本文選題：麻醉藥　切入點(diǎn)：電休克　出處：《重慶醫(yī)科大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：目的研究異丙酚及小劑量氯胺酮對(duì)抑郁大鼠電休克(electroconvulsive shock,ECS)后空間學(xué)習(xí)記憶功能以及海馬內(nèi)谷氨酸N-甲基-D-天冬氨酸受體(N-methyl-D-aspartic acid receptor,NMDAR)和α-氨基-3-羥基-5-甲基-4-異惡唑丙酸受(α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor,AMPAR)蛋白表達(dá)、功能和長時(shí)程增強(qiáng)(long-term potentiation,LTP)的影響,以探討調(diào)控NMDAR介導(dǎo)的突觸再可塑性改變在麻醉藥異丙酚和小劑量氯胺酮保護(hù)抑郁大鼠ECS后學(xué)習(xí)記憶功能中的作用機(jī)制。方法第一部分2~3月齡健康雄性Sprague Dawley(SD)大鼠,抑郁大鼠模型的建立采用慢性輕度不可預(yù)見性應(yīng)激(chronic unpredictable mild stress,CUMS)。80只抑郁模型大鼠隨機(jī)分為8組(n=10):D組、DP組、DE組、DPE組、DK組、DKE組、DPKE組、DPME組。10只同期喂養(yǎng)的健康雄性SD大鼠為對(duì)照組(C組)。D組腹腔注射生理鹽水(normal saline,NS)8ml/kg后給予假ECS處理(雙耳夾電極,但不通電);DP組腹腔注射異丙酚80mg/kg,而后給予行假ECS處理;DE組腹腔注射NS8ml/kg后給予ECS處理;DPE組腹腔注射異丙酚80mg/kg后行ECS;DK組腹腔注射氯胺酮10mg/kg;DKE組腹腔注射氯胺酮10mg/kg后行ECS;DPKE組腹腔注射氯胺酮10mg/kg和異丙酚80mg/kg行ECS;DPME組腹腔注射MK801 1mg/kg和異丙酚80mg/kg后行ECS。上述處理每日1次,連續(xù)7d。C組大鼠不給予任何處理。采用糖水偏好實(shí)驗(yàn)(sucrose preference test,SPT)檢測糖水偏好百分比(sucrose preference percentage,SPP)、曠場實(shí)驗(yàn)(open field test,OFT)檢測直立次數(shù)、穿格次數(shù),評(píng)估大鼠抑郁樣行為;采用Morris水迷宮評(píng)價(jià)大鼠空間學(xué)習(xí)記憶功能;采用western-blot檢測大鼠海馬NR1、NR2A、NR2B、Glu R1蛋白表達(dá)水平。第二部分2~3月齡健康雄性SD大鼠,采用CUMS建立抑郁大鼠模型。72只抑郁模型大鼠隨機(jī)分為6組(n=12):D組、DE組、DK組、DPE組、DPKE組、DPME組。12只同期喂養(yǎng)的健康雄性SD大鼠為C組。各組大鼠處理方法同第一部分所述,C組大鼠不予以處理。采用細(xì)胞外記錄檢測海馬Schaffer側(cè)枝-CA1神經(jīng)通路NMDAR、AMPAR介導(dǎo)的興奮性突觸后場電位(field excitatory postsynaptic potential,f EPSP);檢測各組大鼠海馬Schaffer側(cè)枝-CA1神經(jīng)通路高頻電刺激(protocol1和protocol2,protocol2刺激強(qiáng)度大于protocol1)的LTP誘導(dǎo)反應(yīng)差異;檢測含不同濃度異丙酚和氯胺酮的人工腦脊液灌流期間該神經(jīng)通路f EPSP的變化。結(jié)果第一部分實(shí)驗(yàn)1:(1)實(shí)驗(yàn)處理前,D組、DE組、DKE組SPP、直立次數(shù)和穿格次數(shù)組間差異無統(tǒng)計(jì)學(xué)意義,均低于C組(P0.01)。實(shí)驗(yàn)處理后,與D組相比,DE組和DK組SPP、直立次數(shù)和穿格次數(shù)均升高,(P0.01);DE組SPP、直立次數(shù)高于DK組(P0.01)。(2)實(shí)驗(yàn)處理前,D組、DE組、DK組大鼠逃避潛伏期和目標(biāo)象限探索時(shí)間組間差異無統(tǒng)計(jì)學(xué)意義(P0.05),其逃避潛伏期長于C組而目標(biāo)象限探索時(shí)間短于C組(P0.01)。實(shí)驗(yàn)處理后,與C組、D組、DK組相比,DE組逃避潛伏期延長,目標(biāo)象限探索時(shí)間縮短(P0.01);D組、DK組大鼠逃避潛伏期長于C組,而目標(biāo)象限探索時(shí)間短于C組;D組、DK組兩指標(biāo)組間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。(3)D組、DE組、DK組大鼠Glu R1蛋白表達(dá)水平均低于C組(P0.01);與D組相比,DE組、DK組大鼠海馬Glu R1表達(dá)水平升高(P0.01),而DE組與DK組大鼠海馬Glu R1蛋白表達(dá)水平差異無統(tǒng)計(jì)學(xué)意義(P0.05)。與C組相比,D組、DE組、DK組大鼠海馬的NR1、NR2A、NR2B蛋白表達(dá)水平降低(P0.05);與D組相比,DE組大鼠海馬NR1、NR2A、NR2B蛋白表達(dá)水平降低,而DK組大鼠NR1、NR2B亞基蛋白表達(dá)水平升高,差異有統(tǒng)計(jì)學(xué)意義(P0.05)。D組及DK組大鼠海馬NR2A蛋白表達(dá)水平差異無統(tǒng)計(jì)學(xué)意義(P0.05)。各組大鼠NR2B與NR2A亞基蛋白表達(dá)水平的比例(NR2B/NR2A)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)2:(1)實(shí)驗(yàn)處理前,D組、DP組、DE組、DPE組、DKE組大鼠SPP、直立次數(shù)、穿格次數(shù)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)處理后,與D組相比,DE組、DPE組和DKE組大鼠SPP、直立次數(shù)、穿格次數(shù)明顯升高(P0.01);DPE組大鼠SPP低于DE組和DKE組(P0.01),而直立次數(shù)和穿格次數(shù)與DE組差異無統(tǒng)計(jì)學(xué)意義(P0.05);DE組和DKE組大鼠SPP差異無統(tǒng)計(jì)學(xué)意義,(P0.05);DKE組直立次數(shù)和穿格次數(shù)高于DE組(P0.01)。(2)實(shí)驗(yàn)處理前,各組大鼠逃避潛伏期、目標(biāo)象限探索時(shí)間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)處理后,與D組相比,DE組、DPE組、DKE組逃避潛伏期延長,目標(biāo)象限探索時(shí)間縮短(P0.01);與DE組相比,DPE組、DKE組大鼠逃避潛伏期縮短,而目標(biāo)象限探索時(shí)間延長(P0.01);D組、DP組大鼠逃避潛伏期和目標(biāo)象限探索時(shí)間差異無統(tǒng)計(jì)學(xué)意義(P0.05),DPE組和DKE組組間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。(3)與D組相比,DE組、DPE組以及DKE組大鼠海馬Glu R1蛋白表達(dá)水平明顯升高(P0.01)。D組大鼠海馬Glu R1蛋白表達(dá)水平與DP組無明顯差異(P0.05),DE組、DPE組、DKE組大鼠海馬Glu R1蛋白表達(dá)水平差異無統(tǒng)計(jì)學(xué)意義(P0.05)。與D組相比,DE組、DPE組、DKE組大鼠海馬的NR1、NR2B蛋白表達(dá)水平明顯降低(P0.05);DPE組、DKE組大鼠海馬NR1、NR2B蛋白表達(dá)水平高于DE組(P0.05)。與D組相比,DE組、DKE組大鼠海馬NR2A蛋白表達(dá)水平降低,而DPE組大鼠海馬NR2A表達(dá)水平高于D組和DKE組(P0.01)。與D組相比,DKE組NR2B/NR2A明顯升高,而DPE組降低(P0.05)。D組、DP組、DE組NR2B/NR2A差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)3:(1)實(shí)驗(yàn)處理前,D組、DPE組、DPKE組、DPME組SPP、直立次數(shù)、穿格次數(shù)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)處理后,與D組相比,DPE組、DPKE組和DPME組SPP、直立次數(shù)和穿格次數(shù)升高(P0.01);與DPE組相比,DPKE組和DPME組大鼠SPP、直立次數(shù)升高(P0.01),DPKE組和DPME組大鼠SPP、直立次數(shù)和穿格次數(shù)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。(2)實(shí)驗(yàn)處理前,各組大鼠逃避潛伏期和目標(biāo)象限探索時(shí)間組間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)處理后,與DPE組相比,DPKE組和DPME組逃避潛伏期縮短、目標(biāo)象限探索時(shí)間延長(P0.05);D組、DPKE組、DPME組逃避潛伏期和目標(biāo)象限探索時(shí)間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。(3)與D組相比,DPE組、DPKE組、DPME組大鼠海馬Glu R1蛋白表達(dá)水平升高(P0.01);DPKE組大鼠海馬Glu R1蛋白表達(dá)水平高于DPE組(P0.01);DPE組與DPME組大鼠海馬Glu R1蛋白表達(dá)水平差異無統(tǒng)計(jì)學(xué)意義(P0.05)。各組大鼠海馬NR2A亞基的蛋白表達(dá)水平差異無統(tǒng)計(jì)學(xué)意義(P0.05)。與DPE組相比,DPKE組、DPME組大鼠海馬NR1、NR2B蛋白表達(dá)水平以及NR2B/NR2A明顯升高(P0.05)。與D組相比,DPKE組和DPME組大鼠海馬NR2B蛋白表達(dá)水平升高(P0.05);DPKE組和DPME組大鼠海馬NR2B蛋白表達(dá)水平組間差異無統(tǒng)計(jì)學(xué)意義(P0.05)。D組、DPKE組、DPME組大鼠海馬NR1蛋白表達(dá)水平和NR2B/NR2A差異無統(tǒng)計(jì)學(xué)意義(P0.05)。第二部分實(shí)驗(yàn)1:(1)與C組相比,D組大鼠NMDAR I/O明顯降低;與D組相比,DK組大鼠NMDAR I/O明顯升高,而DE組大鼠NMDAR I/O明顯降低(P0.01)。與C組相比,D組大鼠AMPAR介導(dǎo)的f EPSP降低(P0.01);與D組相比,DK組、DE組大鼠AMPAR介導(dǎo)的f EPSP明顯升高,而DE組高于DK組,差異有統(tǒng)計(jì)學(xué)意義(P0.01)。(2)protocol1高頻刺激后,與C組相比,D組、DE組、DK組大鼠Schaffer側(cè)枝-CA1神經(jīng)通路LTP幅度降低;DE組大鼠LTP幅度低于D組和DK組(P0.01),D組和DK組大鼠LTP幅度差異無統(tǒng)計(jì)學(xué)意義(P0.05)。C組、DE組大鼠海馬Schaffer側(cè)枝-CA1通路protocol2誘導(dǎo)的LTP幅度高于protocol1,(P0.05);D組protocol2誘導(dǎo)的LTP幅度高于protocol1,但差異無統(tǒng)計(jì)學(xué)意義(P0.05)。實(shí)驗(yàn)2:(1)與DE組相比,DPE、DPKE組、DPME組大鼠NMDAR I/O明顯升高(P0.01);與DPE組相比,DPKE組、DPME組大鼠NMDAR I/O升高,DPME組大鼠NMDAR I/O低于DPKE組(P0.01)。(2)與DE組相比,DPE、DPKE組、DPME組大鼠LTP幅度明顯升高(P0.01);與DPE組相比,DPKE組、DPME組大鼠LTP幅度升高(P0.01);DPKE組與DPME組大鼠LTP幅度差異無統(tǒng)計(jì)學(xué)意義(P0.05)。(3)10μM及20μM異丙酚灌流均可顯著抑制大鼠海馬Schaffer側(cè)枝-CA1通路f EPSP;20μM異丙酚抑制作用強(qiáng)于10μM(P0.01),起效更快,抑制程度更高。1μM氯胺酮灌流對(duì)大鼠海馬Schaffer側(cè)枝-CA1通路f EPSP無明顯作用,10μM氯胺酮可輕度增加f EPSP幅度,而兩劑量氯胺酮灌流期間f EPSP差異無統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論(1)ECS抗抑郁機(jī)制與其上調(diào)抑郁大鼠海馬Glu R1表達(dá)和AMPAR功能有關(guān);而其損傷抑郁大鼠空間記憶功能與下調(diào)海馬NMDAR表達(dá)量及Schaffer側(cè)枝CA1神經(jīng)通路NMDAR功能有關(guān)。(2)異丙酚可部分逆轉(zhuǎn)ECS對(duì)NMDAR表達(dá)和功能的下調(diào)作用,減輕ECS所致的學(xué)習(xí)記憶功能損傷;異丙酚麻醉下,NMDAR阻滯劑(氯胺酮和MK801)可進(jìn)一步減輕ECS對(duì)NMDAR表達(dá)和功能的下調(diào)作用,減輕ECS所致的學(xué)習(xí)記憶功能損傷。(3)激活NMDAR所介導(dǎo)的突觸再可塑性改變是ECS下調(diào)NMDAR表達(dá)和功能的機(jī)制;異丙酚及氯胺酮阻滯ECS對(duì)NMDAR的激活,是其逆轉(zhuǎn)ECS下調(diào)NMDAR表達(dá)和功能,進(jìn)而發(fā)揮學(xué)習(xí)記憶功能保護(hù)作用的機(jī)制。
[Abstract]:Objective to study the effect of propofol and ketamine for electroconvulsive therapy in depressed rats (electroconvulsive shock, ECS) after the spatial learning and memory function and hippocampal glutamate N- methyl -D- aspartate receptor (N-methyl-D-aspartic acid receptor, NMDAR -3-) and alpha amino hydroxy -5- methyl isoxazole propionate by -4- (alpha -amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor, AMPAR) the expression, function and long term potentiation (long-term potentiation, LTP) to investigate the effects of NMDAR mediated regulation of synaptic plasticity and mechanisms of learning and memory function in the anesthetic propofol and ketamine protect depression after ECS in rats. Methods the first part of the 2~3 month old male Sprague (Dawley SD) rats, establishment of rat model of depression by chronic unpredictable mild stress (chronic unpredictable mild stress, CUMS).80 suppression Depression model rats were randomly divided into 8 groups (n=10): D group, DP group, DE group, DPE group, DK group, DKE group, DPKE group, healthy male SD rats DPME group.10 rats fed over the same period as control group (group C).D group received intraperitoneal injection of saline (normal saline, NS 8ml/kg) sham ECS treatment (ear clip electrodes, but no electricity); DP group received intraperitoneal injection of propofol 80mg/kg, and then give the sham ECS treatment; group DE received intraperitoneal injection of NS8ml/kg after treated with ECS; DPE group received intraperitoneal injection of propofol 80mg/kg ECS after intraperitoneal injection of ketamine 10mg/kg; DK group; DKE group received intraperitoneal injection of ketamine 10mg/kg after ECS; DPKE group received intraperitoneal injection of ketamine and propofol for 10mg/kg 80mg/kg ECS; DPME group intraperitoneal injection of MK801 1mg/kg and 80mg/kg ECS. after the treatment of propofol 1 times a day, continuous 7d.C group were not given any treatment. The sucrose preference test (sucrose preference test SPT (s) detection of sucrose preference percentage Ucrose preference percentage, SPP), open field test (open field, test, OFT) detection of vertical frequency, the number of crossing line, to evaluate the depression like behavior of rats; to assess learning and memory function of rats by Morris water maze; using Western-blot detection of rat hippocampal NR1, NR2A, NR2B, Glu expression level of R1 protein. The second part 2~3 month old healthy male SD rats, using CUMS to establish the rat model of depression.72 depression model rats were randomly divided into 6 groups (n=12): D group, DE group, DK group, DPE group, DPKE group, healthy male SD rats group DPME.12 synchronous feeding for C group. Each group in the same way as the first part of the treatment, C group rats were treated by extracellular recording. Detection of hippocampal Schaffer collateral -CA1 neural pathway NMDAR, excitatory postsynaptic field potentials mediated by AMPAR (field excitatory postsynaptic potential, f; EPSP) were measured in rat hippocampal Schaffer collateral -CA1 God 緇忛，

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