8）min。与对照组比较，NaCN组、CN-30组和CN-60组小鼠脑血流值分别降低12%，40%和55%（P＜0.01），脑组织含水量均显著增加（P＜0.01），海马组织T-SOD活力和MDA含量均显著增加（P＜0.01），皮质和海马组织表达Iba1+和CD68+小胶质细胞百分比均显著增加；NaCN组Bcl-2和Bax表达显著增加，CN-30组和CN-60组进行性增加。结论NaCN中毒延长小鼠缺氧存活时间，加重缺氧诱发的脑血流下降，引发脑组织水肿，加剧海马组织的氧化应激反应和小胶质细胞炎症反应，促进脑组织损伤。

关键词：氰化钠；脑血流；氧化应激；免疫荧光；凋亡

通讯作者：王丽韫，E-mail:lylywang1103@163.com

T3-20

Comparison between cannabidiol and sertra⁃line in modulation of post-traumatic stress dis⁃order-like behaviors and fear memory in mice

HAN Xiao1＊,SONG Xian-kui1,2＊,SONG Da-ke1,XIE Guan-bo1,GUO Hong-yan2,WU Ning1,LI Jin1

(1.Beijing Key Laboratory of Neuropsychopharma⁃cology,State Key Laboratory of Toxicology and Medical Countermeasures,Beijing Institute of Pharmacology and Toxicology,Beijing 100850,China;2.Shanxi Medical University School and Hospital of Stomatology,Taiyuan 030001,China)

Abstract:OBJECTIVEPost-traumatic stress disorder(PTSD)is characterized by poor adapta⁃tion to a traumatic experience and disturbances in fear memory regulation,and currently lacks effective medication.Cannabidiol(CBD)is the primary component of the Cannabis sativa plant;it does not have any psychoactive effects and has been implicated in modulating fear learning in mammals.The present study investigated the effect of CBD on PTSD-like behaviors in a mouse pre-shock model,the effect of CBD in the modulation of trauma-related fear memory,a crucial process leading to core symptoms of PTSD.METHODSPre-shock model was applied in which mice were submitted to training with two days of 0.8 mA×12 times of foot-shock,and PTSD-like behaviors was evaluated during 3 and 26 d,including freezing time to the conditioned context,open field test,elevated plus maze test and social interaction test.RESULTSCBD(10 mg·kg-1)administration alleviated main PTSD-like symptoms in the mouse pre-shock model by attenuating trauma-related fear memory,decreasing anxiety-like behavior,and increasing social interaction behavior.However,sertraline(15 mg·kg-1)was only effective when adminis⁃tered throughout the test period.Furthermore,CBD reduced the formation,retrieval,and recon⁃solidation of trauma-related fear memory,whereas sertraline only reduced fear-memory retrieval.Neither CBD nor sertraline influenced the acquisi⁃tion of trauma-related fear memory.CONCLU⁃SIONCBD produced anti-PTSD-like actions in mice,and could disrupt trauma-related fear mem⁃ory by interfering with multiple aspects of fear memory processing in mice.These findings indi⁃cate that CBD may be a promising candidate for treating PTSD.

Key words:post-traumatic stress disorder;can⁃nabidiol;sertraline;trauma-related fear memory processing;endocannabinoid system

Corresponding author:HAN Xiao,E-mail:xiaohan6699@yeah.net; WU Ning, E-mail:wuning7671@126.com.

＊Co-first author.

T3-21

Establishment of lipopolysaccharide induced microglial inflammation model

AN Zi-xuan1,SHI Cai-yun1,LI Wei1,2

(1.Department of pharmacy, Hebei North University,Zhangjiakou 075000,China;2.Hebei Provincial Key Laboratory of neuropharmacology,Zhangjiakou 075000,China)

Abstract:OBJECTIVETo establish an in vitro inflammatory model of BV2 by observing the activity,the release amount of NO and the expression of inflammatory factors of microglial cells (BV2) induced by lipopolysaccharides(LPS).METHODSBV2 was routinely cultured in vitro.Cell viability was measured by CCK-8 meth⁃od.And by drew cell growth curve to determine the logarithmic growth cycle of the cells.After 24 h of routine culture,BV2 were induced by adding different concentrations of LPS(0.1,1.0 and 10.0 mg·L-1)for 4,8,12,24 and 48 h,respectively.Meanwhile,the morphological changes of BV2 were observed under inverted microscope to compare the activation degree of microglia at dif⁃ferent time and concentration.Cell activity and nitric oxide(NO)level were determined by CCK-8 and Griess method respectively,which could help to determine the optimal concentration and time of modeling.Finally,It were determined by ELISA that the concentrations of tumor necrosis factor α (TNF-α),interleukin-6(IL-6)and IL-1 β in supernatant of LPS 1 mg·L-1culture for 24 h.RESULTSBV2 were in logarithmic growth phase for 1 to 3 d after subculture.LPS 1 mg·L-1induced BV2 for 24 or 48 h which could increase the release amount of NO significantly(P<0.05).In order to save time,LPS induced BV2 for 24 h were selected for subsequent experiments.Microglial cells in resting state were observed to be elongated spindle shape under inverted micro⁃scope.After LPS activation,the cell body became larger and the branching processes shrank back,presenting an amoeba-like appearance.ELISA results showed that the concentrations of TNF-α,IL-6 and IL-1 β in supernatant of LPS 1 mg·L-1cultured for 24 h were significantly increased which compared with the control group(P<0.05).CONCLUSIONLPS could induce the activation of BV2 and up-regulate the level of inflammatory factors.The optimal condition for establishing stable BV2 microglial inflammatory model was used LPS 1 g·L-1induced for 24 h.

Key words:microglia; lipopolysaccharides;inflammation model

Foundation item:Natural science foundation of Hebei Province(H2020405298)

Corresponding author:LI Wei,E-mail:leewei 318@163.com

T3-22

Neuronal PPARα deficiency-induced axonal mitochondrial transport defects underly isch⁃emic stroke pathology

XU Lan-xi,ZHUO Ren-gong,JIN Xin,YANG Li-chao

(School of Medicine,Xiamen University,Xiamen 361005,China)

Abstract:OBJECTIVEPeroxisome prolifer⁃ator activated receptor alpha(PPAR α)is an important protective factor in neurovascular diseases such as ischemic stroke.Although PPAR α expression is higher in neurons than astrocytes and microglia,the pathophysiological functions of neuronal specific-PPARα in isch⁃emic stroke remains unknown.Here,we report that neuronal PPARα deficiency is a key factor of neuronal injury.PPARα expression markedly decreased in neurons after ischemic stroke.METHODSAND RESULTS Neuronal-specific PPARα knockout(NCKO)exacerbates neuronal damage and brain ischemic injury.PPARα defi⁃ciency disrupts axonal microtubule organization and mitochondrial transport by decreasing the expression of dynein light chain Tctex-type 1(Dynlt1),which is implicated in cytoprotective role with damaged neurons.Furthermore,resto⁃ration of Dynlt1 expression in neurons of NCKO mice rescue mitochondrial transport disorder,cognitive deficits and brain ischemic injury asso⁃ciated with PPARα deletion.CONCLUSIONThese results reveal a critical role for neuronal PPARα in ischemic brain injury by modulating axonal mitochondrial transportation.

Key words:peroxisome proliferator activated receptor alpha;mitochondrial dysfunction;axonal transport;neuronal injury;ischemic stroke

T3-23

血小板介导的神经炎症在中枢神经性疾病中的作用

孔德平，冯兆阳，亓会斌，谭 瑞，高永峰，于海洋，赵晓民

〔山东第一医科大学（山东省医学科学院）药理学研究所，山东 泰安 271016〕

摘要：血小板是无核细胞，血液中数量较多，表面有丰富受体，细胞内有多种颗粒，可释放活性物质，除具有止血、促进血栓形成作用之外，还在炎症和免疫方面发挥重要功能。由于炎症在严重的神经性疾病发生发展中占据重要位置，因而研究中枢神经性疾病需要考虑血小板的因素。在人或动物相关研究中，血小板介导的神经炎症在肌萎缩侧索硬化症、癫痫、创伤性脑损伤、帕金森病、亨廷顿病、舞蹈病、阿尔茨海默病和脑卒中等进展中产生影响，机制涉及血小板参与的血栓，与白细胞、中枢血管内皮细胞、免疫细胞和补体系统的相互作用，分子事件包括血小板表面的受体、释放的活性物质及相应信号途径，还可调节神经元的电活动、突触功能和可塑性。靶向调控血小板介导的神经炎症，可能成为一种辅助治疗中枢神经性疾病的有效手段。

关键词：血小板；神经；炎症；血栓形成

通讯作者：赵晓民，E-mail：xmzhao@sdfmu.edu.cn

T3-24

CZL-80,a novel caspase-1 inhibitor,pro⁃motes functional recovery after progressive ischemic stroke

PAN Ling,TANG Wei-dong,CHEN Zhong,ZHANG Xiang-nan

(Institute of Pharmacology & Toxicology,College of Pharmaceutical Sciences,Key Laboratory of Medical Neurobiology of the Ministry of Health of China,Zhejiang University,Hangzhou,310058,China)

Abstract:OBJECTIVEProgressive isch⁃emic stroke is characterized by aggravation of neurological dysfunction and poor prognosis.Neuroinflammation is involved in the pathological process of cerebral ischemia.Inflammasomesactivated caspase-1 has thus been considered a promising target for stroke therapy.However,it remains not fully understood how caspase-1 ag⁃gravates progressive functional impairment.We previously identified a novel caspase-1 inhibitor CZL-80,the present study is to explore whether CZL-80 protects against progressive ischemic stroke.METHODSMale C57/BL6 mice and cas⁃pase-1-/-mice were subjected to photothrombotic(PT)-induced cerebral ischemia.CZL-80 was in⁃traperitoneally injected daily during 1-7 d,1-4 d,4-7 d after PT.The grid-walking task and the cyl⁃inder task were used to determine the motor function.RESULTSMice developed primary and the secondary neurological dysfunction at 1 d and 4-7 d after PT onset.The activation of cas⁃pase-1 peaked at 7 d after ischemic stroke and caspase-1 was mainly derived from activated microglia.Treatment with CZL-80(30 mg·kg-1)during 1-7 d significantly improved motor func⁃tion.Administration of CZL-80 during 1-4 d could not ameliorate motor function loss while administration during 4-7 d after PT onset signifi⁃cantly reduced foot faults and forelimb symme⁃try.Remarkably,treatment with CZL-80 during 4-7 d showed no significant difference in efficacy compared with the its administration during 1-7 d,which indicated a key therapeutic window.More⁃over,the neuroprotective effect of CZL-80 during 4-7 d was available at least until 43 d after isch⁃emic stroke,indicating CZL-80 can improve the long-term neurological function after cerebral ischemia.Furthermore,administration of CZL-80(30 mg·kg-1)during 4-7 d after PT onset in cas⁃pase-1-/-mice failed to improve the motor func⁃tion,which suggested that the neuroprotective effect of CZL-80 was caspase-1-dependent.The results showed that CZL-80 did not inhibit the expression of GSDMD and failed to reduce neu⁃ronal loss after ischemia.These results indicated the effect of CZL-80 was not attributable to inhib⁃it pyroptosis.We further found that CZL-80 signif⁃icantly reduced the number of activated microglia in the peri-infarct brain cortex after ischemic stroke,which might be involved in its neuropro⁃tective effect.CONCLUSIONCZL-80,a novel caspase-1 inhibitor,improved motor function after progressive ischemic stroke in mice.The effective therapeutic window of CZL-80 would be 4-7 d after ischemia,when the secondary neuro⁃logical dysfunction occurred.Therefore,the inter⁃vention by targeting caspase-1 in this window phase provides a novel strategy for the function⁃al recovery of stroke survivors.

Key words:caspase-1;progressive ischemic stroke;functional recovery;microglia

Corresponding author:CHEN Zhong,E-mail:chenzhong@zju.edu.cn;ZHANG Xiang-nan,E-mail:xiangnan_zhang@zju.edu.cn

T3-25

Lysosomal dysfunction in Schwann cells is involved in bortezomib-induced peripheral neuropathy

WU Zhan-xun1,YAN Wen-ping2,CHEN Zhong1,WU Jia-ying1,ZHANG Xiang-nan1

(1.Institute of Pharmacology & Toxicology,Col⁃lege of Pharmaceutical Sciences,Key Laboratory of Medical Neurobiology of the Ministry of Health of China,Zhejiang University,Hangzhou 310058,China;2.Department of Pharmacy,the First Affili⁃ated Hospital,School of Medicine,Zhejiang Univer⁃sity,Hangzhou 310003,China)

Abstract:OBJECTIVEThe proteasome inhibitor bortezomib(BTZ)is a first-line anti-multi⁃ple myeloma drug.BTZ-induced peripheral neu⁃ropathy(BIPN)is a main adverse effect that char⁃acterized by neuropathic pain.There is still no strategy to prevent or treat BIPN,attributed to the unidentified mechanisms underlying BIPN.Previous studies suggested that BTZ impairs Schwann cells and thus leads to axonal demye⁃lination,whereas it remained not fully understood how BTZ cause Schwann cell death.It was observed that BTZ upregulates the autophagy marker LC3-Ⅱ protein in Schwann cells.However,it remains unclear whether BTZ causes autopha⁃gy-lysosome dysfunction in Schwann cells.METHODSThe male C57BL/6 mice were intra⁃venous injection of BTZ(1 mg·kg-1per day,twice weekly for a total of 4 weeks).The paw withdraw⁃al latency was tested by the Von Frey test and Hargreaves test to reflect the neuropathic pain.The conduction velocity and the action potential amplitude of the tail nerve were tested by neuro⁃physiological assessment to reflect peripheral nerve function.The histomorphology of the sciat⁃ic nerves was detected by immunofluorescence and transmission electron microscopy to reflect the demyelination and axonal degeneration.The RSC96 cells,the Schwann cell-like immortal cells,were cultured and exposed to BTZ.The lysosomal function was determined by Lyso⁃Tracker and DQ-BSA staining.Autophagy-relat⁃ed proteins,including p62 and LC3,and lysosom⁃al hydrolase cathepsin B were determined by Western blotting.RESULTS①BTZ induced mechano-allodynia, neurological conduction abnormalities of the tail nerve,demyelination and axonal degeneration of the sciatic nerves.②BTZ caused lysosomal dysfunction,resulting in the blockade of autophagy flux in Schwann cells and sciatic nerves.③The lysosomal activator Torin1 reversed lysosomal dysfunction caused by BTZ in Schwann cells.④ Torin1 improved BTZ-induced mechano-allodynia and demyelination of sciatic nerves.CONCLUSIONBTZ led to lyso⁃somal dysfunction in Schwann cells and contrib⁃uted to BIPN.Lysosomal activation could be a promising strategy for BIPN intervention.

Key words:bortezomib;peripheral neuropathy;sciatic nerves;demyelination;Schwann cells;lysosome.

Corresponding author:ZHANG Xiang-nan,E-mail:xiangnan_zhang@zju.edu.cn

T3-26

Loss of Hrh2 on dopaminergic neurons leads to mania-like behavior in mice

MA Shi-jia1,ZHANG Xing-xian1,LI Yue1,HU Wei-wei1,CHEN Zhong1,2,ZHANG Xiang-nan1

(1.Institute of Pharmacology & Toxicology,College of Pharmaceutical Sciences,Key Laboratory of Medical Neurobiology of the Ministry of Health of China,Zhejiang University,Hangzhou310058,China;2.Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province,College of Pharmaceutical Sciences,Zhejiang Chinese Medical University,Hangzhou 310058,China)

Abstract:OBJECTIVEDysfunction of the dopaminergic(DA)neurons is implicated in the pathogenesis of bipolar disorder(BPD).Hista⁃mine receptor 2(Hrh2)is highly expressed in DA neurons,and its antagonists have been reported to induce mania phase of BPD.However,whether Hrh2 on DA neurons contributes to BPD patho⁃genesis is unclear.The present study aims to explore the role of hrh2 on DA neurons in the pathology of BPD.METHODSAAV-FLEX-shHrh2 was injected into a targeted brain area of DAT-Cre mice,leading to a selective brainregional loss of Hrh2 on DA neurons.A series of behavior tests were used to measure the sponta⁃neous activity,anxiety and depression level of Hrh2-deficient mice.RESULTS①In the open field test and home-cage activity test,Hrh2-defi⁃cientmice displayed increased spontaneous activity.②Hrh2-deficient mice showed reduced depression level in the tail suspension test,forced swimming test and sucrose preference test.③The anxiety level of Hrh2-deficient mice was decreased in the open field test.CONCLU⁃SIONHrh2 on DA neurons is closely related with mania-like behavior.

Key words:bipolar disorder;mania;histamine receptor 2;dopaminergic neurons

Corresponding author:CHEN Zhong,E-mail:chenzhong@zju.edu.cn;ZHANG Xiang-nan,E-mail:xiangnan_zhang@zju.edu.cn

专题4：离子通道及受体信号与神经精神疾病

T4-1

Ibrutinib alleviated LPS-induced neuroinflam⁃mation and synaptic defects in mouse model of depression

LI Wei-fen,Tahir Ali,HE Kai-wu,LIU Zi-zhen,Fawad Ali Shah,REN Qing-guo,LIU Yan,JIANG An-long,LI Shu-peng

(Laboratory of Chemical Genomics,School of Chemical Biology and Biotechnology,Peking Uni⁃versity Shenzhen Graduate School,Shenzhen 518055,China)

Abstract:OBJECTIVEPrevious studies have demonstrated a close association between an altered immune system and major depressive disorders,and inhibition of neuroinflammation may represent an alternative mechanism to treat depression.Recently,the anti-inflammatory activ⁃ity of ibrutinib has been reported.However,the effect of ibrutinib on neuroinflammation-induced depression and its underlying mechanism has not been comprehensively studied.Therefore,we aimed to elucidate the potential anti-depres⁃sive role and mechanism of ibrutinib against neu⁃roinflammation-induced depression and synaptic defects.METHODS AND RESULTSIbrutinib treatment significantly reduced lipopolysaccha⁃ride(LPS)-induced depressive-like behaviors and neuroinflammation via inhibiting NF-κB acti⁃vation,decreasing proinflammatory cytokine lev⁃els,and normalizing redox signaling and its downstream components,including Nrf2,HO-1,and SOD2,as well as glial cell activation mark⁃ers,such as Iba-1 and GFAP.Further,ibrutinib treatment inhibited LPS-activated inflammasome activation by targeting NLRP3/P38/caspase-1 signaling.Interestingly,LPS reduced the number of dendritic spines and expression of BDNF,and synaptic-related markers, including PSD95,snap25,and synaptophysin,were improved by ibrutinib treatment in the hippocampal area of the mouse brain.CONCLUSIONIbrutinib can allevi⁃ate neuroinflammation and synaptic defects,sug⁃gesting it has antidepressant potential against LPS-induced neuroinflammation and depression.

Key words:ibrutinib;neuroinflammation;depres⁃sion;synaptic defects

Corresponding author:LI Shu-peng,E-mail：2001112065@puk.edu.cn

T4-2

Ginsenoside Rb1 produces antidepressantlike effects in chronic social defeat stress model of depression through BDNF-TrkB sig⁃naling pathway

JIANG Ning1,HUANG Hong1,ZHANG Yi-wen1,LYU Jing-wei1,WANG Qiong2,LIU Xin-min1

(1.Research Center for Pharmacology and Toxi⁃cology,Institute of Medicinal Plant Development,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100193,China;2.Affiliated TCM Hospital/School of Pharmacy/Sino-Portugal TCM International Cooperation Center,Southwest Medical University,Luzhou 646000,China)

Abstract:OBJECTIVEGinsenoside Rb1(Rb1),an important bioactive ingredient of Panax ginseng,has potent neuroprotective effects.The objective of the study is to elucidate the impact of Rb1 treatment on chronic social defeat stress(CSDS)-induced depressive-like behaviors and its related mechanism.METHODS AND RE⁃SULTSThe daily oral administration of Rb1(35 and 70 mg·kg-1)and imipramine(15 mg·kg-1)for 28 d significantly reversed the social avoidance behavior,anhedonia,and behavioral despair via CSDS exposure,as demonstrated by the consid⁃erable elevation in the time in the zone in social interaction test and consumption of sucrose solu⁃tion in sucrose preference test and decrease of immobility time in forced swim test.Moreover,Rb1 obviously restored the CSDS-induced decrease of BDNF-signaling pathway and hippo⁃campal neurogenesis.Rb1 significantly increased the hippocampal levels of ERK,AKT,and CREB phosphorylation and increased the number of DCX+cells in DG.Importantly,the antidepres⁃sant effects of Rb1 were completely blocked in mice by using K252a(the nonselective tyrosine kinase B inhibitor).CONCLUSIONRb1 exerts promising antidepressant-like effects in mice with CSDS-induced depression,and its effects was facilitated by enhancing the BDNF signaling cas⁃cade and up-regulation of hippocampal neuro⁃genesis.

Key words:ginsenoside Rb1;depression;chronic social defeat stress;neurogenesis

Foundation item:Ministry of Science and Tech⁃nology of China(2017ZX09301029);and Space Medical Experiment Project of China Manned Space Program(HYZHXM05003)

Corresponding author:LIU Xin-min,E-mail:liuxinmin@hotmail.com

T4-3

Mechanism of charged interface of stargazin EX1 on AMPAR opening and desensitization

YANG Zu-xiao,ZHANG Wei

(Department of Pharmacology of Chinese Materia Medica,Institution of Chinese Integrative Medicine,the Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University,Shijiazhuang 050017,China)

Abstract:OBJECTIVEAMPA-subtype iono⁃tropic glutamate receptors(iGluRs)mediate fast excitatory synaptic transmission in the mammali⁃an central nervous system(CNS).It plays the key role in many central nerves disorder such as epilepsy,depression and schizophrenia.Star⁃gazin(STZ,also named TARP-γ2),as the first TARPs found in CNS,potentiates AMPAR activity by attenuating deactivation and desensitization,enhancing recovery from desensitization,and facilitating agonist affinity and efficacy.However,it is still not fully understanding how γ -2 modu⁃late AMPAR gating.METHODS AND RESULTSThe desensitization for different mutation of AMPAR and γ-2 was compared.It was shown that the electric attraction was involved in the interaction of AMPAR and γ-2.In addition,the interaction of KGK motif in ligand binding domain and pre-M1 chain of AMPAR and EX1 of γ-2 modulate AMPAR opening and desensitization.Substitution of these charged residues had sur⁃prisingly effects on AMPAR desensitization kinet⁃ics.CONCLUSIONThe electric attraction has two impacts on the channels gating process the first destablizing the receptor closed state and enabling the channel opening,the second pro⁃moting the channels entering desensitization state upon the channel opening.

Key words:ionotropic glutamate receptors;desensitization;stargazin EX1

T4-4

Voltage-dependent potassium channel Kv4.2 promotes neurogenesis and alleviates isch⁃emic stroke impairments

ZHAO Yu-ming

(Department of Pharmacology,School of Basic Medical Science, Capital Medical University,Beijing 100069,China)

Abstract:OBJECTIVEStroke has become the top ten leading cause of death in China.Isch⁃emic stroke accounts for 85% of stroke cases,and insufficiency of cerebral blood supply caused by atherosclerosis is one of the important causes of ischemic stroke.Therefore,it is of posi⁃tive significance to study the molecular mecha⁃nism of stroke injury caused by hypoperfusion in the search for drug targets.Voltage-dependent potassium channels are a family of potassium channels widely expressed in the central ner⁃vous system.However,their roles in neurogene⁃sis after stroke insults have not been clearly illus⁃trated.The purpose of this experiment is to explore the expression changes of different sub⁃families of voltage-dependent potassium chan⁃nels after the occurrence of ischemic stroke and their influence on neuroregeneration,to study the molecular mechanism of stroke injury caused by hypoperfusion,and to find potential targets for drug therapy of ischemic stroke.METHODSC57BL/6 mice aged 7-8 weeks and C17.2 cells were used in vivo and in vitro in the experiment.The mice in the experimental group were suf⁃fered from bilateral common carotid artery occlu⁃sion(BCCO)for 1 h and reperfusion for 7 d.In the control group,bilateral carotid artery was dis⁃sected without occlusion.Behavioral assay of suspension test were performed to assess the motor deficits of the mice.In this assay,the time of the first drop(latency),the number of drops within one minute (frequency),and the final scores were recorded as the results of athletic ability.A lower score indicated more severe motor damage of the mice.TTC staining was used to observe the cerebral infarction areas caused by ligation of bilateral common carotid arteries.After seven days,mice were sacrificed and brain tissue protein samples were collected for real-time quantitative PCR(RT-PCR)and Western blotting test to detect the changes of potassium channel subfamily expression levels in different brain regions.Neuronal injuries in all brain regions were detected using Nissl staining methods 7 d following model establishment.To detect the effects and the underlying mechanism of the related potassium channel on neurogene⁃sis,recombinant plasmids of the potassium chan⁃nels were transfected in cultured C17.2 neural stem cells.Afterwards,oxygen glucose depriva⁃tion experiments were performed.RESULTSBehavioral tests showed that BCCO can cause impaired motor performance.TTC staining showed that cerebral infarction existed in the stri⁃atum region,and the motor function decline caused by the injury in this region was consistent with the behavioral experiment results which veri⁃fied the effectiveness of our surgical operation.Nissl staining revealed a large amount of neuronal cell necrosis in the cortex and striatum regions,and dense neuronal cells in the lateral ventricular limbic region,suggesting that neurogenesis may have occurred in this region.The results of realtime quantitative RT-PCR showed that among the detected potassium channels distributed in the measured nervous system,the expression of voltage-dependent potassium channel Kv4.2 decreased significantly in all brain regions after stroke,suggesting that it may be involved in the pathological process of stroke.Immunohisto⁃chemical staining showed that there was neuro⁃genesis in the subgranular zone(SGZ)and sub⁃ventricular zone(SVZ)of the mice,and Kv4.2 expression was significantly changed in the regions,suggesting that it may be involved in the regulation of neuro regeneration after stroke.The transfected Kv4.2 plasmid enhanced the dif⁃ferentiation of the C17.2 neural stem cells to neu⁃rons and astrocytes under normoxia and the oxy⁃gen-glucose deprivation,suggesting that Kv4.2 may induce the differentiation of neural stem cells after stroke.Kv4.2 could induce the neural stem cells to differentiate into neurons in vitro and in vivo,and Western blotting assay showed that Kv4.2 could up-regulate the expression level of ERK1/2,p-ERK1/2,p-STAT3,NGF,p-TtkA,and BDNF.Moreover,the calcium ions and CAMK Ⅱ was also increased by Kv4.2 in vitro.CONCLUSIONBCCO insults can induce the expressions of the potassium channels in the brains,among which the expression of Kv4.2 is down-regulated in the cerebral cortex,hippocam⁃pus and striatum.In vitro experiments confirmed that Kv4.2 can induce the differentiation of C17.2 neural stem cells into neurons and astrocytes under the condition of normoxia and oxygenglucose deprivation.We concluded that Kv4.2 possibly promoted neurogenesis through ERK1/2/STAT3,NGF/TrkA,and Ca2+/CAMK Ⅱ signal pathways after stroke.Regulating the physiologi⁃cal functions of Kv4.2 channel might contribute to the rehabilitation of neuronal damage after stroke.

Key words:voltage-dependent potassium channels;bilateral common carotid artery occlu⁃sion;neurogenesis

Corresponding author:ZHAO Yu-ming,E-mail:yumingzhao@ccmu.edu.cn

T4-5

山茱萸环烯醚萜苷对tau蛋白转基因小鼠精神症状的影响及其机制

马登磊，罗 艺，李 林，张 兰

（首都医科大学宣武医院药学部，神经变性病教育部重点实验室，北京市神经药物工程研究中心，北京 100053）

摘要：目的微管相关蛋白tau在阿尔茨海默病（AD）发病机制中发挥重要作用。过表达P301L突变tau蛋白的rTg4510转基因小鼠具有明显的tau病理表现，广泛应用于AD的基础研究及药理研究中。山茱萸环烯醚萜苷（CIG）为中药山茱萸的有效部位，可降低tau蛋白的磷酸化。本研究探讨CIG以tau蛋白为靶点治疗AD的潜在价值。方法应用rTg4510转基因小鼠为AD模型，给予CIG处理3个月后进行Y迷宫和自主活动行为学检测，之后分别灌注固定取材及新鲜取材。应用Western印迹法检测蛋白的表达水平，应用免疫组织化学等方法观察病理性tau蛋白表达。结果CIG可以显著降低rTg4510小鼠的过度活跃状态，降低小鼠脑内杏仁核、海马区tau蛋白病变，改善小鼠杏仁核区域的神经元丢失；同时增加小鼠脑内NMDA受体及突触功能蛋白的表达，减少白质胼胝体区域的髓鞘碱性蛋白的丢失。结论CIG可改善rTg4510转基因小鼠的精神行为学表现，改善神经元丢失和白质脱髓鞘病变，其主要机制为CIG降低tau蛋白的过度磷酸化及病理性tau蛋白在杏仁核区域的沉积，增加NMDA受体及突触功能蛋白的表达，提示CIG是一个潜在抗AD药物。

关键词：山茱萸环烯醚萜苷；阿尔茨海默病；tau蛋白

T4-6

啮齿类动物触屏操作实验方法研究进展

张亦文，黄 红，姚彩虹，姜 宁，刘新民

（中国医学科学院北京协和医学院药用植物研究所，北京 100193）

摘要：通过文献检索，探索啮齿类动物触屏操作实验方法在啮齿类动物神经精神药理学研究的应用现状。本文对国内外中英文共6个文献库的近30年文献进行检索筛选和数据提取，系统分析总结了实验中常用动物的种类、训练方法、训练模式和相关参数等，得到相关文献300余篇，近10年来相关文章数量增长较快。大、小鼠和多种转基因动物均适用此实验方法，常用的实验模式多达10余种，其中视觉辨别与逆转最为常用，对常用参数和实验方法等也进行了系统阐述。啮齿类动物触屏操作实验方法基于斯金纳条件反射原理，通过计算机呈现图案信号刺激，啮齿类动物用鼻子直接触碰屏幕对信号做出反应，从而检测动物操作任务下对不同模式的图案信号的兴趣、解决问题的能力和速度等，反应动物的注意力、工作记忆、长期记忆、空间记忆和决策等高级认知功能。齿类动物触屏操作实验在临床干预转化中有着较高的转化潜力，在神经精神类疾病以及操作任务下学习记忆的研究中有着巨大的应用前景。

关键词：触屏操作；神经精神药理；啮齿类动物

基金项目：国家自然科学基金（81773930）；空间站工程航天医学实验项目（HYZHXM05003）

通讯作者：刘新民，E-mail：liuxinmin@hotmail.com；姜 宁，E-mail：jiangning0603@163.com

T4-7

Histamine H2receptor deletion in glutamatergic neurons causes schizophrenia-like pheno⁃types

JIANG Lei,MA Qian-yi,CHEN Zhong,HU Wei-wei

(Department of Pharmacology,NHC and CAMS Key Laboratory of Medical Neurobiology,School of Basic Medical Sciences,College of Pharmaceuti⁃cal Sciences,Zhejiang University,Hangzhou 310058,China)

Abstract:OBJECTIVEGenetic variation in histamine H2receptor(H2R)and H2R ligands are linked to schizophrenia,however little is known about how H2R contributes to pathogenesis of schizophrenia.Here,we detected a decreased expression of H2R in medial prefrontal cortex(mPFC)glutamatergic neurons in schizophrenia patients.METHODS AND RESULTSSelective knockout of H2R gene(Hrh2)in glutamatergic neurons induced schizophrenia-like behaviors including sensorimotor gating deficit,increased locomotor activity,social withdrawal and anhedo⁃nia in behavior tests,as well as reduced sponta⁃neous firing of mPFC glutamatergic neurons in electrophysiological tests.Selective deletion of the Hrh2 in mPFC glutamatergic neurons but not hippocampus glutamatergic neurons also induced such schizophrenia-like behaviors.Patch-clamp electrophysiology establishes that H2R deficiency reduced the intrinsic excitability of glutamatergic neurons by up-regulation of hyperpolarization activated cyclic nucleotide-gated channels.Fur⁃thermore,either overexpression of H2R in gluta⁃matergic neurons or activation of H2R in the mPFC reversed the MK-801-induced schizophrenia-like symptoms.CONCLUSIONH2R can serve as a novel drug target given that functional deficiency of this receptor in mPFC glutamatergic neurons may be crucial for the pathogenesis of schizo⁃phrenia.H2R agonists can be viewed as drug candidates for the treatment of schizophrenia.

Key words:histamine H2receptor;glutamater⁃gic neurons;schizophrenia-like behaviors

Foundation item:National Natural Science Foundation of China(81973302)

Corresponding author:HU Wei-wei,E-mail：huww@zju.edu.cn

T4-8

铃蟾肽致小鼠瘙痒下丘脑响应及PKA/creb信号参与机制

张晶鑫1，2，周 琥1，石京山2，王永安1，王丽韫1

（1.北京毒物药物研究所，北京 100850；2.遵义医科大学基础药理国家重点实验室，贵州 遵义 563000）

摘要：目的研究铃蟾肽（BN）致瘙痒药理学作用特点，确定下丘脑参与其中枢调控瘙痒关键分子机制。方法观察KM小鼠经侧脑室注射不同剂量BN（0.04，0.40和4.00 μg）诱发的瘙痒效应；构建耦联荧光标签NBD（7-nitrobenz-2-oxa-1，3-diazole）的BN（NBD-BN），并通过检测荧光信号示踪NBDBN在脑内的分布；利用免疫组化和Western印迹法检测下丘脑BN受体神经介素B受体（NMBR）、胃泌素释放肽受体（GRPR）及受体信号PKA/CREB相关蛋白的表达，鉴定下丘脑内特异神经元c-FOS蛋白响应。结果KM小鼠经侧脑室注射BN（0.04，0.40和4.00 μg）后，30 min观察期内产生显著瘙痒抓挠行为，抓挠次数呈剂量依赖性增加。与对照组相比，BN 3个剂量组抓挠次数均显著增加（P＜0.05，P＜0.01）；荧光示踪结果显示，侧脑室给予BN后NBD-BN在30 min内在下丘脑特异性分布，且10 min时荧光强度达到峰值。免疫组化和Western印迹结果显示，BN 2种受体亚型GRPR和NMBR在下丘脑特异性分布；侧脑室给予BN 4.00 μg后，与对照组相比，下丘脑PKA，p-CREB和c-FOS蛋白均显著增加（P＜0.01），且c-FOS蛋白在下丘脑背内侧核密集分布。结论BN具有强的中枢致瘙痒作用，侧脑室注射BN可致小鼠瘙痒并激活下丘脑BN受体依赖的PKA/CREB信号通路，诱导下丘脑神经元响应，表明下丘脑参与调控BN中枢致瘙痒作用。

关键词：铃蟾肽；痒；下丘脑背内侧核；胃泌素释放肽受体；NMBR神经介素B受体

T4-9

M4 muscarinic receptors regulates dopamine/DARPP-32 signaling and glutamate transmis⁃sion to balance dopaminergic D1function in mouse dorsal striatum

ZHOU Hu,ZHANG Jing-xin,LI Xing,SHI Hua-xiang,SUI Xin,WANG Yong-an,LI Jin,WANG Li-yun

(Beijing Institute of Pharmacology and Toxicology,Beijing 100850,China)

Abstract:OBJECTIVEAbnormal striatal dopaminergic and glutamatergic neurotransmis⁃sion is central to the pathophysiology of schizo⁃phrenia.In this study,we investigated the roles of M4receptor interplay with D1signaling in stria⁃tal neurotransmission that affect glutamatergic transmission to control the etiology of neuropsy⁃chiatric disorders.METHODSTo study dorsal striatum(DS)region-specific neuronal and behav⁃ioral responses modulated by M4receptors,we used clustered regularly interspaced short palin⁃dromic repeats-associated protein 9 technology to generate mice lacking M4in the dorsal stria⁃tum(DS-M4-KD).The M4positive allosteric modu⁃lator,VU0467154,were used to study the phar⁃macologically profiles with M4receptor stimula⁃tion in WT mice.Oxotremorine M(Oxo-M),a no subtype-selective muscarinic agonist,was used to show that mAchRs activation,in order to dissect the particular function of M4,in DS-M4-KD mice.Open filed test and forced swim test were used to assess the change of psychiatric-like behav⁃iors.Western blotting and immunohistochemistry were used to detect protein levels of phosphory⁃lation site of dopamine-and cAMP-regulated phosphoprotein of 32 ku(DARPP-32).Wholecell patch-clamp recording was used to assess M4-mediated cholinergic inhibition of glutamater⁃gic synaptic input transmission.RESULTSWest⁃ern blotting and immunohistochemistry assay showed VU0467154(5 mg·kg-1,ip)promoted phosphorylation of DARPP-32 at Thr75,and atten⁃uated D1-dependent phosphorylation of DARPP-32 at Thr34 within the mouse DS.Consistently,the Oxo-M(4 μg,icv)also increased DARPP-32 phosphorylation at site Thr75 to reversed phos⁃phorylation at site Thr34 in WT mice,but not in DS-M4-KD mice.In parallel with altered DARPP-32 responses,VU0467154 or Oxo-M evoked a psychological stress response and reversed D1-induced hyperlocomotion in mice in open field test and force swim tests.However,Oxo-M sup⁃pression of D1-depengdeng behavioral respons⁃es was impaired in DS-M4-KD mice.Whole-cell patch recording showed that VU0467154 or Oxo-M mediated endogenous cholinergic inhibition of miniature excitatory postsynaptic currents through M4receptors,which in turn suppressed D1-depen⁃dent glutamatergic synaptic transmission in the DS.CONCLUSIONThis study provides evidence for the role of M4receptors in regulation of dopa⁃mine/DARPP-32 signaling and glutamate respons⁃es in the DS,and therefore modulation of psychi⁃atric behaviors associated with D1signaling.This results indicate the mechanisms of treatments targeting M4in psychiatric disorders.

Key words:dorsal striatum;dopamine receptor 1;muscarinic acetylcholine M4receptor;dopamineand cAMP-regulated phosphoprotein of 32 ku

Corresponding author:WANG Li-yun,E-mail:lylywang1103@163.com

T4-10

Takeda G protein-coupled receptor 5 modu⁃lates depression-like behaviors via hippocam⁃pal CA3 pyramidal neurons afferent to dorso⁃lateral septum

WANG Hao,TAN Yuan-zhi,MU Rong-hao,TANG Su-su,LIU Xiao,XING Shu-yun,LONG Yan,YUAN Dan-hua,HONG Hao

(Department of Pharmacology,Key Laboratory of Neuropsychiatric Diseases,China Pharmaceutical University,Nanjing 211198,China)

Abstract:OBJECTIVETakeda G proteincoupled receptor 5(TGR5)is recognized as a promising target for type 2 diabetes and metabolic syndrome;its expression has been demonstrat⁃ed in the brain and is thought to be neuroprotec⁃tive.Here,we hypothesize that dysfunction of central TGR5 may contribute to the pathogene⁃sis of depression.METHODSIn well-established chronic social defeat stress(CSDS)and chronic restraint stress(CRS)models of depression,we investigated the functional roles of TGR5 in CA3 pyramidal neurons(PyNs)and underlying mech⁃anisms of the neuronal circuit in depression(for in vivo studies,n=10;for in vitro studies,n=5-10)using fiber photometry;optogenetic,chemoge⁃netic,pharmacological,and molecular profiling techniques;and behavioral tests.RESULTSBoth CSDS and CRS most significantly reduced TGR5 expression of hippocampal CA3 PyNs.Genetic overexpression of TGR5 in CA3 PyNs or intra-CA3 infusion of INT-777,a specific agonist,protected against CSDS and CRS,exerting sig⁃nificant antidepressant-like effects that were mediated via CA3 PyN activation.Conversely,genetic knockout or TGR5 knockdown in CA3 facilitated stress-induced depression-like behav⁃iors.Re-expression of TGR5 in CA3 PyNs rather than infusion of INT-777 significantly improved depression-like behaviors in Tgr5 knockout mice exposed to CSDS or CRS.Silencing and stimula⁃tion of CA3 PyNs→somatostatin-GABAergic(gamma-aminobutyric acidergic)neurons of the dorsolateral septum circuit bidirectionally regulat⁃ed depression-like behaviors,and blockade of this circuit abrogated the antidepressant-like effects from TGR5 activation of CA3 PyNs.CON⁃CLUSIONTGR5 can regulate depression via CA3 PyNs→somatostatin-GABAergic neurons of dorsolateral septum transmission,suggesting that TGR5 could be a novel target for developing antidepressants.

Key words:depression;dorsolateral septum;GABAergic neuron;hippocampus;pyramidal neuron;takeda G protein-coupled receptor 5

Corresponding author:HONG Hao,E-mail:honghao@cpu.edu.cn

T4-11

神经营养因子异常与抑郁症发病机制研究进展

王惠芹1，2，陈乃宏1，2

（1.湖南中医药大学药学院，湖南 长沙 410208；2.中国医学科学院北京协和医学院药物研究所，北京 100050）

摘要：抑郁症已成为“二十一世纪的流行病”，对我国乃至国际公民和社会的危害日益凸显。神经营养因子是神经可塑性的关键介质，抑郁症的发病涉及到神经营养因子及其基因异常。抗抑郁药物治疗后神经营养因子分泌增加，从而促进神经元的存活，保护神经元免受应激损伤。脑源性神经营养因子、血管内皮生长因子、胶质细胞源性神经营养因子和神经生长因子等与抑郁症关系的研究表明神经营养因子有望成为抗抑郁治疗的有效靶点。因此，研究与神经营养因子有关的抑郁症发病机制，对寻找快速有效的抗抑郁治疗方法至关重要。

关键词：抑郁症；神经营养因子；发病机制

T4-12

白芷的化学成分及对神经系统疾病的治疗作用

庞心悦1，景永帅2，郑玉光1，吴兰芳1

（1.河北中医学院药学院，河北 石家庄 050200；2.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：白芷为伞形科植物白芷或杭白芷的干燥根，始载于东汉《神农本草经》，味辛，性温，具有解表散寒、宣通鼻窍、消肿排脓等功效。白芷含挥发油、香豆素类、多糖类、黄酮类、生物碱类等成分。研究表明白芷及其活性成分对神经系统疾病有一定的缓解或治疗作用。①抗惊厥：白芷总香豆素类成分具有抗惊厥作用，可明显延长戊四氮和3-巯基丙酸所致模型小鼠的惊厥潜伏期，缩短惊厥持续时间，从而降低小鼠的死亡率；白芷的二氯甲烷萃取物在中、大剂量条件下也具有抗惊厥的作用。②缓解偏头痛：白芷能改善偏头痛动物行为学表现，减少NO分泌，增加5-羟色胺(5-HT)含量，降低降钙素基因相关肽的阳性率；另外，白芷的香豆素和挥发油具有明显的镇痛作用，其机制可能是减少模型动物脑内和血中NO的合成。③止疼作用：白芷总挥发油、水煎液对疼痛模型大鼠有明显的镇痛作用，其中水合氧化前胡素、佛手柑内酯可能是白芷镇痛的效应物质基础；另外，白芷挥发油可促进β-内啡肽的前体物质前阿黑皮素信使核糖核酸的表达、调整体内单胺类神经递质含量而发挥镇痛作用。④抗抑郁：白芷冰片方在多个动物模型上具有显著的抗抑郁作用，其机制与增强中枢5-HT能神经系统功能，上调海马BDNF/TrkB/p-CREB神经营养通路有关。本文对白芷在神经系统相关疾病中的治疗作用进行归纳总结，进一步明确白芷发挥药效的物质基础，可为白芷治疗神经系统相关疾病的临床应用及新药开发提供依据。

关键词：白芷；活性成分；神经系统疾病

基金项目：河北省省级科技计划（H2021423057）

通讯作者：吴兰芳，E-mail：wulanfang757@163.com

T4-13

Activation of MRGPRB2 receptor in mouse peritoneal mast cells depends on PLC-IP3-ORAI1 pathway

YE Fan, JIANG Yu-cui, ZONG Yin-xin,TANG Zong-xiang

(Nanjing University of Chinese Medicine,Nanjing,210023,China)

Abstract:OBJECTIVEA novel mast cellspecific G-protein-coupled receptor (GPCR),known as mas-related GPCR-B2(MRGPRB2),plays important roles in the immune response.The opening of ion channels mediated by MRGPRB2 activation remains unclear.METHODS AND RE⁃SULTSCalcium influx induced by activation of MRGPRB2 receptor in mouse peritoneal mast cells was related to the concentrations of calcium ions in the extracellular solution.Similarly,the volt⁃age-dependent current generated by MRGPRB2 activation was also correlated with the extracellu⁃lar calcium concentration.In addition,the in⁃creased of calcium influx or voltage-dependent current caused by activation of MrgprB2 could be blocked by U73122(PLC blocker)or 2-APB(IP3-ORAI1blocker).Meanwhile,calcium-activated chlorine channel(TMEM16A)was involved in the generation of voltage-dependent currents in⁃duced by MRGPRB2 activation in mouse perito⁃neal mast cells.Furthermore,the degranulation of mouse peritoneal mast cells mediated by MRGPRB2 receptor could also be inhibited by U73122or2-APB.CONCLUSIONPLC-IP3-ORAI1-TMEM16A signaling pathway was involved in MRGPRB2-mediated mast cell activation.

Key words:mast cells;mas-related GPCR-B2;ORAI1;calcium-activated chlorine channel

T4-14

Tenuifolin attenuates schizophrenia-like be⁃haviors induced by MK-801 via enhancing GABA function in prefrontal cortex

CUI Su-ying,CAO Qing,ZHANG Yong-he

(Department of pharmacology,Peking University,School of Basic Medical Science,Beijing 100191,China)

Abstract:OBJECTIVETenuifolin is a natu⁃ral neuroprotective compound isolated from Polygala tenuifolia Willd,which have a long history of being used in mental illness therapy in China.Acute or chronic administration of NMDA recep⁃tor blocker MK-801 is a validated animal model for the positive,negative and cognitive symp⁃toms of schizophrenia.This study aimed to inves⁃tigate the antipsychotic effects and possible mechanism of tenuifolin in MK-801-treated mice.METHODSSensorimotor gating deficit and hyper⁃locomotion,which implying positive symptoms,were assessed by prepulse inhibition(PPI)test and open field test.Social withdrawal,a main character of negative symptoms,was assessed by social interaction tests.Cognitive deficit was assessed by novel object recognition(NOR)test.The level of glutamate decarboxylase(GAD),GABA and monoamines were detected by West⁃ern blotting or HPLC-ECD.RESULTSAcute administration of MK-801(0.1 mg·kg-1,sc)could significantly induce hyperlocomotion and prepulse inhibition(PPI)deficit in mice,which could be reversed by the treatment of tenuifolin(40 and 80 mg·kg-1,ig).Moreover,tenuifolin ameliorated social withdrawal and cognitive deficit induced by the chronic administration of MK-801(0.3 mg·kg-1per day,sc,14 d followed by 7 d washout period).In addition,tenuifolin reversed the reduction in GAD and GABA induced by MK-801 and normal⁃ized dopamine,serotonin and norepinephrine in the prefrontal cortex.CONCLUSIONThese results provided persuasive evidence that tenui⁃folin has multiple antipsychotic-like effects in ani⁃mal model of schizophrenia and its underlying mechanism may be associated with its potential effects on GABA function in the prefrontal cortex.

Key words:tenuifolin;schizophrenia;GABA function

Corresponding author:ZHANG Yong-he,E-mail:zhyh@hsc.pku.edu.cn

T4-15

Bidirectional regulation of intravenous anes⁃thetic etomidate on TREK-1 potassium channel

WANG Wei-ping,SHAN Jin-feng,WANG Ling,WANG Xiao-liang

(State Key laboratory of Bioactive Substances and Functions of Natural Medicines,Department of Pharmacology, Institute of materia Medica,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100050,China)

Abstract:OBJECTIVETwo-pore domain potassium channel subtype TREK-1 was widely proved to be activated by inhalational anesthet⁃ics such as chloroform,diethyl ether,halothane,and isoflurane.But little is known about whether TREK-1 was also a potentially important target of intravenous anesthetics.Etomidate is a popularly used intravenous anesthetic with good safety in clinic.The action of etomidate on TREK-1 was seldom reported.METHODS AND RESULTSBy using patch-clamp whole-cell recording tech⁃niques,we found for the first time that etomidate could bidirectionally regulate the TREK-1 potassi⁃um channel in CHO/TREK-1 cells.TREK-1 current amplitudes were observed after the administra⁃tion of etomidate at concentrations ranging from 3 to 100 μmol· L-1.Etomidate activated TREK-1 current at concentrations of 3,10,and 15 μmol·L-1with maximum activation at 10 μmol·L-1.Interest⁃ingly,at higher concentrations from 20 to 100 μmol·L-1,etomidate inhibited TREK-1 current in a concentration-dependent way.According to the concentration-response curve,the fitted criti⁃cal concentration of etomidate between TREK-1 activation and inhibition was 20.7 μmol·L-1,which close to the result that etomidate had no obvious effect on TREK-1 at 20 μmol· L-1.In addition,etomidate 10 μmol·L-1induced a significant mem⁃brane potential hyperpolarization while etomidate 30 μmol·L-1showed obvious membrane potential depolarization.Furthermore,the bidirectional regulation still existed when the extracellular pH of CHO/TREK-1 cells was decreased.CONCLUSIONTREK-1 is activated by etomi⁃date at clinically relevant concentrations but inhib⁃ited by supraclinical concentrations of etomidate,which is different to other volatile anesthetics.TREK-1 might be a potential target for anesthetic such as etomidate and the complicated bidirec⁃tional regulation mechanism of etomidate needed to be fully studied in the future.

Keywords:TREK-1;etomidate;intravenous anesthetic;bidirectional regulation

Corresponding author:WANG Xiao-liang,E-mail:wangxl@imm.ac.cn

T4-16

Flumazenil-insensitive benzodiazepine effects in vitro and in vivo

WANG Na,LIAN Jing-jing,CAO Yan-qing,YU Gang,SU Rui-bin

(State Key Laboratory of Toxicology and Medical Countermeasures,Beijing Key Laboratory of Neu⁃ropsychopharmacology,Beijing Institute of Phar⁃macology and Toxicology,Beijing 100850,China)

Abstract:OBJECTIVETo identify benzodi⁃azepine(BZD)effects that are insensitive to the classical BZD binding site antagonist,flumazenil.Whether the flumazenil-insensitive BZD effects have selectivity on different GABAAreceptor sub⁃types was also investigated.METHODSThe high-concentration effects of BZDs and their sen⁃sitivity to flumazenil were determined on recombi⁃nant synaptic(α1β2γ2, α2β2γ2, α5β2γ2)and extrasynaptic(α4β2δ)GABAAreceptors using the voltageclamp electrophysiology technique.The in vivo evaluation of flumazenil-insensitive BZD effects was conducted in mice loss of right reflex(LORR)test.RESULTSDiazepam induced a biphasic potentiation for the α1β2γ2, α2β2γ2and α5β2γ2receptor channels,but did not affect the α4β2δ receptor.In contrast to the nanomolar com⁃ponent of potentiation,the second potentiation elicited by micromolar diazepam was insensitive to flumazenil.Midazolam,clonazepam,and loraz⁃epam at 200 μmol·L-1exhibited similar flumaze⁃nil-insensitive effects on α1β2γ2, α2β2γ2and α5β2γ2receptors,whereas the potentiation induced by 200 μmol· L-1zolpidem or triazolam was abol⁃ished by flumazenil.Consistent with the in vitro results,flumazenil antagonized the zolpidem(50 mg·kg-1)-induced LORR,but not those induced by 50 mg·kg-1diazepam or 100 mg·kg-1midazolam.CONCLUSIONThe existence of non-classical BZD binding sites on certain GABAAreceptor subtypes and the flumazenil-insensitive effects depend on the chemical structures of the allosteric modulators.

Key words:GABAAreceptor;benzodiazepine;non-classical binding sites;flumazenil

Foundation item:Institutional funding from Beijing Institute of Pharmacology and Toxicology

Corresponding author:YU Gang,E-mail:yg1st@163.com;Su Rui-Bin,E-mail:ruibinsu@126.com

T4-17

慢性束缚应激对小鼠疲劳发生的动态影响及机制

王 智，闫明珠，夏天吉，靳苏维，刘永广，常 琪

（中国医学科学院&北京协和医学院药用植物研究所，北京 100094）

摘要：目的当今，更多人群因常常需要在狭小环境中久坐工作，长期作息不规律且缺乏活动而导致疲劳。本研究希望通过慢性束缚应激（CRS）模拟以上人群疲劳的发生情况，并对CRS小鼠疲劳模型的动态发生及机制进行探索。方法将48只4周龄雄性ICR小鼠按体重和前肢抓力随机分成4组，分别束缚0，5，10和15 d，8 h·d-1。束缚结束后进行空场实验、抓力实验和负重游泳实验。次日各组小鼠强迫游泳30 min后休息30 min，戊巴比妥钠麻醉后从腹主动脉取血，脱颈处死取肝和后肢腓肠肌组织用于生化指标检测。结果与空白组相比，CRS小鼠体重和腓肠肌明显减轻；自主活动路程与时间均有减少，且15 d组均变化显著；抓力均显著降低；束缚10和15 d组负重游泳力竭时间显著缩短。CRS小鼠的血糖和肝糖原水平均显著降低，肌糖原水平均显著升高，血清乳酸堆积量、乳酸脱氢酶和谷草转氨酶活性显著升高，这些指标的变化与束缚时长成正相时效关系。腓肠肌组织的蛋白印迹测定显示，CRS小鼠LC3，p-AMPK/AMPK，PGC-1α，TFAM和OXPHOS表达降低，Parkin，p-AKT/AKT，p-mTOR/mTOR，p-ULK1(Ser757)和P62表达升高，表明线粒体发生和自噬受阻。结论在CRS下，小鼠疲劳逐渐发生，10 d（8 h·d-1）即可诱导疲劳，15 d可造成小鼠稳定疲劳。CRS致小鼠腓肠肌线粒体发生和功能受损，线粒体保护性自噬受阻，进而无法维持线粒体功能平衡，并且这与AMPK信号通路受抑制相关。

关键词：慢性束缚；疲劳；线粒体功能障碍；线粒体自噬；AMPK

基金资助：科技部国家重点研发计划“食品安全关键技术研发”专项（2018YFC1602105）

T4-18

Effect and regulation of α-dstroglycan glyco⁃sylation on chronic social defeat induced depressive-like behaviors of mice

LI Yu-ke,WANG Fang

(Department of pharmacology,School of basic medicine,Tongji Medical College,Huazhong Uni⁃versity of Science and Technology,Wuhan 430030,China)

Abstract:OBJECTIVEα-Dstroglycan(α-DG)is a predominant component in the dystrophinglycoprotein complex(DGC)and a recently char⁃acterized receptor for several extracellular matrix components with high affinity.Recent research⁃es have reported that hypoglycosylation of α-DG is associated with the pathophysiology of diseas⁃es,especially muscular dystrophy,but little is known about major depressive disorder(MDD).Like-acetylglucosaminyl transferase(Large)is a key enzyme for glycosylation of α-DG,which mainly modifies two points in the middle domain of α-DG:Thr-317 and Thr-319.Glycosylated α-DG(GLY-α-DG)can bind with high affinity to extracellular matrix(ECM)molecules that con⁃tain laminin globular(LG)domains,including per⁃lecan,agrin and neurexin.Agrin is mainly derived from neurons rather than glial cells.In cultured hippocampal neurons,it was found that agrin could regulate the homeostatic plasticity of inhibi⁃tory neurons by acting on GLY-α-DG.Mdx mice are transgenetic models for the investigation of Duchenne muscular dystrophy. Many studies have shown that the expression of GLY-α-DG in the peripheral and brain tissues of Mdx mice is significantly down-regulated.Mdx mice show cognitive impairment and high levels of anxiety.In this study,we employed chronic social defeat stress(CSDS)to establish an animal model of depression and detected the expression of GLY-α-DG among the brain areas associated with the pathophysiology of depression.METHODSSo⁃cial interaction test(SIT)and sucrose preference test(SPT)were used to evaluate depressive-like behavior.Open field(OF)and elevated plus maze(EPM)test were used to determine the anxiety-like behavior of Mdx mice.Novelty-sup⁃pressed feeding test(NSFT)forced swim test(FST)and tail suspension test(TST)were used to detect the depressive-like behavior of Mdx mice.Novel object recognition test(NOR)was applied to evaluate the cognition of Mdx mice.Subthreshold social defeat stress was used to explore the susceptibility to stress in Mdx mice.Stereotactic infusion of agrin into the ventral hippocampus(vHip),FST and TST were used to investigate the antidepressant effects of agrin.Adeno-associated virus(AAV)-mediated overex⁃pression techniques,behavior tests and wholecell path-clamp technique were conducted to determine the impact of Large overexpression on CSDS susceptible mice.RESULTSThe expres⁃sion of α -DG and GLY-α-DG were significantly decreased in the vHip of CSDS susceptible mice.Mdx mice showed decreased expression of GLY-α-DG and increased anxiety-like behav⁃iors.Mdx mice displayed some depressive-like behaviors,and the susceptibility to stress was significantly increased.Downregulation of the expression α-DG in the vHip by lentivirus increased the susceptibility to stress.Administra⁃tion of agrin to CSDS susceptible mice exerted antidepressant effects,and this effect could par⁃tially sustain for a week.The expression of Large was decreased in vHip.Overexpression of Large through AAV-Large reversed the depressive-like behaviors and restored the decreased frequency and amplitude of mIPSC.CONCLUSIONGLY-α-DG and its glycosylase are significantly decreased in CSDS susceptible mice.Adminis⁃tration of agrin and overexpression of Large displays antidepressant effect,which may be related to its promotion of inhibitory synaptic transmission.

Key words:α-dstroglycan;depressive-like behav⁃iors;social defeat

T4-19

海马CD39(ENTPD1)对小鼠抑郁样行为的影响及机制

董婉婷，胡壮丽，陈建国

（华中科技大学同济医学院基础医学院药理学系，湖北 武汉 430030）

摘要：目的ATP是一种快速兴奋性神经调质，在中枢神经系统富集。最新的研究表明，细胞外间隙低水平的ATP可能促进重度抑郁症（MDD）形成。细胞外ATP的浓度受其水解酶——外核苷酸三（二）磷酸水解酶（ENTPD）的调节，但有关ENTPD在抑郁症中的作用尚不清楚。因此，本文探讨CD39（称为ENTPD1）在慢性社会挫败应激（CSDS）诱导的小鼠抑郁样模型中的作用。方法和结果 CSDS增强海马组织中CD39的表达和活性。进一步给予CD39功能类似物三磷酸腺苷双磷酸酶，也可诱导小鼠抑郁样行为。有趣的是，再次补充ATP可逆转上述抑郁样行为。最后，通过药理学抑制和基因沉默的方法增加细胞外ATP浓度，亦可发挥抗抑郁作用，且同时增加海马神经发生和树突棘数量。结论海马CD39通过水解细胞外ATP参与诱导抑郁样行为，提示CD39可作为抑郁症的一个有前景的靶点。

关键词：海马；抑郁；CD39

专题5：新技术与新方法在神经精神药理学研究中的应用

T5-1

基于苯乙醇苷类天然分子探针的抗脑缺血新靶点发现

曾克武

（北京大学药学院天然药物及仿生药物国家重点实验室，北京 100191）

摘要：目的缺血性脑卒中是威胁人类生命健康的重大疾病，目前严重缺乏有效的药物靶点。肉苁蓉是我国著名的补益中药，被称为“沙漠人参”。研究发现，肉苁蓉具有良好的脑保护作用，其中苯乙醇苷是其主要药效成分。我们设想：是否可以利用代表性的苯乙醇苷类成分——松果菊苷（ECH）作为活性分子探针，从神经细胞中“钩钓”其直接作用靶点，并揭示其发挥神经保护作用的分子药理机制。方法以ECH作为分子探针，利用键合有ECH的固相载体从神经细胞裂解液中捕获其直接作用靶点并进行高分辨质谱鉴定，同时结合化学生物学及分子药理学方法探究其调控神经细胞缺血性损伤的分子信号通路。结果揭示了ECH的作用靶点为酪氨酸激酶α′催化亚基（CK2α′），且CK2α′通过激活Wnt/β-catenin信号通路促进线粒体融合，发挥抗神经细胞缺血性损伤的作用。此外，在CK2α′WT和CK2α′+/-小鼠MCAO模型上也证实了ECH通过靶点CK2α′发挥改善脑缺血的作用。结论揭示了ECH的直接作用靶点为CK2α′，同时也提示CK2α′是一个基于天然活性分子探针发现的新颖的脑缺血治疗靶点，为今后缺血性脑卒中先导药物的发现提供了新思路。

关键词：脑缺血；治疗靶点；苯乙醇苷类分子；活性分子探针；靶点钩钓

通讯作者：曾克武，E-mail：ZKW@bjmu.edu.cn

T5-2

中脑导水管周围灰质腹外侧部星形胶质细胞对糖尿病神经病理性疼痛及痛相关负性情绪的调控作用

杨 澜1，陈 理1，2，俞昌喜1，2

（1.福建医科大学药学院药理学系神经药理课题组，福建 福州 350122；2.福建省天然药物药理学重点实验室，福建 福州 350122）

摘要：目的探索中脑导水管周围灰质腹外侧部（vlPAG）星形胶质细胞活化在糖尿病神经病理性疼痛（DNP）及痛相关负性情绪发生、发展及维持过程中的具体作用及特征。方法使用链脲佐菌素（STZ）诱导SD大鼠Ⅰ型糖尿病，复制DNP模型；免疫荧光法观察、Sholl分析评价vlPAG星形胶质细胞在DNP进展过程中的形态和数量变化；采用化学遗传学技术定点定量向vlPAG注射携有星形胶质细胞特异性启动子gfaABC1D及修饰后的人毒蕈碱受体（hM3Dq或hM4Di）基因片段的腺相关病毒；待病毒稳定转染表达后，利用受体的外源性配体氯氮平-N-氧化物（CNO）靶向调控vlPAG星形胶质细胞活性；以机械性异常性疼痛阈值为观察指标，评价星形胶质细胞活化在DNP中的作用；同时，采用旷场试验、高架十字迷宫试验、条件性位置偏爱试验，评价vlPAG星形胶质细胞活化在DNP相关焦虑样、厌恶样情绪中的作用。结果STZ成功诱导高血糖症状后，SD大鼠机械痛阈进行性下降，提示DNP模型复制成功。随着DNP症状加剧，vlPAG星形胶质细胞数量逐渐增多、胞体面积逐渐增大、形态复杂性逐渐增强。化学遗传学手段激活正常大鼠vlPAG星形胶质细胞可诱导神经病理性疼痛样症状（机械痛阈下降）；化学遗传学手段抑制DNP模型大鼠vlPAG星形胶质细胞可改善DNP症状（机械痛阈升高）。同时，化学遗传学手段抑制vlPAG星形胶质细胞可减少DNP模型大鼠痛相关焦虑样行为，改善模型动物痛相关厌恶情绪。结论vlPAG星形胶质细胞活化与DNP的发生、发展及维持密切相关，vlPAG星形胶质细胞可能参与了DNP及痛相关负性情绪的编码和调控。

关键词：糖尿病神经病理性疼痛；中脑导水管灰质腹外侧部；星形胶质细胞；化学遗传学；痛相关负性情绪

基金项目：国家自然科学基金（81973309）

通讯作者：俞昌喜，E-mail：changxiyu@mail.fjmu.edu.cn

T5-3

Bioinformatics and system biology approach to identify influences of BDNF on depression model

WEI Song-ren

(Department of Neuropharmacology and Drug Dis⁃covery,School of Pharmaceutical Sciences,South⁃ern Medical University,Guangzhou 510515,China)

Abstract:OBJECTIVEThe classic animal model of depression is CUMS,and one of the hy⁃potheses is related to the decrease of brain-de⁃rived neurotrophic factor(BNDF)expression in the brain,and its corresponding animal model is BDNF knockdown mice.BDNF can promote neu⁃ronal differentiation,maintain neuronal growth,and repair neuronal damage.The reduction of BDNF is related to the severity of depression.In contrast,the deletion of IDO-1 could promote the maintain the homeostasis of neurotransmitters in the brain.Therefore,the construction of BDNF and IDO-1 models for sequencing comparison can reveal the neurotransmitter hypothesis of de⁃pression from the transcriptome level.METH⁃ODSIn order to obtain its transcriptome expres⁃sion profile,firstly construct BDNF knockdown,IDO-1 knockout and CUMS model.The differen⁃tial expression of mRNAs was done by sequenc⁃ing provider BGI,conducting R for analyze.Clus⁃terproflier was used to enrich GO and KEGG pathway annotations.The ceRNA package was used search database LncRNA2Target to dig potential lncRNAs.Finally,the Stringdb was used to construct the PPI network.RESULTSWe identified a large number of mRNAs in mice normal and depression-like tissues from diverse genomic locations,and these mRNAs were col⁃lected from medial prefrontal cortex(mPFC)manner.Based on the analysis of the enrichment pathway and ceRNA network,we found numer⁃ous abundant mRNAs are specifically expressed in gene editing group,and differentially expressed in depression-like compared with normal tissues or depression-like antagonism group.Indicates that they serve a specific function in specific pathways.We focused on the mPFC sequencing of gene modification mice especially in BDNF+/-and IDO-/-.There is a relatively large difference in expression matrix.Certain mRNA,like Ptbp1,are predominantly expressed in comparison of BDNF+/-and other groups,and present at sub⁃stantial levels,that suggest these mRNAs are purposefully produced.Furtherly analysis of cor⁃relation map,Ptbp1 are related to the protection of vulnerable neuronal circuits and pathways are clustered in nervous system development and synapse organization.CONCLUSIONWe described the potential enrichment DEGs and its pathway in three model mice related to depres⁃sion.We propose gene regulation by the ceRNA network in the progression of depression mod⁃els,which could help in revealing the new mech⁃anisms and understranding of neurotransmitter hypothesis of depression.

Key words:depression;brain-derived neuro⁃trophic factors;neurotransmitter hypothesis

T5-4

胶质细胞离子通道功能调控与癫痫药物靶标

卢应梅

（南京医科大学基础医学院，江苏 南京 211166）

摘要：目的癫痫以反复发作为特征，世界范围内有6500～7000万癫痫患者。目前认为神经元兴奋-抑制的失衡是构成癫痫发作的根本原因，但是直接针对神经元兴奋性的抗癫痫的策略已经被证明部分癫痫患者中不能很好的发挥抗癫痫作用。因此，进一步深入探究癫痫的发病机制显得尤为必要。方法采用基因小鼠，结合活体双光子、在体电生理记录、膜片钳记录、药理学/基因调控等技术，逐层深入地开展胶质细胞离子通道功能异常介导神经元功能改变的机制。结果①血管癫痫发生过程中，神经元、胶质细胞及脑微血管的异常改变。②脑血管内皮Cdk5缺失导致脑血管内皮细胞CX3CL1分泌过多。脑血管内皮源性CXCL1/CXCR2信号介导了星形胶质细胞GLT1的谷氨酸重摄取功能异常，导致锥体神经元活性增加，进而介导癫痫发生。③导致靶向Cdk5或CXCL1抗体或基因治疗，明显改善胶质细胞离子通道功能和神经元活性，改善癫痫的发生。结论脑血管源性的相关炎症因子是细胞间通讯的重要媒介分子，间接调控神经元功能，内皮细胞Cdk5以及它下游的信号通路为临床治疗癫痫提供了新思路和药物新靶点。

关键词：癫痫；胶质细胞；离子通道

T5-5

Optogenetic activation of glutamatergic neu⁃rons in somatosensory cortex promotes remy⁃elination in ischemic vascular dementia

ZHOU Yi-ting1,2＊,AN Da-dao1＊,XU Yi-xin1,ZHOU Ying4,LI Qing-qing4,ZHANG Xiang-nan1,WANG Yi1,3,LOU Min4,CHEN Zhong1,3,4,HU Wei-wei1

(1.Department of Pharmacology and Department of Pharmacy of the Second Affiliated Hospital,NHC and CAMS Key Laboratory of Medical Neuro⁃biology,School of Basic Medical Sciences,Col⁃lege of Pharmaceutical Sciences,Zhejiang Univer⁃sity,Hangzhou 310012,China;2.Department of Pharmacy,Sir Run Run Shaw Hospital,Hangzhou 310020,China;3.Key Laboratory of Neurophar⁃macology and Translational Medicine of Zhejiang Province,School of Pharmaceutical Sciences,Zhejiang Chinese Medical University,Hangzhou 310053,China;4.Department of Neurology,the Second Affiliated Hospital of Zhejiang University,School of Medicine,Hangzhou 310003,China)

Abstract:OBJECTIVEChronic cerebral hy⁃poperfusion can lead to progressive demyelin⁃ation and ischemic vascular dementia,yet there are no effective treatments.METHODSMagnetic resonance imaging was employed in patients with white matter damage,and optogenetics and skin stroking were exerted to activate glutamater⁃gic neurons in the somatosensory cortex in a clas⁃sical mouse model of ischemia vascular dementia.RESULTSWhite matter damage was correlated with disrupted cortical structure from MRI results.In a mouse model,activating glutamatergic neu⁃rons in the somatosensory cortex promotes prolif⁃eration of OPCs and remyelination to rescue cog⁃nitive impairment after chronic cerebral hypoper⁃fusion.Such therapeutic action was limited to stimulation with moderate intensity at the upper layers of the cortex,but was achieved over a wide time window after ischemia.Mechanistically,enhanced glutamatergic neuron-OPC functional synaptic connections are required for protection from activation of cortical glutamatergic neurons.Finally,skin stroking activation of the somatosen⁃sory cortex,an easier approach for clinical trans⁃lation,promoted OPC proliferation and remyelin⁃ation as well as cognitive recovery after cerebral hypoperfusion.CONCLUSIONActivation of gluta⁃matergic neurons in the somatosensory cortex may serve as novel approaches for treating isch⁃emic vascular dementia through precise modula⁃tion of glutamatergic neuron-OPC circuits.

Key words:optogenetics;glutamatergic neurons;ischemic vascular dementia

Corresponding author:HU Wei-wei,E-mail:huww@zju.edu.cn; CHEN Zhong,E-mail:chenzhong@zju.edu.cn

＊Co-first author.

T5-6

人诱导型多能干细胞来源多巴胺能神经元在精神神经性疾病研究中的应用

李 红，李 斐，卢关伊，周详斌，方 婷，吴 宁，李 锦

（军事科学院军事医学研究院毒物药物研究所，北京 100850）

摘要：目的人脑多巴胺系统功能异常可引起多种精神神经性疾病，但因为人脑组织样品极难获得，研究手法有限，大大限制其研究。采用人诱导型多能干细胞（iPSC）来源的多巴胺（DA）能神经元建立人脑发育/疾病模型，以增加对相关精神神经性疾病的认识。方法利用临床吸脂术后的脂肪组织，提取脂肪干细胞，将其重编程为iPSC，并进一步分化为DA能神经元，对包含变异基因的神经元功能进行检测。结果成功利用仙台病毒将人脂肪干细胞重编程为iPSC。现有胚胎干细胞诱导DA能神经元的方案并不适合iPSC；重新调整分化方案后的最佳的小分子化合物组合为Puromophine 3 μmol·L-1和CHIR-99021 0.8 μmol·L-1，可将iPSC高效向DA能神经元分化；神经元具有自发Pace-making动作电位的能力，分化后25 d细胞注射帕金森病模型大鼠纹状体内，可改善大鼠的行为学指标。在此基础上进一步发现Parkin缺失的iPSC来源的DA能神经元过早成熟和快速老化；对比FKBP5 rs1360780 CC/TT iPSC源DA能神经元的发育和功能，发现TT型来源的DA能神经元在基因和蛋白水平上AADC,TH和NURR1表达均显著减低，且多巴胺的分泌能力下降，在地塞米松处理建立的应激模型中表现为较强的应激反应性。结论人iPSC来源的神经元可建立人脑发育或疾病模型，在增加人类精神神经性疾病认识及促进高效靶向药物的开发中发挥重大作用。

关键词：诱导型多能干细胞；多巴胺；神经元；应激

T5-7

电刺激小脑齿状核对大鼠运动皮层缺血后神经发生的作用

孙芳玲，吴 铮，李子洁，刘 敏，田 欣，刘婷婷，王文

（首都医科大学宣武医院，北京 100053；北京市老年病医疗研究中心，北京 100053）

摘要：目的脑卒中是全球死亡和残疾的主要原因，约占全球5%的伤残调整年和10%的死亡率。目前临床上脑卒中的治疗存在有效治疗手段的局限性，除溶栓外仍缺乏有效的神经保护及再生修复治疗手段，且治疗时间窗窄，临床预后改善率低。脑卒中后存活患者不同程度丧失劳动力，其中上肢运动功能障碍在首次脑卒中后生存的患者中占73%～88%，在慢性脑卒中中占到55%～75%。寻找有效的神经再生修复治疗新方法，打破脑卒中治疗的瓶颈，是脑卒中后康复治疗的重点目标，对于最大程度促进患者功能恢复、减轻脑卒中负担有极为重要的意义。脑深部电刺激（DBS）作为一种新型的治疗方法，具有靶点明确、安全、可调等优点，已用于帕金森病、癫痫和肌张力障碍等疾病患者神经功能改善治疗上。但DBS在脑卒中治疗上的应用主要集中在神经疼痛方面，只有很少几个临床试验在研究其他脑卒中后遗症时关注到对运动功能的改善。因此本研究选用了小脑齿状核（LCN）作为DBS的靶点，观察电刺激是否能够激活小脑-丘脑-皮层通路，进而改善局灶性脑缺血大鼠的运动功能；并且观察了电刺激LCN后室管膜下区（SVZ）神经干细胞的增殖和分化情况，以期为脑卒中后的神经再生修复治疗提供新方法。方法采用内皮素-1定点注射诱导大鼠运动皮层缺血，之后在对侧LCN安置刺激电极。术前分别进行面条矩阵任务和水平梯子行走任务训练。其中面条矩阵抓取任务行为学评价，国内尚无相关实验方法报道，使用的动物实验装置为我们自行制备。实验中训练大鼠完成对食物-干面条的抓取任务，通过矩阵对其前肢运动的距离和和方向进行测量，定量和定性评估后前肢运动能力，为脑卒中后感觉-运动神经功能评估提供可靠的新方法。本研究在手术后第8天开始每天8 h的LCN DBS治疗，且每天配合进行15 min面条矩阵抓取训练，治疗持续14 d。尼氏染色计算梗死体积，免疫荧光双标检测SVZ神经发生。结果LCN DBS能有效改善缺血大鼠的神经功能损伤，缩小皮质梗死体积，促进运动功能的恢复。LCN DBS能够显著增加缺血侧和健侧室管膜下区BrdU+Nestin+神经干细胞和BrdU+DCX+成神经细胞的数量，并增加梗死周边BrdU+NeuN+的数量。还对脑卒中后第29天开始LCN DBS治疗大鼠的神经发生情况进行了检测，发现LCN DBS能够显著增加缺血侧和健侧室管膜下区BrdU+DCX+成神经细胞的数量，以及梗死周边BrdU+NeuN+的数量。结论LCN DBS促进神经功能恢复的机制可能与内源性神经干细胞增殖和分化相关。

关键词：脑深部电刺激；小脑齿状核；脑卒中；神经干细胞；运动功能

T5-8

基于神经再生修复策略的药物及电刺激康复治疗手段

王文

（首都医科大学宣武医院，北京 100053；北京市老年病医疗研究中心，北京 100053）

摘要：脑卒中等神经系统疾病的关键临床问题之一就是神经再生修复，以及相应的促进功能修复的治疗方法研究。有效的康复治疗能减轻脑损伤患者功能上的残疾，加速康复进程，降低潜在的治疗费用，减轻社会和家庭负担。以脑卒中为例，目前除溶栓外仍缺乏有效的神经保护及再生修复治疗手段，存活患者中有70%～80%的病人不同程度丧失劳动力，其中40%为中度功能障碍，15%～30%为重度残疾，而临床的康复治疗效果受限于现有康复机制、康复理念以及康复治疗技术，亟待寻找有效的脑卒中再生修复治疗靶点和新方法。我们课题组总结十多年的研究工作，针对内源性干细胞再生修复治疗缺血性脑损伤，提出了“神经血管稳态重构”假说：脑卒中等损伤能诱发自体干细胞跃迁增殖，但该反应持续时间较短，增殖逐渐回落到正常水平，不足以修复大脑功能；通过给予外源性药物或电刺激等物理治疗手段增强神经发生、血管新生，以及血脑屏障重构过程，促进神经血管单元稳态恢复，最终达到脑损伤后神经功能修复的作用。在该理论基础上，我们首次发现了单体化合物莫诺苷能显著促进脑卒中后神经发生、血管新生过程，维持血脑屏障完整性并促进微循环网络重构，进而改善缺血侧皮层的神经元功能，最终达到再生修复治疗脑卒中的作用。通过表面等离子共振结合实验和转录组测序，以及体内外基因敲除分子生物学技术研究手段，验证了莫诺苷促内源性神经干细胞增殖、分化为神经元的关键靶点。

随着科技的进步，脑机接口、人工智能等为医学发展带来了无限的可能。2018年首都医科大学宣武医院正式成立了中国国际神经科学研究所类脑智能临床转化研究中心，并启动了国际“重拾行走计划”在我国的首个项目。类脑智能中心依托于首都医科大学宣武医院强大的神经学科优势，立足于类脑智能，致力于通过类脑智能的手段研究和解决神经系统相关的疑难问题。以脑电信号分析技术、神经影像技术和深度学习技术为桥梁，寻求在高级脑功能保护、神经血管、癫痫、认知疾病和神经肿瘤等方面的重大原创突破。在这个背景下，我们与神经外科密切合作，开始探索脑深部电刺激对内源性神经干细胞的作用及调控机制，发现了脑深部电刺激小脑齿状核对侧脑室下层神经干细胞增殖分化的调控作用及对缺血性脑损伤神经功能的修复作用，部分研究数据已在Scientific Reports发表。该研究进一步验证了“神经血管稳态重构”假说，并对内源性干细胞治疗脑卒中提供一定的指导意义。而对于老年痴呆、帕金森病等神经退行性疾病，电刺激或许能重新激活大脑内的神经干细胞，配合药物干预和康复训练，给慢性神经系统疾病患者的神经功能再生修复治疗带来希望。

关键词：神经血管稳态重构；内源性神经干细胞；莫诺苷；电刺激；脑卒中；神经退行性疾病；神经再生修复

T5-9

基于CRISPR系统的人工改造外泌体在药理学研究中的应用

姚新刚

（南方医科大学药学院，广东省新药筛选重点实验室，广东 广州 510515）

摘要：目的CRISPR介导的基因编辑对基因治疗带来了巨大的前景，目前已有多项基于CRISPR的研究用于阿尔茨海默症的治疗。然而目前将CRISPR递送到相应的细胞甚至体内应用依然缺乏安全有效的手段，在细胞层面目前主要依赖于电转手段将CRISPR系统运送到细胞中，操作上比较复杂。而基于慢病毒和AAV的CRISPR递送体系不仅会造成免疫原性，而且往往容易整合到基因组中长期表达，引发不良后果。本研究拟基于外泌体的递送策略改善该问题。方法本论文采用构建工程化的外泌体平台，采用基于com和配体结合蛋白Com的富集系统，可以高效富集CRISPR系统并降低其脱靶效应和免疫原性。结果构建了多种RNP验证其效果，在细胞水平发现其富集RNP（spCas9，saCas9以及ABE系统）效率可以达到10倍以上，同时其脱靶效率显著降低。该系统不仅可以单独包装，也可以将多个RNP进行共包装，而且发现共包装外泌体系统优于单独的外泌体相加。同时也在体内水平验证了该系统的基因编辑效率，在del52hDMD/mdx小鼠模型中，注射包装了DMD exon 53 RNP的外泌体，通过confocal也观察到了肌肉dystrophin蛋白的表达。结论提供了一种免疫原性低于病毒系统的广泛性的RNP递送系统，且其效率显著高于同类型的外泌体系统。基于外泌体独特的免疫原性和归巢特性，以及穿过血脑屏障等特点，该系统对于神经性疾病的治疗中具有潜在的意义。

关键词：CRISPR；人工改造外泌体；基因编辑

通讯作者：姚新刚，E-mail:yaoxingang@smu.edu.cn