周围神经损伤晚期修复后脊髓后角P物质和降钙素基因相关肽变化的定量研究

目的：探讨周围神经损伤晚期修复后脊髓后角内P物质（SP）、降钙素基因相关肽（CGRP）变化，以反映脊髓后角的功能状态。方法：新西半兔16只腓总神经切断1年后修复，术后1，3，5月取腰骶段脊髓，切片作SP、CGRP免疫组化染色。采用计算机图像分析对后角内SP、CGRP含量进行定量分析研究。结果：失神经1年后，脊髓后角内SP、CGRP含量分别为正常的70％和86％。腓总神经晚期修复后，后角内SP、CGRP先丧失，随后逐步恢复，至术后5月，后角内SP、CGRP含量接近正常。结论：周围神经损伤晚期修复后，脊髓后角有再生表现，从神经肽角度证明晚期神经损伤有修复价值。     

晚期周围神经损伤后感觉功能恢复中：脊髓后角P物质及降钙素基因相关肽的变化

背景：晚期周围神经损伤有无修复价值?如果脊髓神经元中P物质和降钙素基因相关肽的变化发生了不可逆的变化，其修复后也预示着感觉功能的缺失。目的：定量研究周围神经损伤24周后，脊髓后角中P物质和降钙素基因相关肽的变化。设计：建立以大鼠坐骨神经损伤为研究对象的实验模型，损伤后24周为最远期观察点，自身对照(对侧空白组)，定量化研究。单位：第四军医大学骨科研究所。材料：实验于2002—10／2003—05在第四军医大学骨科研究所完成。SD大鼠55只，分成11组，即坐骨神经切断1，2，3，4，6，8，10，12，16，20．24周各组。干预：切断大鼠一侧坐骨神经并结扎其近端的方法建立周围神经损伤模型；另一侧为对照侧。应用计算机图像分析技术测试P物质和降钙素基因相关肽免疫反应区的面积。主要观察指标：各组大鼠脊髓后角中P物质和降钙素基因相关肽阳性纤维的终末分布面积的变化。结果：55只大鼠均进入结果分析。①P物质时间序列表示：周围神经损伤后2．6周，P物质在脊髓后角免疫反应区面积下降至最低，随之回升，至16周恢复正常，20，24周无明显的进一步变化。②脊髓后角降钙素基因相关肽阳性纤维和终末分布面积损伤与自身对照侧的比值：1周时1．14，6周时1．13，24周时0．29，各时间点基本相似(P&;gt;0．05)结论：周围神经损伤至晚期，脊髓后角及后根神经节细胞合成和分泌P物质及降钙素基因相关肽的功能尚未受到破坏，脊髓后角已处于一种稳定的平衡状态，仍有恢复感觉功能的神经学基础。     

低频电疗对周围神经损伤后白鼠的脊髓后角神经细胞活动电位的影响

目的:应用周围神经损伤的动物模型,探讨低频电疗对被机械性刺激和温度刺激所引发的脊髓后角神经细胞活动电位(膜电位)的影响。方法:用Sprague-Dawley雄性白鼠,制作出周围神经损伤疼痛模型作为实验组。用行动学和电生理学方法测定后腿的躲避反应和脊髓后角神经细胞的膜电位,并与对照组进行比较。结果:①实验组被机械性刺激和温度刺激所引发的躲避反应频率明显高于对照组(P<0.01)。②实验组被机械性刺激和温度刺激所引发的脊髓后角神经细胞膜电位明显高于对照组(P<0.01)。③实验组经低频电刺激10min后,其脊髓后角神经细胞的膜电位明显低于低频电刺激前脊髓后角神经细胞的膜电位(P<0.01)。④在实验组低频电刺激的同时静脉注射纳络酮10min后,其脊髓后角神经细胞的膜电位明显高于未注射纳络酮组(P<0.01)。结论:低频电缓解周围神经损伤所致疼痛的作用机理可能与其促进分泌内啡呔等中枢鸦片样物质,作用于脊髓后角神经细胞,降低其活性有关。     

晚期周围神经损伤后感觉功能恢复中:脊髓后角P物质及降钙素基因相关肽的变化

背景晚期周围神经损伤有无修复价值?如果脊髓神经元中P物质和降钙素基因相关肽的变化发生了不可逆的变化,其修复后也预示着感觉功能的缺失. 目的定量研究周围神经损伤24周后,脊髓后角中P物质和降钙素基因相关肽的变化.设计建立以大鼠坐骨神经损伤为研究对象的实验模型,损伤后24周为最远期观察点,自身对照(对侧空白组),定量化研究.单位第四军医大学骨科研究所.材料实验于2002-10/2003-05在第四军医大学骨科研究所完成.SD大鼠55只,分成11组,即坐骨神经切断1,2,3,4,6,8,10,12,16,20,24周各组.干预切断大鼠一侧坐骨神经并结扎其近端的方法建立周围神经损伤模型;另一侧为对照侧.应用计算机图像分析技术测试P物质和降钙素基因相关肽免疫反应区的面积.主要观察指标各组大鼠脊髓后角中P物质和降钙素基因相关肽阳性纤维的终末分布面积的变化.结果55只大鼠均进入结果分析.①P物质时间序列表示周围神经损伤后2～6周,P物质在脊髓后角免疫反应区面积下降至最低,随之回升,至16周恢复正常,20,24周无明显的进一步变化.②脊髓后角降钙素基因相关肽阳性纤维和终末分布面积损伤与自身对照侧的比值1周时1.14,6周时1.13,24周时0.29,各时间点基本相似(P＞0.05)结论周围神经损伤至晚期,脊髓后角及后根神经节细胞合成和分泌P物质及降钙素基因相关肽的功能尚未受到破坏,脊髓后角已处于一种稳定的平衡状态,仍有恢复感觉功能的神经学基础.     

低功率半导体激光照射对大鼠神经损伤后脊髓降钙素基因相关肽表达的影响

目的：研究１５ｍＷ半导体激光照射对大鼠坐骨神经损伤后脊髓降钙素基因相关肽（ＣＧＲＰ）的表达的影响。方法：９６只成年雄性ＳＤ大鼠制成坐骨神经损伤模型,半导体激光照射损伤的坐骨神经,能量密度为３１．８５Ｊ／ｃｍ＾２,应用免疫组化方法观察脊髓内ＣＧＲＰ的变化。结果：神经损伤后１天脊髓内ＣＧＲＰ表达即增加;第２周及第４周时激光照射组ＣＧＲＰ的表达高于对照组,阳性表达主要集中于脊髓前角运动神经元和背根感觉纤     

臂丛神经损伤后疼痛患者脊髓后角自发场电位的监测及意义

目的:应用脊髓电图技术记录臂丛神经损伤后疼痛患者受损脊髓节段后角的自发场电位,观察脊髓后根入髓区(Dorsal Root Entry Zone,DREZ)切开前、后的电位变化,对结果进行描述性研究,并分析其临床意义.方法:使用Medcare Da Vinci数字信号采集系统,在每个患者记录3段脊髓电图,分别在DREZ切开前的健侧和患侧脊髓、DREZ切开后的患侧脊髓记录1段.共采集到15例患者的可供分析的脊髓电图,对其频率和特殊放电形式进行分析.结果:DREZ切开前的患侧和健侧脊髓以及DREZ切开后患侧脊髓的后角自发场电位背景电活动频率之间无统计学差异.DREZ切开前的患侧脊髓后角的脊髓电图比健侧更多出现尖波,存在统计学差异.DREZ切开后,患侧脊髓这种电活动全部消失.术后止痛疗效优秀率为93.3％.结论:尖波是患侧脊髓相对特异的异常电活动,是相对特异的监测指标;成功的DREZ切开术后,患侧脊髓的这种特殊电活动不再出现,尖波消失可以提示术后止痛效果良好.     

低强度超声促进周围神经损伤后的再生

目的：探讨低强度超声对周围神经损伤后再生的作用。方法：将64只大鼠右侧坐骨神经重度钳夹伤制作神经损伤模型，随机分为2组各32只。超声组造模侧神经损伤处以声强250mW／cm^2、频率1．0MHz的超声进行体外治疗，隔天1次；对照组相应部位予以未启动治疗系统的假治疗。术后不同时期进行电生理、坐骨神经功能指数等指标测定及组织学检查。结果：超声组损伤神经远侧Wallerian变性进程加速、雪旺细胞增殖、变性组织吸收，轴索及髓鞘再生、感觉传导速度及坐骨神经功能的恢复等与对照组比较均提前（P〈0．01或P〈0．05）。结论：低强度超声通过影响神经再生的多个环节而促进周围神经的再生和功能恢复。     

降钙素基因相关肽在面神经切断后SD大鼠面神经核中的表达变化

目的：研究降钙素基因相关肽（calcitonin gene—related peptide，CGRP）在面神经损伤后再生过程中面神经核中的表达变化。方法：健康SD大鼠36只分别于茎乳孔处行左侧面神经切断术，而右侧于相应部位仅行面神经游离术；随机均分为术后3、7、14、21、28、35d6组。分别于相应时间点取出含面神经核团脑干部分，用免疫组化及图像分析技术，观察大鼠两侧面神经核中CGRP表达的变化。结果：CGRP分布于正常SD大鼠面神经各亚核，损伤侧的面神经核中CGRP表达比对照侧增强（P〈0．05）；图像分析CGRP灰度值两侧比较，差异有统计学意义（P〈0．05）。结论：周围性面神经损伤导致CGRP在面神经核中的表达增加，提示CGRP在面神经再生修复过程中发挥调理作用。     

周围神经损伤后不同时间的修复比较

背景:大量实验证明,Bungner带-许旺细胞-基底膜结构是神经再生理想的微环境.这一结构在神经损伤两三周后形成.而在神经损伤数小时后,近断端的神经纤维就开始发芽再生神经纤维开始再生与所需微环境的形成并不同步.目的:观察周围神经损伤后不同时间进行修复的最佳效果.设计、时间及地点:随机对照动物实验,于2007-06/2008-06在哈尔滨医科大学动物实验中心完成.材料:新西兰大白兔20只,随机数字表法分为4组:2周后神经修复组、4周后神经修复组、3个月后神经修复组、即时神经修复组.方法:建立成年新西兰大白兔周围神经损伤模型,即时修复组立即缝合伤口,2周后神经修复组、4周后神经修复组、3个月后神经修复组采用神经两断端分别固定于肌膜上,逐层缝合伤口,2周,4周,3个月后重新打开伤口,在手术显微镜下用10-0无损伤尼龙针线进行外膜缝合修复坐骨神经,缝合伤口.主要观察指标:各组缝合神经段的神经电生理、轴突数、光镜及电镜观察结果.结果:2周后神经修复组神经传导速度慢于4周后神经修复组、3个月后神经修复组(P＜0.01);即时神经修复组与2周后神经修复组差异无显著性意义(P＞0.05).2周后神经修复组效果最好,神经纤维走行正常、排列完好,神经纤维可见血管增生,髓鞘结构较好,许旺细胞功能活跃,新生轴突内微缝密集排列.4周后神经修复组最差,神经纤维数量少、排列紊乱,髓鞘轴突变性明显,大部分神经纤维脱髓鞘崩解,轴突消失,未见再生神经纤维.3个月后神经修复组效果较差,可见较多神经纤维结构破坏,捧列略紊乱,髓鞘和轴突变性较明显,仅见少量再生神经纤维,许旺细胞略少,胞质不发达.即时进行神经修复组效果较好,神经纤维结构破坏不明显,排列整齐,髓鞘和轴突变性轻,神经纤维内见有大量再生髓鞘,许旺细胞明显增多,胞质较发达.2周后神经修复组轴突计数优于其他3组(P＜0.05),4周后神经修复组最少.结论:神经损伤2周后进行神经修复效果优于其他时间点,是周围神经损伤后修复的最佳时机.     

干细胞及神经相关因子对运动性周围神经损伤再生的影响

背景:当前干细胞技术在心脑血管病、中枢神经损伤和外周神经损伤等都具有十分重要的作用.但是在对运动引起周围神经损伤方面的研究还处于基础阶段.目的:论文旨在研究当前干细胞技术运用于运动性周围神经损伤的研究现状,为运动性周围神经损伤的康复奠定科学基础.方法:通过计算机检索PubMed数据库1991-01/2010-01的相关文献,文献所述内容与干细胞和运动性周围神经损伤的研究密切相关.共选取26篇文献,结果与结论:运动训练中经常出现所谓的周围神经损伤性疾病,而依靠干细胞移植和诱导分化技术可以促进周围神经损伤的再生,且对促进运动训练中周围神经损伤的防治具有十分重要的作用.但由于目前尚处于基础研究阶段,还有待进一步的深入研究.     

带血管蒂周围神经在联合移植修复脊髓损伤中的作用

目的探讨带血管蒂周围神经促进胚胎脊髓修复成鼠脊髓损伤的能力。方法成鼠胸段脊髓损伤后,分别移植带血管蒂正中神经(VPN组)、孕14天胚胎脊髓(FSC组)、带血管蒂正中神经加胚胎脊髓(V+F组)。术后8周行组织学及电生理检查。结果V+F组移植神经与脊髓连接紧密,有较多新生轴突长入,雪旺氏细胞大量存活和增殖,神经丝蛋白、100％饱和硫蛋白阳性反应均明显高于VPN组(P＜0.01);胚胎移植物与受体融合佳,体积增长速度、神经纤维和神经元数目显著高于FSC组(P＜0.01),大部分细胞分化较好,突触较成熟;SEP检查示P1、N1波波伏期显著缩短(P＜0.01)。结论带血管蒂周围神经与胚胎脊髓联合移植,对FSC的生长发育、对损伤神经元的再生能力均有一定的促进作用。     

脊髓损伤后慢性中枢性疼痛与脊髓背角P物质关系研究

目的探讨脊髓损伤 (SCI)后慢性中枢性疼痛 (CCP)与P物质的关系。方法选取SD大鼠 2 8只 ,分为正常组 (A组 )、假手术组 (B组 ) ,以及用WADE法复制出SCI后无CCP组 (C组 )和CCP组 (D组 )。取大鼠T13 和L2 脊髓节段 ,采用免疫荧光组织化学染色法结合激光共聚焦显微镜技术观察脊髓背角P物质 (SP)的变化。结果各组大鼠T13 和L2 节段脊髓背角SP含量比较为 :D组较C组减少 (P <0 .0 5 ) ,较A组和B组明显减少 (P <0 .0 1) ;C组较A组和B组减少 (P <0 .0 5 ) ;A组与B组无显著性差异。结论SCI后CCP大鼠脊髓背角SP可能对CCP有某种程度的抑制作用。     

Peripheral nerve injury produces a sustained shift in the balance between glutamate release and uptake in the dorsal horn of the spinal cord

Peripheral nerve injury provokes heightened excitability of primary sensory afferents including nociceptors, and elicits ectopic activity in lesioned and neighboring intact nerve fibers. The major transmitter released by sensory afferents in the superficial dorsal horn of the spinal cord is glutamate. Glutamate is critically involved in nociceptive signaling and the development of neuropathic pain. We recorded miniature excitatory postsynaptic currents (mEPSCs) from neurons in lamina II of the rat dorsal horn to assess spontaneous synaptic activity after spared nerve injury (SNI), a model of chronic neuropathic pain. Following SNI, the frequency of mEPSCs doubled, indicating heightened glutamate release from primary afferents or spinal interneurons. Consistent with this finding, glutamate concentrations in the cerebrospinal fluid were elevated at 1 and 4 weeks after SNI. Transmitter uptake was insufficient to prevent the rise in extracellular glutamate as the expression of glutamate transporters remained unchanged or decreased. 2-Methyl-6-(phenylethynyl)pyridine hydrochloride, an antagonist of metabotropic glutamate receptor 5 (mGluR5), reduced the frequency of mEPSCs to its preinjury level, suggesting a positive feedback mechanism that involves facilitation of transmitter release by mGluR5 activation in the presence of high extracellular glutamate. Treatment with the β-lactam antibiotic ceftriaxone increased the expression of glutamate transporter 1 (Glt1) in the dorsal horn after SNI, raised transmitter uptake, and lowered extracellular glutamate. Improving glutamate clearance prevented the facilitation of transmitter release by mGluR5 and attenuated neuropathic pain-like behavior. Balancing glutamate release and uptake after nerve injury should be an important target in the management of chronic neuropathic pain.     

颈部脊髓损伤对患者双下肢神经传导功能的影响

目的:观察颈部脊髓损伤(SCI)后双下肢周围神经传导功能的变化,并比较完全性颈部SCI和不完全性颈部SCI之间的差异。方法:分别检测20例完全性颈部SCI患者、20例不完全性颈部SCI患者以及20例正常成年男性的胫神经、腓总神经、腓肠神经、隐神经的神经传导潜伏期、波幅和传导速度。结果:(1)运动神经:完全性损伤组和不完全性损伤组的末端运动神经潜伏期(DML)延长、运动神经传导速度(MCV)降低,与正常组比较有显著性差异(P0.05),但其异常率都低于10%;完全性损伤组和不完全性损伤组的复合肌肉动作电位(CAMP)降低,与正常组比较有显著性差异(P0.05),其异常率都高于20%,且完全性损伤组CAMP异常率高于不完全性损伤组(P0.05)。(2)感觉神经:完全性损伤组和不完全性损伤组的感觉神经动作电位(SNAP)波幅降低、感觉神经传导速度(SNCV)降低,与正常组比较有显著性差异(P0.05),但其异常率为0。结论:颈部SCI患者双下肢运动神经存在轴索变性,完全性损伤比不完全性损伤更重,感觉神经无明显异常。     

Functional magnetic resonance imaging within the rat spinal cord following peripheral nerve injury

Functional magnetic resonance imaging (fMRI) was used to detect the effects of graded peripheral nerve injury at the spinal level. Graded peripheral nerve injury in rats was accomplished by transection of nerves entering the spinal cord at the L3 and L4 levels of the spinal cord segments. Electrical stimulation of the hindpaw was used to elicit activity within the spinal cord. The stimulation experimental paradigm consisted of 62 functional images, 5 slices each, with a total of 3 rest and 2 stimulation periods. A 9.4 T MRI system and a quadrature volume rf coil covering the lumbar spinal cord were used for the fMRI study. Sets of fast spin echo images were acquired repeatedly following sham preparatory surgery under control conditions and in rats following sham surgery (pre nerve cut), followed by L3 nerve and then L4 nerve section. In rats with sham surgery, there was a significant activation within the dorsal horn of slices corresponding to L3 and L4 spinal cord segments. Following section of the L3 nerve, there was a reduction in the number of active voxels in the L3 and L4 spinal cord segments. The activation was reduced further by sectioning of the L4 nerve. Thus, following an increasing loss of axonal connections to the spinal cord, there was a decreasing number of active voxels within the spinal cord. The results demonstrate that spinal fMRI in the rat has sufficient sensitivity to detect within the spinal cord the effects of a graded reduction in peripheral connectivity.     

Rapid transneuronal destruction following peripheral nerve transection in the medullary dorsal horn is enhanced by strychnine, picrotoxin and bicuculline

The effects of systemic administration of strychnine (1 mg/kg), picrotoxin (0.5 mg/kg) and bicuculline (2 mg/kg) on acute transsynaptic destruction of medullary dorsal horn neurons following transection of the inferior alveolar nerve were assessed in rats. Single intraperitoneal injections of the above drugs were given without, l min before or l min after the nerve transection. The effect of transection without drug administration was also examined. Eighteen hours after nerve transection without drug, approximately 7 dark neurons were found in a single toluidine blue stained l μm section of the rostral medullary dorsal horn ipsilateral to the nerve transection. Administration of the drugs l min before the nerve transection significantly increased the number of dark neurons in a single section to about 17 (strychnine), 46 (picrotoxin) and 20 (bicuculline). These dark neurons were found mainly in the dorsal half of medullary dorsal horn. Delivery of any of the drugs l min after the nerve transection did not increase the number of dark neurons. The data thus indicate that the transneuronal effect of transection of the nerve was enhanced by antagonism of glycinergic and GABAergic inhibition of dorsal horn neurons. In view of the short latency and duration of transsynaptic destructive activity, a massive injury discharge of primary afferent neurons and the subsequent release of excitatory neurotransmitters appear to be the direct cause of convulsant-enhanced rapid transsynaptic destruction which follows the peripheral nerve transection.     

Activation of glia and microglial p38 MAPK in medullary dorsal horn contributes to tactile hypersensitivity following trigeminal sensory nerve injury

Glial activation is known to contribute to pain hypersensitivity following spinal sensory nerve injury. In this study, we investigated mechanisms by which glial cell activation in medullary dorsal horn (MDH) would contribute to tactile hypersensitivity following inferior alveolar nerve and mental nerve transection (IAMNT). Activation of microglia and astrocytes was monitored at 2 h, 1, 3, 7, 14, 28, and 60 days using immunohistochemical analysis with OX-42 and GFAP antibodies, respectively. Tactile hypersensitivity was significantly increased at 1 day, and this lasted for 28 days after IAMNT. Microglial activation, primarily observed in the superficial laminae of MDH, was initiated at 1 day, maximal at 3 days, and maintained until 14 days after IAMNT. Astrocytic activation was delayed compared to that of microglia, being more profound at 7 and 14 days than at 3 days after IAMNT. Both tactile hypersensitivity and glial activation appeared to gradually reduce and then return to the basal level by 60 days after IAMNT. There was no significant loss of trigeminal ganglion neurons by 28 days following IAMNT, suggesting that degenerative changes in central terminals of primary afferents might not contribute to glial activation. Minocycline, an inhibitor of microglial activation, reduced microglial activation, inhibited p38 mitogen-activated protein kinase (MAPK) activation in microglia, and significantly attenuated the development of pain hypersensitivity in this model. These results suggest that glial activation in MDH plays an important role in the development of neuropathic pain and activation of p38 MAPK in hyperactive microglia contributes to pain hypersensitivity in IAMNT model.     

大鼠脊髓背角TRPV4在背根神经节持续受压致神经病理性疼痛中的作用

目的:探讨大鼠背根神经节(dorsal root ganglion,DRG)持续受压(chronic compression of right side dorsal root ganglion,CCD)后脊髓背角瞬时感受器电位离子通道4(TRPV4)基因及蛋白变化,明确脊髓背角TRPV4在CCD致神经病理性疼痛中的作用。方法:采用健康成年雄性Wistar大鼠,共36只,随机分为3组,分别为空白对照组、CCD手术组、CCD+钌红组。制备大鼠背根神经节持续受压模型,于术前1天、术后第7天、给药前及给药2h后,测量大鼠机械刺激缩爪反应阈值,观察机械痛阈的变化;利用RT-PCR及Western Blot技术检测各组大鼠手术侧脊髓背角TRPV4基因及蛋白表达的变化。结果:与空白对照组相比,术后第7天,CCD组大鼠术侧机械痛阈值明显下降(P0.001),同侧脊髓背角TRPV4基因及蛋白表达升高(P0.05);与给药前相比,给予钌红2h后,术侧机械痛阈值明显升高(P0.001),同侧脊髓背角TRPV4基因及蛋白表达下降(P0.05)。结论:CCD后大鼠术侧机械痛阈下降,脊髓背角TRPV4基因及蛋白表达升高;钌红可部分逆转CCD后痛觉过敏,部分降低脊髓背角TRPV4基因及蛋白表达。脊髓背角TRPV4参与CCD后大鼠神经病理性疼痛形成。