本文關(guān)鍵詞： 心理應(yīng)激 束縛應(yīng)激 咬肌 能量代謝 自由基代謝 三叉神經(jīng)運(yùn)動(dòng)核 神經(jīng)遞質(zhì)　出處：《第四軍醫(yī)大學(xué)》2014年博士論文　論文類型：學(xué)位論文

【摘要】：隨著我國經(jīng)濟(jì)的快速發(fā)展、社會(huì)的劇烈變革、人們生活方式的轉(zhuǎn)變，來自于學(xué)業(yè)、工作、家庭等各方面的精神壓力不斷困擾著人們，由此造成的心理疾病和心身疾病的發(fā)病率逐年攀升。負(fù)面性的生活事件產(chǎn)生的心理應(yīng)激對(duì)人體造成的影響不僅體現(xiàn)在精神疾病方面，大量的證據(jù)也表明心理應(yīng)激刺激在顳下頜關(guān)節(jié)紊亂病、夜磨牙癥等口頜系統(tǒng)疾病的發(fā)生和發(fā)展過程中都扮演了重要的角色；研究也證實(shí)長時(shí)間的應(yīng)激刺激不僅會(huì)引起焦慮、抑郁等心理改變，還會(huì)導(dǎo)致口頜肌肉的病理變化，以及咀嚼肌的緊張度升高。心理應(yīng)激環(huán)境下出現(xiàn)的咀嚼肌肌緊張度顯著上升，可能不僅僅是應(yīng)激觸發(fā)的一種機(jī)體反應(yīng)，同時(shí)也可能是心理應(yīng)激導(dǎo)致顳下頜關(guān)節(jié)紊亂病產(chǎn)生或加重的內(nèi)在病理過程。然而到目前為止，學(xué)界對(duì)這一現(xiàn)象內(nèi)在機(jī)制的認(rèn)識(shí)仍為空白，了解這一現(xiàn)象的內(nèi)在機(jī)制，并據(jù)此建立有效的預(yù)防措施和合理的治療方法對(duì)心理應(yīng)激相關(guān)口頜系統(tǒng)疾病的治療將產(chǎn)生深遠(yuǎn)的影響。 基于以上原因，我們建立了大鼠慢性束縛應(yīng)激模型，并采用肌肉電生理技術(shù)、高效液相色譜技術(shù)、Elisa、Western Blot、qRT-PCR等分子生物學(xué)技術(shù)，對(duì)長期束縛應(yīng)激對(duì)大鼠咬肌及腦內(nèi)相關(guān)核團(tuán)的影響，及相應(yīng)變化的內(nèi)在機(jī)制進(jìn)行了研究，同時(shí)對(duì)拉莫三嗪在上述病態(tài)變化中的治療效果進(jìn)行了評(píng)估。 本研究共分為2大部分，具體內(nèi)容如下： 第一部分心理應(yīng)激動(dòng)物模型的建立及評(píng)價(jià) 采用慢性束縛應(yīng)激（chronic restraint stress，CRS）方法建立心理應(yīng)激模型，將大鼠隨機(jī)分入對(duì)照組、心理應(yīng)激組、20mg/kg低劑量藥物組、30mg/kg中劑量藥物組和40mg/kg高劑量藥物組，在束縛應(yīng)激3周后，對(duì)大鼠體重進(jìn)行檢測，同時(shí)采用糖水偏好實(shí)驗(yàn)、曠場實(shí)驗(yàn)等行為學(xué)評(píng)估方法以及促腎上腺皮質(zhì)激素（adrenocorticotropichormone, ACTH）、皮質(zhì)酮（corticosterone, CORT）血清學(xué)激素測定對(duì)慢性束縛應(yīng)激模型進(jìn)行評(píng)估。實(shí)驗(yàn)結(jié)果顯示，心理應(yīng)激后大鼠體重較應(yīng)激前顯著下降（p＜0.05）；糖水偏好程度顯著降低（p＜0.005），應(yīng)激大鼠在曠場中移動(dòng)總距離及移動(dòng)速率顯著降低（p＜0.005），中央停留時(shí)間顯著延長（p＜0.005）；應(yīng)激大鼠血清CORT（p＜0.005）和ACTH的含量顯著升高（p＜0.005）。以上結(jié)果證實(shí)，本實(shí)驗(yàn)所建立的慢性束縛應(yīng)激大鼠模型存在明顯的應(yīng)激相關(guān)的行為學(xué)和血清學(xué)特征，可以滿足后續(xù)實(shí)驗(yàn)的要求。 第二部分心理應(yīng)激對(duì)大鼠咬肌肌緊張度及腦內(nèi)神經(jīng)遞質(zhì)變化的影響及藥物干預(yù)研究 1.心理應(yīng)激對(duì)大鼠咬肌能量代謝的影響及藥物干預(yù)研究 應(yīng)激后大鼠咬肌內(nèi)肌酸激酶（creatine kinase, CK）、乳酸脫氫酶（lactatedehydrogenase, LDH）和乳酸（lactate, LD）的含量顯著升高（p0.005），磷酸肌酸（creatine phosphate, CP）含量顯著降低（p0.005）；30和40mg/kg劑量的拉莫三嗪可以有效逆轉(zhuǎn)應(yīng)激引起的上述變化（p0.005）；20mg/kg劑量的拉莫三嗪則無明顯效果(p＞0.005)。 2.心理應(yīng)激對(duì)大鼠咬肌肌電圖(electromyography, EMG)的影響及藥物干預(yù)研究 不論在清醒安靜狀態(tài)還是清醒活動(dòng)狀態(tài)下，應(yīng)激后大鼠咬肌肌緊張度較應(yīng)激前均顯著升高(p＜0.05)；30和40mg/kg劑量的拉莫三嗪可以有效逆轉(zhuǎn)應(yīng)激后出現(xiàn)的肌緊張水平上升（p0.005），20mg/kg劑量的拉莫三嗪則無明顯效果(p＞0.005)。 3.心理應(yīng)激對(duì)大鼠腦內(nèi)自由基代謝的影響及藥物干預(yù)研究 應(yīng)激后大鼠腦中中央杏仁核(central amygdaloid nucleus, CeA)、小細(xì)胞網(wǎng)狀結(jié)構(gòu)(parvocellular reticular nucleus, PCRt)及三叉神經(jīng)運(yùn)動(dòng)核（motor nucleus of trigeminalnerve, Vm)內(nèi)谷胱甘肽過氧化物酶（glutathione peroxidase, GSH-Px）、過氧化氫酶（catalase, CAT）、超氧化物歧化酶（superoxide dismutase, SOD）的活性較對(duì)照組均出現(xiàn)顯著下降（p0.005）；丙二醛（malondialdehyde, MDA）的含量則顯著升高（p0.005）；30和40mg/kg劑量的拉莫三嗪可以有效逆轉(zhuǎn)心理應(yīng)激引起的上述變化（p0.005）；20mg/kg劑量的拉莫三嗪則無明顯心理應(yīng)激對(duì)抗效果(p＞0.005)。 4.心理應(yīng)激對(duì)大鼠腦內(nèi)神經(jīng)遞質(zhì)變化的影響及藥物干預(yù)研究 應(yīng)激后大鼠腦中上述核團(tuán)內(nèi)谷氨酸（glutamate，，Glu）含量、谷氨酰胺酶（glutaminase，GLS）活性、囊泡谷氨酸轉(zhuǎn)運(yùn)體蛋白1(vesicular glutamate transporter1，VGluT1)及囊泡谷氨酸轉(zhuǎn)運(yùn)體蛋白2(vesicular glutamate transporter2，VGluT2)的含量等均顯著升高（p0.005），谷氨酰胺合成酶（glutamine synthetase，GS）的活性顯著下降（p0.005）；杏仁核內(nèi)NMDAR2a（N-methyl-D-aspartate receptor2a）表達(dá)水平顯著升高（p0.005）；三叉神經(jīng)運(yùn)動(dòng)核內(nèi)NMDAR1（N-methyl-D-aspartate receptor1）表達(dá)顯著升高（p0.005）；20、30和40mg/kg劑量的拉莫三嗪對(duì)上述變化無明顯的治療效果（p0.005）。 通過本研究我們發(fā)現(xiàn)：慢性束縛應(yīng)激會(huì)造成大鼠咬肌肌緊張度增高、咬肌能量代謝異常；而咬肌肌緊張度增高是由三叉神經(jīng)運(yùn)動(dòng)核內(nèi)谷氨酸的水平增高所引起；GLS和GS的活性改變可能是導(dǎo)致應(yīng)激后三叉神經(jīng)運(yùn)動(dòng)核內(nèi)谷氨酸濃度升高的主要原因，VGluT1和VGluT2的表達(dá)上調(diào)可能對(duì)細(xì)胞外谷氨酸水平產(chǎn)生了影響，而NMDAR1的表達(dá)增加可能導(dǎo)致了NMDA受體的表達(dá)上調(diào)，繼而導(dǎo)致神經(jīng)元細(xì)胞對(duì)谷氨酸濃度的增加更為敏感，這些因素共同作用引發(fā)了三叉神經(jīng)運(yùn)動(dòng)核內(nèi)運(yùn)動(dòng)神經(jīng)元的異常神經(jīng)沖動(dòng)，最終導(dǎo)致了咬肌肌緊張度的升高；此外應(yīng)激還導(dǎo)致了三叉神經(jīng)運(yùn)動(dòng)核組織的氧化應(yīng)激損傷；應(yīng)激后三叉神經(jīng)運(yùn)動(dòng)核內(nèi)谷氨酸濃度增加的部分原因可能源于來自于其上級(jí)神經(jīng)元—小細(xì)胞網(wǎng)狀結(jié)構(gòu)及杏仁核的興奮性神經(jīng)遞質(zhì)的增加；30、40mg/kg劑量的拉莫三嗪針對(duì)谷氨酸濃度濃度的干預(yù)效應(yīng)并不依賴于其對(duì)谷氨酸代謝酶、囊泡谷氨酸轉(zhuǎn)運(yùn)體蛋白及谷氨酸受體亞基的調(diào)節(jié)作用，而可能與拉莫三嗪抑制谷氨酸向突觸間隙內(nèi)的釋放有關(guān)。 本研究初步闡明了心理應(yīng)激導(dǎo)致咬肌肌緊張度增高現(xiàn)象的內(nèi)在原因，并且證實(shí)了拉莫三嗪對(duì)抗心理應(yīng)激負(fù)效應(yīng)的潛在治療效果，豐富了心理應(yīng)激導(dǎo)致口頜系統(tǒng)疾病的致病機(jī)理，為未來制定應(yīng)激相關(guān)疾病的治療方法提供了實(shí)驗(yàn)參考依據(jù)。
[Abstract]:The psychological stress caused by psychological stress is not only caused by psychological changes such as anxiety , depression and so on , but also can be the intrinsic pathological process of psychological stress stimulation in the development and development of oral and maxillofacial diseases such as temporo - mandibular joint disorder and night grinding . Based on the above reasons , we established the rat chronic restraint stress model , and adopted molecular biology techniques such as muscle electrophysiology technique , high performance liquid chromatography ( HPLC ) , ELISA ( Western Blot ) , qRT - PCR ( Western Blot ) and qRT - PCR . This study is divided into 2 parts , the concrete contents are as follows : Establishment and Evaluation of the Animal Model of Psychological Stress in Part I The rats were randomly divided into control group , psychological stress group , 20 mg / kg low - dose group , 30 mg / kg medium - dose group and 40 mg / kg high - dose drug group . Effect of second partial psychological stress on muscle tone and neurotransmitter in rats and study on drug intervention 1 . Effect of psychological stress on energy metabolism in rats and study on drug intervention The contents of creatine kinase ( CK ) , lactate dehydrogenase ( LDH ) and lactate ( LD ) in rats after stress were significantly increased ( p > 0.005 ) , creatine phosphate ( CP ) content was significantly lower ( p > 0.005 ) , 30 and 40 mg / kg of lamotrigine could effectively reverse the above - mentioned changes induced by stress ( p > 0.005 ) ; and 20 mg / kg of lamotrigine had no significant effect ( p > 0.005 ) . 2 . Effect of psychological stress on EMG and drug intervention in rats The muscle tone after stress was significantly increased ( p & lt ; 0.05 ) , 30 and 40 mg / kg of lamotrigine can effectively reverse the increase of muscle tone after stress ( p & lt ; 0 . 005 ) , and 20 mg / kg of lamotrigine has no obvious effect ( p & gt ; 0.005 ) , whether in the conscious and quiet state or in the conscious active state . 3 . Effect of psychological stress on free radical metabolism in rat brain and drug intervention The activities of glutathione peroxidase ( GSH - Px ) , catalase ( GSH - Px ) , catalase ( catalase , CAT ) and superoxide dismutase ( SOD ) in the brain of the rats after stress were significantly lower than those in the control group ( p > 0.005 ) ; the content of malondialdehyde ( MDA ) increased significantly ( p > 0.005 ) ; 30 and 40 mg / kg of lamotrigine could effectively reverse the above - mentioned changes caused by psychological stress ( p > 0.005 ) ; 30 and 40 mg / kg of lamotrigine had no obvious psychological stress - antagonistic effect ( p > 0.005 ) . 4 . Effect of psychological stress on neurotransmitters in brain of rats and study on drug intervention The contents of glutamate ( Glu ) , glutamine ( GLS ) , glutamate transporter 1 ( VGluT1 ) and glutamate transporter 2 ( VGluT2 ) in the brain of rats after stress were significantly increased ( p . 005 ) . The expression level of NMDAR2a ( N - methyl - D - aspartate receptor1 ) increased significantly ( p . 005 ) . The increase of glutamate concentration in the nucleus of trigeminal nerve was caused by the increase of the expression of GLS and GS , which could result in the increase of glutamate concentration in the nucleus of trigeminal nerve . In this study , the intrinsic reason of stress induced by psychological stress and the potential therapeutic effects of lamotrigine against psychological stress were demonstrated , and the mechanism of psychological stress induced by psychological stress was enriched , which provided an experimental reference for the future development of stress - related diseases .

【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R782

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本文編號(hào)：1501926