本文選題：腰椎間盤突出癥 + 神經(jīng)病理性疼痛　； 參考：《蘇州大學》2015年碩士論文

【摘要】：目的1.探究P2X3受體在大鼠腰椎間盤突出誘導的神經(jīng)病理性疼痛中的作用;2.探討H2S信號分子在調(diào)節(jié)L5/L6背根神經(jīng)節(jié)P2X3受體表達和功能中的作用特征。方法1.采用200-220g SD雄性大鼠,取出自體尾椎的正常髓核組織,置于手術(shù)顯露后的腰5,6左側(cè)神經(jīng)根處,建立腰椎間盤突出癥動物模型(LDH)。分別在術(shù)前1天,術(shù)后3,7,14,21,28和35天運用VFF(Von Frey filaments)及雙足平衡實驗(Weight Bearing)的方法來評測SD大鼠的行為學疼痛閾值反應。2.運用逆行標記的熒光素(Dil)來標記支配后肢的L5/L6背根神經(jīng)節(jié)(DRG)中、小神經(jīng)元,運用鈣成像方法研究腰椎間盤突出癥(LDH)大鼠L5/L6 DRG神經(jīng)元中ATP誘導的細胞內(nèi)鈣信號反應及神經(jīng)元的興奮性變化。3.運用Western Blotting方法檢測LDH模型大鼠L5/L6 DRG神經(jīng)元中內(nèi)源性硫化氫合酶CBS和CSE以及嘌呤受體P2XRs的蛋白表達情況。4.運用免疫組織化學方法檢測LDH大鼠L5/L6 DRG神經(jīng)元中CBS和P2X3R蛋白共表達情況。結(jié)果1.自體尾椎髓核(NP)移植能夠顯著降低LDH模型大鼠后肢的機械性疼痛閾值,并且這一現(xiàn)象是與L5/L6 DRG神經(jīng)元中硫化氫合酶CBS蛋白的表達上調(diào)相關聯(lián)的。2.與假手術(shù)組(Sham)相比,LDH組增加了L5/L6背根神經(jīng)節(jié)(DRG)中P2X3R的蛋白表達量,而P2X2R,P2X1R的蛋白表達量并沒有發(fā)生變化。3.對LDH大鼠鞘內(nèi)注射硫化氫合酶CBS抑制劑AOAA后,發(fā)現(xiàn)AOAA能夠顯著的翻轉(zhuǎn)LDH大鼠的疼痛閾值,并呈現(xiàn)劑量和時間上的依賴性。而作為對照,AOAA并不能夠?qū)偈中g(shù)組SD大鼠的疼痛閾值產(chǎn)生作用。4.對LDH大鼠鞘內(nèi)注射P2X3R的選擇性拮抗劑A-317491后,發(fā)現(xiàn)A-317491能夠顯著的翻轉(zhuǎn)LDH大鼠的疼痛閾值,并呈現(xiàn)劑量和時間上的依賴性。5.運用免疫組化的實驗方法檢測后發(fā)現(xiàn)硫化氫合酶CBS與P2X3R在L5/L6神經(jīng)元中能夠共表達。而且運用Western Blotting的方法進行檢測后發(fā)現(xiàn),連續(xù)給予LDH大鼠AOAA能夠顯著翻轉(zhuǎn)NP引起的L5/L6神經(jīng)元中的P2X3R表達的上調(diào)。6.全細胞膜片鉗記錄顯示,與Sham組相比,在LDH大鼠L5/L6 DRG上,NP能顯著增強ATP誘導的細胞內(nèi)鈣信號。鞘內(nèi)連續(xù)給與硫化氫合酶CBS抑制劑AOAA后可以抑制LDH大鼠中ATP誘導的鈣信號變化。7.通過單次或連續(xù)給與LDH大鼠足趾注射硫化氫合酶CBS抑制劑AOAA后,發(fā)現(xiàn)AOAA不能夠翻轉(zhuǎn)LDH大鼠的疼痛閾值,而且運用Western Blotting的方法進行檢測后發(fā)現(xiàn),連續(xù)足趾給藥后大鼠L5/L6神經(jīng)元中P2X3R表達未發(fā)生變化。結(jié)論腰椎間盤突出癥模型(LDH)中髓核(NP)激活了內(nèi)源性硫化氫(H2S)合酶CBS表達的上調(diào),通過大鼠DRG中初級感覺神經(jīng)元,調(diào)節(jié)了L5/L6神經(jīng)元中P2X3受體蛋白的表達,從而增加L5/L6神經(jīng)元的興奮性進而產(chǎn)生神經(jīng)病理性疼痛;我們的研究結(jié)果表明,內(nèi)源性硫化氫(H2S)合酶CBS表達的上調(diào)可能在LDH大鼠椎間盤源性疼痛過程中扮演著重要的角色。因此,本研究能夠在一定程度上揭示根源性疼痛產(chǎn)生的分子機制,從而為神經(jīng)病理性疼痛的治療提供更為有效的治療方案。
[Abstract]:Objective 1. To explore the role of P2X3 receptor in neuropathic pain induced by lumbar disc herniation in rats. To investigate the role of H _ 2S signaling molecules in regulating the expression and function of P2X3 receptor in L5/L6 dorsal root ganglion. Method 1. The normal nucleus pulposus of the autologous tail vertebrae was removed from 200-220g SD male rats and placed at the left nerve root of the left lumbar vertebrae after operation. The animal model of lumbar disc herniation was established. The behavioral pain threshold responses of SD rats were evaluated by VFF(Von Frey filaments and weight balance test 1 day before operation, 3 days after operation, and 35 days after operation. Small neurons in the dorsal root ganglion of L5/L6, which innervate the hind limbs, were labeled with retrograde fluorescein Dil. Calcium imaging was used to study the intracellular calcium signal response induced by ATP and the changes of neuronal excitability in L5/L6 DRG neurons of rats with lumbar disc herniation. The expression of endogenous hydrogen sulfide synthase (CBS) CSE and purine receptor P2XRs (P2XRs) in L5/L6 DRG neurons of LDH model rats was detected by Western Blotting method. The co-expression of CBS and P2X3R in L5/L6 DRG neurons of LDH rats was detected by immunohistochemical method. Result 1. Autologous transplantation of nucleus pulposus of caudal vertebrae can significantly reduce the threshold of mechanical pain in the hind limbs of LDH model rats, and this phenomenon is associated with the up-regulation of CBS protein expression in L5/L6 DRG neurons. Compared with sham group, the expression of P2X3R protein was increased in L5/L6 dorsal root ganglion (DRG), but the protein expression of P2X2RnP P2X1R did not change. After intrathecal injection of AOAA, a CBS inhibitor of hydrogen sulfide synthase, in LDH rats, it was found that AOAA could significantly reverse the pain threshold of LDH rats in a dose-and time-dependent manner. AOAA, as a control group, had no effect on the pain threshold of SD rats in sham-operated group. After intrathecal injection of P2X3R selective antagonist A-317491 to LDH rats, it was found that A-317491 could significantly reverse the pain threshold of LDH rats, and showed a dose-and time-dependent dependence. It was found that CBS and P2X3R co-expressed in L5/L6 neurons by immunohistochemistry. Moreover, by using Western Blotting method, it was found that the up-regulation of P2X3R expression in NP-induced L5/L6 neurons could be significantly reversed by continuous administration of AOAA in LDH rats, and the up-regulation of P2X3R expression in NP-induced L5/L6 neurons was significantly reversed. Whole cell patch clamp recording showed that NP on L5/L6 DRG of LDH rats could significantly enhance intracellular calcium signal induced by ATP compared with Sham group. Intrathecal administration of AOAA, a CBS inhibitor of hydrogen sulfide synthase, inhibited the changes of calcium signal induced by ATP in LDH rats. After a single or continuous injection of AOAA, a CBS inhibitor of hydrogen sulfide synthase, into the toes of LDH rats, it was found that AOAA could not reverse the pain threshold of LDH rats, and that AOAA could not reverse the pain threshold of LDH rats. There was no change in P2X3R expression in L5/L6 neurons of rats after continuous toe administration. Conclusion NPN in nucleus pulposus of lumbar disc herniation model can activate the up-regulation of CBS expression of endogenous hydrogen sulfide (H2S) synthase, and regulate the expression of P2X3 receptor protein in L5/L6 neurons through primary sensory neurons in rat DRG. Our results suggest that the up-regulation of CBS expression of endogenous hydrogen sulfide (H2S) synthase may play an important role in the process of discogenic pain in LDH rats. Therefore, to some extent, this study can reveal the molecular mechanism of root pain, thus providing a more effective treatment for neuropathic pain.
【學位授予單位】：蘇州大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R681.5

【相似文獻】

相關期刊論文 前10條

1 陳同度,張昌穎;素食大鼠的貧血現(xiàn)象[J];營養(yǎng)學報;1957年04期

2 陳偉強;趙善廣;;自制注射用大鼠固定裝置[J];上海實驗動物科學;1992年04期

3 肖柳英,林培英,馮昭明,張丹;不同周齡的SD大鼠生理、生化及體重的正常值測定[J];中藥新藥與臨床藥理;1996年03期

4 李淑云;簡易大鼠灌胃器的制作[J];錦州醫(yī)學院學報;2001年04期

5 楊明智,陳積圣;一種大鼠抓取與固定的新工具介紹[J];上海實驗動物科學;2001年03期

6 戴英,陸群;復方H_(505)對Wistar大鼠外周血的血液流變學指標的影響[J];中國血液流變學雜志;2001年01期

7 韋應波,孫喜慶,曹新生,姚永杰,馮岱雅,楊長斌;+Gz暴露時間對大鼠記憶功能和行為的影響[J];航天醫(yī)學與醫(yī)學工程;2003年01期

8 呂學軍,郭俊生,李敏,周利梅,張永娟;暈船大鼠體內(nèi)鐵含量的變化[J];中國職業(yè)醫(yī)學;2003年04期

9 湯仁仙,王迎偉,王慧,周峰;201A中藥合劑對大鼠抗腎小球基底膜腎炎病變的影響[J];徐州醫(yī)學院學報;2003年06期

10 孫同柱,付小兵,翁立新,梁雪梅,陳偉;介紹一種簡易的大鼠保定方法[J];上海實驗動物科學;2004年01期

相關會議論文 前10條

1 尹音;孫振宇;胡敏;李冬霞;;持續(xù)性高正加速度對大鼠顳頜關節(jié)損傷的作用[A];第八屆全國顳下頜關節(jié)病學及（牙合）學大會論文匯編[C];2011年

2 祝~=驤;iJ梊霞;洃克琴;崔素英;文允摪;，

本文編號：1934445