内吗啡肽2和μ型阿片受体在正常猕猴脊髓背角和背根神经节中的分布

目的:观察内吗啡肽2(endomorphin-2,EM2)和μ型阿片受体(MOR)在猕猴脊髓腰段第4-6节段(L4-L6)背角及相应节段背根神经节(dorsal root ganglion,DRG)中的形态学分布特点。方法:取1只成年猕猴的L4-L6段脊髓并留取相应节段的DRG置于固定液中。脊髓冷冻水平切片(30μm)平均分为2组,分别行Nissl染色和荧光免疫组织化学染色,再于光学显微镜下观察;DRG平均分为4组,分别行荧光免疫组织化学染色、免疫组织化学染色和对照组处理,再于光学显微镜下观察。结果:(1)在猕猴脊髓背角浅层,EM2和MOR阳性终末密集分布,且EM2和MOR大量共存;(2)在猕猴DRG中,EM2和MOR集中分布在中型(20μm直径≤40μm)、小型神经元(直径≤20μm),且EM2和MOR大量共存。结论:本研究为进一步研究猕猴在内的灵长类动物中EM2和MOR的作用机制提供了形态学依据。     

神经生长因子在成年猴脊髓的表达--免疫组织化学研究

目的探讨神经生长因子(NGF)在成年猴脊髓的表达分布情况,为成年猴脊髓存在此生长因子的表达提供免疫组织化学证据。方法云南成年健康雄性猕猴5只,平均体重6kg左右,氯氨酮(0．25ml／kg)肌肉注射麻醉后,Zamboni液经心脏主动脉插管灌注固定动物,取T8脊髓节段入4％多聚甲醛后固定(4℃)6小时,在-20℃恒冷切片机下制作20μm厚的连续冰冻切片,间隔取片后分别以抗NGF抗体行免疫组化ABC染色。在Olympus光学显微镜下观察NGF在猴脊髓的表达分布情况。结果1．NGF的免疫阳性反应产物在成年猴脊髓腹角、背角神经元均有分布。2．NGF的免疫阳性反应产物在猴脊髓背角Li ssauer束有分布,表达NGF的Li ssauer束因染色而呈明显的毛刷状／发束状。3．NGF阳性神经膨体分布于猴脊髓背角Ⅰ、Ⅱ板层。4．NGF的免疫阳性反应产物在成年猴脊髓中央管(X板层)均有分布。结论成年猴脊髓存在NGF的表达,提示其可能在成年猴脊髓的生理过程中发挥作用。     

NT-4在成年猴脊髓的表达

目的探讨NT-4在成年猴脊髓的表达分布情况,为成年猴脊髓中该生长因子的表达提供可靠的免疫组织化学证据。方法云南成年健康雄性猕猴5只,平均体重6 kg左右,氯氨酮(0.25 ml/kg)肌肉注射麻醉后,Zamboni液经心脏主动脉插管灌注固定动物,取T8脊髓节段入4%多聚甲醛后固定(4℃)6小时。制作20μm厚的连续冰冻切片,间隔取片后,分别以抗NT-4抗体行免疫组化ABC染色。观察各生长因子在猴脊髓的表达分布情况。结果NT-4的免疫阳性反应产物在成年猴脊髓腹角神经元、脊髓背角神经元和脊髓中央管(Ⅹ板层)均有分布。结论成年猴脊髓存在多肽生长因子NT-4的表达,提示多肽生长因子在成年猴脊髓的生理过程中发挥作用。     

NGF及NGF mRNA在去部分背根猫脊髓和背根节表达的变化

用成年猫单侧备用根模型 (切断一侧 L1～ 5 、L7～ S2 节段脊髓背根 ,保留 L6 背根 ) ,用免疫组织化学和原位杂交技术 ,对脊髓背角 板层、背核和 L6 背根节中神经生长因子的表达进行了动态观察。结果证明 ,部分背根去传入后 :(1)脊髓各节段双侧背角 板层的神经生长因子和神经生长因子 m RNA阳性神经元的数量及体积显著增高、增大 ;(2 )手术侧背核神经生长因子和神经生长因子 m RNA阳性神经元的增高强于非手术侧 ,但术后两时间组的手术侧之间无差异 ;(3 )备用根背根节新出现了神经生长因子 m RNA阳性神经元 ,且神经生长因子阳性神经元数量明显高于正常组 ,且随术后时间增加而递增。以上结果表明 :部分背根去传入猫脊髓背角 板层及背核内神经生长因子水平提高 ,尤其备用根背根节神经元神经生长因子的增加 ,为备用背根传入纤维侧支生芽和突触重建提供了良好的条件 ,有助于解释脊髓初级传入纤维损伤溃变后代偿性机能恢复的机制     

GDNF在成年猴脊髓表达的免疫组织化学研究

目的探讨多肽生长因子GDNF在成年猴脊髓的表达分布情况。方法取云南成年健康雄性猕猴T8脊髓节段制作20μm厚的连续冰冻切片,间隔取片后以抗GDNF抗体行免疫组化ABC染色。光学显微镜下观察GDNF样免疫阳性反应物在脊髓的分布。结果阳性反应物GDNF-IR在成年猴脊髓腹角神经元、背角神经元、Clarke核神经元、背角Lissauer束、背角Ⅰ、Ⅱ板层和中央管(Ⅹ板层)均有分布。结论成年猴脊髓存在多肽生长因子GDNF的表达,提示其在成年猴脊髓的生理过程中发挥作用,而且这种作用具有复杂性和多面性。     

阿片μ受体在猫脊髓内的分布

应用免疫组织化学方法观察了阿片μ受体在猫脊髓和脊神经节内的分布。在脊髓内，ＭＯＲ样免疫反应产物主要分布于神经毯内，偶尔可见阳性胞体。致密的ＭＯＲ样免疫反应产物主要分布于背角的Ⅰ，Ⅱ层。在胸腰髓和骶髓的中间带外侧核，中央管周围灰质，骶髓节段的后连合核和Ｏｎｕｆ核内发现有中等密度的ＭＯＲ样免疫反应产物。     

前列腺酸性磷酸酶在慢性痛大鼠脊髓背角和背根神经节的表达变化

目的:观察前列腺酸性磷酸酶(prostatic acid phosphatase,PAP)在多种慢性痛大鼠脊髓背角(spinaldorsal horn,SDH)和背根神经节(dorsal root ganglion,DRG)内的表达变化。方法:应用免疫组织化学染色法以及免疫荧光多重染色技术在多种慢性痛模型大鼠观察PAP的表达变化。结果:在正常大鼠,PAP阳性反应产物主要位于DRG的中、小型的非肽能神经元,PAP阳性神经元约占DRG神经元总数的64±4.3%;在脊髓背角,PAP阳性纤维和终末主要位于Ⅱ层。在神经病理性痛模型大鼠,术侧脊髓背角Ⅱ层的PAP阳性初级传入终末较对侧减少甚至消失,DRG内PAP阳性神经元较对侧明显减少。在慢性炎性痛模型大鼠,双侧脊髓背角和DRG内PAP的表达未见明显改变。结论:PAP特异地定位于DRG神经元以及脊髓背角Ⅱ层,可能与神经病理性痛信号的传递和加工密切相关。     

IGF和EGF在成年猴脊髓的表达

目的探讨多肽生长因子IGF和EGF在成年猴脊髓的表达分布情况,为成年猴脊髓存在IGF和EGF的表达提供可靠的免疫组织化学证据。方法云南成年健康雄性猕猴5只,平均体重6kg左右,氯氨酮(0.25ml/kg)肌肉注射麻醉后,Zamboni液经心脏主动脉插管灌注固定动物,取T8脊髓节段入4%多聚甲醛后固定(4℃)6小时,制作20μm厚的连续冰冻切片,间隔取片后分别以抗IGF和EGF抗体行免疫组化ABC染色。其后在-20℃恒冷切片机下,观察各生长因子在猴脊髓的表达分布情况。结果IGF和EGF的免疫阳性反应产物在成年猴脊髓腹角神经元、背角神经元、脊髓背角Lissauer束和脊髓中央管(Ⅹ板层)均有分布,表达IGF和EGF的Lissauer束因染色而呈明显的毛刷状或发束状。结论成年猴脊髓存在多肽生长因子IGF和EGF的表达,提示IGF和EGF在成年猴脊髓的生理过程中发挥作用,而且这种作用具有复杂性和多面性。     

孤儿受体酪氨酸激酶在成年大鼠腰段脊髓和背根节中的分布

目的　观察孤儿受体酪氨酸激酶 (Ret)在成年大鼠腰段脊髓和背根节中的分布 ,探讨胶质细胞源性神经营养因子 (GDNF)对大鼠脊髓和背根节神经元的作用。　方法　采用Ret抗体免疫组织化学ABC法。　结果　Ret免疫反应存在于成年大鼠腰段脊髓神经元和背根节神经元中。　结论　Ret在大鼠腰段脊髓和背根节神经元中有一定的分布     

人胎儿和新生儿脊髓和DRG内nNOS阳性神经元的表达和分布

目的：观察24～27w人胎儿和新生儿的脊髓和背根神经节(DRG)内神经元型一氧化氮合酶(nNOS)阳性神经元的表达和分布。方法：ABC免疫细胞化学方法。结果：(1)24～27w人胎儿胸髓和腰1～3节段的中间带外侧核和前角Ⅷ层和Ⅸ层内可观察到nNOS阳性神经元。颈、胸、腰各段DRG内nNOS免疫阳性神经元占DRG细胞总数的84％～88％。(2)新生儿脊髓和DRG内nNOS免疫阳性神经元的分布情况与上述胎儿相似。但DRG内nNOS免疫阳性神经元的体积有所增加，数量明显减少，约占DRG细胞总数的70％～73％。结论：人胎儿脊髓和DRG在发育的24～27w至出生时，nNOS阳性神经元的表达在定位分布上无差异，但随胚胎发育阳性神经元的数量明显减少。     

日本猴脑内阿片μ-受体样免疫反应物质的分布:Ⅱ.颈、胸、腰、骶段脊髓及后根节(英)

通过应用识别大鼠阿片μ-受体（MOR）C末端30个氨基酸残基特异性位点的豚鼠抗体，本文对日本猴颈、胸、腰、骶段脊髓和背根节内MOR-样免疫反应物质的分布形式和特点进行了免疫组织化学研究。结果如下：强染色MOR－Li密集分布于脊髓吻尾全长的背角浅层，主要以Rexed第二层内侧部为主。在脊髓背角深层和中央导水管周围区域内也可见中到低等数量的MOR-Li标记。在脊髓背角浅层，MOR-Li广泛分布于Ⅰ、Ⅱ层内神经毯和神经元突起，但似乎只分布于第Ⅱ层神经元胞体。背角深层（Ⅳ－Ⅵ层）有些神经元的胞体和树突也有较强的MOR－Li染色。MOR－Li在背根入髓区和Lissauer's氏束中分布密集且染色强。在后根节，MOR－Li主要分布于中、小细胞，其中MOR－Li阳性小细胞约占65.71％（1018／1562），直径平均为29.50±0．11μm（17．31～34．88μm），中等细胞约占34．19％（534／1562），直径平均为39．04±0．14μm（24.01～49．86μm），而MOR－Li阳性的大细胞只占0．64％（10／1562），直径平均为59.00±2.35μm（51．09～71．39μm）。本结果揭示猴脊髓内阿片肽主要通过突触前和突触后两种方式发挥镇痛作用，除此之外，还可能通过作用于外周伤害性感受器部位的MOR发挥镇痛作用。     

Effects of electro-acupuncture on NT-4 expression in spinal dorsal root ganglion and associated segments of the spinal dorsal horn in cats subjected to adjacent dorsal root ganglionectomy

It is well known that neuroplasticity occurs in the central nervous system in response to injury. Electro-acupuncture (EA) may also promote neuroplasticity. But little is known about the underlying molecular mechanisms for the beneficial effects of EA. This study investigated the effects of EA on neurotrophin-4 (NT-4) expression in L(6) spinal dorsal root ganglion (DRG) and associated segments of the spinal dorsal horn in cats subjected to unilateral removal of L(1)-L(5) and L(7)-S(2) DRG. NT-4 protein was normally present in the cytoplasm of the L(6) DRG neurons and L(3) and L(6) spinal dorsal horn neurons and glia. Adjacent ganglionectomy leads to a significant decrease in NT-4 expression in the L(6) DRG, but no change in the spinal dorsal horn. Following EA treatment a significant increase occurred in the L(6) DRG at 14 days post-operation (dpo) as well as the L(6) cord segment at 7 and 14 dpo. These findings pointed to a possible association between NT-4 expression and EA promoted spinal cord plasticity in adult cats subjected to partial ganglionectomy.     

Fifth lumbar spinal nerve injury causes neurochemical changes in corresponding as well as adjacent spinal segments: A possible mechanism underlying neuropathic pain

Previous investigations of the anatomical basis of the neuropathic-like manifestations in the spinal nerve ligation animal model have shown that the central terminations of the unmyelinated primary afferents of L5 spinal nerve are not restricted to the corresponding L5 spinal segment, and rather extend to two spinal segments rostrally and one segment caudally where they intermingle with primary afferents of the adjacent L4 spinal nerve. The aim of the present study was to investigate the neurochemical changes in the dorsal horn of the spinal cord and DRGs after L5 nerve injury in rats. In the first experiment, the right L5 nerve was ligated and sectioned for 14 days, and isolectin B4 (IB4, a tracer for unmyelinated primary afferents) was injected into the left L5 nerve. The results showed that the vasoactive intestinal peptide (VIP) was up-regulated in laminae I–II of L3–L6 spinal segments on the right side in exactly the same areas where IB4 labelled terminals were revealed on the left side. In the second experiment, L5 was ligated and sectioned and the spinal cord and DRGs were stained immunocytochemically with antibodies raised against various peptides known to be involved in pain transmission and hyperalgesia. The results showed that L5 nerve lesion caused down-regulation of substance P, calcitonin-gene related peptide and IB4 binding and up-regulation of neuropeptide Y and neurokinin-1 receptor in the dorsal horn of L4 and L5 spinal segments. Similar neurochemical changes were observed only in the corresponding L5 DRG with minimal effects observed in L3, L4 and L6 DRGs. Although, L5 nerve injury caused an up-regulation in NPY, no change in SP and CGRP immunoreactivity was observed in ipsilateral garcile nucleus. These neuroplastic changes in the dorsal horn of the spinal cord, in the adjacent uninjured territories of the central terminations of the adjacent uninjured nerves, might explain the mechanism of hyperalgesia after peripheral nerve injury.     

Changes in PDGF expression in spared dorsal root ganglia and associated spinal dorsal horns in cats subjected to partial dorsal root ganglionectomy

Changes in the platelet derived growth factor (PDGF) in the spared dorsal root ganglia (DRG) and associated spinal dorsal horns were evaluated in cats subjected to unilateral removal of L1-L5 and L7-S2 DRG, sparing the L6 DRG. The number of PDGF immunopositive neurons and protein expression decreased significantly in the spared DRG and associated dorsal horns of the L3 and L6 cord segments at 3 days post-operation (dpo). It bottomed to the lowest level at 7 dpo in the DRG, then returned to the control level at 14 dpo; while in the L6 dorsal horn, it rapidly increased at 7 dpo and exceeded the control level at 14 dpo. This showed a significant upregulation in the spared DRG and associated spinal dorsal horns, especially in the L6 cord segment following a transient decrease. Meanwhile, a significant upregulation of PDGF mRNA was also seen in L6 DRG and L3 and L6 dorsal horns at 3 dpo. The upregulation of the endogenous PDGF in the said structures indicated a potential role of this factor in spinal cord plasticity after partial dorsal root ganglia removal in cats.     

Increased neuropeptide Y (NPY)-like immunoreactivity in rat sensory neurons following peripheral axotomy.

The effects of peripheral axotomy (sciatic nerve transection) on the presence and distribution of neuropeptide Y (NPY) in rat dorsal root ganglion (DRG) and spinal grey matter were examined using immunocytochemistry. In normal rats and on the sham-operated side of experimental rats, NPY-like immunoreactivity (NPYir) was observed in all laminae of the lumbar spinal cord, with an especially dense concentration of immunostained axons and axonal varicosities in laminae I-II of the dorsal horn. There was no detectable NPYir in L4-L5 DRG cells from normal rats or from the sham-operated side of experimental rats. At 14 days after axotomy, there was a large ipsilateral increase in the density of NPYir axons and varicosities in the lumbar spinal cord on the side of the nerve injury; this was especially apparent in laminae III-V. In the same rats, NPYir was observed in many small, medium, and large neurons in the L4-L5 DRGs on the side of the severed nerve.     

部分背根切断及针刺对备用背根节神经元血小板源性生长因子表达的影响

目的:探讨部分去背根及部分去背根后针刺对备用背根节神经元血小板源性生长因子(PDGF)表达的影响。方法:制备猫双侧备用背根节模型,并针刺位于右侧备用根支配区的两组穴位,针刺7d及14d后分别取针刺、非针刺侧备用背根节,行免疫组织化学ABC法显色。分别计数各组阳性神经元总数以及大、中小阳性神经元的数量。结果:PDGF在备用背根节的大、中小神经元均有表达。部分背根切断后,针刺侧、非针刺侧PDGF阳性神经元总数及中小阳性神经元数在7d时较正常显著减少,针刺后7d时,PDGF阳性总数及大神经元数针刺多于非针刺侧,针刺14d时,阳性总数及中小神经元数多于非针刺侧。结论:部分背根切断及针刺可影响PDGF在备用背根节的表达,这种作用显效的时间因不同时相、不同类型的背根节神经元而异。     

部分去背根猫备用背根节和脊髓Ⅱ板层NT-3及其mRNA的表达变化

采用免疫组化和原位杂交技术探讨了部分去背根猫备用背根节 (L6 )和 L3、L5脊髓 II板层 NT-3及其 m RNA的表达变化。结果发现 ,正常组 NT-3及其 m RNA阳性产物主要分布于背根节的大型神经元和少数中、小型神经元。部分去背根后 ,3 d和10 d两时相 NT-3 m RNA大型神经元阳性数明显减少 ,而 NT-3阳性大型神经元数术后 10 d时方明显减少 (P<0 .0 1) ;NT-3及其 m RNA阳性小型细胞数在术后两时相均较正常组者增多 (P<0 .0 1) ;而在中型神经元只有 NT-3阳性神经元数有增加。相对地 ,在脊髓 板层 ,两时相 NT-3阳性神经元及胶质细胞百分数均较正常者明显增加 (P<0 .0 1) ,且以 3 d组者为最明显 ,但均未见 NT-3 m RNA阳性信号。结果表明 ,部分去背根不仅导致背根节各类神经元中 NT-3的表达发生了变化 ,且对 板层 NT-3阳性神经元及胶质细胞数量也有明显影响。提示 NT-3可能在脊髓 板层可塑性中发挥作用     

Effects of electro-acupuncture on the expression of c-jun and c-fos in spared dorsal root ganglion and associated spinal laminae following removal of adjacent dorsal root ganglia in cats

This study evaluated the plastic changes of c-jun andc-fos in the right sixth lumbar dorsal root ganglion (L₆ DRG), Rexed’s lamina II in representative spinal segments L₃, L₅, and L₆ and in the nucleus dorsalis (ND) at L₃ segments after electro-acupuncture (EA) in cats subjected to removal of L₁–L₅ and L₇–S₂ DRG. Following dorsal root ganglionectomy, there was a significant increase in the density of c-jun immunoreactivity in the neurons and glia in spinal lamina II and in the ND; there was also marked elevation in the expression of c-fos in ND. In both cases there was no change in the c-jun and c-fos immunoreactivity in the DRG. After EA in the operated animals, there was an up-regulation in the expression of c-jun in the L₆ DRG and the associated spinal lamina II; however, increased c-fos expression was detected only in the L₆ DRG. Western blot and RT-PCR were also performed to quantitatively explore the mRNA and protein expression changes in the spinal dorsal horn and associated DRG. Following partial deafferentation, there was a significant increase in the protein level of both c-jun and c-fos in the dorsal horn, while, in both cases there was no change in c-jun and c-fos protein and mRNA in the DRG. After EA in the operated animals, both c-jun protein and its mRNA in the L₆ DRG as well as the associated dorsal horn of L6 spinal segment were upregulated, but increased c-fos protein and its mRNA was observed only in the L₆ DRG. These findings suggested that c-jun and c-fos might be related to the acupuncture promoted spinal cord plasticity as reported previously.     

Expression of nerve growth factor in spinal dorsal horn following crushed spinal cord injury

Expression of nerve growth factor in spinal dorsal horn following crushed spinal cord injury     

背根切断对小鼠脊髓内S100A4表达的影响

目的:采用免疫组织化学技术探讨S100A4在正常以及背根切断小鼠脊髓内的表达情况。方法:成年小鼠行单侧L4、L5背根切断术(对侧为非手术侧),术后一周取L4-L5段脊髓制作14μm厚冰冻切片,用S100A4、GFAP、NG-2、Mac1抗体进行免疫荧光组化染色。观察S100A4、GFAP、NG-2、Mac1免疫阳性反应物在脊髓内的分布。结果:正常小鼠脊髓内S100A4免疫阳性反应主要分布在Lissauer束(后外侧束)。背根切断一周后,小鼠脊髓手术侧背索内星形胶质细胞表达S100A4明显增加,同时GFAP、NG-2、Mac1免疫反应也增强。结论:小鼠脊髓中S100A4的表达与其在大鼠脊髓中的相似;该蛋白在背根切断后,在白质星形胶质细胞内的表达增强。