• 
    

    
    

      99热精品在线国产_美女午夜性视频免费_国产精品国产高清国产av_av欧美777_自拍偷自拍亚洲精品老妇_亚洲熟女精品中文字幕_www日本黄色视频网_国产精品野战在线观看

      ?

      第十九屆全國神經(jīng)精神藥理學(xué)學(xué)術(shù)會(huì)議論文摘要

      2021-03-28 21:49:472021年10月1517日廣東廣州
      關(guān)鍵詞:姜黃素阿爾茨海默病作用機(jī)制

      2021年10月15-17日,廣東廣州

      專題1:神經(jīng)退行性疾病

      T1-1

      NMDA受體對中樞神經(jīng)系統(tǒng)的影響

      孫麗叢,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:N-甲基-D-門冬氨酸(NMDA)受體是中樞神經(jīng)系統(tǒng)重要的興奮性神經(jīng)遞質(zhì),主要分布于大腦和脊髓等。研究發(fā)現(xiàn),NMDA參與許多神經(jīng)系統(tǒng)疾病的過程如腦卒中、癲癇、抑郁癥和強(qiáng)迫癥等。在正常機(jī)體的中樞神經(jīng)系統(tǒng)內(nèi),NMDA受體被激活后,Na+,K+和Ca2+的通透性增加,可產(chǎn)生一系列生化改變,影響突觸效率,進(jìn)而促進(jìn)學(xué)習(xí)和記憶能力的形成,并促進(jìn)海馬神經(jīng)干細(xì)胞增殖。但NMDA受體對中樞神經(jīng)系統(tǒng)也存在不利影響,表現(xiàn)為以下2個(gè)方面。①興奮性增高:有研究表明,在急性腦缺血中,NMDA受體亞基NR2A和NR2B的mRNA及蛋白表達(dá)均明顯上調(diào),導(dǎo)致細(xì)胞內(nèi)鈣超載,引起細(xì)胞毒性作用,甚至可導(dǎo)致細(xì)胞凋亡;當(dāng)NMDA受體被過度激活時(shí),過多的神經(jīng)元突觸后膜發(fā)生同步性去極化,持續(xù)性放電,引發(fā)癲癇;當(dāng)炎性疼痛產(chǎn)生時(shí),NMDA受體被激活,NMDA受體磷酸化水平升高,使神經(jīng)元的興奮作用放大,最終導(dǎo)致神經(jīng)元敏化。②興奮性降低:NMDA受體可調(diào)控γ-氨基丁酸(GABA)能中間神經(jīng)元的功能,其活性異常及不同亞基缺失可導(dǎo)致GABA能中間神經(jīng)元功能異常,從而引發(fā)抑郁癥和強(qiáng)迫癥等。本文通過對NMDA受體在中樞神經(jīng)系統(tǒng)中的影響進(jìn)行綜述,以期為NMDA受體異常導(dǎo)致疾病的治療提供參考。

      關(guān)鍵詞:N-甲基-D-門冬氨酸受體;中樞神經(jīng)系統(tǒng);興奮性神經(jīng)遞質(zhì);γ-氨基丁酸;神經(jīng)干細(xì)胞;學(xué)習(xí)能力;記憶能力

      基金項(xiàng)目:河北省自然科學(xué)基金(H2020208032)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T1-2

      姜黃素對阿爾茨海默病治療作用及其機(jī)制

      孫 婷,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:姜黃素是從姜科或天南星科一些植物的根莖中提取的一種二酮類化合物,是中藥姜黃發(fā)揮藥理作用的主要活性成分。近年來國內(nèi)外研究發(fā)現(xiàn),姜黃素的藥理活性廣泛,具有抗氧化、抗炎、神經(jīng)保護(hù)、降血脂、抗腫瘤等作用。阿爾茨海默?。ˋD)即老年癡呆癥,是一種常見的中樞神經(jīng)退行性疾病,其臨床特征主要表現(xiàn)為學(xué)習(xí)記憶能力減退和認(rèn)知功能障礙,其發(fā)病機(jī)制復(fù)雜,神經(jīng)遞質(zhì)乙酰膽堿缺失、氧化應(yīng)激、炎癥反應(yīng)及細(xì)胞凋亡等均可導(dǎo)致AD。姜黃素治療AD有顯著的療效,本文主要從其作用機(jī)制方面進(jìn)行綜述。①抗氧化應(yīng)激:姜黃素可以通過提高SOD和GSH-Px活性、降低MDA含量,抑制自由基生成并加強(qiáng)其清除,減輕氧化應(yīng)激;②抗炎癥反應(yīng):姜黃素可以降低1L-1β和TNF-α炎癥因子的表達(dá),減少炎癥介質(zhì)的產(chǎn)生,發(fā)揮神經(jīng)保護(hù)作用;③抑制細(xì)胞凋亡:姜黃素通過激活NF-κB信號通路,調(diào)控細(xì)胞凋亡相關(guān)基因Bax和Bcl-2表達(dá),降低促凋亡因子Bax表達(dá),增強(qiáng)抑制凋亡因子Bcl-2表達(dá)而阻止細(xì)胞凋亡;還可通過抑制糖原合成酶激酶3β活性抑制細(xì)胞凋亡;④調(diào)節(jié)膽堿能系統(tǒng)損傷:乙酰膽堿(ACh)是與學(xué)習(xí)記憶相關(guān)的一種重要的神經(jīng)遞質(zhì),姜黃素通過降低乙酰膽堿酯酶活性,增加膽堿乙酰轉(zhuǎn)移酶活性,增加ACh含量而改善ACh的代謝平衡。綜上所述,姜黃素可通過抗氧化應(yīng)激、抗炎癥反應(yīng)、抑制細(xì)胞凋亡及調(diào)節(jié)膽堿能系統(tǒng)損傷等治療AD。本文對姜黃素在治療AD作用及機(jī)制進(jìn)行綜述,以期為其臨床應(yīng)用及研究提供參考。

      關(guān)鍵詞:阿爾茨海默??;姜黃素;作用機(jī)制

      基金項(xiàng)目:河北省自然科學(xué)基金(H2020208032)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T1-3

      氨基酸類神經(jīng)遞質(zhì)對神經(jīng)系統(tǒng)疾病的影響及可能機(jī)制

      楊靜涵,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 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)是人體內(nèi)重要的氨基酸類神經(jīng)遞質(zhì),在人體內(nèi)屬低水平游離氨基酸,參與維系腦部的血液循環(huán),調(diào)控人腦的功能,神經(jīng)系統(tǒng)疾病如阿爾茨海默?。ˋD)與腦內(nèi)氨基酸關(guān)系密切。氨基酸之間比例失衡會(huì)導(dǎo)致多種神經(jīng)系統(tǒng)疾病的發(fā)生。有研究表明,AD和腦缺血等中樞神經(jīng)系統(tǒng)疾病模型動(dòng)物中的興奮性氨基酸類神經(jīng)遞質(zhì)Glu和Asp有不同程度的升高;抑制性氨基酸類神經(jīng)遞質(zhì)GABA和Gly則發(fā)生了不同程度的降低,而神經(jīng)系統(tǒng)疾病的治療藥物可以抑制這種明顯的改變,提示一個(gè)治療腦部疾病的新方向。氨基酸類神經(jīng)遞質(zhì)與神經(jīng)元信息傳遞、營養(yǎng)發(fā)育和學(xué)習(xí)記憶等過程有著緊密的聯(lián)系,Glu/GABA比值在一定范圍內(nèi)的升高可提高學(xué)習(xí)記憶能力,比值過高則產(chǎn)生抑制作用,蛇床子素可降低AD大鼠腦內(nèi)Glu和Glu/GABA比值,這可能是其改善認(rèn)知的機(jī)制。神經(jīng)系統(tǒng)疾病是一類病因復(fù)雜、容易復(fù)發(fā)且難以被及時(shí)診斷的疾病,多種神經(jīng)系統(tǒng)疾病僅能被緩解而無法治愈。因此,深入了解氨基酸類神經(jīng)遞質(zhì)在此類疾病中的變化,有助于揭示該類疾病發(fā)生發(fā)展進(jìn)程及藥物作用機(jī)制,為該類疾病的藥物研發(fā)提供新靶點(diǎn)。

      關(guān)鍵詞:氨基酸;神經(jīng)遞質(zhì);神經(jīng)系統(tǒng)疾??;學(xué)習(xí)記憶

      基金項(xiàng)目:河北省自然科學(xué)基金(H2020208032)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T1-4

      小膠質(zhì)細(xì)胞在治療神經(jīng)退行性疾病中的作用

      張智超,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:小膠質(zhì)細(xì)胞(MG)來源于卵黃囊,在血腦屏障形成之前進(jìn)入腦組織,成為大腦中的免疫細(xì)胞。MG參與一系列神經(jīng)退行性疾?。∟D)的發(fā)生,神經(jīng)系統(tǒng)紊亂可導(dǎo)致炎癥和MG的激活,不同信號刺激MG可呈現(xiàn)不同的活化表型與功能。MG在ND發(fā)生中起雙重作用,靜息態(tài)MG被激活,可釋放促炎因子,而活化的MG還可釋放抗炎因子,對神經(jīng)元起修復(fù)和保護(hù)作用。腦卒中和腦缺氧等急性ND中,MG釋放炎癥介質(zhì)對神經(jīng)細(xì)胞有益,可減輕繼發(fā)性損傷;阿爾茨海默病和帕金森病等慢性ND,慢性炎癥使得MG被長時(shí)程激活,持續(xù)釋放一系列炎癥因子,導(dǎo)致氧化應(yīng)激,造成神經(jīng)組織損傷。本文從MG的角度對治療ND的策略進(jìn)行綜述。①調(diào)節(jié)MG代謝:MG可通過腦內(nèi)微環(huán)境的代謝變化,改變轉(zhuǎn)錄表型和功能,當(dāng)受到炎癥刺激時(shí),MG代謝方式會(huì)從氧化磷酸化減少轉(zhuǎn)為糖酵解增加。對糖酵解代謝進(jìn)行靶向性治療有望改善神經(jīng)炎癥反應(yīng)。②抑制MG分泌促炎因子:MG類似巨噬細(xì)胞有吞噬、清除作用,可清除掉凋亡神經(jīng)細(xì)胞,疾病早期MG可自行清除,晚期神經(jīng)毒性作用抑制MG的吞噬作用,分泌的促炎細(xì)胞因子會(huì)加速疾病的發(fā)展。抑制MG分泌促炎因子,激發(fā)MG吞噬功能可緩解ND。

      關(guān)鍵詞:小膠質(zhì)細(xì)胞;神經(jīng)退行性疾?。恢委煵呗?/p>

      基金項(xiàng)目:河北省自然科學(xué)基金(H2020208032)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T1-5

      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

      T1-6

      歐前胡素對Aβ1-42致阿爾茨海默病模型小鼠神經(jīng)細(xì)胞凋亡Smac/DIABLO信號通路的影響

      羅 麗,萬航娟,何 蔚

      (贛南醫(yī)學(xué)院,江西省腦血管藥理重點(diǎn)實(shí)驗(yàn)室,心腦血管疾病防治教育部重點(diǎn)實(shí)驗(yàn)室,江西 贛州 341000)

      摘要:目的研究歐前胡素(Imp)對Aβ1-42致阿爾茨海默?。ˋD)模型小鼠Smac/DIABLO介導(dǎo)神經(jīng)細(xì)胞凋亡線粒體途徑的影響。方法雄性小鼠隨機(jī)分為正常對照組、AD模型組、AD模型+Imp 2.5 mg·kg-1組和AD模型+Imp 5.0 mg·kg-1組,每組10只。通過腦室內(nèi)注射Aβ1-42制備小鼠AD模型,Imp(2.5和5.0 mg·kg-1)在手術(shù)后當(dāng)天開始ip給藥,每天1次,連續(xù)給藥13 d。給藥后第14天,分離小鼠大腦皮質(zhì)組織,通過Western印跡法檢測皮質(zhì)組織中Smac/DIABLO、X染色體連鎖凋亡抑制蛋白(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的蛋白表達(dá)。通過胱天蛋白酶3和胱天蛋白酶9的活性檢測試劑盒檢測胱天蛋白酶3和胱天蛋白酶9蛋白活性。結(jié)果與正常對照組比較,模型組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蛋白表達(dá)及胱天蛋白酶3和胱天蛋白酶9蛋白活性均顯著升高(P<0.05),XIAP、胱天蛋白酶原3、胱天蛋白酶原9和PARP蛋白表達(dá)均顯著下降(P<0.05);與AD模型組相比,Imp(2.5 和 5.0 mg·kg-1)藥物治療組Smac/DIABLO和活化的胱天蛋白酶3蛋白表達(dá)及胱天蛋白酶3和胱天蛋白酶9蛋白活性均顯著下降(P<0.05),活化的胱天蛋白酶9和活化的PARP蛋白表達(dá)無明顯變化,XIAP和胱天蛋白酶原3的蛋白表達(dá)均明顯升高(P<0.05),胱天蛋白酶原9和PARP無明顯變化。結(jié)論Imp可能通過抑制Smac/DIABLO介導(dǎo)的線粒體途徑降低Aβ1-42致AD模型小鼠神經(jīng)細(xì)胞凋亡,從而發(fā)揮對AD的神經(jīng)保護(hù)作用。關(guān)鍵詞:歐前胡素;阿爾茨海默病;細(xì)胞凋亡;線粒體途徑

      T1-7

      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

      雌激素介導(dǎo)線粒體途徑的神經(jīng)保護(hù)作用

      戴月英,趙雨薇,甄艷杰,沈麗霞

      (河北北方學(xué)院藥學(xué)系,河北 張家口 075000)

      摘要:阿爾茨海默?。ˋD)是多因素引起的神經(jīng)退行性疾病,以認(rèn)知障礙、執(zhí)行力障礙為主要的臨床表現(xiàn),發(fā)病機(jī)制有Aβ級聯(lián)和Tau蛋白異常學(xué)說。諸多研究表明,Aβ沉積和Tau過度磷酸化之間存在密切的聯(lián)系。除此之外,由于線粒體被視為細(xì)胞的能量體,因而AD線粒體機(jī)制也成為AD發(fā)病機(jī)制研究領(lǐng)域。Aβ可直接與線粒體膜結(jié)合,從而改變線粒體動(dòng)力學(xué)和功能,導(dǎo)致能量代謝異常,最終引起細(xì)胞凋亡等一系列鏈?zhǔn)椒磻?yīng)。研究表明,線粒體功能障礙會(huì)加劇Aβ沉積和Tau蛋白異常磷酸化,而這2種病理變化反過來又能促進(jìn)線粒體損傷,通過線粒體依賴性凋亡通路來誘導(dǎo)細(xì)胞凋亡。流行病學(xué)調(diào)查顯示,絕經(jīng)后婦女較同齡男性AD發(fā)病率高1.5~3.0倍,認(rèn)為與女性絕經(jīng)后雌激素降低相關(guān)。研究表明,雌激素有神經(jīng)元保護(hù)作用。雌激素通過線粒體途徑發(fā)揮神經(jīng)保護(hù)作用及其機(jī)制的研究,為AD防治提供一定的思路。

      關(guān)鍵詞:阿爾茨海默病;雌激素;線粒體

      基金項(xiàng)目:河北省自然科學(xué)基金(H2019405057);河北省高等學(xué)??茖W(xué)技術(shù)研究項(xiàng)目(ZD2020136)

      通訊作者:沈麗霞,E-mail:shenlixiacn@163.com

      T1-9

      槲皮素神經(jīng)保護(hù)作用研究進(jìn)展

      趙雨薇,戴月英,甄艷杰,沈麗霞

      (河北北方學(xué)院藥學(xué)系,河北 張家口 075000)

      摘要:阿爾茨海默?。ˋD)主要病理特征有β-淀粉樣蛋白沉積形成的斑塊、胞內(nèi)Tau蛋白異常磷酸化導(dǎo)致的神經(jīng)纖維纏結(jié)、神經(jīng)元突觸缺失與凋亡。雌激素替代治療在防治和延遲AD發(fā)生的同時(shí),也增加了致癌的風(fēng)險(xiǎn),限制了其在臨床上的應(yīng)用。槲皮素作為一種植物雌激素,與雌激素結(jié)構(gòu)特征相似,可發(fā)揮雌激素樣神經(jīng)保護(hù)作用,且無致畸、致癌和致死作用,因此受到了廣泛關(guān)注。研究表明,槲皮素能調(diào)控核因子E2相關(guān)因子2易位,維持氧化還原平衡和線粒體功能的穩(wěn)定;介導(dǎo)MAPK信號通路,抑制對氧磷酶2的表達(dá),抵抗氧化應(yīng)激損傷;抑制β-分泌酶(BACE1)基因表達(dá),促進(jìn)非淀粉樣蛋白途徑,減少Aβ積聚;激活PI3K/AKT/GSK-3β信號通路,抑制Tau蛋白異常磷酸化;降低一氧化氮和一氧化氮合酶蛋白水平,減少核轉(zhuǎn)錄因子κB的核轉(zhuǎn)位,發(fā)揮抗炎特性;還可與雌激素受體結(jié)合,對神經(jīng)元凋亡起到保護(hù)作用。槲皮素的神經(jīng)保護(hù)作用機(jī)制復(fù)雜,涉及多條信號通路相互作用。

      關(guān)鍵詞:阿爾茨海默病;槲皮素;植物雌激素;神經(jīng)保護(hù)

      基金項(xiàng)目:河北省自然科學(xué)基金(H2019405057);河北省高等學(xué)??茖W(xué)技術(shù)研究項(xiàng)目(ZD2020136)

      通訊作者:沈麗霞,E-mail:shenlixiacn@163.com

      T1-10

      姜黃素類似物促神經(jīng)分化活性及其作用機(jī)制

      房紅志1,吳 敏1,陽澤界1,石 浩1,許文博1,張 雙1,李莎莉1,唐根云1,2

      (湖南醫(yī)藥學(xué)院1.基礎(chǔ)醫(yī)學(xué)院,2.腦與神經(jīng)內(nèi)分泌疾病湖南省高等學(xué)校重點(diǎn)實(shí)驗(yàn)室,湖南 懷化 418000)

      摘要:目的對姜黃素類似物進(jìn)行促神經(jīng)分化活性篩選,發(fā)現(xiàn)活性最佳的化合物后進(jìn)行分子作用機(jī)制研究。方法利用Neuro-2a細(xì)胞進(jìn)行藥物毒性和促神經(jīng)分化活性篩選,用MTT法進(jìn)行細(xì)胞毒性實(shí)驗(yàn);藥物誘導(dǎo)Neuro-2a分化后,采用神經(jīng)突起專一性抗體β-tubulin Ⅲ進(jìn)行免疫熒光染色,拍照后用Image J軟件對細(xì)胞分化率和神經(jīng)突的長度進(jìn)行測量,進(jìn)行統(tǒng)計(jì)分析。用Western印跡法對活性最佳化合物激活的分子信號通路關(guān)鍵蛋白進(jìn)行檢測,并用藥理學(xué)抑制劑進(jìn)行驗(yàn)證。結(jié)果通過對12個(gè)姜黃素類似物的促神經(jīng)分化活性篩選,其中雙去氧基姜黃素具有較好的促神經(jīng)分化活性,在很低的濃度就能夠促進(jìn)Neuro-2a細(xì)胞分化,1,2和4 μmol·L-1均能夠促進(jìn)細(xì)胞分化率和神經(jīng)突起長度顯著增加,并且成濃度依賴性。隨后采用4 μmol·L-1進(jìn)行分子作用機(jī)制研究,發(fā)現(xiàn)雙去氧基姜黃素通過激活ERK/MEK和PI3K/AKT通路誘導(dǎo)Neuro-2a細(xì)胞分化,并且用抑制劑U0126和LY200942得到驗(yàn)證。結(jié)論雙去氧基姜黃素具有較好的促神經(jīng)分化活性,對治療神經(jīng)退行性疾病具有潛在的研究價(jià)值。

      關(guān)鍵詞:姜黃素;神經(jīng)分化;抑制劑

      基金項(xiàng)目:國家自然科學(xué)基金(81703821);湖南省教育廳項(xiàng)目(2018B530);湖南醫(yī)藥學(xué)院自然科學(xué)基金(2015ZKB01);國家級大學(xué)生創(chuàng)新創(chuàng)業(yè)訓(xùn)練計(jì)劃(S202012214012);國家級大學(xué)生創(chuàng)新創(chuàng)業(yè)訓(xùn)練計(jì)劃(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抑制劑羅氟普蘭通過促進(jìn)自噬抑制小膠質(zhì)細(xì)胞活化作用機(jī)制

      劉 璐,汪海濤,徐江平

      (南方醫(yī)科大學(xué)藥學(xué)院,廣東 廣州 510515)

      摘要:目的神經(jīng)炎癥參與神經(jīng)退行性疾病的發(fā)展進(jìn)程,而小膠質(zhì)細(xì)胞活化是神經(jīng)炎癥的特征之一。羅氟普蘭(ROF)是一種新型致嘔吐潛能低的磷酸二酯酶4(PDE4)抑制劑。我們之前發(fā)現(xiàn),ROF可能通過促進(jìn)自噬從而抑制活化的小膠質(zhì)細(xì)胞中促炎因子的產(chǎn)生,但其機(jī)制尚不明確。本研究旨在探究ROF促進(jìn)自噬抑制小膠質(zhì)細(xì)胞活化作用及機(jī)制。方法使用Aβ25-35誘導(dǎo)BV-2細(xì)胞活化,采用ELISA試劑盒檢測白細(xì)胞介素6(IL-6)、IL-1β和腫瘤壞死因子α(TNF-α)含量;建立神經(jīng)炎癥細(xì)胞及炎癥小體活化細(xì)胞模型,模型條件下用自噬激活劑雷帕霉素誘導(dǎo)自噬,用自噬特異性抑制劑3-MA等抑制自噬及轉(zhuǎn)染PDE4質(zhì)粒,實(shí)現(xiàn)敲低PDE4基因;使用AMPK抑制劑和ULK1抑制劑評價(jià)是否阻斷藥物作用,上述情況下檢測自噬相關(guān)信號通路中關(guān)鍵蛋白及激酶的變化。在C57小鼠側(cè)腦室定位注射Aβ25-35構(gòu)建小鼠炎癥模型,通過新物體識別等行為學(xué)實(shí)驗(yàn)方法觀察模型小鼠學(xué)習(xí)記憶能力的損傷及PDE4抑制劑能否改善學(xué)習(xí)記憶的能力。取腦組織,采用免疫熒光雙標(biāo)法檢測海馬和皮質(zhì)中小膠質(zhì)細(xì)胞自噬相關(guān)蛋白的表達(dá)。用ELISA和Western印跡法分別檢測小膠質(zhì)細(xì)胞炎癥因子的水平及M1和M2表型標(biāo)志物的水平。結(jié)果①Aβ25-35處理BV-2小膠質(zhì)細(xì)胞顯著增加細(xì)胞的活化水平,ROF可以有效減輕Aβ25-35誘導(dǎo)的小膠質(zhì)細(xì)胞活化,并降低Iba1的表達(dá)水平;同時(shí),給予ROF或者敲低PDE4B能顯著降低模型條件下炎癥因子的水平。②Aβ25-35處理BV-2小膠質(zhì)細(xì)胞后自噬水平下降,給予ROF,顯著上升;在給予ULK1抑制劑后,ROF的促自噬作用則被阻斷。③小鼠側(cè)腦室注射凝聚態(tài)的Aβ25-35,明顯影響小鼠的學(xué)習(xí)記憶能力;ig給予ROF改善其學(xué)習(xí)記憶能力,海馬和皮質(zhì)中小膠質(zhì)細(xì)胞活化水平也明顯下降,促炎因子降低,其自噬蛋白水平升高;ig給予ROF和ULK1抑制劑組逆轉(zhuǎn)這些現(xiàn)象。結(jié)論P(yáng)DE4抑制劑ROF可能通過AMPK/ULK1信號轉(zhuǎn)導(dǎo)通路促進(jìn)自噬,從而抑制Aβ25-35誘導(dǎo)BV-2小膠質(zhì)細(xì)胞炎癥因子,減輕海馬和皮質(zhì)中小膠質(zhì)活化,也可能依賴ULK1蛋白,從而產(chǎn)生神經(jīng)保護(hù)作用。

      關(guān)鍵詞:磷酸二酯酶4;羅氟普蘭;神經(jīng)炎癥;小膠質(zhì)細(xì)胞

      基金項(xiàng)目:國家自然科學(xué)基金(81773698);國家自然科學(xué)基金(81974501)

      通訊作者: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

      甜菜堿通過RAS影響大鼠高血壓蓄積致學(xué)習(xí)記憶障礙

      韓文東,陳姜艷,劉麗媛,王 浩,高 山

      〔山東第一醫(yī)科大學(xué)(山東省醫(yī)學(xué)科學(xué)院),山東泰安 271016〕

      摘要:目的高血壓蓄積會(huì)導(dǎo)致體內(nèi)氧自由基增多和炎癥反應(yīng)加劇,從而誘發(fā)許多心腦疾病的發(fā)生,其中包括學(xué)習(xí)記憶障礙。本研究探討甜菜堿(Bet)是否通過抗氧化對高血壓導(dǎo)致學(xué)習(xí)記憶障礙大鼠產(chǎn)生保護(hù)作用。方法雄性SD大鼠,160~180 g,制備2K1C高血壓模型。造模成功后,隨機(jī)分為6組:對照組、模型組、Bet 10 和 30 mg·kg-1、抑制劑組(A779+Bet 30 mg·kg-1)和A779組,給藥4周。采用水迷宮、新物體識別和Y迷宮檢測大鼠學(xué)習(xí)記憶能力;用Western印跡法檢測海馬中APP,Aβ,PS-1,Catalase和HO-1蛋白表達(dá);ELISA檢測血漿中Ang1-7含量。結(jié)果①行為學(xué)實(shí)驗(yàn)結(jié)果表明,穿越平臺次數(shù)、目標(biāo)象限停留時(shí)間、總探索時(shí)間、認(rèn)知指數(shù)、總進(jìn)臂次數(shù)和交替比模型組顯著低于對照組,給予Bet可抑制此作用,并呈劑量依賴性。②Western印跡法結(jié)果顯示,與對照組相比,模型組大鼠海馬中APP,Aβ和PS-1含量均增加,鹽酸Bet可抑制此變化,并呈劑量依賴性;模型組海馬中Catalase和HO-1含量均低于對照組,鹽酸Bet可抑制此變化并呈劑量依賴性。③ELISA結(jié)果顯示,模型組血漿中Ang1-7含量均低于對照組,鹽酸Bet可抑制此變化,并呈劑量依賴性;A799可逆轉(zhuǎn)上述鹽酸Bet的作用。結(jié)論鹽酸Bet對高血壓蓄積致學(xué)習(xí)記憶障礙的保護(hù)可能是通過作用于Ang1-7/Mas軸,增強(qiáng)抗氧化作用,從而改善大鼠學(xué)習(xí)記憶障礙。

      關(guān)鍵詞:甜菜堿;高血壓蓄積;學(xué)習(xí)記憶障礙;氧化應(yīng)激

      基金項(xiàng)目:國家自然科學(xué)基金青年科學(xué)基金(81400182);山東省博士基金(BS2014YY045)

      通訊作者:高 山,E-mail:gshan84117@163.com

      T1-19

      組胺及其受體在阿爾茨海默病發(fā)病中的生物學(xué)作用

      劉麗媛,張 猛,王 浩

      (山東第一醫(yī)科大學(xué)藥理學(xué)研究所,山東 泰安 271016)

      摘要:阿爾茨海默?。ˋD)為中樞退行性疾病。組胺為內(nèi)源性生物胺,與其受體構(gòu)成中樞組胺能系統(tǒng),在腦穩(wěn)態(tài)的維持和高級腦功能中發(fā)揮重要作用。近年發(fā)現(xiàn),組胺及其受體在神經(jīng)調(diào)控及AD發(fā)生發(fā)展中發(fā)揮關(guān)鍵作用,可能是治療AD的潛在靶點(diǎn)。本文從組胺受體的腦內(nèi)分布及功能、組胺在AD發(fā)生發(fā)展中的作用及機(jī)制對組胺及其受體在AD發(fā)病中的生物學(xué)作用進(jìn)行綜述。組胺通過激動(dòng)其受體發(fā)揮作用。組胺受體包括H1~H4亞型,4種亞型在腦內(nèi)均有分布,發(fā)揮多種生物學(xué)功能,其中目前研究最廣泛的是組胺H3受體(H3R),其分布于下丘腦、大腦皮質(zhì)、海馬、杏仁核、紋狀體和基底神經(jīng)節(jié)等多個(gè)部位。H3R通過對組胺合成的反饋抑制來調(diào)控組胺的釋放。同時(shí),其也可作為異源受體,參與細(xì)胞信號轉(zhuǎn)導(dǎo)、調(diào)控其他神經(jīng)遞質(zhì)的釋放。H3R可能通過膽堿能通路、Aβ?lián)p傷、Tau蛋白通路和海馬神經(jīng)發(fā)生等多種機(jī)制參與AD的發(fā)生。總之,組胺與其受體組成的組胺能系統(tǒng)廣泛分布于中樞神經(jīng)系統(tǒng),與AD發(fā)生密切相關(guān),其中組胺H3R拮抗劑研究廣泛,可通過多種機(jī)制改善AD,可成為AD的治療的潛在靶點(diǎn)。

      關(guān)鍵詞:阿爾茨海默??;組胺;膽堿能通路;Aβ?lián)p傷;Tau蛋白

      基金項(xiàng)目:國家自然科學(xué)基金(81441111);國家自然科學(xué)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

      山茱萸有效成分對老年快速老化小鼠的藥理作用及其機(jī)制

      李雅莉,馬登磊,張 蘭,李 林

      (首都醫(yī)科大學(xué)宣武醫(yī)院藥學(xué)部,神經(jīng)變性病教育部重點(diǎn)實(shí)驗(yàn)室,北京市神經(jīng)藥物工程研究中心,北京 100053)

      摘要:目的阿爾茨海默病(AD)是一種與年齡相關(guān),以神經(jīng)元退行性病變導(dǎo)致個(gè)體行為異常為主要特征的神經(jīng)系統(tǒng)疾病??焖倮匣∈螅⊿AM)是具有快速老化特征的小鼠品系,SAMP8小鼠表現(xiàn)不可逆的快速老化,表現(xiàn)與老年人和AD患者類似的老化癥狀。山茱萸環(huán)烯醚萜苷(CIG)是本室從中藥山茱萸中提取的有效部位。方法給予12月齡SAMP8小鼠ig CIG治療至14月齡后,觀察SAMP8小鼠的行為學(xué)改變和老化狀態(tài)等變化及相應(yīng)的作用機(jī)制。結(jié)果①CIG能降低14月齡SAMP8小鼠的老化指數(shù)評分(根據(jù)小鼠反應(yīng)性、皮毛等程度進(jìn)行評分,指數(shù)越高表明小鼠的老化程度越嚴(yán)重),增高存活率,改善轉(zhuǎn)棒實(shí)驗(yàn)中運(yùn)動(dòng)協(xié)調(diào)能力。②CIG能降低SAMP8小鼠紋狀體區(qū)tau蛋白在Thr205和Ser396位點(diǎn)的過度磷酸化。③CIG能降低APP相關(guān)代謝蛋白的表達(dá)。④CIG增加突觸相關(guān)蛋白表達(dá)。⑤GIG降低壞死性凋亡相關(guān)蛋白表達(dá)。結(jié)論CIG具有延緩衰老的作用,有利于預(yù)防和治療AD,并且對于晚期AD患者可能起到改善生活質(zhì)量的作用。

      關(guān)鍵詞:山茱萸;快速老化小鼠;tau蛋白

      T1-22

      sPINK1抑制泛素依賴蛋白酶體的活性損傷神經(jīng)元

      陳 聰,易華偉,沈 晨,高童謠,唐 淳,張緯萍

      (浙江大學(xué)醫(yī)學(xué)院,浙江 杭州 310043)

      摘要:目的PINK1有線粒體膜定位的fPINK1和胞漿分布的sPINK1,2種PINK1在老年人、阿爾茨海默病患者腦內(nèi)的神經(jīng)元均顯著增加,它們都能夠磷酸化泛素Ser65位點(diǎn)。fPINK1通過磷酸化泛素誘導(dǎo)線粒體自噬,發(fā)揮神經(jīng)保護(hù)作用,而sPINK1的作用尚不清楚。本文揭示sPINK1在神經(jīng)元中的作用及機(jī)制。方法通過噬神經(jīng)元的腺相關(guān)病毒(AAV)在小鼠海馬CA1神經(jīng)元,分別過表達(dá)GFP、sPINK1和sPINK1及S65A突變的泛素(Ub/S65A,不能被磷酸化的泛素)和Ub/S65E(模擬磷酸化的泛素),通過分析小鼠神經(jīng)行為、形態(tài)和生化等變化,揭示sPINK1的作用,并在蛋白質(zhì)水平分析sPINK1的作用機(jī)制。結(jié)果過表達(dá)sPINK1可損傷小鼠海馬依賴的學(xué)習(xí)記憶功能,導(dǎo)致神經(jīng)元損傷,包括線粒體腫脹、神經(jīng)突起發(fā)生退行性變化和突觸減少,以及神經(jīng)元內(nèi)的蛋白質(zhì)聚集增加、星形膠質(zhì)細(xì)胞和小膠質(zhì)細(xì)胞反應(yīng)性激活。sPINK1的作用能夠被Ub/S65A抑制,而Ub/S65E能夠模擬sPINK1的作用。在蛋白質(zhì)水平,泛素化的GFP能夠被純化的蛋白酶體降解;而泛素被sPINK1磷酸化后,GFP被蛋白酶體降解的速率顯著降低。結(jié)論sPINK1可通過磷酸化泛素,抑制泛素依賴的蛋白質(zhì)降解,損傷神經(jīng)元。

      關(guān)鍵詞:PINK1;蛋白酶體;神經(jīng)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

      鮮天麻對慢性束縛應(yīng)激誘導(dǎo)小鼠學(xué)習(xí)記憶損傷的改善作用

      黃 紅,姜 寧,張亦文,姚彩虹,劉新民

      (中國醫(yī)學(xué)科學(xué)院北京協(xié)和醫(yī)學(xué)院藥用植物研究所,北京 100193)

      摘要:目的現(xiàn)代科技發(fā)展使得人類生活模式、生存空間發(fā)生了重大轉(zhuǎn)變,也對人類神經(jīng)精神活動(dòng)產(chǎn)生重大影響。這種內(nèi)外源性應(yīng)激會(huì)導(dǎo)致學(xué)習(xí)記憶的功能性損傷(亞健康狀態(tài)),嚴(yán)重降低人們的生活質(zhì)量。我國傳統(tǒng)中藥對于應(yīng)激所致認(rèn)知功能性損傷的亞健康狀態(tài)的防護(hù)具有獨(dú)特優(yōu)勢。鮮天麻(FG)是一種藥食同源中藥,現(xiàn)代藥理學(xué)研究表明,它具有良好的改善學(xué)習(xí)記憶作用。本研究采用慢性束縛應(yīng)激(CRS)誘導(dǎo)小鼠學(xué)習(xí)記憶障礙模型,探討FG對CRS所致學(xué)習(xí)記憶障礙的改善作用。方法除對照組外,其余各組ICR雄鼠采用本實(shí)驗(yàn)室自制的束縛器連續(xù)造模35 d,每天10 h(22∶00~8∶00),F(xiàn)G汁(每天ig 0.5和1.0 g·kg-1),陽性藥組給予多奈哌齊(Don,每天ig 1.6 mg·kg-1),給藥3周。新物體識別實(shí)驗(yàn)(NORT)和物體位置識別實(shí)驗(yàn)(OLRT),分別用于評價(jià)小鼠的辨別能力和空間學(xué)習(xí)記憶能力,避暗實(shí)驗(yàn)評價(jià)小鼠的被動(dòng)回避能力。實(shí)驗(yàn)數(shù)據(jù)采用SPSS軟件21.0(ANOVA,±s)統(tǒng)計(jì)分析,P<0.05認(rèn)為差異具有統(tǒng)計(jì)學(xué)意義。結(jié)果CRS能顯著性降低OLRT和NORT的辨別指數(shù)(P<0.05),縮短避暗實(shí)驗(yàn)動(dòng)物進(jìn)入暗室的潛伏期(P<0.05)。FG和Don治療組可逆轉(zhuǎn)上述變化(P<0.05)。結(jié)論FG可改善CRS所致的學(xué)習(xí)記憶障礙。FG是可供長期服用的治療應(yīng)激相關(guān)記憶障礙的藥食同源中藥,本研究結(jié)果為研發(fā)天麻改善記憶減退類產(chǎn)品提供了依據(jù)。

      關(guān)鍵詞:天麻;學(xué)習(xí)記憶能力;應(yīng)激

      通訊作者:劉新民,E-mail:liuxinmin@hotmail.com

      T1-26

      左旋丁苯酞對肌萎縮性側(cè)索硬化癥細(xì)胞模型的保護(hù)作用和機(jī)制

      蘭嘉琦,彭雨晨,彭 英

      (中國醫(yī)學(xué)科學(xué)院藥物研究所,北京 100050)

      摘要:目的肌萎縮性側(cè)索硬化(ALS)是一種復(fù)雜的多因素慢性神經(jīng)元疾病,TAR DNA結(jié)合蛋白43(TDP-43)在細(xì)胞中形成的包涵體是其特征性病理標(biāo)志,并與其另一個(gè)病理特征氧化應(yīng)激密切相關(guān)。本研究探討了神經(jīng)保護(hù)劑左旋丁苯酞(L-NBP)對突變TDP-43所致的ALS相關(guān)損傷的可能保護(hù)作用和機(jī)制。方法建立了穩(wěn)定轉(zhuǎn)染TDP-43 M337V突變的NSC-34細(xì)胞模型以考察L-NBP對細(xì)胞損傷的改善作用,對氧化應(yīng)激的影響,對線粒體的可能作用,對TDP-43 M337V聚集的改善作用,對DNA損傷的保護(hù)作用以及對相應(yīng)蛋白的調(diào)控作用,并探討其可能的作用機(jī)制。結(jié)果在轉(zhuǎn)染TDP-43 M337V的小鼠運(yùn)動(dòng)神經(jīng)元細(xì)胞模型上,L-NBP濃度依賴地提升細(xì)胞的存活率,并顯著降低TDP-43 M337V造成的氧化應(yīng)激,可能與其激活Nrf2/HO-1軸的作用有關(guān)。L-NBP對于TDP-43 M337V在胞漿內(nèi)的聚集也有顯著的改善作用,但對泛素-蛋白酶體通路和自噬-溶酶體通路均無明顯影響。L-NBP顯著升高穩(wěn)轉(zhuǎn)細(xì)胞的線粒體膜電位,并對線粒體片段化有一定改善作用,可能與其降低線粒體分裂蛋白FIS1的表達(dá),并促進(jìn)融合相關(guān)蛋白MFN2的表達(dá)有關(guān)。結(jié)論L-NBP可能通過激活Nrf2/HO-1抗氧化信號通路抑制細(xì)胞內(nèi)氧化應(yīng)激和相關(guān)損傷,并降低TDP-43 M337V在胞漿內(nèi)的聚集,改善線粒體片段化,提高膜電位,進(jìn)而發(fā)揮其抗ALS的作用,提示L-NBP可能成為治療ALS的新策略。

      關(guān)鍵詞:左旋丁苯酞;氧化應(yīng)激;線粒體膜電位

      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

      大黃對潰瘍性結(jié)腸炎治療作用研究進(jìn)展

      郭子霞1,張丹參1,2,李 煒1

      (1.河北北方學(xué)院藥學(xué)系,河北省神經(jīng)藥理學(xué)重點(diǎn)實(shí)驗(yàn)室,河北 張家口 075000;2.河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:潰瘍性結(jié)腸炎(UC)是一種慢性非特異性腸道疾病。該病病程漫長,易反復(fù)發(fā)作,嚴(yán)重影響患者身心健康及生活品質(zhì)。UC具體病因不明,與機(jī)體遺傳、免疫反應(yīng)、腸道菌群、環(huán)境、精神心理狀態(tài)等多種因素有關(guān)。臨床癥狀有腹痛腹瀉、結(jié)直腸因潰瘍出血出現(xiàn)黏液膿血便。此外患者血液高凝、血小板易聚集造成微循環(huán)障礙。中藥大黃具有通經(jīng)解瘀、清熱解毒之功效。大黃的多種藥理活性對應(yīng)UC的多種臨床癥狀及病理特點(diǎn)。大黃中鞣質(zhì)具有收斂止瀉作用,當(dāng)小劑量使用時(shí),鞣質(zhì)的收斂作用蓋過蒽醌類物質(zhì)的瀉下作用,可減輕患者腹瀉癥狀。大黃全成分活血止血,臨床研究已證明其可有效減輕上消化道出血癥狀,其中的沒食子酸等成分可促進(jìn)血小板在創(chuàng)傷部位聚集,促進(jìn)凝血進(jìn)而止血。大黃的活血止血功效可緩解UC患者腸道潰瘍出血、血便癥狀。大黃黃酮類成分可通過升高血漿滲透壓增加血容量來改善微循環(huán)、大黃酚-8-葡萄糖苷可抑制血小板聚集從而抑制血栓形成。此外,大黃蒽醌類物質(zhì)如蘆薈大黃素、大黃酚、大黃酸等可減少炎癥因子的釋放,緩解UC免疫功能異常引起的炎癥反應(yīng)。綜上,大黃可多方面、多角度地對UC起到緩解甚至治療的作用。

      關(guān)鍵詞:大黃;潰瘍性結(jié)腸炎;治療作用

      基金項(xiàng)目:河北省高等學(xué)??茖W(xué)技術(shù)研究項(xiàng)目(ZD2020117)

      通訊作者:張丹參,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

      斯皮諾素通過減輕炎癥改善阿爾茨海默病

      杜會(huì)枝

      (山西大學(xué)分子科學(xué)研究所,山西 太原 030006)

      摘要:目的阿爾茨海默病(AD)是以記憶障礙和學(xué)習(xí)認(rèn)知損傷為特征的慢性神經(jīng)退行性疾病。目前為止,AD的確切發(fā)病機(jī)制并未完全闡明,主要有β-淀粉樣蛋白(Aβ)沉積學(xué)說、氧化應(yīng)激學(xué)說、神經(jīng)炎癥學(xué)說和細(xì)胞凋亡學(xué)說等。近年來大量研究發(fā)現(xiàn),神經(jīng)炎癥在AD癥的發(fā)生發(fā)展中起著重要作用,腦內(nèi)慢性炎癥反應(yīng)是AD的病理特征之一,Aβ沉積激活神經(jīng)膠質(zhì)細(xì)胞釋放促炎因子,如白細(xì)胞介素6(IL-6)、腫瘤壞死因子α(TNF-α)和一氧化氮(NO)等,進(jìn)而損傷周圍神經(jīng)元,引起突觸丟失和細(xì)胞凋亡,最終導(dǎo)致學(xué)習(xí)記憶功能受損。減輕炎癥被認(rèn)為是改善甚至是治療AD癥的重要途經(jīng)之一。斯皮諾素(spinosin),屬于黃酮類化合物,是酸棗仁(Ziziphi Spinosae Semen)中主要活性成分之一。本文先在化學(xué)角度研究了斯皮諾素對Aβ聚集的影響,又在小鼠海馬腦片上探討了其改善AD的機(jī)制。方法和結(jié)果 37℃孵育96 h產(chǎn)生的Aβ25-35聚集體與新鮮配制的Aβ25-35單體相比,粒徑顯著增大;斯皮諾素(終濃度為30 μmol·L-1)與Aβ單體一起孵育后,粒徑明顯降低,表明斯皮諾素緩解Aβ25-35的聚集。Aβ25-35(5 μmol·L-1)組炎癥因子IL-1β和PGE-2含量顯著升高;在腦片上預(yù)給藥斯皮諾素0.5 h(終濃度為 30 μmol·L-1),該 2種炎癥因子的含量降低至接近對照組。COX-2蛋白可以降解2-花生四烯酸甘油從而產(chǎn)生PGE-2。本研究檢測了各組COX-2蛋白表達(dá)。結(jié)果顯示,斯皮諾素抑制Aβ25-35引起的COX-2蛋白的過表達(dá)。隨后研究了斯皮諾素對促凋亡蛋白Bax的作用。Aβ25-35處理海馬腦片后,上調(diào)Bax蛋白表達(dá),增加細(xì)胞凋亡;用斯皮諾素預(yù)保護(hù)后,Bax蛋白表達(dá)顯著下降,抑制Aβ25-35引起的細(xì)胞凋亡。長時(shí)程增強(qiáng)(LTP)是反應(yīng)學(xué)習(xí)記憶的主要指標(biāo)之一。采用膜片鉗技術(shù),記錄興奮性突觸后場電位,研究斯皮諾素對Aβ25-35引起的學(xué)習(xí)記憶障的改善作用。用斯皮諾素預(yù)保護(hù)后,LTP比例從Aβ25-35組的108%回升至與對照組的147%,對照組為142%。表明斯皮諾素對Aβ25-35誘導(dǎo)的學(xué)習(xí)記憶障礙有改善作用。結(jié)論斯皮諾素通過抑制COX-2蛋白過表達(dá),減輕炎癥,減少細(xì)胞凋亡,改善AD癥狀。

      關(guān)鍵詞:斯皮諾素;Aβ25-35;神經(jīng)炎癥;阿爾茨海默病

      基金項(xiàng)目:山西省自然科學(xué)基金(201801D121290);山西省回國留學(xué)人員科研項(xiàng)目(2017-021);山西醫(yī)科大學(xué)細(xì)胞生理學(xué)教育部重點(diǎn)實(shí)驗(yàn)室開放基金(KLMEC/SXMU-201911)

      通訊作者:杜會(huì)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

      黃精多糖對阿爾茨海默病模型斑馬魚p38MAPK/N-cadherin的影響

      陳毅飛1,劉凱菲1,吳世敏1,李訊怡1,黃 瑤1,黃典鳳1,蔣才武2

      (1.桂林醫(yī)學(xué)院藥學(xué)院,廣西 桂林 541199;2.廣西中醫(yī)藥大學(xué)藥學(xué)院,廣西 南寧 530200)

      摘要:目的研究黃精多糖對阿爾茨海默病模型斑馬魚的影響。方法將斑馬魚置于T迷宮中訓(xùn)練7 d,將訓(xùn)練成功的40條斑馬魚分為4組:空白組、模型組、陽性組的和黃精多糖組。模型組、陽性組和黃精多糖組在100 μg·L-1的AlCl3中造模6 d;陽性組暴露在4 μg·L-1的石杉堿甲藥液,黃精多糖組暴露在6 g·L-1黃精多糖藥液,均給藥6 d,模型組不予以給藥,空白組不做任何處理,并用錄像的方法對斑馬魚各階段進(jìn)行記錄,對各組在EC區(qū)的時(shí)間進(jìn)行分析。給藥結(jié)束后取出腦組織,采用Western印跡法測定N-cadherin,P38和p-P38蛋白因子的表達(dá)。結(jié)果在行為學(xué)上,分析在EC區(qū)所待時(shí)間,空白組、陽性組和黃精多糖組分別與模型組相比均有統(tǒng)計(jì)學(xué)差異(P<0.05)。在蛋白水平上,與模型組比較,陽性組和黃精多糖組P38和p-P38蛋白下調(diào),而N-cadherin蛋白上調(diào)(P<0.05)。結(jié)論黃精多糖通過上調(diào)N-cadherin蛋白水平,阻礙P38磷酸化,改善阿爾茨海默病模型斑馬魚的學(xué)習(xí)記憶能力。

      關(guān)鍵詞:阿爾茨海默??;斑馬魚;黃精多糖;N-cadherin;P38蛋白

      基金項(xiàng)目:國家自然科學(xué)基金(8216140711);廣西壯族自治區(qū)自然科學(xué)基金(2017GXNSFAA198255),廣西壯族自治區(qū)自然科學(xué)基金(2018GXNSFBA13 8028);廣西腦與認(rèn)知神經(jīng)科學(xué)重點(diǎn)實(shí)驗(yàn)室開放課題(GKLBCN-20180105-03);2019大學(xué)生創(chuàng)新創(chuàng)業(yè)項(xiàng)目訓(xùn)練計(jì)劃(201910601038);第四期廣西高等學(xué)校千名中青年骨干教師培育計(jì)劃

      通訊作者:蔣才武,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

      專題2:藥物成癮與精神疾病

      T2-1

      熱休克蛋白70功能抑制劑亞甲藍(lán)緩解嗎啡依賴小鼠納洛酮催促戒斷癥狀

      宮 琦1,魏守鵬1,李語玲2,梁 慧1,梁建輝1,3

      (北京大學(xué)1.基礎(chǔ)醫(yī)學(xué)院藥理學(xué)系,3.藥學(xué)院分子與細(xì)胞藥理學(xué)系,北京 100083;2.同濟(jì)大學(xué)醫(yī)學(xué)院附屬上海市東方醫(yī)院藥學(xué)部,上海 200120)

      摘要:目的探討熱休克蛋白70(Hsp70)抑制劑亞甲藍(lán)對嗎啡依賴小鼠納洛酮催促戒斷癥狀的藥理作用。方法采用自發(fā)活動(dòng)視頻分析系統(tǒng)(Dig Behv-LG)測定亞甲藍(lán)2.5~10 mg·kg-1對雄性昆明小鼠自主活動(dòng)的影響。給雄性昆明小鼠連續(xù)2 d sc給予嗎啡,第1天嗎啡劑量為每次30 mg·kg-1,第2天嗎啡劑量為每次60 mg·kg-1,每天2次,建立嗎啡依賴小鼠模型。每次給予嗎啡前5 min,ip給予不同劑量的亞甲藍(lán)(0,2.5,5和10 mg·kg-1)進(jìn)行干預(yù)試驗(yàn)。第3天,sc給予嗎啡60 mg·kg-1后3 h,ip給予納洛酮8 mg·kg-1,并觀察和記錄30 min內(nèi)小鼠的跳躍次數(shù)以及注射納洛酮前后30 min小鼠體重的變化。結(jié)果亞甲藍(lán)在2.5~10 mg·kg-1劑量范圍內(nèi),對小鼠自主活動(dòng)無明顯影響。嗎啡依賴小鼠納洛酮催促戒斷試驗(yàn)顯示,亞甲藍(lán)可以劑量依賴性地抑制小鼠的催促戒斷跳躍,并明顯抑制納洛酮催促戒斷小鼠的體重丟失。結(jié)論作為Hsp70抑制劑,亞甲藍(lán)對嗎啡依賴小鼠納洛酮催促戒斷癥狀具有明確的抑制作用。

      關(guān)鍵詞:嗎啡;納洛酮;催促戒斷;熱休克蛋白70;亞甲藍(lán)

      基金項(xiàng)目:國家自然科學(xué)基金(81773705)

      通訊作者:梁建輝,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

      鹽酸羥哌吡酮(YL-0919)的快速抗抑郁作用及其電生理機(jī)制

      尹勇玉1,張黎明1,李云峰1,2

      (軍事科學(xué)院軍事醫(yī)學(xué)研究院1.毒物藥物研究所,2.軍事認(rèn)知與腦科學(xué)研究所,北京 100850)

      摘要:目的探究鹽酸羥哌吡酮(YL-0919)快速抗抑郁的行為學(xué)效應(yīng)及其電生理機(jī)制。方法采用慢性應(yīng)激模型、開窗測驗(yàn)、強(qiáng)迫游泳測驗(yàn)、行為類別分析等評價(jià)YL-0919的抗抑郁活性,采用在體多通道電生理記錄技術(shù)、在體誘導(dǎo)神經(jīng)場電位技術(shù)探究YL-0919快速抗抑郁作用的電生理機(jī)制。結(jié)果①連續(xù)8周的慢性應(yīng)激可使食蟹猴表現(xiàn)出顯著的抑郁樣行為,包括蜷縮行為、自我撓體行為等顯著增加,環(huán)境探索、自主運(yùn)動(dòng)等顯著下降;開窗潛伏期顯著增加,開窗時(shí)間顯著下降;慢性給予YL-0919(1.2 mg·kg-1,ig)9 d即可顯著逆轉(zhuǎn)由慢性應(yīng)激所導(dǎo)致的系列抑郁樣行為,具體表現(xiàn)為顯著降低蜷縮行為、自我撓體行為等消極行為,顯著增加環(huán)境探索、自主運(yùn)動(dòng)等積極行為,顯著增加食蟹猴在開窗測驗(yàn)上的探索興趣;氟西汀(FLX)卻需要慢性ig給予17 d才會(huì)表現(xiàn)出抗抑郁行為學(xué)效應(yīng),提示YL-0919在該模型上具有較快速的抗抑郁活性。②在體多通道電生理研究表明單次ip給予氯胺酮(Ket,10 mg·kg-1)在給藥1.5和24 h后均可顯著減少大鼠在強(qiáng)迫游泳測驗(yàn)上的不動(dòng)時(shí)間,表現(xiàn)出顯著的抗抑郁行為學(xué)效應(yīng),同時(shí)可以顯著增加錐體神經(jīng)元的放電活動(dòng)并顯著抑制中間神經(jīng)元的放電活動(dòng),對mPFC中的E∶I平衡產(chǎn)生快速調(diào)節(jié)作用;而在相同實(shí)驗(yàn)條件下,F(xiàn)LX卻無上述作用。進(jìn)一步研究發(fā)現(xiàn)慢性ig給予FLX(10 mg·kg-1)21 d,末次給藥24 h后表現(xiàn)出顯著的抗抑郁行為學(xué)效應(yīng),同時(shí)對mPFC中E∶I平衡產(chǎn)生顯著性的調(diào)節(jié)作用。③ YL-0919(2.5 mg·kg-1,ip)可顯著增加mPFC中錐體神經(jīng)元的放電活動(dòng)并抑制中間神經(jīng)元的放電活動(dòng),可對mPFC中E∶I平衡產(chǎn)生顯著性的調(diào)節(jié)作用,而5-HT1A受體拮抗劑WAY-100635(0.3 mg·kg-1,ip)可顯著阻斷YL-0919 對mPFC中E∶I功能性平衡的影響,而5-HT6受體拮抗劑SB271046(10 mg·kg-1,ip)不能阻斷 YL-0919對mPFC中E∶I平衡的影響,提示5-HT1A受體在YL-0919對mPFC中E∶I平衡的影響中發(fā)揮著重要的調(diào)節(jié)作用。④在體誘導(dǎo)神經(jīng)場電位記錄結(jié)果表明,ig給予YL-0919(2.5 mg·kg-1)7 d即可顯著增加大鼠海馬齒狀回群峰電位幅值,WAY-100635(0.3 mg·kg-1,ip)可顯著阻斷YL-0919對海馬神經(jīng)可塑性的影響,SB271046(10 mg·kg-1,ip)不能阻斷YL-0919對海馬神經(jīng)可塑性的影響,提示5-HT1A受體在YL-0919對海馬神經(jīng)可塑性的影響中發(fā)揮著重要的調(diào)節(jié)作用。結(jié)論YL-0919在食蟹猴慢性應(yīng)激模型上具有快速的抗抑郁行為學(xué)效應(yīng);mPFC中E∶I功能性平衡可能是抗抑郁藥起效的重要限速環(huán)節(jié),YL-0919通過5-HT1A受體快速調(diào)節(jié)mPFC中E∶I平衡并增強(qiáng)海馬神經(jīng)可塑性,這可能是其抗抑郁效應(yīng)快速起效的關(guān)鍵機(jī)制之一。

      關(guān)鍵詞:YL-0919;抑郁癥;E∶I平衡;慢性不可預(yù)知應(yīng)激;快速起效

      通訊作者:張黎明,E-mail:zhanglm0308@163.com;李云峰,E-mail:lyf619@aliyun.com

      T2-5

      基于膠質(zhì)細(xì)胞18 ku轉(zhuǎn)位蛋白的快速起效抗抑郁新藥研究

      張黎明1,姚俊琪1,姚如夢1,劉 暢1,袁 瑾1,李云峰1,2

      (軍事科學(xué)院軍事醫(yī)學(xué)研究院1.毒物藥物研究所,2.軍事認(rèn)知與腦科學(xué)研究所,北京 100850)

      摘要:目的腦內(nèi)18 ku轉(zhuǎn)位蛋白(TSPO)主要位于膠質(zhì)細(xì)胞上,是介導(dǎo)神經(jīng)類固醇(如別孕烯醇酮)合成的關(guān)鍵蛋白。本研究擬在動(dòng)物行為、細(xì)胞和分子水平探討TSPO在精神-認(rèn)知障礙疾病治療中的靶標(biāo)價(jià)值,并研究自主研發(fā)的TSPO激活劑YL-IPA08的抗精神-認(rèn)知障礙的藥理學(xué)作用及其機(jī)制。方法①采用TSPO敲除小鼠進(jìn)行YL-IPA08給藥后的強(qiáng)迫游泳實(shí)驗(yàn)、懸尾實(shí)驗(yàn)和高架十字迷宮實(shí)驗(yàn)檢測抗抑郁和抗焦慮行為效應(yīng)。②采用PCPA和5,7-DHT等藥理學(xué)技術(shù)干擾大腦5-HT系統(tǒng)后,探索5-HT系統(tǒng)在YL-IPA08抗抑郁和抗焦慮中的作用。③采用大鼠慢性不可預(yù)知應(yīng)激模型、小鼠慢性束縛應(yīng)激模型和小鼠皮質(zhì)酮飲水模型,以糖水偏好實(shí)驗(yàn)檢測糖水偏嗜度這一核心指標(biāo)評價(jià)YLIPA08抗抑郁起效時(shí)程。采用Western印跡以及高爾基染色進(jìn)行相應(yīng)的神經(jīng)生物學(xué)機(jī)制研究。④用大鼠在體多通道電生理記錄研究YL-IPA08對內(nèi)側(cè)前額皮層(mPFC)部位谷氨酸能錐體神經(jīng)元和γ-氨基丁酸(GABA)能中間神經(jīng)元放電頻率的影響。結(jié)果①TSPO敲除可取消YL-IPA08的抗抑郁、抗焦慮效應(yīng),進(jìn)一步確證了YL-IPA08的靶標(biāo)特異性。②5-HT系統(tǒng)被干擾之后可以阻斷YL-IPA08的行為學(xué)效應(yīng),進(jìn)一步微透析實(shí)驗(yàn)研究發(fā)現(xiàn),YL-IPA08可以快速增加大鼠海馬細(xì)胞間隙5-HT水平,而TSPO拮抗劑PK11195可以完全阻斷YL-IPA08誘導(dǎo)的海馬5-HT升高,上述結(jié)果提示5-HT系統(tǒng)的完整性可能參與了YL-IPA08的抗抑郁、抗焦慮效應(yīng)。③YL-IPA08給藥1周可顯著發(fā)揮抗抑郁效應(yīng),而氟西汀則需3周,提示YL-IPA08具有快速起效的優(yōu)勢。YL-IPA08的快速抗抑郁效應(yīng)可能與快速增加mPFC神經(jīng)元樹突復(fù)雜性有關(guān)。④YL-IPA08單次給藥使大鼠mPFC中谷氨酸能錐體神經(jīng)元放電頻率增加,GABA能中間神經(jīng)元放電頻率降低,提示YL-IPA08抗抑郁抗焦慮效應(yīng)可能與興奮錐體神經(jīng)元,抑制中間神經(jīng)元有關(guān),即YL-IPA08可快速調(diào)節(jié)mPFC的E∶I功能性再平衡,這可能是其快速起效的重要啟動(dòng)機(jī)制。結(jié)論①TSPO選擇性配體YL-IPA08在多種抑郁動(dòng)物模型上具有快速起效的抗抑郁作用(1周);② YL-IPA08快速調(diào)節(jié)mPFC中谷氨酸錐體神經(jīng)元和GABA中間神經(jīng)元的E∶I功能性再平衡,進(jìn)而快速增強(qiáng)錐體神經(jīng)元樹突復(fù)雜性,這可能是其快速抗抑郁的重要機(jī)制。本研究為前期課題組提出的“單胺(5-HT)-谷氨酸/GABA長神經(jīng)環(huán)路”候選假說提供了有力證據(jù),為快速起效抗抑郁新藥的發(fā)現(xiàn)及靶標(biāo)機(jī)制研究提供了有益的借鑒。

      關(guān)鍵詞:18 ku轉(zhuǎn)位蛋白;抑郁癥;焦慮癥;神經(jīng)可塑性;YL-IPA08

      通訊作者:李云峰,E-mail:lyf619@aliyun.com

      T2-6

      大麻二酚通過Sigma1R/Akt/GSK3 β/CREB信號通路減輕甲基苯丙胺誘導(dǎo)的大鼠復(fù)吸和PC12細(xì)胞神經(jīng)毒性

      劉 柳1,李 娟2,王 嬋1,許 悅1,楊根夢1,林紓丞1,張樹威1,澹 憶1,張慧潔1,王浩偉1,曾曉鋒1,劉建興1

      (昆明醫(yī)科大學(xué)1.法醫(yī)學(xué)院,2.基礎(chǔ)醫(yī)學(xué)院,云南 昆明 650500)

      摘要:目的甲基苯丙胺(METH),俗稱“冰毒”,是一種精神依賴性極強(qiáng)的新型合成毒品,在全球范圍內(nèi)被廣泛濫用。防治METH濫用主要通過減少吸食者對METH渴求和防止METH復(fù)吸發(fā)生2條途徑。大麻二酚(CBD)因其在治療藥物依賴和防止藥物復(fù)吸方面具有潛在藥理學(xué)價(jià)值而受到廣泛關(guān)注。在我們的前期研究中,通過建立METH誘導(dǎo)的大鼠條件位置偏愛(CPP)模型來研究CBD的藥理學(xué)作用。方法和結(jié)果 研究證實(shí),ip給予CBD 20,40和80 mg·kg-1可顯著減少M(fèi)ETH(2 mg·kg-1)誘導(dǎo)的大鼠CPP,且該作用可能涉及CBD對Sigma1R/Akt/GSK3β/CREB信號通路的調(diào)節(jié)。在此基礎(chǔ)上,本研究繼續(xù)采用METH誘導(dǎo)的大鼠CPP模型來探討CBD在METH戒斷和復(fù)吸中的作用。研究發(fā)現(xiàn),對大鼠ip給予METH 2 mg·kg-1可以誘導(dǎo)CPP產(chǎn)生,待大鼠經(jīng)歷5 d的METH戒斷后,再次注射METH 0.5 mg·kg-1以恢復(fù)METH誘導(dǎo)的CPP;而在戒斷期內(nèi)連續(xù)5 d注射CBD 20,40和80 mg·kg-1不僅能促進(jìn)大鼠CPP滅絕,還能有效防止METH誘導(dǎo)的大鼠CPP恢復(fù),并且Sigma1R和Akt/GSK3β/CREB信號通路的表達(dá)水平變化參與該過程。為了進(jìn)一步驗(yàn)證上述調(diào)控機(jī)制,本研究建立了體外PC12細(xì)胞模型,在METH暴露前,分別采用CBD、Sigma1R拮抗劑BD1047、Sigma1R基因敲低的慢病毒預(yù)處理細(xì)胞,隨后使用Western印跡法檢測Sigma1R,Akt,GSK3β,CREB等相關(guān)蛋白的表達(dá)。結(jié)果表明,CBD可減輕METH誘導(dǎo)的PC12細(xì)胞神經(jīng)毒性并且通過拮抗Sigma1R的功能或表達(dá)抑制METH對Akt/GSK3β/CREB信號通路的激活作用。結(jié)論CBD可能是一種防止METH復(fù)吸和神經(jīng)毒性的藥物,而Sigma1R可能是CBD藥理學(xué)作用的重要靶點(diǎn),但其具體的干預(yù)作用和機(jī)制還需要更多的研究證明。

      關(guān)鍵詞:大麻二酚;甲基苯丙胺;復(fù)吸;神經(jīng)毒性;Sigma1R

      基金項(xiàng)目:國家自然科學(xué)基金地區(qū)基金項(xiàng)目(8196070173);云南省科技廳-昆明醫(yī)科大學(xué)聯(lián)合專項(xiàng)(202001AY070001015)

      通訊作者:曾晚鋒,E-mail:zxf2004033@163.com;劉建興,E-mail:444319658@qq.com

      T2-7

      甲基苯丙胺和HIV-Tat蛋白通過Nrf2/NQ01/HO-1通路協(xié)同誘導(dǎo)小膠質(zhì)細(xì)胞氧化應(yīng)激和自噬

      楊根夢,沈?qū)氂?,?嬋,許 悅,林紓丞,洪仕君,李利華,曾曉鋒

      (昆明醫(yī)科大學(xué)法醫(yī)學(xué)院,云南 昆明 650500)

      摘要:目的探討Nrf2通路在甲基苯丙胺(METH)和HIV-Tat蛋白協(xié)同誘導(dǎo)小膠質(zhì)細(xì)胞自噬中的作用。方法以濫用METH和濫用METH同時(shí)感染HIV死亡者,C57BL/6J小鼠,BV2細(xì)胞和原代小膠質(zhì)細(xì)胞為研究對象。分別通過激活Nrf2和敲除或沉默Nrf2后,觀察人體腦組織,小鼠紋狀體組織和小膠質(zhì)細(xì)胞內(nèi)LC3-Ⅱ,Beclin1,ATG5,ATG7,T-Nrf2,N-Nrf2,HO-1和NQ01的變化和小膠質(zhì)細(xì)胞內(nèi)氧化應(yīng)激水平。結(jié)果人體腦組織:METH組和METH+AIDS組紋狀體內(nèi)小膠質(zhì)細(xì)胞被激活,同時(shí)Nrf2和LC3-Ⅱ表達(dá)水平明顯升高,且METH+AIDS組表達(dá)水平比METH組高。體內(nèi)實(shí)驗(yàn):METH和HIV-Tat蛋白協(xié)同激活紋狀體內(nèi)小膠質(zhì)細(xì)胞并誘導(dǎo)自噬水平明顯升高。激活Nrf2后,兩者協(xié)同誘導(dǎo)自噬水平降低。敲除Nrf2后,兩者協(xié)同誘導(dǎo)自噬水平升高。與WT小鼠相比,Nrf2-KO后,自噬表達(dá)水平更高。體外實(shí)驗(yàn):METH和HIV-Tat蛋白協(xié)同誘導(dǎo)小膠質(zhì)細(xì)胞自噬和氧化應(yīng)激水平明顯升高,且具有METH和HIV-Tat蛋白濃度依賴性。激活Nrf2后,兩者協(xié)同誘導(dǎo)自噬和氧化應(yīng)激水平降低。沉默Nrf2后,兩者協(xié)同誘導(dǎo)自噬和氧化應(yīng)激水平明顯升高。結(jié)論濫用METH同時(shí)感染HIV對激活小膠質(zhì)細(xì)胞和Nrf2以及誘導(dǎo)自噬具有協(xié)同作用。METH和HIVTat蛋白可協(xié)同誘導(dǎo)小膠質(zhì)細(xì)胞自噬和氧化應(yīng)激,Nrf2通路可通過調(diào)節(jié)氧化應(yīng)激來調(diào)控兩者協(xié)同誘導(dǎo)的自噬作用。該研究可為毒品濫用的HIV感染者尋找有效的藥物干預(yù)靶點(diǎn)提供理論依據(jù)和新思路。

      關(guān)鍵詞:甲基苯丙胺;氧化應(yīng)激;自噬

      通訊作者:曾曉鋒,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

      尼古丁對大腦獎(jiǎng)賞系統(tǒng)的調(diào)控作用及腦源性神經(jīng)營養(yǎng)因子相關(guān)機(jī)制

      常 芮,黃澤怡,陳運(yùn)凡,譚日東,廖海琳,譚思杰

      (南華大學(xué)衡陽醫(yī)學(xué)院,湖南 衡陽 421001)

      摘要:目的煙草依賴是嚴(yán)重的公共衛(wèi)生問題,尼古丁是煙草中的主要精神活性物質(zhì)。本研究運(yùn)用顱內(nèi)自我刺激技術(shù)(ICSS)和條件性位置偏愛(CPP)模型,研究尼古丁對大腦獎(jiǎng)賞系統(tǒng)功能的影響,并進(jìn)一步探討腦源性神經(jīng)營養(yǎng)因子(BDNF)相關(guān)信號通路在尼古丁成癮中的作用。方法通過立體定位手術(shù)在大鼠腹側(cè)被蓋區(qū)-伏隔核通路上方植入刺激微電極,sc給予不同劑量的尼古丁,檢測ICSS的電流閾值以衡量大腦獎(jiǎng)賞系統(tǒng)的功能。結(jié)果尼古丁≥0.1 mg·kg-1能顯著降低ICSS的閾值;此外,與雄性大鼠相比,雌性大鼠的閾值下降更顯著,表明尼古丁的獎(jiǎng)賞效應(yīng)存在性別差異。在尼古丁戒斷時(shí),ICSS電流閾值顯著升高,表明尼古丁戒斷能降低大腦獎(jiǎng)賞系統(tǒng)的功能。在CPP實(shí)驗(yàn)中發(fā)現(xiàn),尼古丁0.5 mg·kg-1組小鼠的CPP得分顯著高于生理鹽水組,表明尼古丁0.5 mg·kg-1能誘導(dǎo)小鼠形成CPP,而BDNF高親和力受體TrkB的阻斷劑可以降低小鼠CPP得分。單次尼古丁0.5 mg·kg-1注射對小鼠伏隔核和前額葉皮質(zhì)BDNF表達(dá)無影響,但可顯著降低海馬BDNF的表達(dá);連續(xù)7 d給小鼠注射尼古丁0.5 mg·kg-1,停藥2 h后,海馬BDNFTrkB信號通路表達(dá)顯著上調(diào),下游AKT表達(dá)顯著升高,且Hip CA3和DG內(nèi)GluA1蛋白表達(dá)增加。結(jié)論急性尼古丁能激活大腦獎(jiǎng)賞系統(tǒng),而尼古丁戒斷時(shí)大腦獎(jiǎng)賞系統(tǒng)功能下調(diào);尼古丁對獎(jiǎng)賞系統(tǒng)的調(diào)控作用可能與BDNF-TrkB信號通路有關(guān)。

      關(guān)鍵詞:尼古?。荒X源性神經(jīng)營養(yǎng)因子;獎(jiǎng)賞系統(tǒng)

      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.RESULTS①M(fèi)AM 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

      海馬神經(jīng)發(fā)生對甲苯丙胺成癮小鼠成癮記憶消退及再喚起的作用

      車曉航,白亦君,蔡佳伶,吳春福,楊靜玉

      (沈陽藥科大學(xué)生命科學(xué)與生物制藥學(xué)院,遼寧 沈陽 110016)

      摘要:目的最近研究表明海馬神經(jīng)發(fā)生(HN)可能對記憶產(chǎn)生雙向調(diào)節(jié)作用,即促進(jìn)HN不但能夠促進(jìn)記憶的形成,而且同時(shí)可能誘導(dǎo)記憶的遺忘。目前,包括嗎啡、可卡因等藥物所導(dǎo)致的成癮記憶被證明可能與HN有關(guān),但HN在甲苯丙胺(METH)相關(guān)成癮記憶中的作用還有待研究。方法通過METH誘導(dǎo)的條件性位置偏愛模型模擬小鼠對METH的成癮記憶,然后進(jìn)一步通過給予7,8-二羥基黃酮(DHF)與替莫唑胺(TMZ)分別在METH成癮記憶獲得前及獲得后促進(jìn)或抑制HN,同時(shí)結(jié)合神經(jīng)發(fā)生標(biāo)志物雙皮質(zhì)素(DCX)及5-溴脫氧尿嘧啶核苷(BrdU)考察HN在METH成癮記憶中的作用。結(jié)果條件性位置偏愛實(shí)驗(yàn)結(jié)果表明,METH能夠顯著誘導(dǎo)小鼠的成癮記憶;BrdU和DCX的免疫組化及免疫熒光結(jié)果表明DHF及TMZ分別能夠促進(jìn)或抑制小鼠HN。在METH誘導(dǎo)小鼠成癮記憶獲得后,通過DHF促進(jìn)HN能夠顯著縮短其成癮記憶的消退時(shí)間并抑制成癮記憶的再喚起程度,而通過TMZ抑制HN則起到相反的結(jié)果;而在METH誘導(dǎo)成癮記憶獲得前,無論給予DHF或TMZ,均能延長小鼠成癮記憶的消退時(shí)間,并增加其再喚起程度。結(jié)論在METH成癮記憶獲得后,促進(jìn)HN能夠顯著誘導(dǎo)METH成癮記憶的遺忘,提示基于干預(yù)HN的方式可能為METH成癮行為的治療提供新的手段與策略。

      關(guān)鍵詞:甲苯丙胺;成癮記憶;神經(jīng)發(fā)生

      通訊作者:楊靜玉,E-mail:yangjingyu2006@gmail.com;吳春福,E-mail:wucf@syphu.edu.cn

      T2-15

      梔子厚樸湯抗抑郁作用及機(jī)制

      邢 航,張 闊

      (沈陽藥科大學(xué)生藥教研室,遼寧 沈陽 110016)

      摘要:目的抑郁癥是由各種原因引起以抑郁為主要特征的心境或情感性精神障礙,具有患病率高、復(fù)發(fā)率高、致殘率高等特點(diǎn)。梔子厚樸湯出自于《傷寒論》,由梔子、枳實(shí)和厚樸3味藥組成。主治傷寒下后,心煩腹?jié)M,臥起不安?,F(xiàn)代研究表明,梔子厚樸湯具有抗抑郁的作用。本文研究梔子厚樸湯對慢性不可預(yù)見溫和應(yīng)激所致抑郁大鼠的保護(hù)作用及相關(guān)機(jī)制。方法采用成年雄性SD大鼠,通過慢性不可預(yù)見溫和刺激建立大鼠抑郁模型,連續(xù)ig給予梔子厚樸湯(3.66,7.32和14.64 g·kg-1)21 d,結(jié)合大體打分、糖水偏愛、強(qiáng)迫游泳和曠場等多種行為學(xué)方法,明確梔子厚樸湯對慢性不可預(yù)見溫和應(yīng)激抑郁大鼠的保護(hù)作用;進(jìn)一步利用酶聯(lián)免疫和免疫熒光等技術(shù),研究梔子厚樸湯對慢性不可預(yù)見溫和應(yīng)激抑郁大鼠的保護(hù)機(jī)制。結(jié)果行為學(xué)結(jié)果表明,與慢性不可預(yù)見溫和刺激模型組相比,梔子厚樸湯給藥組大鼠外貌評分顯著降低,糖水偏愛顯著增加,游泳不動(dòng)時(shí)間顯著縮短,在曠場中心區(qū)域和非邊緣區(qū)域停留時(shí)間顯著增加,在邊緣區(qū)域停留時(shí)間顯著減少。神經(jīng)內(nèi)分泌結(jié)果表明,梔子厚樸湯給藥組大鼠血清皮質(zhì)酮抑制率顯著增加,血清皮質(zhì)酮水平顯著降低。免疫組織化學(xué)結(jié)果表明梔子厚樸湯給藥組大鼠海馬中腦源性神經(jīng)營養(yǎng)因子(BDNF)表達(dá)顯著增加,海馬齒狀回DCX和BrdU/NeuN陽性細(xì)胞數(shù)顯著增加。結(jié)論梔子厚樸湯連續(xù)灌胃21 d對慢性不可預(yù)見溫和應(yīng)激所致抑郁大鼠具有保護(hù)作用,其機(jī)制可能與增加海馬BDNF和增強(qiáng)海馬神經(jīng)發(fā)生有關(guān)。

      關(guān)鍵詞:梔子厚樸湯;抑郁;腦源性神經(jīng)營養(yǎng)因子;神經(jīng)發(fā)生

      通訊作者:張 闊,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

      賴右苯丙胺對前額葉皮質(zhì)執(zhí)行功能的影響

      陳健民,王志媛,吳世軒,吳 寧,李 錦

      (軍事科學(xué)院軍事醫(yī)學(xué)研究院毒物藥物研究所,北京 100850)

      摘要:目的研究賴右苯丙胺(Lis)與傳統(tǒng)認(rèn)知增強(qiáng)劑右旋苯丙胺(Amp)對前額葉皮質(zhì)執(zhí)行功能(工作記憶、持續(xù)性主動(dòng)注意、抑制控制和辨別能力)的藥效學(xué)差異,為把Lis作為安全高效且低成癮的認(rèn)知增強(qiáng)候選化合物提供研究依據(jù)。方法建立反映空間工作記憶的Y迷宮連續(xù)自發(fā)交替任務(wù),反映持續(xù)性主動(dòng)注意及抑制控制的五項(xiàng)選擇連續(xù)反應(yīng)時(shí)間任務(wù),反映空間信息辨別的Y迷宮延遲自發(fā)交替任務(wù)以及視覺信息辨別的視覺辨別學(xué)習(xí)任務(wù),在此基礎(chǔ)上研究Lis與Amp的藥效學(xué)差異。結(jié)果在Y迷宮連續(xù)自發(fā)交替任務(wù)中,Lis 0.2~1.5 mg·kg-1和Amp均不影響大鼠自發(fā)交替比,Lis 4.5 mg·kg-1顯著提高大鼠自發(fā)交替比率(P<0.05),而Amp 4.5 mg·kg-1顯著降低大鼠自發(fā)交替比率(P<0.01),進(jìn)一步增加劑量至13.5 mg·kg-1,Amp顯著抑制大鼠自發(fā)交替比(P<0.01),而賴右苯丙胺對此無顯著影響。在5項(xiàng)選擇連續(xù)反應(yīng)時(shí)間任務(wù)中,在長時(shí)程高注意任務(wù)情況下,Amp 0.05~0.5 mg·kg-1及 Lis 0.1~0.3 mg·kg-1均無提高正確率的作用,而Lis 1.0 mg·kg-1則顯著提高大鼠反應(yīng)正確率(P<0.05),縮短正確反應(yīng)潛伏期(P<0.05)。在高沖動(dòng)狀態(tài)下,Amp 0.5 mg·kg-1顯著增加大鼠沖動(dòng)行為(P<0.01),而Lis 1.0 mg·kg-1并無明顯影響。在空間信息辨別的Y迷宮延遲自發(fā)交替任務(wù),Amp 0.2~4.5 mg·kg-1均無增加大鼠新臂進(jìn)臂次數(shù)和停留時(shí)間百分比,而Lis 4.5 mg·kg-1顯著提高新臂進(jìn)臂次數(shù)(P<0.05)和停留時(shí)間百分比(P<0.01),增加劑量至13.5 mg·kg-1后,Amp顯著降低該指標(biāo)(P<0.01)而Lis對此無明顯影響。在視覺辨別學(xué)習(xí)任務(wù)中,Lis 1.0 mg·kg-1急性給藥顯著提高大鼠的辨別正確率(P<0.05),而Amp 0.05~0.5 mg·kg-1無明顯影響。結(jié)論相比于Amp,Lis能提高空間工作記憶,在長時(shí)程高強(qiáng)度注意力任務(wù)中能提高持續(xù)性主動(dòng)注意,急性給藥能促進(jìn)視覺信息的辨別學(xué)習(xí)能力,提示Lis能提高前額葉皮質(zhì)的執(zhí)行功能,且安全性較好。本研究為把賴右苯丙胺能作為安全高效且低成癮的認(rèn)知增強(qiáng)候選化合物提高研究基礎(chǔ)和研究依據(jù)。

      關(guān)鍵詞:賴右苯丙胺;右旋苯丙胺;前額葉皮質(zhì);執(zhí)行功能

      通訊作者:吳 寧,E-mail:wuning7671@126.com;李 錦,E-mail:jinli9802@163.com

      專題3:腦卒中與腦損傷

      T3-1

      星形膠質(zhì)細(xì)胞對腦缺血再灌注損傷的保護(hù)作用

      耿宇涵1,張丹參1,2,李 煒1

      (1.河北北方學(xué)院藥學(xué)系,河北省神經(jīng)藥理學(xué)重點(diǎn)實(shí)驗(yàn)室,河北 張家口 075000;2.河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:在缺血性腦損傷中,神經(jīng)元和星形膠質(zhì)細(xì)胞會(huì)迅速耗盡能量和氧氣,這個(gè)過程最終導(dǎo)致神經(jīng)元失去存活的能力,發(fā)生不可逆轉(zhuǎn)的組織損傷和神經(jīng)損傷。星形膠質(zhì)細(xì)胞在中樞神經(jīng)系統(tǒng)中有著十分重要的作用,如調(diào)控神經(jīng)系統(tǒng)的發(fā)育、腦的物質(zhì)代謝和血流。在正常情況下,正常健康的星形膠質(zhì)細(xì)胞會(huì)保護(hù)神經(jīng)元,避免氧化應(yīng)激和興奮性毒性。星形膠質(zhì)細(xì)胞擁有十分豐富的縫隙連接,且有研究表明,在傳導(dǎo)細(xì)胞的凋亡信號和細(xì)胞間信息交流時(shí),縫隙連接有很重要的作用。連接蛋白是縫隙連接的重要組成部分。在大腦中,縫隙連接作為細(xì)胞間通信(GJIC)可能允許能量代謝物和危險(xiǎn)分子的傳遞。星形膠質(zhì)細(xì)胞GJIC和神經(jīng)元共培養(yǎng)可增強(qiáng)神經(jīng)元細(xì)胞抵抗氧化應(yīng)激的能力;阻塞星形膠質(zhì)細(xì)胞間隙連接,增加共培養(yǎng)時(shí)神經(jīng)元對谷氨酸細(xì)胞毒性的敏感性。

      關(guān)鍵詞:星形膠質(zhì)細(xì)胞;腦缺血;保護(hù)作用

      基金項(xiàng)目:河北省重點(diǎn)研發(fā)計(jì)劃項(xiàng)目生物醫(yī)藥專項(xiàng)(20372509D)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T3-2

      星形膠質(zhì)細(xì)胞拯救缺血性腦卒中

      蘇曉梅1,張丹參1,2

      (1.河北醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院藥理學(xué)教研室,河北 石家莊 050017;2.河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:星形膠質(zhì)細(xì)胞(AS)是中樞神經(jīng)系統(tǒng)內(nèi)含量最豐富的神經(jīng)膠質(zhì)細(xì)胞,約占腦總體積的50%,其數(shù)量約為神經(jīng)元的10倍。AS與神經(jīng)元之間存在廣泛而復(fù)雜的信息傳遞,以直接或間接作用的方式與神經(jīng)元發(fā)生聯(lián)系。在生理情況下,AS對神經(jīng)元發(fā)揮營養(yǎng)支持的作用,可參與神經(jīng)元的糖、脂肪和體液代謝過程。在缺血性腦卒中發(fā)生時(shí),由于AS線粒體內(nèi)可以儲(chǔ)存大量糖原,使得其對低氧環(huán)境具有彈性和適應(yīng)性,AS可以通過自身生物能量和線粒體動(dòng)力學(xué)的變化為損傷神經(jīng)元提供能量支持。不僅如此,在缺血性腦卒中發(fā)生時(shí),AS可以通過隧道納米管或細(xì)胞外囊泡與周圍的受損神經(jīng)元進(jìn)行線粒體交換,釋放功能性線粒體。同時(shí),神經(jīng)元也可以釋放受損線粒體并將其轉(zhuǎn)移至AS內(nèi),進(jìn)行內(nèi)吞和降解,實(shí)現(xiàn)線粒體的循環(huán)利用,使得線粒體在健康細(xì)胞和受損細(xì)胞之間發(fā)生串?dāng)_。此外,AS的終足與中樞神經(jīng)系統(tǒng)中的微動(dòng)脈和毛細(xì)血管接觸,成為神經(jīng)元網(wǎng)絡(luò)和血管網(wǎng)絡(luò)之間的橋梁,通過影響血管直徑來調(diào)節(jié)腦血流。綜上,AS可以作為治療缺血性腦卒中的重要靶點(diǎn)之一,改善缺血性腦卒中的臨床治療效果。

      關(guān)鍵詞:星形膠質(zhì)細(xì)胞;缺血性腦卒中;生物能量;線粒體轉(zhuǎn)移;腦血流

      基金項(xiàng)目:河北省重點(diǎn)研發(fā)計(jì)劃項(xiàng)目生物醫(yī)藥專項(xiàng)(20372509D)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T3-3

      姜黃素對腦缺血再灌注損傷的神經(jīng)保護(hù)作用及其機(jī)制

      謝佳佳,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:姜黃素是從姜黃根莖中提取出來的一種脂溶性多酚類化合物,是姜黃發(fā)揮藥理作用的主要活性成分。研究表明,姜黃素可減輕多種器官的缺血再灌注損傷,容易透過血腦屏障,可用于治療中樞神經(jīng)系統(tǒng)疾病。腦缺血再灌注損傷是在發(fā)生缺血性腦卒中后的短時(shí)間內(nèi),壞死腦組織周圍的缺血神經(jīng)元細(xì)胞在恢復(fù)血供時(shí),發(fā)生加重缺血級聯(lián)反應(yīng),造成腦組織更為嚴(yán)重的再灌注損傷。國內(nèi)外研究表明,姜黃素可通過抑制腦缺血再灌注損傷的多種病理機(jī)制而發(fā)揮顯著的神經(jīng)保護(hù)作用,本文主要從其作用機(jī)制方面進(jìn)行總結(jié)。①激活SOD,加速體內(nèi)超氧陰離子和羥自由基等ROS的清除。姜黃素作為一種強(qiáng)抗氧化劑,主要通過激活SOD,GSHPx,CAT和GST等抗氧化酶,從而穩(wěn)定大腦的抗氧化酶系統(tǒng),加速體內(nèi)自由基的清除;②降低IL-6和TNF-α的組織濃度,下調(diào)NF-κB的表達(dá),抑制炎癥反應(yīng)。姜黃素可以通過介導(dǎo)TLR-4/P38/MAPK途徑,降低IL-6,TNF-α和NOS等炎癥因子的水平,減少炎癥反應(yīng),從而改善神經(jīng)功能;③抑制凋亡。姜黃素可通過激活絲裂原活化蛋白激酶信號通路,降低胱天蛋白酶3活性,抑制凋亡蛋白表達(dá),最終抑制腦缺血再灌注損傷誘導(dǎo)的細(xì)胞凋亡;④抑制內(nèi)質(zhì)網(wǎng)應(yīng)激。姜黃素可通過GADD153和胱天蛋白酶12途徑抑制內(nèi)質(zhì)網(wǎng)應(yīng)激,CADD153作為一種信號分子,通過影響細(xì)胞內(nèi)的Ca2+代謝和下調(diào)Bcl-2等多種途徑抑制內(nèi)質(zhì)網(wǎng)應(yīng)激,促進(jìn)神經(jīng)功能恢復(fù)。綜上所述,姜黃素能通過多種分子機(jī)制抑制腦缺血再灌注損傷,發(fā)揮神經(jīng)保護(hù)作用,具有補(bǔ)充現(xiàn)有缺血性腦卒中治療方法的潛力。

      關(guān)鍵詞:姜黃素;腦缺血再灌注損傷;神經(jīng)保護(hù)

      基金項(xiàng)目:河北省重點(diǎn)研發(fā)計(jì)劃項(xiàng)目生物醫(yī)藥專項(xiàng)(20372509D)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T3-4

      PI3K/Akt信號通路對腦缺血再灌注損傷的作用機(jī)制

      袁鑫茹,張丹參,景永帥

      (河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:PI3K/Akt信號通路是經(jīng)典的抗凋亡和促存活的信號轉(zhuǎn)導(dǎo)通路之一,在腦卒中、腫瘤、糖尿病和抑郁等疾病的發(fā)生中具有重要作用。腦缺血是目前世界上繼心臟病和癌癥之后的第三大致死性疾病,其最常用的治療手段是再灌注,但可能引起缺血細(xì)胞損傷加重,即腦缺血再灌注損傷(CIRI),其機(jī)制非常復(fù)雜。大量研究表明,通過激活PI3K/Akt信號通路,可減輕CIRI。因此,PI3K/Akt信號通路在CIRI中發(fā)揮重要作用,其機(jī)制主要有以下4個(gè)方面:①抑制細(xì)胞凋亡。CIRI后多數(shù)受損細(xì)胞最終會(huì)死亡或凋亡。據(jù)報(bào)道,PI3K/Akt信號通路可通過調(diào)節(jié)mTOR和Bcl-2表達(dá),進(jìn)而控制凋亡和促凋亡蛋白的合成。此外,PI3K/Akt信號通路還可減少線粒體介導(dǎo)的細(xì)胞凋亡;②促進(jìn)腦血管新生。血管內(nèi)皮生長因子(VEGF)能夠促進(jìn)神經(jīng)元和新生血管的生成,激活VEGF介導(dǎo)的PI3K/Akt信號通路可促進(jìn)腦血管的新生;③促進(jìn)神經(jīng)生長因子釋放。大腦周圍神經(jīng)損傷后神經(jīng)營養(yǎng)因子釋放減少,神經(jīng)生長因子與酪氨酸激酶結(jié)合后激活PI3K/Akt信號通路,抑制神經(jīng)元的損傷,從而促使神經(jīng)營養(yǎng)因子釋放增加;④抑制自噬。缺血損傷后可導(dǎo)致自噬過度活化,細(xì)胞損傷死亡,PI3K/Akt信號通路的激活可抑制過度活化的自噬水平,減輕細(xì)胞損傷。本文總結(jié)PI3K/Akt信號通路在CIRI中的作用機(jī)制,為治療CIRI提供參考依據(jù)。

      關(guān)鍵詞:PI3K/Akt信號通路;腦缺血再灌注損傷;作用機(jī)制

      基金項(xiàng)目:河北省重點(diǎn)研發(fā)計(jì)劃項(xiàng)目生物醫(yī)藥專項(xiàng)(20372509D)

      通訊作者:張丹參,E-mail:zhangds2011@126.com

      T3-5

      TIMP1通過與CD63/整合素β1復(fù)合物相互作用并調(diào)節(jié)下游FAK/RhoA信號保護(hù)血腦屏障

      唐婧姝,康鈺瑩,黃龍艦,吳 鐳,彭 英

      (中國醫(yī)學(xué)科學(xué)院&北京協(xié)和醫(yī)學(xué)院藥物研究所,北京 100050)

      摘要:目的創(chuàng)傷性腦外傷(TBI)是青壯年致殘和死亡的首位原因。TBI后血腦屏障(BBB)明顯受損,而改善BBB功能障礙、維持其完整性可能是TBI治療的潛在策略。TIMP1參與調(diào)節(jié)血管重塑以及穩(wěn)定血管通透性,但分子機(jī)制尚不明確。本研究采用整體和離體TBI模型,探討TIMP1對TBI導(dǎo)致BBB功能失調(diào)的改善作用及具體調(diào)控機(jī)制,以期為TBI的臨床治療和藥物研發(fā)提供更多策略。方法首先,建立TBI模型小鼠,給予rTIMP1,采用平衡木和轉(zhuǎn)棒實(shí)驗(yàn)評估神經(jīng)功能;采用伊文思蘭實(shí)驗(yàn)評估BBB通透性。其次,利用缺氧和外源加入IL-1β,在人腦微血管內(nèi)皮細(xì)胞中建立離體TBI模型,分別敲減、過表達(dá)TIMP1或外源給予rTIMP1,應(yīng)用熒光素酶滲漏實(shí)驗(yàn)和TEER實(shí)驗(yàn)檢測細(xì)胞屏障功能,利用生物素標(biāo)記法提取膜蛋白和IF實(shí)驗(yàn)檢測連接復(fù)合體的表達(dá)和定位。在機(jī)制探討部分,比較rTIMP1和Ala-rTIMP1(無MMP抑制活性的rTIMP1)對內(nèi)皮屏障功能的影響。最后采用Co-IP檢測TIMP1與CD63/整合素β1相互作用,利用FAK抑制劑明確TIMP1調(diào)控BBB的關(guān)鍵通路,采用Rhotekin pulldown檢測RhoA活化水平,以及應(yīng)用IF檢測F-actin聚集程度。結(jié)果rTIMP1可以改善TBI小鼠神經(jīng)功能損傷,減少腦組織伊文思藍(lán)滲漏和連接復(fù)合體表達(dá)缺失。敲減TIMP1增加熒光素酶滲漏、降低TEER、破壞連接復(fù)合體的表達(dá)和定位,而給予rTIMP1或過表達(dá)TIMP1均能逆轉(zhuǎn)上述損傷,表明TIMP1是調(diào)控內(nèi)皮屏障完整性的關(guān)鍵分子。rTIMP1和Ala-rTIMP1均能改善TBI小鼠神經(jīng)功能損傷和內(nèi)皮細(xì)胞屏障功能損傷,二者作用無顯著性差異,表明TIMP1對BBB調(diào)控作用可以不依賴于其MMP抑制活性。TIMP1與CD63/整合素β1結(jié)合并激活下游FAK信號轉(zhuǎn)導(dǎo),F(xiàn)AK抑制劑可逆轉(zhuǎn)TIMP1對TBI小鼠血腦屏障的保護(hù)作用以及連接復(fù)合體的調(diào)控作用,表明FAK激活是TIMP1發(fā)揮作用的關(guān)鍵節(jié)點(diǎn)。此外,TIMP1抑制RhoA活化和F-actin解聚,參與維持細(xì)胞骨架穩(wěn)定性。結(jié)論TIMP1與血管內(nèi)皮細(xì)胞表面CD63/整合素β1結(jié)合,以受體介導(dǎo)的細(xì)胞因子活性調(diào)節(jié)RhoA活化、抑制細(xì)胞骨架重構(gòu)從而調(diào)控連接復(fù)合體的表達(dá)和分布,減輕BBB損傷進(jìn)而在TBI中發(fā)揮保護(hù)作用。本研究提示針對TIMP1及其下游通路的干預(yù)可能是TBI治療的潛在策略。

      關(guān)鍵詞:TIMP1;創(chuàng)傷性腦外傷;血腦屏障;連接復(fù)合體

      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

      大麻二酚抗高熱驚厥的作用機(jī)制

      于永洲,張 煒

      (河北醫(yī)科大學(xué)中西醫(yī)結(jié)合研究所,河北 石家莊 050013)

      摘要:目的明確大麻二酚是否對高熱驚厥小鼠具有保護(hù)作用,及其可能的分子機(jī)制和可能的作用靶點(diǎn)。方法應(yīng)用HE染色觀察大麻二酚對高熱驚厥小鼠海馬神經(jīng)元的形態(tài)學(xué)改變;膜片鉗觀察記錄大麻二酚對海馬神經(jīng)元的興奮性的影響。結(jié)果①行為學(xué)實(shí)驗(yàn)結(jié)果表明,大麻二酚能夠顯著延長高熱驚厥小鼠的潛伏期,且具有劑量依賴性。②HE染色結(jié)果顯示,大麻二酚對高熱驚厥誘發(fā)的小鼠海馬CA1區(qū)神經(jīng)元損傷具有保護(hù)作用。③急性海馬腦片膜片鉗實(shí)驗(yàn)結(jié)果顯示,大麻二酚能夠顯著降低小鼠海馬CA1區(qū)錐體細(xì)胞的興奮性,且呈劑量依賴性。結(jié)論大麻二酚具有抑制小鼠高熱驚厥的作用,其機(jī)制可能為通過抑制AMPA受體激活,使得AMPA受體內(nèi)流進(jìn)入細(xì)胞的陽離子減少,繼而降低海馬神經(jīng)元興奮性,從而發(fā)揮對高熱驚厥狀態(tài)下的海馬神經(jīng)元的保護(hù)作用。

      關(guān)鍵詞:大麻二酚;高熱驚厥;作用機(jī)制;作用靶點(diǎn)

      通訊作者:張 煒,E-mail:zhangwei_wz59@163.com

      T3-10

      山藥多糖對腦缺血再灌注損傷的保護(hù)作用機(jī)制

      李朋月1,2,景永帥3,吳蘭芳1,2,鄭玉光1,2

      (1.河北中醫(yī)學(xué)院藥學(xué)院,河北 石家莊 050200;2.河北省中藥炮制技術(shù)創(chuàng)新中心,河北 石家莊 050091;3.河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:山藥是常用藥食兩用資源,含有蛋白質(zhì)、多糖、薯蕷皂苷、尿囊素及微量元素等成分。多糖作為山藥的主要活性成分之一,具有抗腫瘤、降血脂、降血糖、調(diào)節(jié)免疫、抗炎、抗氧化和神經(jīng)保護(hù)等作用。腦缺血再灌注損傷可導(dǎo)致一系列嚴(yán)重的腦疾病,導(dǎo)致死亡或不同程度的殘疾,研究表明,山藥多糖可能通過多種機(jī)制發(fā)揮對腦缺血再灌注損傷的保護(hù)作用。①改善腦組織抗氧化能力。氧化應(yīng)激是腦缺血再灌注損傷的主要原因之一,在氧化應(yīng)激條件下,超氧化物歧化酶(SOD)和谷胱甘肽(GSH)等抗氧化酶和氧化應(yīng)激標(biāo)志物丙二醛(MDA)在神經(jīng)細(xì)胞對活性氧(ROS)的抵抗中起關(guān)鍵作用——誘導(dǎo)細(xì)胞死亡。相關(guān)研究結(jié)果表明,山藥多糖可使細(xì)胞抗氧化劑SOD和GSH含量顯著增高、MDA含量顯著降低。此外,山藥多糖也可以通過調(diào)節(jié)CaMMKβ的表達(dá)進(jìn)而影響Nrf2/HO-1信號通路的激活,減少ROS的積累。②抑制神經(jīng)元細(xì)胞凋亡。當(dāng)缺血再灌注誘導(dǎo)腦組織產(chǎn)生并積累ROS時(shí),這些自由基會(huì)激活線粒體、死亡受體和內(nèi)質(zhì)網(wǎng)應(yīng)激,進(jìn)而誘導(dǎo)神經(jīng)元凋亡。研究發(fā)現(xiàn),山藥多糖能顯著提高BV2細(xì)胞的活性,調(diào)節(jié)BV2細(xì)胞誘導(dǎo)的神經(jīng)細(xì)胞凋亡;另外可調(diào)節(jié)腦缺血再灌注誘導(dǎo)的小膠質(zhì)細(xì)胞凋亡,降低腦梗死面積。③抑制炎性細(xì)胞因子過度表達(dá)。氧化應(yīng)激損傷產(chǎn)生的ROS會(huì)誘導(dǎo)外周免疫系統(tǒng)和中樞神經(jīng)系統(tǒng)的炎癥反應(yīng),從而產(chǎn)生大量的促炎因子如IL-6,IL-1β和TNF-α,促炎因子通過引起血管收縮、微血管阻塞、分泌細(xì)胞毒素、破壞血腦屏障的完整性和促進(jìn)ROS的產(chǎn)生,加劇氧化應(yīng)激損傷,最終導(dǎo)致神經(jīng)細(xì)胞凋亡。山藥多糖能夠顯著降低腦缺血再灌注損傷模型的IL-6,IL-1β和TNF-α的水平,減少炎癥因子對神經(jīng)元的損傷。本文綜述了山藥多糖在神經(jīng)保護(hù)方面的作用及機(jī)制,以期為山藥及其多糖成分的開發(fā)利用提供依據(jù)。

      關(guān)鍵詞:山藥;多糖;腦損傷再灌注;作用機(jī)制

      基金項(xiàng)目:河北省中醫(yī)藥管理局中醫(yī)藥科研計(jì)劃項(xiàng)目(2021104)

      通訊作者:鄭玉光,E-mail:zyg314@163.com

      T3-11

      SMU-X通過Mfn2/JNK通路發(fā)揮抗炎作用對抗實(shí)驗(yàn)性自身免疫性腦脊髓炎疾病進(jìn)程發(fā)展

      李夢凡,汪海濤,徐江平

      (南方醫(yī)科大學(xué)藥學(xué)院,廣東 廣州 510515)

      摘要:目的SMU-X是治療多發(fā)性硬化的潛在藥物,但SMU-X的作用靶點(diǎn)較多,其治療多發(fā)性硬化的具體機(jī)制還未明確。線粒體融合蛋白2(Mfn2)是一種線粒體外膜GTP酶,調(diào)節(jié)線粒體融合,影響內(nèi)質(zhì)網(wǎng)與線粒體的相互作用,并且被認(rèn)為密切參與炎癥小體的形成。本研究旨在確定SMU-X對實(shí)驗(yàn)性自身免疫性腦脊髓炎(EAE)小鼠的治療作用是否與Mfn2有關(guān)。方法在BV2小膠質(zhì)細(xì)胞系上構(gòu)建脂多糖(LPS)刺激模型。采用Western印跡法觀察炎癥相關(guān)蛋白(iNOS和COX-2)變化,ELISA法檢測炎癥因子(IL-6和TNF-α)水平以及線粒體氧化應(yīng)激的變化。Western印跡法觀察Mfn2蛋白水平、炎癥通路蛋白(p38 MAPK和JNK)磷酸化水平的變化。用C57小鼠構(gòu)建EAE模型,進(jìn)行臨床評分,通過Western印跡法檢測髓鞘堿性蛋白(MBP)的表達(dá),采用HE染色和Western印跡法觀察脊髓組織炎癥細(xì)胞浸潤及小膠質(zhì)細(xì)胞激活相關(guān)蛋白的表達(dá)。結(jié)果①LPS處理后,BV2細(xì)胞激活,炎癥因子IL-6和TNF-α及炎癥相關(guān)蛋白iNOS和COX2顯著上調(diào),而給予SMU-X可逆轉(zhuǎn)LPS誘導(dǎo)的BV2細(xì)胞激活以及炎癥因子和蛋白上調(diào)。②LPS處理BV2細(xì)胞后,線粒體膜電位降低,超氧陰離子活性氧的生成明顯上調(diào),線粒體氧化應(yīng)激水平上升,而給予SMU-X后可以改善線粒體氧化應(yīng)激水平。③LPS處理BV2細(xì)胞后,Mfn2表達(dá)下調(diào),P38和JNK磷酸化水平明顯增加,給予SMU-X后可以逆轉(zhuǎn)Mfn2的下調(diào)及P38和JNK磷酸化水平的增高。④敲低Mfn2可對抗SMU-X對LPS誘導(dǎo)的BV2細(xì)胞的抗炎保護(hù)作用。⑤敲低Mfn2可對抗SMU-X對BV2線粒體氧化應(yīng)激水平的降低作用。⑥敲低Mfn2可對抗SMU-X對LPS誘導(dǎo)的BV2細(xì)胞的JNK通路的抑制作用。⑦SMU-X可降低EAE小鼠的臨床評分及脊髓白質(zhì)區(qū)炎癥細(xì)胞浸潤,增加脊髓組織結(jié)構(gòu)蛋白MBP的表達(dá)。⑧EAE小鼠發(fā)病后,小膠質(zhì)細(xì)胞激活標(biāo)志物Iba1和炎癥相關(guān)蛋白COX-2的表達(dá)明顯上調(diào),而給予SMU-X后可以改善這一現(xiàn)象。⑨EAE小鼠發(fā)病后,小鼠脊髓白質(zhì)區(qū)Mfn2的表達(dá)明顯降低,CD11b的表達(dá)明顯升高,給予SMU-X后可以明顯逆轉(zhuǎn)這些變化。結(jié)論SMU-X可通過Mfn2/JNK通路發(fā)揮抗炎作用,從而對抗EAE小鼠疾病進(jìn)程的發(fā)展。

      關(guān)鍵詞:多發(fā)性硬化;SMU-X;炎癥;線粒體融合蛋白2

      基金項(xiàng)目:國家自然科學(xué)基金(81773698);國家自然科學(xué)基金(81974501)

      通訊作者:徐江平,E-mail:jpx@smu.edu.cn

      T3-12

      抑制磷酸二酯酶4通過調(diào)控內(nèi)質(zhì)網(wǎng)應(yīng)激對抗缺血性腦損傷

      許炳鈿,汪海濤,徐江平

      (南方醫(yī)科大學(xué)藥學(xué)院,廣東 廣州 510515)

      摘要:目的抑制磷酸二酯酶4(PDE4)可減少缺血性腦損傷導(dǎo)致的神經(jīng)元死亡,然而機(jī)制還不清楚。本研究旨在確定抑制PDE4對缺血性腦損傷保護(hù)作用的機(jī)制。方法在HT-22細(xì)胞及原代皮質(zhì)神經(jīng)元上構(gòu)建OGD模型,Western印跡法觀察相關(guān)蛋白水平的變化,Co-IP實(shí)驗(yàn)觀察肌醇依賴酶1α(IRE1α)以及腫瘤壞死因子受體相關(guān)因子2(TRAF2)的相互作用。在SD大鼠上構(gòu)建MCAO導(dǎo)致的局灶性腦缺血?jiǎng)游锬P停捎肨TC染色檢測腦梗死體積,F(xiàn)luoro-Jade C染色實(shí)驗(yàn)觀察缺血半暗帶區(qū)神經(jīng)元凋亡,滾軸實(shí)驗(yàn)和膠帶黏附實(shí)驗(yàn)觀察大鼠的運(yùn)動(dòng)功能障礙。結(jié)果抑制PDE4可減少SD大鼠腦梗死的體積,減少缺血半暗帶區(qū)神經(jīng)元凋亡以及改善SD大鼠的運(yùn)動(dòng)功能障礙。抑制PDE4或敲低PDE4B可逆轉(zhuǎn)OGD誘導(dǎo)的HT-22細(xì)胞凋亡。抑制PDE4可降低SD大鼠缺血半暗帶區(qū)內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)蛋白的表達(dá)。過表達(dá)PDE4B可上調(diào)內(nèi)質(zhì)網(wǎng)應(yīng)激,而敲低PDE4B或抑制PDE4可對抗OGD誘導(dǎo)的HT-22細(xì)胞內(nèi)質(zhì)網(wǎng)應(yīng)激。抑制PDE4可促進(jìn)OGD誘導(dǎo)的核因子紅系2相關(guān)因子2(NFE2L2,Nrf-2)核轉(zhuǎn)位,并降低氧化應(yīng)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,而過表達(dá) GSK3β組成性激活質(zhì)粒GSK3β(S9A)可消除抑制PDE4對Nrf-2核轉(zhuǎn)位、氧化應(yīng)激和內(nèi)質(zhì)網(wǎng)應(yīng)激的影響。Nrf-2抑制劑ML385、敲低Nrf-2以及下游靶蛋白HO-1均可阻斷抑制PDE4的保護(hù)作用。抑制PDE4可減少IRE1α與TRAF2的相互作用,并減弱c-Jun N端激酶的磷酸化水平。抑制PDE4在原代皮質(zhì)神經(jīng)元中產(chǎn)生了與上述一致的保護(hù)作用。結(jié)論抑制PDE4可以通過激活Nrf-2保護(hù)內(nèi)質(zhì)網(wǎng)應(yīng)激誘導(dǎo)的損傷,從而恢復(fù)內(nèi)質(zhì)網(wǎng)穩(wěn)態(tài)并防止神經(jīng)元死亡。

      關(guān)鍵詞:缺血性腦損傷;磷酸二酯酶4;內(nèi)質(zhì)網(wǎng)應(yīng)激

      基金項(xiàng)目:國家自然科學(xué)基金(81773698);國家自然科學(xué)基金(81974501)

      通訊作者:汪海濤,E-mail:wht821@smu.edu.cn;徐江平,E-mail:jpx@smu.edu.cn

      T3-13

      神經(jīng)血管耦合功能障礙與腦損傷藥物靶標(biāo)研究

      韓峰

      (南京醫(yī)科大學(xué)藥學(xué)院,江蘇 南京 211166)

      摘要:目的大腦神經(jīng)元活動(dòng)依賴于腦血流持續(xù)供應(yīng)的氧氣和葡萄糖等營養(yǎng)物質(zhì),其中神經(jīng)-血管偶聯(lián)保障與局部大腦活動(dòng)變化進(jìn)行時(shí)空高度匹配的局部腦血流供應(yīng),從而為大腦信息傳遞和處理保證了充分的能量供應(yīng)。眾多腦損傷相關(guān)疾病源于大腦的神經(jīng)-血管偶聯(lián)障礙。因此,闡明神經(jīng)-血管偶聯(lián)的調(diào)控機(jī)制對發(fā)現(xiàn)精準(zhǔn)藥物靶標(biāo)有重要意義。方法利用藥理學(xué)/基因調(diào)控、電生理記錄、組學(xué)等技術(shù),探究血管源性的分泌蛋白Sema3G缺失介導(dǎo)的神經(jīng)血管耦合功能障礙在神經(jīng)系統(tǒng)疾病中的作用。結(jié)果①腦血管內(nèi)皮源性Sema3G基因敲除導(dǎo)致認(rèn)知功能下降,Sema3G/Nrp2/PlexinA4參與神經(jīng)元樹突棘的成熟及LTP。②Sema3G基因敲除導(dǎo)致血管叢的密度下降,血管發(fā)育延遲。靶向Sema3G/Nrp2/PlexinA4的基因調(diào)控,可以促進(jìn)健康血管網(wǎng)絡(luò)的形成,并促進(jìn)病變血管的退化,同時(shí)改善認(rèn)知功能。結(jié)論腦血管源性Sema3G信號分子介導(dǎo)的跨細(xì)胞類型通訊模式參與神經(jīng)血管耦合障礙和腦損傷病理過程。因此,腦血管源性Sema3G及其下游信號通路為精確調(diào)控神經(jīng)血管耦合提供新思路和藥物新靶點(diǎn)。

      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

      星形膠質(zhì)細(xì)胞和小膠質(zhì)細(xì)胞對中樞神經(jīng)系統(tǒng)的作用

      孔德平,亓?xí)?,馮兆陽,譚 瑞,趙曉民

      (山東第一醫(yī)科大學(xué)藥理研究所,山東 泰安 271016)

      摘要:星形膠質(zhì)細(xì)胞與小膠質(zhì)細(xì)胞的作用以及相互作用是一直處于神經(jīng)膠質(zhì)細(xì)胞研究的前沿且推動(dòng)著中樞神經(jīng)系統(tǒng)疾病的全面認(rèn)識。星形膠質(zhì)細(xì)胞是神經(jīng)系統(tǒng)中數(shù)量最多的細(xì)胞,當(dāng)其正常功能喪失時(shí)會(huì)引發(fā)一系列中樞神經(jīng)系統(tǒng)疾病,如阿爾茨海默癥、肝性腦病等。星形膠質(zhì)細(xì)胞被激活后形成反應(yīng)性星形膠質(zhì)細(xì)胞A1或A2表型,對中樞神經(jīng)系統(tǒng)發(fā)揮著神經(jīng)毒性和神經(jīng)保護(hù)性的作用。小膠質(zhì)細(xì)胞是中樞神經(jīng)系統(tǒng)微環(huán)境變化的傳感器,它與星形膠質(zhì)細(xì)胞相比,較為敏感。研究表明小膠質(zhì)細(xì)胞與中樞神經(jīng)系統(tǒng)損傷和炎癥密切相關(guān)。另外,小膠質(zhì)細(xì)胞也可以被激活形成M1或M2表型從而發(fā)揮促炎和抗炎的作用。星形膠質(zhì)細(xì)胞與小膠質(zhì)細(xì)胞不僅可以單獨(dú)作用于中樞神經(jīng)系統(tǒng),還可以相互作用調(diào)節(jié)中樞神經(jīng)系統(tǒng)。小膠質(zhì)細(xì)胞決定了反應(yīng)性星形膠質(zhì)細(xì)胞在中樞神經(jīng)系統(tǒng)的功能。相反,星形膠質(zhì)細(xì)胞通過其分泌的分子調(diào)節(jié)改變小膠質(zhì)細(xì)胞的表型和功能以及吞噬作用。本綜述討論了目前星形膠質(zhì)細(xì)胞與小膠質(zhì)細(xì)胞以及兩者相互作用對于中樞神經(jīng)系統(tǒng)的作用,為中樞神經(jīng)系統(tǒng)疾病提供參考依據(jù)。

      關(guān)鍵詞:星形膠質(zhì)細(xì)胞;小膠質(zhì)細(xì)胞;神經(jīng)系統(tǒng)

      通訊作者:趙曉民,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

      擬人參皂苷F-11通過激活calcineurin介導(dǎo)的TFEB核轉(zhuǎn)位緩解腦缺血后神經(jīng)元自噬-溶酶體通路的功能障礙

      劉月陽,付曉曉,楊靜玉,吳春福

      (沈陽藥科大學(xué)生命科學(xué)與生物制藥學(xué)院,遼寧 沈陽 110016)

      摘要:目的轉(zhuǎn)錄因子-EB(transcription factor EB,TFEB)是一種調(diào)控自噬和溶酶體生物合成的重要轉(zhuǎn)錄因子。課題組前期研究已證明,TFEB可作為腦缺血治療的新靶標(biāo)。擬人參皂苷F-11(PF11)具有顯著的腦缺血保護(hù)作用,其機(jī)制可能與其調(diào)節(jié)ALP功能相關(guān)。為此,本研究深入探究PF11調(diào)節(jié)腦缺血后ALP功能的作用是否與其調(diào)節(jié)TFEB的核轉(zhuǎn)位相關(guān)及其調(diào)節(jié)機(jī)制。方法采用大鼠永久性腦缺血(pMCAO)及原代皮質(zhì)神經(jīng)元氧糖剝奪(OGD)模型,結(jié)合慢病毒敲低技術(shù)等觀察PF11對腦缺血后TFEB核轉(zhuǎn)位的調(diào)節(jié)。結(jié)果PF11(6和12 mg·kg-1,iv)顯著增加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。與在體結(jié)果一致,PF11(30 和 100 μmol·L-1)可顯著增加OGD后12 h原代神經(jīng)元TFEB總蛋白表達(dá)及核蛋白表達(dá),顯著促進(jìn)TFEB入核,改善溶酶體功能,減少自噬小體及底物的不正常積累。此外,慢病毒敲低原代神經(jīng)元中的TFEB能夠逆轉(zhuǎn)PF11對OGD后12 h ALP功能障礙的緩解作用。進(jìn)一步研究表明,calcineurin抑制劑環(huán)孢菌素(10 μmol·L-1)可顯著逆轉(zhuǎn)PF11對神經(jīng)元OGD后TFEB核轉(zhuǎn)位的促進(jìn)作用及其對ALP功能的調(diào)節(jié)。結(jié)論P(yáng)F11可能通過促進(jìn)calcineurin介導(dǎo)的TFEB核轉(zhuǎn)位來緩解腦缺血后ALP功能障礙,進(jìn)一步明確了PF11抗腦缺血的自噬相關(guān)機(jī)制。

      關(guān)鍵詞:轉(zhuǎn)錄因子-EB;腦缺血;氧糖剝奪;溶酶體;自噬

      T3-19

      氰化鈉加重缺氧誘發(fā)腦損傷機(jī)制

      石華香,周夢瑋,李鵬飛,周 琥,王麗韞

      (軍事科學(xué)院軍事醫(yī)學(xué)研究院毒物藥物研究所,抗毒藥物與毒理學(xué)國家重點(diǎn)實(shí)驗(yàn)室,北京 100850)

      摘要:目的探究急性氰化鈉(NaCN)中毒合并缺氧對小鼠腦損傷的作用。方法觀察生理鹽水對照組、NaCN 0.26,0.38 和 0.51 mg·kg-1(ip)中毒組小鼠缺氧存活時(shí)間。小鼠隨機(jī)分為對照組、NaCN(3.8 mg·kg-1)處理組、NaCN 復(fù)合缺氧30 min(CN-30)組和NaCN復(fù)合缺氧60 min(CN-60)組,采用激光散斑成像儀檢測小鼠腦血流變化;組織含水量測定腦組織水腫影響;酶標(biāo)法檢測海馬組織氧化應(yīng)激反應(yīng)指標(biāo)T-SOD活力和MDA含量變化;免疫熒光法檢測皮質(zhì)和海馬組織小膠質(zhì)細(xì)胞炎性標(biāo)志物Iba1和CD68信號表達(dá)變化;Western印跡法測定Iba1和CD68及凋亡信號分子Bcl-2和Bax蛋白表達(dá)變化。結(jié)果與對照組比較,NaCN中毒顯著延長缺氧小鼠存活時(shí)間(P<0.01),0.38 mg·kg-1延長(74.7±17.8)min。與對照組比較,NaCN組、CN-30組和CN-60組小鼠腦血流值分別降低12%,40%和55%(P<0.01),腦組織含水量均顯著增加(P<0.01),海馬組織T-SOD活力和MDA含量均顯著增加(P<0.01),皮質(zhì)和海馬組織表達(dá)Iba1+和CD68+小膠質(zhì)細(xì)胞百分比均顯著增加;NaCN組Bcl-2和Bax表達(dá)顯著增加,CN-30組和CN-60組進(jìn)行性增加。結(jié)論NaCN中毒延長小鼠缺氧存活時(shí)間,加重缺氧誘發(fā)的腦血流下降,引發(fā)腦組織水腫,加劇海馬組織的氧化應(yīng)激反應(yīng)和小膠質(zhì)細(xì)胞炎癥反應(yīng),促進(jìn)腦組織損傷。

      關(guān)鍵詞:氰化鈉;腦血流;氧化應(yīng)激;免疫熒光;凋亡

      通訊作者:王麗韞,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

      血小板介導(dǎo)的神經(jīng)炎癥在中樞神經(jīng)性疾病中的作用

      孔德平,馮兆陽,亓?xí)螅T 瑞,高永峰,于海洋,趙曉民

      〔山東第一醫(yī)科大學(xué)(山東省醫(yī)學(xué)科學(xué)院)藥理學(xué)研究所,山東 泰安 271016〕

      摘要:血小板是無核細(xì)胞,血液中數(shù)量較多,表面有豐富受體,細(xì)胞內(nèi)有多種顆粒,可釋放活性物質(zhì),除具有止血、促進(jìn)血栓形成作用之外,還在炎癥和免疫方面發(fā)揮重要功能。由于炎癥在嚴(yán)重的神經(jīng)性疾病發(fā)生發(fā)展中占據(jù)重要位置,因而研究中樞神經(jīng)性疾病需要考慮血小板的因素。在人或動(dòng)物相關(guān)研究中,血小板介導(dǎo)的神經(jīng)炎癥在肌萎縮側(cè)索硬化癥、癲癇、創(chuàng)傷性腦損傷、帕金森病、亨廷頓病、舞蹈病、阿爾茨海默病和腦卒中等進(jìn)展中產(chǎn)生影響,機(jī)制涉及血小板參與的血栓,與白細(xì)胞、中樞血管內(nèi)皮細(xì)胞、免疫細(xì)胞和補(bǔ)體系統(tǒng)的相互作用,分子事件包括血小板表面的受體、釋放的活性物質(zhì)及相應(yīng)信號途徑,還可調(diào)節(jié)神經(jīng)元的電活動(dòng)、突觸功能和可塑性。靶向調(diào)控血小板介導(dǎo)的神經(jīng)炎癥,可能成為一種輔助治療中樞神經(jīng)性疾病的有效手段。

      關(guān)鍵詞:血小板;神經(jīng);炎癥;血栓形成

      通訊作者:趙曉民,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

      專題4:離子通道及受體信號與神經(jīng)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

      山茱萸環(huán)烯醚萜苷對tau蛋白轉(zhuǎn)基因小鼠精神癥狀的影響及其機(jī)制

      馬登磊,羅 藝,李 林,張 蘭

      (首都醫(yī)科大學(xué)宣武醫(yī)院藥學(xué)部,神經(jīng)變性病教育部重點(diǎn)實(shí)驗(yàn)室,北京市神經(jīng)藥物工程研究中心,北京 100053)

      摘要:目的微管相關(guān)蛋白tau在阿爾茨海默?。ˋD)發(fā)病機(jī)制中發(fā)揮重要作用。過表達(dá)P301L突變tau蛋白的rTg4510轉(zhuǎn)基因小鼠具有明顯的tau病理表現(xiàn),廣泛應(yīng)用于AD的基礎(chǔ)研究及藥理研究中。山茱萸環(huán)烯醚萜苷(CIG)為中藥山茱萸的有效部位,可降低tau蛋白的磷酸化。本研究探討CIG以tau蛋白為靶點(diǎn)治療AD的潛在價(jià)值。方法應(yīng)用rTg4510轉(zhuǎn)基因小鼠為AD模型,給予CIG處理3個(gè)月后進(jìn)行Y迷宮和自主活動(dòng)行為學(xué)檢測,之后分別灌注固定取材及新鮮取材。應(yīng)用Western印跡法檢測蛋白的表達(dá)水平,應(yīng)用免疫組織化學(xué)等方法觀察病理性tau蛋白表達(dá)。結(jié)果CIG可以顯著降低rTg4510小鼠的過度活躍狀態(tài),降低小鼠腦內(nèi)杏仁核、海馬區(qū)tau蛋白病變,改善小鼠杏仁核區(qū)域的神經(jīng)元丟失;同時(shí)增加小鼠腦內(nèi)NMDA受體及突觸功能蛋白的表達(dá),減少白質(zhì)胼胝體區(qū)域的髓鞘堿性蛋白的丟失。結(jié)論CIG可改善rTg4510轉(zhuǎn)基因小鼠的精神行為學(xué)表現(xiàn),改善神經(jīng)元丟失和白質(zhì)脫髓鞘病變,其主要機(jī)制為CIG降低tau蛋白的過度磷酸化及病理性tau蛋白在杏仁核區(qū)域的沉積,增加NMDA受體及突觸功能蛋白的表達(dá),提示CIG是一個(gè)潛在抗AD藥物。

      關(guān)鍵詞:山茱萸環(huán)烯醚萜苷;阿爾茨海默?。籺au蛋白

      T4-6

      嚙齒類動(dòng)物觸屏操作實(shí)驗(yàn)方法研究進(jìn)展

      張亦文,黃 紅,姚彩虹,姜 寧,劉新民

      (中國醫(yī)學(xué)科學(xué)院北京協(xié)和醫(yī)學(xué)院藥用植物研究所,北京 100193)

      摘要:通過文獻(xiàn)檢索,探索嚙齒類動(dòng)物觸屏操作實(shí)驗(yàn)方法在嚙齒類動(dòng)物神經(jīng)精神藥理學(xué)研究的應(yīng)用現(xiàn)狀。本文對國內(nèi)外中英文共6個(gè)文獻(xiàn)庫的近30年文獻(xiàn)進(jìn)行檢索篩選和數(shù)據(jù)提取,系統(tǒng)分析總結(jié)了實(shí)驗(yàn)中常用動(dòng)物的種類、訓(xùn)練方法、訓(xùn)練模式和相關(guān)參數(shù)等,得到相關(guān)文獻(xiàn)300余篇,近10年來相關(guān)文章數(shù)量增長較快。大、小鼠和多種轉(zhuǎn)基因動(dòng)物均適用此實(shí)驗(yàn)方法,常用的實(shí)驗(yàn)?zāi)J蕉噙_(dá)10余種,其中視覺辨別與逆轉(zhuǎn)最為常用,對常用參數(shù)和實(shí)驗(yàn)方法等也進(jìn)行了系統(tǒng)闡述。嚙齒類動(dòng)物觸屏操作實(shí)驗(yàn)方法基于斯金納條件反射原理,通過計(jì)算機(jī)呈現(xiàn)圖案信號刺激,嚙齒類動(dòng)物用鼻子直接觸碰屏幕對信號做出反應(yīng),從而檢測動(dòng)物操作任務(wù)下對不同模式的圖案信號的興趣、解決問題的能力和速度等,反應(yīng)動(dòng)物的注意力、工作記憶、長期記憶、空間記憶和決策等高級認(rèn)知功能。齒類動(dòng)物觸屏操作實(shí)驗(yàn)在臨床干預(yù)轉(zhuǎn)化中有著較高的轉(zhuǎn)化潛力,在神經(jīng)精神類疾病以及操作任務(wù)下學(xué)習(xí)記憶的研究中有著巨大的應(yīng)用前景。

      關(guān)鍵詞:觸屏操作;神經(jīng)精神藥理;嚙齒類動(dòng)物

      基金項(xiàng)目:國家自然科學(xué)基金(81773930);空間站工程航天醫(yī)學(xué)實(shí)驗(yàn)項(xiàng)目(HYZHXM05003)

      通訊作者:劉新民,E-mail:liuxinmin@hotmail.com;姜 寧,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

      鈴蟾肽致小鼠瘙癢下丘腦響應(yīng)及PKA/creb信號參與機(jī)制

      張晶鑫1,2,周 琥1,石京山2,王永安1,王麗韞1

      (1.北京毒物藥物研究所,北京 100850;2.遵義醫(yī)科大學(xué)基礎(chǔ)藥理國家重點(diǎn)實(shí)驗(yàn)室,貴州 遵義 563000)

      摘要:目的研究鈴蟾肽(BN)致瘙癢藥理學(xué)作用特點(diǎn),確定下丘腦參與其中樞調(diào)控瘙癢關(guān)鍵分子機(jī)制。方法觀察KM小鼠經(jīng)側(cè)腦室注射不同劑量BN(0.04,0.40和4.00 μg)誘發(fā)的瘙癢效應(yīng);構(gòu)建耦聯(lián)熒光標(biāo)簽NBD(7-nitrobenz-2-oxa-1,3-diazole)的BN(NBD-BN),并通過檢測熒光信號示蹤NBDBN在腦內(nèi)的分布;利用免疫組化和Western印跡法檢測下丘腦BN受體神經(jīng)介素B受體(NMBR)、胃泌素釋放肽受體(GRPR)及受體信號PKA/CREB相關(guān)蛋白的表達(dá),鑒定下丘腦內(nèi)特異神經(jīng)元c-FOS蛋白響應(yīng)。結(jié)果KM小鼠經(jīng)側(cè)腦室注射BN(0.04,0.40和4.00 μg)后,30 min觀察期內(nèi)產(chǎn)生顯著瘙癢抓撓行為,抓撓次數(shù)呈劑量依賴性增加。與對照組相比,BN 3個(gè)劑量組抓撓次數(shù)均顯著增加(P<0.05,P<0.01);熒光示蹤結(jié)果顯示,側(cè)腦室給予BN后NBD-BN在30 min內(nèi)在下丘腦特異性分布,且10 min時(shí)熒光強(qiáng)度達(dá)到峰值。免疫組化和Western印跡結(jié)果顯示,BN 2種受體亞型GRPR和NMBR在下丘腦特異性分布;側(cè)腦室給予BN 4.00 μg后,與對照組相比,下丘腦PKA,p-CREB和c-FOS蛋白均顯著增加(P<0.01),且c-FOS蛋白在下丘腦背內(nèi)側(cè)核密集分布。結(jié)論BN具有強(qiáng)的中樞致瘙癢作用,側(cè)腦室注射BN可致小鼠瘙癢并激活下丘腦BN受體依賴的PKA/CREB信號通路,誘導(dǎo)下丘腦神經(jīng)元響應(yīng),表明下丘腦參與調(diào)控BN中樞致瘙癢作用。

      關(guān)鍵詞:鈴蟾肽;癢;下丘腦背內(nèi)側(cè)核;胃泌素釋放肽受體;NMBR神經(jīng)介素B受體

      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

      神經(jīng)營養(yǎng)因子異常與抑郁癥發(fā)病機(jī)制研究進(jìn)展

      王惠芹1,2,陳乃宏1,2

      (1.湖南中醫(yī)藥大學(xué)藥學(xué)院,湖南 長沙 410208;2.中國醫(yī)學(xué)科學(xué)院北京協(xié)和醫(yī)學(xué)院藥物研究所,北京 100050)

      摘要:抑郁癥已成為“二十一世紀(jì)的流行病”,對我國乃至國際公民和社會(huì)的危害日益凸顯。神經(jīng)營養(yǎng)因子是神經(jīng)可塑性的關(guān)鍵介質(zhì),抑郁癥的發(fā)病涉及到神經(jīng)營養(yǎng)因子及其基因異常??挂钟羲幬镏委熀笊窠?jīng)營養(yǎng)因子分泌增加,從而促進(jìn)神經(jīng)元的存活,保護(hù)神經(jīng)元免受應(yīng)激損傷。腦源性神經(jīng)營養(yǎng)因子、血管內(nèi)皮生長因子、膠質(zhì)細(xì)胞源性神經(jīng)營養(yǎng)因子和神經(jīng)生長因子等與抑郁癥關(guān)系的研究表明神經(jīng)營養(yǎng)因子有望成為抗抑郁治療的有效靶點(diǎn)。因此,研究與神經(jīng)營養(yǎng)因子有關(guān)的抑郁癥發(fā)病機(jī)制,對尋找快速有效的抗抑郁治療方法至關(guān)重要。

      關(guān)鍵詞:抑郁癥;神經(jīng)營養(yǎng)因子;發(fā)病機(jī)制

      T4-12

      白芷的化學(xué)成分及對神經(jīng)系統(tǒng)疾病的治療作用

      龐心悅1,景永帥2,鄭玉光1,吳蘭芳1

      (1.河北中醫(yī)學(xué)院藥學(xué)院,河北 石家莊 050200;2.河北科技大學(xué)化學(xué)與制藥工程學(xué)院,河北 石家莊 050018)

      摘要:白芷為傘形科植物白芷或杭白芷的干燥根,始載于東漢《神農(nóng)本草經(jīng)》,味辛,性溫,具有解表散寒、宣通鼻竅、消腫排膿等功效。白芷含揮發(fā)油、香豆素類、多糖類、黃酮類、生物堿類等成分。研究表明白芷及其活性成分對神經(jīng)系統(tǒng)疾病有一定的緩解或治療作用。①抗驚厥:白芷總香豆素類成分具有抗驚厥作用,可明顯延長戊四氮和3-巰基丙酸所致模型小鼠的驚厥潛伏期,縮短驚厥持續(xù)時(shí)間,從而降低小鼠的死亡率;白芷的二氯甲烷萃取物在中、大劑量條件下也具有抗驚厥的作用。②緩解偏頭痛:白芷能改善偏頭痛動(dòng)物行為學(xué)表現(xiàn),減少NO分泌,增加5-羥色胺(5-HT)含量,降低降鈣素基因相關(guān)肽的陽性率;另外,白芷的香豆素和揮發(fā)油具有明顯的鎮(zhèn)痛作用,其機(jī)制可能是減少模型動(dòng)物腦內(nèi)和血中NO的合成。③止疼作用:白芷總揮發(fā)油、水煎液對疼痛模型大鼠有明顯的鎮(zhèn)痛作用,其中水合氧化前胡素、佛手柑內(nèi)酯可能是白芷鎮(zhèn)痛的效應(yīng)物質(zhì)基礎(chǔ);另外,白芷揮發(fā)油可促進(jìn)β-內(nèi)啡肽的前體物質(zhì)前阿黑皮素信使核糖核酸的表達(dá)、調(diào)整體內(nèi)單胺類神經(jīng)遞質(zhì)含量而發(fā)揮鎮(zhèn)痛作用。④抗抑郁:白芷冰片方在多個(gè)動(dòng)物模型上具有顯著的抗抑郁作用,其機(jī)制與增強(qiáng)中樞5-HT能神經(jīng)系統(tǒng)功能,上調(diào)海馬BDNF/TrkB/p-CREB神經(jīng)營養(yǎng)通路有關(guān)。本文對白芷在神經(jīng)系統(tǒng)相關(guān)疾病中的治療作用進(jìn)行歸納總結(jié),進(jìn)一步明確白芷發(fā)揮藥效的物質(zhì)基礎(chǔ),可為白芷治療神經(jīng)系統(tǒng)相關(guān)疾病的臨床應(yīng)用及新藥開發(fā)提供依據(jù)。

      關(guān)鍵詞:白芷;活性成分;神經(jīng)系統(tǒng)疾病

      基金項(xiàng)目:河北省省級科技計(jì)劃(H2021423057)

      通訊作者:吳蘭芳,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

      慢性束縛應(yīng)激對小鼠疲勞發(fā)生的動(dòng)態(tài)影響及機(jī)制

      王 智,閆明珠,夏天吉,靳蘇維,劉永廣,常 琪

      (中國醫(yī)學(xué)科學(xué)院&北京協(xié)和醫(yī)學(xué)院藥用植物研究所,北京 100094)

      摘要:目的當(dāng)今,更多人群因常常需要在狹小環(huán)境中久坐工作,長期作息不規(guī)律且缺乏活動(dòng)而導(dǎo)致疲勞。本研究希望通過慢性束縛應(yīng)激(CRS)模擬以上人群疲勞的發(fā)生情況,并對CRS小鼠疲勞模型的動(dòng)態(tài)發(fā)生及機(jī)制進(jìn)行探索。方法將48只4周齡雄性ICR小鼠按體重和前肢抓力隨機(jī)分成4組,分別束縛0,5,10和15 d,8 h·d-1。束縛結(jié)束后進(jìn)行空場實(shí)驗(yàn)、抓力實(shí)驗(yàn)和負(fù)重游泳實(shí)驗(yàn)。次日各組小鼠強(qiáng)迫游泳30 min后休息30 min,戊巴比妥鈉麻醉后從腹主動(dòng)脈取血,脫頸處死取肝和后肢腓腸肌組織用于生化指標(biāo)檢測。結(jié)果與空白組相比,CRS小鼠體重和腓腸肌明顯減輕;自主活動(dòng)路程與時(shí)間均有減少,且15 d組均變化顯著;抓力均顯著降低;束縛10和15 d組負(fù)重游泳力竭時(shí)間顯著縮短。CRS小鼠的血糖和肝糖原水平均顯著降低,肌糖原水平均顯著升高,血清乳酸堆積量、乳酸脫氫酶和谷草轉(zhuǎn)氨酶活性顯著升高,這些指標(biāo)的變化與束縛時(shí)長成正相時(shí)效關(guān)系。腓腸肌組織的蛋白印跡測定顯示,CRS小鼠LC3,p-AMPK/AMPK,PGC-1α,TFAM和OXPHOS表達(dá)降低,Parkin,p-AKT/AKT,p-mTOR/mTOR,p-ULK1(Ser757)和P62表達(dá)升高,表明線粒體發(fā)生和自噬受阻。結(jié)論在CRS下,小鼠疲勞逐漸發(fā)生,10 d(8 h·d-1)即可誘導(dǎo)疲勞,15 d可造成小鼠穩(wěn)定疲勞。CRS致小鼠腓腸肌線粒體發(fā)生和功能受損,線粒體保護(hù)性自噬受阻,進(jìn)而無法維持線粒體功能平衡,并且這與AMPK信號通路受抑制相關(guān)。

      關(guān)鍵詞:慢性束縛;疲勞;線粒體功能障礙;線粒體自噬;AMPK

      基金資助:科技部國家重點(diǎn)研發(fā)計(jì)劃“食品安全關(guān)鍵技術(shù)研發(fā)”專項(xiàng)(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

      海馬CD39(ENTPD1)對小鼠抑郁樣行為的影響及機(jī)制

      董婉婷,胡壯麗,陳建國

      (華中科技大學(xué)同濟(jì)醫(yī)學(xué)院基礎(chǔ)醫(yī)學(xué)院藥理學(xué)系,湖北 武漢 430030)

      摘要:目的ATP是一種快速興奮性神經(jīng)調(diào)質(zhì),在中樞神經(jīng)系統(tǒng)富集。最新的研究表明,細(xì)胞外間隙低水平的ATP可能促進(jìn)重度抑郁癥(MDD)形成。細(xì)胞外ATP的濃度受其水解酶——外核苷酸三(二)磷酸水解酶(ENTPD)的調(diào)節(jié),但有關(guān)ENTPD在抑郁癥中的作用尚不清楚。因此,本文探討CD39(稱為ENTPD1)在慢性社會(huì)挫敗應(yīng)激(CSDS)誘導(dǎo)的小鼠抑郁樣模型中的作用。方法和結(jié)果 CSDS增強(qiáng)海馬組織中CD39的表達(dá)和活性。進(jìn)一步給予CD39功能類似物三磷酸腺苷雙磷酸酶,也可誘導(dǎo)小鼠抑郁樣行為。有趣的是,再次補(bǔ)充ATP可逆轉(zhuǎn)上述抑郁樣行為。最后,通過藥理學(xué)抑制和基因沉默的方法增加細(xì)胞外ATP濃度,亦可發(fā)揮抗抑郁作用,且同時(shí)增加海馬神經(jīng)發(fā)生和樹突棘數(shù)量。結(jié)論海馬CD39通過水解細(xì)胞外ATP參與誘導(dǎo)抑郁樣行為,提示CD39可作為抑郁癥的一個(gè)有前景的靶點(diǎn)。

      關(guān)鍵詞:海馬;抑郁;CD39

      專題5:新技術(shù)與新方法在神經(jīng)精神藥理學(xué)研究中的應(yīng)用

      T5-1

      基于苯乙醇苷類天然分子探針的抗腦缺血新靶點(diǎn)發(fā)現(xiàn)

      曾克武

      (北京大學(xué)藥學(xué)院天然藥物及仿生藥物國家重點(diǎn)實(shí)驗(yàn)室,北京 100191)

      摘要:目的缺血性腦卒中是威脅人類生命健康的重大疾病,目前嚴(yán)重缺乏有效的藥物靶點(diǎn)。肉蓯蓉是我國著名的補(bǔ)益中藥,被稱為“沙漠人參”。研究發(fā)現(xiàn),肉蓯蓉具有良好的腦保護(hù)作用,其中苯乙醇苷是其主要藥效成分。我們設(shè)想:是否可以利用代表性的苯乙醇苷類成分——松果菊苷(ECH)作為活性分子探針,從神經(jīng)細(xì)胞中“鉤釣”其直接作用靶點(diǎn),并揭示其發(fā)揮神經(jīng)保護(hù)作用的分子藥理機(jī)制。方法以ECH作為分子探針,利用鍵合有ECH的固相載體從神經(jīng)細(xì)胞裂解液中捕獲其直接作用靶點(diǎn)并進(jìn)行高分辨質(zhì)譜鑒定,同時(shí)結(jié)合化學(xué)生物學(xué)及分子藥理學(xué)方法探究其調(diào)控神經(jīng)細(xì)胞缺血性損傷的分子信號通路。結(jié)果揭示了ECH的作用靶點(diǎn)為酪氨酸激酶α′催化亞基(CK2α′),且CK2α′通過激活Wnt/β-catenin信號通路促進(jìn)線粒體融合,發(fā)揮抗神經(jīng)細(xì)胞缺血性損傷的作用。此外,在CK2α′WT和CK2α′+/-小鼠MCAO模型上也證實(shí)了ECH通過靶點(diǎn)CK2α′發(fā)揮改善腦缺血的作用。結(jié)論揭示了ECH的直接作用靶點(diǎn)為CK2α′,同時(shí)也提示CK2α′是一個(gè)基于天然活性分子探針發(fā)現(xiàn)的新穎的腦缺血治療靶點(diǎn),為今后缺血性腦卒中先導(dǎo)藥物的發(fā)現(xiàn)提供了新思路。

      關(guān)鍵詞:腦缺血;治療靶點(diǎn);苯乙醇苷類分子;活性分子探針;靶點(diǎn)鉤釣

      通訊作者:曾克武,E-mail:ZKW@bjmu.edu.cn

      T5-2

      中腦導(dǎo)水管周圍灰質(zhì)腹外側(cè)部星形膠質(zhì)細(xì)胞對糖尿病神經(jīng)病理性疼痛及痛相關(guān)負(fù)性情緒的調(diào)控作用

      楊 瀾1,陳 理1,2,俞昌喜1,2

      (1.福建醫(yī)科大學(xué)藥學(xué)院藥理學(xué)系神經(jīng)藥理課題組,福建 福州 350122;2.福建省天然藥物藥理學(xué)重點(diǎn)實(shí)驗(yàn)室,福建 福州 350122)

      摘要:目的探索中腦導(dǎo)水管周圍灰質(zhì)腹外側(cè)部(vlPAG)星形膠質(zhì)細(xì)胞活化在糖尿病神經(jīng)病理性疼痛(DNP)及痛相關(guān)負(fù)性情緒發(fā)生、發(fā)展及維持過程中的具體作用及特征。方法使用鏈脲佐菌素(STZ)誘導(dǎo)SD大鼠Ⅰ型糖尿病,復(fù)制DNP模型;免疫熒光法觀察、Sholl分析評價(jià)vlPAG星形膠質(zhì)細(xì)胞在DNP進(jìn)展過程中的形態(tài)和數(shù)量變化;采用化學(xué)遺傳學(xué)技術(shù)定點(diǎn)定量向vlPAG注射攜有星形膠質(zhì)細(xì)胞特異性啟動(dòng)子gfaABC1D及修飾后的人毒蕈堿受體(hM3Dq或hM4Di)基因片段的腺相關(guān)病毒;待病毒穩(wěn)定轉(zhuǎn)染表達(dá)后,利用受體的外源性配體氯氮平-N-氧化物(CNO)靶向調(diào)控vlPAG星形膠質(zhì)細(xì)胞活性;以機(jī)械性異常性疼痛閾值為觀察指標(biāo),評價(jià)星形膠質(zhì)細(xì)胞活化在DNP中的作用;同時(shí),采用曠場試驗(yàn)、高架十字迷宮試驗(yàn)、條件性位置偏愛試驗(yàn),評價(jià)vlPAG星形膠質(zhì)細(xì)胞活化在DNP相關(guān)焦慮樣、厭惡樣情緒中的作用。結(jié)果STZ成功誘導(dǎo)高血糖癥狀后,SD大鼠機(jī)械痛閾進(jìn)行性下降,提示DNP模型復(fù)制成功。隨著DNP癥狀加劇,vlPAG星形膠質(zhì)細(xì)胞數(shù)量逐漸增多、胞體面積逐漸增大、形態(tài)復(fù)雜性逐漸增強(qiáng)?;瘜W(xué)遺傳學(xué)手段激活正常大鼠vlPAG星形膠質(zhì)細(xì)胞可誘導(dǎo)神經(jīng)病理性疼痛樣癥狀(機(jī)械痛閾下降);化學(xué)遺傳學(xué)手段抑制DNP模型大鼠vlPAG星形膠質(zhì)細(xì)胞可改善DNP癥狀(機(jī)械痛閾升高)。同時(shí),化學(xué)遺傳學(xué)手段抑制vlPAG星形膠質(zhì)細(xì)胞可減少DNP模型大鼠痛相關(guān)焦慮樣行為,改善模型動(dòng)物痛相關(guān)厭惡情緒。結(jié)論vlPAG星形膠質(zhì)細(xì)胞活化與DNP的發(fā)生、發(fā)展及維持密切相關(guān),vlPAG星形膠質(zhì)細(xì)胞可能參與了DNP及痛相關(guān)負(fù)性情緒的編碼和調(diào)控。

      關(guān)鍵詞:糖尿病神經(jīng)病理性疼痛;中腦導(dǎo)水管灰質(zhì)腹外側(cè)部;星形膠質(zhì)細(xì)胞;化學(xué)遺傳學(xué);痛相關(guān)負(fù)性情緒

      基金項(xiàng)目:國家自然科學(xué)基金(81973309)

      通訊作者: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

      膠質(zhì)細(xì)胞離子通道功能調(diào)控與癲癇藥物靶標(biāo)

      盧應(yīng)梅

      (南京醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院,江蘇 南京 211166)

      摘要:目的癲癇以反復(fù)發(fā)作為特征,世界范圍內(nèi)有6500~7000萬癲癇患者。目前認(rèn)為神經(jīng)元興奮-抑制的失衡是構(gòu)成癲癇發(fā)作的根本原因,但是直接針對神經(jīng)元興奮性的抗癲癇的策略已經(jīng)被證明部分癲癇患者中不能很好的發(fā)揮抗癲癇作用。因此,進(jìn)一步深入探究癲癇的發(fā)病機(jī)制顯得尤為必要。方法采用基因小鼠,結(jié)合活體雙光子、在體電生理記錄、膜片鉗記錄、藥理學(xué)/基因調(diào)控等技術(shù),逐層深入地開展膠質(zhì)細(xì)胞離子通道功能異常介導(dǎo)神經(jīng)元功能改變的機(jī)制。結(jié)果①血管癲癇發(fā)生過程中,神經(jīng)元、膠質(zhì)細(xì)胞及腦微血管的異常改變。②腦血管內(nèi)皮Cdk5缺失導(dǎo)致腦血管內(nèi)皮細(xì)胞CX3CL1分泌過多。腦血管內(nèi)皮源性CXCL1/CXCR2信號介導(dǎo)了星形膠質(zhì)細(xì)胞GLT1的谷氨酸重?cái)z取功能異常,導(dǎo)致錐體神經(jīng)元活性增加,進(jìn)而介導(dǎo)癲癇發(fā)生。③導(dǎo)致靶向Cdk5或CXCL1抗體或基因治療,明顯改善膠質(zhì)細(xì)胞離子通道功能和神經(jīng)元活性,改善癲癇的發(fā)生。結(jié)論腦血管源性的相關(guān)炎癥因子是細(xì)胞間通訊的重要媒介分子,間接調(diào)控神經(jīng)元功能,內(nèi)皮細(xì)胞Cdk5以及它下游的信號通路為臨床治療癲癇提供了新思路和藥物新靶點(diǎn)。

      關(guān)鍵詞:癲癇;膠質(zhì)細(xì)胞;離子通道

      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

      人誘導(dǎo)型多能干細(xì)胞來源多巴胺能神經(jīng)元在精神神經(jīng)性疾病研究中的應(yīng)用

      李 紅,李 斐,盧關(guān)伊,周詳斌,方 婷,吳 寧,李 錦

      (軍事科學(xué)院軍事醫(yī)學(xué)研究院毒物藥物研究所,北京 100850)

      摘要:目的人腦多巴胺系統(tǒng)功能異??梢鸲喾N精神神經(jīng)性疾病,但因?yàn)槿四X組織樣品極難獲得,研究手法有限,大大限制其研究。采用人誘導(dǎo)型多能干細(xì)胞(iPSC)來源的多巴胺(DA)能神經(jīng)元建立人腦發(fā)育/疾病模型,以增加對相關(guān)精神神經(jīng)性疾病的認(rèn)識。方法利用臨床吸脂術(shù)后的脂肪組織,提取脂肪干細(xì)胞,將其重編程為iPSC,并進(jìn)一步分化為DA能神經(jīng)元,對包含變異基因的神經(jīng)元功能進(jìn)行檢測。結(jié)果成功利用仙臺病毒將人脂肪干細(xì)胞重編程為iPSC。現(xiàn)有胚胎干細(xì)胞誘導(dǎo)DA能神經(jīng)元的方案并不適合iPSC;重新調(diào)整分化方案后的最佳的小分子化合物組合為Puromophine 3 μmol·L-1和CHIR-99021 0.8 μmol·L-1,可將iPSC高效向DA能神經(jīng)元分化;神經(jīng)元具有自發(fā)Pace-making動(dòng)作電位的能力,分化后25 d細(xì)胞注射帕金森病模型大鼠紋狀體內(nèi),可改善大鼠的行為學(xué)指標(biāo)。在此基礎(chǔ)上進(jìn)一步發(fā)現(xiàn)Parkin缺失的iPSC來源的DA能神經(jīng)元過早成熟和快速老化;對比FKBP5 rs1360780 CC/TT iPSC源DA能神經(jīng)元的發(fā)育和功能,發(fā)現(xiàn)TT型來源的DA能神經(jīng)元在基因和蛋白水平上AADC,TH和NURR1表達(dá)均顯著減低,且多巴胺的分泌能力下降,在地塞米松處理建立的應(yīng)激模型中表現(xiàn)為較強(qiáng)的應(yīng)激反應(yīng)性。結(jié)論人iPSC來源的神經(jīng)元可建立人腦發(fā)育或疾病模型,在增加人類精神神經(jīng)性疾病認(rèn)識及促進(jìn)高效靶向藥物的開發(fā)中發(fā)揮重大作用。

      關(guān)鍵詞:誘導(dǎo)型多能干細(xì)胞;多巴胺;神經(jīng)元;應(yīng)激

      T5-7

      電刺激小腦齒狀核對大鼠運(yùn)動(dòng)皮層缺血后神經(jīng)發(fā)生的作用

      孫芳玲,吳 錚,李子潔,劉 敏,田 欣,劉婷婷,王文

      (首都醫(yī)科大學(xué)宣武醫(yī)院,北京 100053;北京市老年病醫(yī)療研究中心,北京 100053)

      摘要:目的腦卒中是全球死亡和殘疾的主要原因,約占全球5%的傷殘調(diào)整年和10%的死亡率。目前臨床上腦卒中的治療存在有效治療手段的局限性,除溶栓外仍缺乏有效的神經(jīng)保護(hù)及再生修復(fù)治療手段,且治療時(shí)間窗窄,臨床預(yù)后改善率低。腦卒中后存活患者不同程度喪失勞動(dòng)力,其中上肢運(yùn)動(dòng)功能障礙在首次腦卒中后生存的患者中占73%~88%,在慢性腦卒中中占到55%~75%。尋找有效的神經(jīng)再生修復(fù)治療新方法,打破腦卒中治療的瓶頸,是腦卒中后康復(fù)治療的重點(diǎn)目標(biāo),對于最大程度促進(jìn)患者功能恢復(fù)、減輕腦卒中負(fù)擔(dān)有極為重要的意義。腦深部電刺激(DBS)作為一種新型的治療方法,具有靶點(diǎn)明確、安全、可調(diào)等優(yōu)點(diǎn),已用于帕金森病、癲癇和肌張力障礙等疾病患者神經(jīng)功能改善治療上。但DBS在腦卒中治療上的應(yīng)用主要集中在神經(jīng)疼痛方面,只有很少幾個(gè)臨床試驗(yàn)在研究其他腦卒中后遺癥時(shí)關(guān)注到對運(yùn)動(dòng)功能的改善。因此本研究選用了小腦齒狀核(LCN)作為DBS的靶點(diǎn),觀察電刺激是否能夠激活小腦-丘腦-皮層通路,進(jìn)而改善局灶性腦缺血大鼠的運(yùn)動(dòng)功能;并且觀察了電刺激LCN后室管膜下區(qū)(SVZ)神經(jīng)干細(xì)胞的增殖和分化情況,以期為腦卒中后的神經(jīng)再生修復(fù)治療提供新方法。方法采用內(nèi)皮素-1定點(diǎn)注射誘導(dǎo)大鼠運(yùn)動(dòng)皮層缺血,之后在對側(cè)LCN安置刺激電極。術(shù)前分別進(jìn)行面條矩陣任務(wù)和水平梯子行走任務(wù)訓(xùn)練。其中面條矩陣抓取任務(wù)行為學(xué)評價(jià),國內(nèi)尚無相關(guān)實(shí)驗(yàn)方法報(bào)道,使用的動(dòng)物實(shí)驗(yàn)裝置為我們自行制備。實(shí)驗(yàn)中訓(xùn)練大鼠完成對食物-干面條的抓取任務(wù),通過矩陣對其前肢運(yùn)動(dòng)的距離和和方向進(jìn)行測量,定量和定性評估后前肢運(yùn)動(dòng)能力,為腦卒中后感覺-運(yùn)動(dòng)神經(jīng)功能評估提供可靠的新方法。本研究在手術(shù)后第8天開始每天8 h的LCN DBS治療,且每天配合進(jìn)行15 min面條矩陣抓取訓(xùn)練,治療持續(xù)14 d。尼氏染色計(jì)算梗死體積,免疫熒光雙標(biāo)檢測SVZ神經(jīng)發(fā)生。結(jié)果LCN DBS能有效改善缺血大鼠的神經(jīng)功能損傷,縮小皮質(zhì)梗死體積,促進(jìn)運(yùn)動(dòng)功能的恢復(fù)。LCN DBS能夠顯著增加缺血側(cè)和健側(cè)室管膜下區(qū)BrdU+Nestin+神經(jīng)干細(xì)胞和BrdU+DCX+成神經(jīng)細(xì)胞的數(shù)量,并增加梗死周邊BrdU+NeuN+的數(shù)量。還對腦卒中后第29天開始LCN DBS治療大鼠的神經(jīng)發(fā)生情況進(jìn)行了檢測,發(fā)現(xiàn)LCN DBS能夠顯著增加缺血側(cè)和健側(cè)室管膜下區(qū)BrdU+DCX+成神經(jīng)細(xì)胞的數(shù)量,以及梗死周邊BrdU+NeuN+的數(shù)量。結(jié)論LCN DBS促進(jìn)神經(jīng)功能恢復(fù)的機(jī)制可能與內(nèi)源性神經(jīng)干細(xì)胞增殖和分化相關(guān)。

      關(guān)鍵詞:腦深部電刺激;小腦齒狀核;腦卒中;神經(jīng)干細(xì)胞;運(yùn)動(dòng)功能

      T5-8

      基于神經(jīng)再生修復(fù)策略的藥物及電刺激康復(fù)治療手段

      王文

      (首都醫(yī)科大學(xué)宣武醫(yī)院,北京 100053;北京市老年病醫(yī)療研究中心,北京 100053)

      摘要:腦卒中等神經(jīng)系統(tǒng)疾病的關(guān)鍵臨床問題之一就是神經(jīng)再生修復(fù),以及相應(yīng)的促進(jìn)功能修復(fù)的治療方法研究。有效的康復(fù)治療能減輕腦損傷患者功能上的殘疾,加速康復(fù)進(jìn)程,降低潛在的治療費(fèi)用,減輕社會(huì)和家庭負(fù)擔(dān)。以腦卒中為例,目前除溶栓外仍缺乏有效的神經(jīng)保護(hù)及再生修復(fù)治療手段,存活患者中有70%~80%的病人不同程度喪失勞動(dòng)力,其中40%為中度功能障礙,15%~30%為重度殘疾,而臨床的康復(fù)治療效果受限于現(xiàn)有康復(fù)機(jī)制、康復(fù)理念以及康復(fù)治療技術(shù),亟待尋找有效的腦卒中再生修復(fù)治療靶點(diǎn)和新方法。我們課題組總結(jié)十多年的研究工作,針對內(nèi)源性干細(xì)胞再生修復(fù)治療缺血性腦損傷,提出了“神經(jīng)血管穩(wěn)態(tài)重構(gòu)”假說:腦卒中等損傷能誘發(fā)自體干細(xì)胞躍遷增殖,但該反應(yīng)持續(xù)時(shí)間較短,增殖逐漸回落到正常水平,不足以修復(fù)大腦功能;通過給予外源性藥物或電刺激等物理治療手段增強(qiáng)神經(jīng)發(fā)生、血管新生,以及血腦屏障重構(gòu)過程,促進(jìn)神經(jīng)血管單元穩(wěn)態(tài)恢復(fù),最終達(dá)到腦損傷后神經(jīng)功能修復(fù)的作用。在該理論基礎(chǔ)上,我們首次發(fā)現(xiàn)了單體化合物莫諾苷能顯著促進(jìn)腦卒中后神經(jīng)發(fā)生、血管新生過程,維持血腦屏障完整性并促進(jìn)微循環(huán)網(wǎng)絡(luò)重構(gòu),進(jìn)而改善缺血側(cè)皮層的神經(jīng)元功能,最終達(dá)到再生修復(fù)治療腦卒中的作用。通過表面等離子共振結(jié)合實(shí)驗(yàn)和轉(zhuǎn)錄組測序,以及體內(nèi)外基因敲除分子生物學(xué)技術(shù)研究手段,驗(yàn)證了莫諾苷促內(nèi)源性神經(jīng)干細(xì)胞增殖、分化為神經(jīng)元的關(guān)鍵靶點(diǎn)。

      隨著科技的進(jìn)步,腦機(jī)接口、人工智能等為醫(yī)學(xué)發(fā)展帶來了無限的可能。2018年首都醫(yī)科大學(xué)宣武醫(yī)院正式成立了中國國際神經(jīng)科學(xué)研究所類腦智能臨床轉(zhuǎn)化研究中心,并啟動(dòng)了國際“重拾行走計(jì)劃”在我國的首個(gè)項(xiàng)目。類腦智能中心依托于首都醫(yī)科大學(xué)宣武醫(yī)院強(qiáng)大的神經(jīng)學(xué)科優(yōu)勢,立足于類腦智能,致力于通過類腦智能的手段研究和解決神經(jīng)系統(tǒng)相關(guān)的疑難問題。以腦電信號分析技術(shù)、神經(jīng)影像技術(shù)和深度學(xué)習(xí)技術(shù)為橋梁,尋求在高級腦功能保護(hù)、神經(jīng)血管、癲癇、認(rèn)知疾病和神經(jīng)腫瘤等方面的重大原創(chuàng)突破。在這個(gè)背景下,我們與神經(jīng)外科密切合作,開始探索腦深部電刺激對內(nèi)源性神經(jīng)干細(xì)胞的作用及調(diào)控機(jī)制,發(fā)現(xiàn)了腦深部電刺激小腦齒狀核對側(cè)腦室下層神經(jīng)干細(xì)胞增殖分化的調(diào)控作用及對缺血性腦損傷神經(jīng)功能的修復(fù)作用,部分研究數(shù)據(jù)已在Scientific Reports發(fā)表。該研究進(jìn)一步驗(yàn)證了“神經(jīng)血管穩(wěn)態(tài)重構(gòu)”假說,并對內(nèi)源性干細(xì)胞治療腦卒中提供一定的指導(dǎo)意義。而對于老年癡呆、帕金森病等神經(jīng)退行性疾病,電刺激或許能重新激活大腦內(nèi)的神經(jīng)干細(xì)胞,配合藥物干預(yù)和康復(fù)訓(xùn)練,給慢性神經(jīng)系統(tǒng)疾病患者的神經(jīng)功能再生修復(fù)治療帶來希望。

      關(guān)鍵詞:神經(jīng)血管穩(wěn)態(tài)重構(gòu);內(nèi)源性神經(jīng)干細(xì)胞;莫諾苷;電刺激;腦卒中;神經(jīng)退行性疾??;神經(jīng)再生修復(fù)

      T5-9

      基于CRISPR系統(tǒng)的人工改造外泌體在藥理學(xué)研究中的應(yīng)用

      姚新剛

      (南方醫(yī)科大學(xué)藥學(xué)院,廣東省新藥篩選重點(diǎn)實(shí)驗(yàn)室,廣東 廣州 510515)

      摘要:目的CRISPR介導(dǎo)的基因編輯對基因治療帶來了巨大的前景,目前已有多項(xiàng)基于CRISPR的研究用于阿爾茨海默癥的治療。然而目前將CRISPR遞送到相應(yīng)的細(xì)胞甚至體內(nèi)應(yīng)用依然缺乏安全有效的手段,在細(xì)胞層面目前主要依賴于電轉(zhuǎn)手段將CRISPR系統(tǒng)運(yùn)送到細(xì)胞中,操作上比較復(fù)雜。而基于慢病毒和AAV的CRISPR遞送體系不僅會(huì)造成免疫原性,而且往往容易整合到基因組中長期表達(dá),引發(fā)不良后果。本研究擬基于外泌體的遞送策略改善該問題。方法本論文采用構(gòu)建工程化的外泌體平臺,采用基于com和配體結(jié)合蛋白Com的富集系統(tǒng),可以高效富集CRISPR系統(tǒng)并降低其脫靶效應(yīng)和免疫原性。結(jié)果構(gòu)建了多種RNP驗(yàn)證其效果,在細(xì)胞水平發(fā)現(xiàn)其富集RNP(spCas9,saCas9以及ABE系統(tǒng))效率可以達(dá)到10倍以上,同時(shí)其脫靶效率顯著降低。該系統(tǒng)不僅可以單獨(dú)包裝,也可以將多個(gè)RNP進(jìn)行共包裝,而且發(fā)現(xiàn)共包裝外泌體系統(tǒng)優(yōu)于單獨(dú)的外泌體相加。同時(shí)也在體內(nèi)水平驗(yàn)證了該系統(tǒng)的基因編輯效率,在del52hDMD/mdx小鼠模型中,注射包裝了DMD exon 53 RNP的外泌體,通過confocal也觀察到了肌肉dystrophin蛋白的表達(dá)。結(jié)論提供了一種免疫原性低于病毒系統(tǒng)的廣泛性的RNP遞送系統(tǒng),且其效率顯著高于同類型的外泌體系統(tǒng)?;谕饷隗w獨(dú)特的免疫原性和歸巢特性,以及穿過血腦屏障等特點(diǎn),該系統(tǒng)對于神經(jīng)性疾病的治療中具有潛在的意義。

      關(guān)鍵詞:CRISPR;人工改造外泌體;基因編輯

      通訊作者:姚新剛,E-mail:yaoxingang@smu.edu.cn

      猜你喜歡
      姜黃素阿爾茨海默病作用機(jī)制
      姜黃素干預(yù)耐吉非替尼肺腺癌細(xì)胞上皮間質(zhì)轉(zhuǎn)化的影響及相關(guān)機(jī)制的研究
      基于內(nèi)容分析法對阿爾茨海默病患者居家照護(hù)概念的解析
      瑣瑣葡萄多糖對阿爾茨海默病模型大鼠行為學(xué)和形態(tài)學(xué)的影響
      通過技術(shù)創(chuàng)新促進(jìn)我鎮(zhèn)農(nóng)業(yè)結(jié)構(gòu)調(diào)整
      HSP70敲低對AD轉(zhuǎn)基因果蠅的神經(jīng)保護(hù)作用
      云南山地仿野生撫育姜黃后姜黃素的含量變化研究
      冠心丹參方及其有效成分治療冠心病的研究進(jìn)展
      功能磁共振成像在輕度認(rèn)知障礙患者中的應(yīng)用研究進(jìn)展
      大數(shù)據(jù)對高等教育發(fā)展的推動(dòng)研究
      人間(2016年26期)2016-11-03 17:51:57
      帕羅西汀治療腦卒中后抑郁癥的臨床療效和安全性及作用機(jī)制
      华阴市| 云林县| 天津市| 大连市| 廉江市| 涞源县| 青海省| 辽阳县| 西城区| 信宜市| 林西县| 浙江省| 闻喜县| 石门县| 祁门县| 澄迈县| 锡林郭勒盟| 都匀市| 白朗县| 循化| 柞水县| 廉江市| 泰兴市| 广元市| 靖安县| 石首市| 九龙县| 丰宁| 新闻| 临江市| 通江县| 阿拉尔市| 彰武县| 临澧县| 那坡县| 临高县| 本溪| 新乐市| 黑龙江省| 望江县| 呼玛县|