本文選題：P物質(zhì)　切入點：神經(jīng)激肽-1受體　出處：《第四軍醫(yī)大學(xué)》2007年碩士論文　論文類型：學(xué)位論文

【摘要】： 延髓吻段腹內(nèi)側(cè)部(RVM)是中樞內(nèi)源性下行痛抑制系統(tǒng)的一個重要組成部分,RVM內(nèi)的神經(jīng)元不但接受多個腦區(qū)的上位調(diào)節(jié),還向脊髓背角淺層發(fā)出下行投射,參與外周傷害性信息向中樞傳遞的調(diào)控。 動物行為學(xué)實驗研究表明,電刺激中縫大核(RMg)可以觀察到動物對熱刺激和機械性刺激的反應(yīng)降低,反應(yīng)閾值增大,對傷害性信息的傳遞有抑制作用。破壞RMg對傷害性信息傳遞的抑制作用降低或消失。RVM內(nèi)神經(jīng)元的活動受到多種機制的調(diào)節(jié),其中最經(jīng)典的例子就是RVM內(nèi)局部給予嗎啡后觀察到RVM內(nèi)神經(jīng)元的活動水平升高,并產(chǎn)生鎮(zhèn)痛效應(yīng)。已經(jīng)證明,多種神經(jīng)活性物質(zhì)和受體均可以通過調(diào)節(jié)RVM內(nèi)向脊髓背角投射的神經(jīng)元參與中樞內(nèi)源性鎮(zhèn)痛,而對這些物質(zhì)及受體的研究和探討,無疑對進一步揭示中樞內(nèi)源性下行抑制系統(tǒng)發(fā)揮鎮(zhèn)痛效應(yīng)的機制具有重要的意義。 P物質(zhì)(SP)是一種重要的,與傷害性信息調(diào)節(jié)相關(guān)的神經(jīng)激肽,主要通過神經(jīng)激肽-1受體(NK-1R)在中樞發(fā)揮調(diào)節(jié)作用。盡管SP在外周對傷害性信息向中樞傳遞的過程中起易化作用,但在中樞神經(jīng)系統(tǒng)內(nèi)卻主要發(fā)揮鎮(zhèn)痛作用。免疫組織化學(xué)研究表明有大量的SP免疫陽性纖維和終末分布于RVM內(nèi),與該處的神經(jīng)元形成復(fù)雜的突觸聯(lián)系;局部給予SP可以使RVM內(nèi)的神經(jīng)元發(fā)放動作電位的頻率升高,這些研究結(jié)果提示SP可能參與了RVM內(nèi)神經(jīng)元活動的調(diào)節(jié)。但是,RVM內(nèi)SP能纖維和終末的來源、SP能纖維和終末與NK-1R陽性神經(jīng)元的聯(lián)系及RVM內(nèi)可能受到SP調(diào)節(jié)的NK-1R陽性神經(jīng)元向脊髓背角的下行投射狀況等問題均尚未闡明。 本研究采用免疫熒光組織化學(xué)染色、熒光束路追蹤以及免疫電鏡技術(shù),觀察了小鼠RVM內(nèi):①SP能纖維和終末的來源,②SP能終末與NK-1R陽性神經(jīng)元之間的突觸聯(lián)系,③NK-1R陽性神經(jīng)元的下行投射狀況。本研究的結(jié)果將為SP參與中樞內(nèi)源性下行鎮(zhèn)痛系統(tǒng)的機能活動提供形態(tài)學(xué)證據(jù)。 第一部分延髓吻段腹內(nèi)側(cè)部內(nèi)P物質(zhì)陽性纖維和終末的來源 本研究采用順行、逆行束路追蹤與免疫熒光組織化學(xué)染色相結(jié)合的方法,觀察了分布于RVM內(nèi)的SP能纖維和終末的來源部位。 將逆行性束路追蹤劑熒光金(FG)注射到中縫大核(RMg)后,結(jié)合SP的免疫熒光組織化學(xué)染色,我們觀察到FG逆行標(biāo)記的SP陽性神經(jīng)元主要分布于楔形核(CnF),中縫背核(DR)與中腦導(dǎo)水管周圍灰質(zhì)外側(cè)部(lPAG)。將順行示蹤劑生物素化葡聚糖胺(BDA)注射到相應(yīng)的CnF、DR和lPAG后,SP、NK-1R和BDA的免疫熒光組織化學(xué)三重染色結(jié)果顯示,在RMg和旁巨細胞核(PGi)內(nèi),BDA標(biāo)記的SP陽性終末與表達NK-1R的神經(jīng)元形成密切接觸。 結(jié)果提示:RVM內(nèi)的SP能纖維和終末主要來源于CnF、DR和lPAG,SP能終末能夠與相應(yīng)的表達NK-1R的神經(jīng)元細胞發(fā)生密切接觸,鑒于RVM內(nèi)神經(jīng)元是內(nèi)源性下行抑制系統(tǒng)的主要組成,該結(jié)果從形態(tài)學(xué)上提示了SP參與中樞內(nèi)源性下行痛抑制系統(tǒng)的可能性。 第二部分中腦導(dǎo)水管周圍灰質(zhì)下行投射的SP能終末與中縫大核內(nèi)神經(jīng)激肽-1受體陽性神經(jīng)元的突觸聯(lián)系 本研究采用順行追蹤與免疫組化雙重染色相結(jié)合的三標(biāo)方法,觀察了起源于PAG的SP能陽性終末與RMg內(nèi)NK-1R陽性神經(jīng)元之間的突觸聯(lián)系。結(jié)果顯示:將BDA注入小鼠PAG的外側(cè)區(qū)后,BDA順行標(biāo)記終末可見于腦干許多區(qū)域,主要位于RMg內(nèi),其中的部分BDA順行標(biāo)記終末呈SP陽性。RMg內(nèi)可見到散在分布的NK-1R陽性神經(jīng)元。在電鏡下可見來自PAG的SP/BDA雙標(biāo)終末與RMg內(nèi)的NK-1R陽性神經(jīng)元的胞體和樹突形成以非對稱性為主的突觸聯(lián)系。 本研究的結(jié)果提示,起源于PAG的下行投射終末所釋放的SP可能參與中樞內(nèi)源性鎮(zhèn)痛系統(tǒng),通過激活RMg內(nèi)表達NK-1R的下行投射神經(jīng)元發(fā)揮鎮(zhèn)痛作用,在電鏡水平進一步證實了第一部分的實驗結(jié)果。 第三部分延髓吻段腹內(nèi)側(cè)部內(nèi)神經(jīng)激肽-1受體陽性神經(jīng)元向脊髓背角的投射 本研究采用FG逆行束路追蹤和NK-1R免疫熒光組織化學(xué)相結(jié)合的方法,觀察了RVM內(nèi)NK-1R陽性神經(jīng)元向脊髓背角的下行投射狀況。 將FG注射入脊髓背角后,FG逆行標(biāo)記的神經(jīng)元主要分布于RVM內(nèi)。其中,部分神經(jīng)元同時呈NK-1R免疫陽性。 以上結(jié)果提示:在RVM內(nèi),部分表達NK-1R的神經(jīng)元向脊髓背角發(fā)出下下行投射,參與脊髓背角淺層神經(jīng)元活動的調(diào)節(jié)。該下行投射可能與傷害性信息的傳入調(diào)控有關(guān)。
[Abstract]:The rostral ventromedial part of the medulla oblongata (RVM) is an important part of the central endogenous descending pain suppression system. The neurons in RVM not only receive superordinate regulation in multiple brain regions, but also send down projections to the superficial part of the spinal dorsal horn, and participate in the regulation and control of peripheral nociceptive information to the central nervous system.
Experimental results show that the animal behavior, electrical stimulation of nucleus raphe magnus (RMg) can be observed in animal to reduce the heat and mechanical stimulus response, response threshold increases, has inhibitory effect on the transmission of nociceptive information. RMg damage is regulated by multiple mechanisms of inhibition of nociceptive transmission decreased or disappeared in.RVM the activity of neurons, one of the most classic example is the RVM in the local administration of morphine was observed after RVM in neuronal activity levels, and produce analgesic effect. It has been proven, neuroactive substances and receptors are mediated by the RVM spinal projecting neurons in the central endogenous pain, and study of the materials and their receptors the discussion is important to further reveal the central endogenous descending inhibitory system play analgesic effect mechanism.
Substance P (SP) is a kind of important, and nociceptive information related to modulation of neurokinin, mainly through neurokinin -1 receptor (NK-1R) play a regulatory role in the CNS. Although SP in the peripheral of the afferent nociceptive information in facilitation, but in the central nervous system is mainly play analgesic. Immunohistochemical studies show that a large number of SP immunoreactive fibers and terminals are distributed in the RVM, make complicate synaptic connections with the neurons; local administration of SP can make RVM neurons within the spike frequency increases, these results suggest that SP may be involved in the modulation of neurons in the RVM but. RVM, SP fibers and terminals, SP NK-1R may be regulated by SP positive neurons to the spinal dorsal horn of the descending projection of contact and ask RVM fibers and terminals and NK-1R positive neurons The questions have not been clarified.
This study used immunofluorescence staining, fluorescence tracing and immunoelectron microscopy, observed in RVM mice: SP fibers and terminals of the source, the SP synaptic connections between terminals and NK-1R positive neurons and NK-1R positive neurons in the descending projection condition. The results of this study will provide morphological evidence for that SP participates in the central endogenous descending pain control system.
The source of substance P positive fibers and terminals in the ventromedial ventromedial part of the medulla oblongata
In this study, anterograde retrograde tracer tracing and immunofluorescence histochemical staining were used to observe the location of SP energy fibers and terminals in RVM.
The retrograde tract tracing of fluoro gold (FG) injected into the nucleus raphe magnus (RMg), combined with immunohistochemical SP staining, we observed that FG labeled SP positive neurons were mainly distributed in the cuneate nucleus (CnF), dorsal raphe nucleus (DR) and lateral (periaqueductal gray lPAG). The anterograde tracer biotinylated dextran amine (BDA) was injected into the corresponding CnF, DR and lPAG, SP, NK-1R and BDA immunofluorescence histochemical staining showed that in three, RMg and paragigantocellular nucleus (PGi), BDA labeled SP positive terminals and NK-1R expressing neurons the formation of close contact.
The results suggest that RVM in SP fibers and terminals mainly derived from CnF, DR and lPAG, SP neurons and the corresponding expression of NK-1R has close contact to the end, in view of the neurons in RVM is a major component of endogenous descending inhibitory system, the results suggest the possibility that SP participates in the central endogenous descending inhibitory system from the morphology.
The second part of the periaqueductal gray of the descending projection terminals and SP synapses in the nucleus raphe magnus neurokinin -1 receptor positive neurons
Three standard methods used in this study anterograde tracing and immunohistochemical double staining combined with observation, originated from the PAG SP to synaptic connections between NK-1R positive terminals and RMg positive neurons. The results showed that BDA was injected into the lateral region of mouse PAG, BDA anterogradely labeled terminals can be found in many regions of the brain stem. Mainly located in the RMg, BDA anterogradely labeled terminal which were positive for SP.RMg can be seen scattered in the distribution of NK-1R positive neurons in the electron microscope. From PAG SP/BDA double labeled terminals and RMg in NK-1R positive cell bodies and dendrites of neurons form synapses with non symmetry based.
The results of this study suggest that the SP released from the descending projection terminals originating from PAG may participate in the central endogenous analgesia system. It plays an analgesic role by activating the down projection neurons expressing NK-1R in RMg, and further confirms the experimental results in the first part at the electron microscope level.
The third part of neurokinin -1 receptor positive neurons in the medulla oblongata kiss section within the Department of the ventral part of the medial projection to the dorsal horn of spinal cord
In this study, FG retrograde tracing and NK-1R immunofluorescence histochemistry were used to observe the descending projection of NK-1R positive neurons in RVM to the dorsal horn of spinal cord.
After injecting FG into the dorsal horn of the spinal cord, the FG retrograde labeled neurons were mainly distributed in RVM, and some of the neurons were NK-1R immunoreactive at the same time.
The above results suggest that in RVM, some neurons expressing NK-1R are descending projections to the dorsal horn of the spinal cord, which are involved in the regulation of neurons in the superficial dorsal horn of spinal cord. This downward projection may be related to the afferent regulation of noxious information.

【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2007
【分類號】：R322

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