大鼠腰神经根受压后背根神经节和脊髓后角含生长抑素mRNA阳性神经元的变化

目的探讨大鼠腰神经根受压后背根神经节和脊髓后角含生长抑素mRNA阳性神经元的变化.方法选用12只Wistar大鼠,随机分为实验组和正常组,实验组用硅胶管压迫右L4脊神经后根,采用原位杂交法结合图像分析进行研究.结果大鼠腰神经根受压4周后,实验组背根神经节和脊髓后角内生长抑素mRNA阳性神经元神经元的数量和平均面积明显增多,与正常组相比均有显著性差异.神经元以中、小型神经元为主.结论大鼠腰神经根受压后可使背根神经节和脊髓后角内神经元生长抑素mRNA的表达上调,提示生长抑素可能与根性腰腿痛的发生有关.     

MEF2C调控大鼠脊髓背根节感觉神经元P物质和低分子量神经丝微管蛋白的表达

目的:研究肌细胞增强因子2C(MEF2C)在大鼠脊髓背根神经节神经元细胞内的表达及其与P物质和低分子量神经丝微管蛋白合成的关系。方法:分离培养背根神经节神经元细胞,然后暴露于不同浓度的神经生长因子下24h,最后用实时聚合酶链反应技术检测P物质与低分子量神经丝微管蛋白基因在背根神经节神经元细胞内的表达。通过化学转染方法将合成的3条siRNA-MEF2C分别转染PC12细胞株,并用实时聚合酶链反应技术筛选干扰效率最高的siRNA。采用化学转染方法干扰背根神经节神经元细胞内MEF2C的表达,在高浓度神经生长因子刺激背根神经节神经元细胞后采用实时聚合酶链反应技术检测干扰后背根神经节神经元细胞内P物质与低分子量神经丝微管蛋白基因的表达。结果:P物质及低分子量神经丝微管蛋白基因表达随刺激用神经生长因子浓度的升高而升高。使用化学转染方法成功地干扰背根神经节神经元细胞内MEF2C的表达,MEF2C较对照组下降52%,同时没有检测到对cAMP反应元件结合蛋白表达的影响。实验组背根神经节神经元细胞内P物质在RNA水平较对照组下降了39%,而低分子量神经丝微管蛋白在RNA水平较对照组下降了62%。结论:神经生长因子促进大鼠脊髓背根节感觉神经元内P物质与低分子量神经丝微管蛋白的合成。大鼠背根神经节神经元细胞内P物质及低分子量神经丝微管蛋白基因表达受MEF2C调控。     

中老年雄性大鼠心内神经节雄激素受体及其mRNA的表达变化

用免疫组织化学和原位杂交的方法观察成年(3月龄)大鼠和中老年(15月龄)大鼠心内神经节雄激素受体(androgenreceptor,AR)蛋白及其mRNA的表达和变化。结果显示:成年及中老年大鼠心内神经节均存在AR阳性神经元和AR mRNA阳性神经元,但中老年大鼠心内神经节AR阳性神经元和AR mRNA阳性神经元数目明显减少,表达明显降低。这些结果为AR在心脏的存在提供了形态学依据,并且证实心内神经节部分神经元能够合成AR,提示心内神经节可能受雄激素影响。     

神经再生素促大鼠背根神经节生长作用的研究

目的研究神经再生素（NRF）对体外培养新生大鼠背根神经节生长及NF—H表达的影响。方法体外大鼠背根神经节（DRG）培养;免疫荧光细胞化学、实时荧光定量PCR和Western blot等方法。结果免疫荧光细胞化学结果提示,NRF能促进背根神经节神经突起的生长,浓度为2．0mg／L时生长状况最佳;实时荧光定量PCR和Western blot结果提示,NRF能增加体外培养的背根神经节NF-H mRNA和蛋白的表达,在浓度为2．0mg／L时表达最高。结论NRF能促进体外培养背根神经节神经突起的生长和NF—H的表达,表明NRF对发育期感觉神经元具有神经营养作用。     

肿瘤抑制因子PTEN蛋白在成年大鼠腰段脊髓和背根节中的表达与分布

目的　观察肿瘤抑制因子PTEN蛋白在成年大鼠腰段脊髓和背根节中的表达和分布。　方法　免疫组织化学ABC法显示大鼠脊髓和背根神经节中PTEN阳性免疫反应产物。　结果　背根节神经元及穿越背根节神经纤维轴突有阳性免疫反应产物存在 ;PTEN阳性免疫反应产物在脊髓主要位于灰质神经元。　结论　PTEN在成年大鼠腰段脊髓和背根节神经元及神经纤维轴突中表达。     

移植坐骨神经大鼠相应背根神经节中生长相关蛋白43的表达

背景:周围神经移植后,相应的背根神经节感觉神经元胞体内生长相关蛋白43(GAP-43)表达的时程与规律尚未明确。目的:观察大鼠坐骨神经移植术后GAP-43在相应背根神经节中的表达变化。方法:取健康成年雄性Wistar大鼠,随机分为2组,对照组行假手术组;实验组行坐骨神经移植,分别于术后3d,1,2,4,6,8周6个时间点处死后取其手术侧L4~L5背根神经节,应用RT-PCR和Western Blot技术检测GAP-43mRNA及其蛋白表达。结果与结论:对照组大鼠背根神经节中GAP-43mRNA表达量较低,且随时间无明显改变;实验组术后1周时GAP-43mRNA在相应背根神经节中即有表达增强,2周时达到高峰,6~8周表达逐渐减弱。两组大鼠背根神经节中GAP-43蛋白与GAP-43mRNA表达的变化规律相同。结果表明神经损伤后相应神经元的再生能力存在损伤反应性变化。     

神经营养因子-3对大鼠脊髓半横断后背根神经节c-Jun表达的影响

目的：研究神经营养因子-3(NT-3)对脊髓半横断后背根神经节c-Jun表达的影响，探索NT-3促进脊髓修复的作用机制。方法：将实验动物分为：对照组，损伤组和NT-3注射组，应用荧光免疫组化法结合激光扫描共聚焦显微镜，观察各组背根神经节c-Jun的表达，并计数细胞核完整的神经元数目。结果：脊髓损伤后，背根神经节的细胞内c-Jun的表达上调；NT-3注射组脊髓损伤侧背根神经节神经元的c-Jun表达明显上调，背根神经节内细胞核呈完整状态的神经元数量明显增多。结论：(1)c-Jun在轴突损伤后表达上调。(2)NT-3对轴突损伤后的神经元有保护作用。(3)NT-3可能通过使c-Jun表达上调而发挥其促神经再生作用。     

香草酸受体亚型Ⅰ与降钙素基因相关肽和植物凝集素结合位点在大鼠脊神经节内感觉神经元的共存

目的:研究香草酸受体亚型Ⅰ(VR1)在大鼠脊神经节(DRG)内感觉神经元的表达.及与降钙素基因相关肽(CGRP)、植物凝集素(IB4)结合位点的共存,为探讨VR1在伤害性感觉刺激信号传导中的作用提供形态学依据.方法:应用免疫荧光组织化学三标方法结合激光共聚焦扫描显微镜技术观察VR1与CGRP和IB4结合位点在DRG的分布及相互关系.结果:背根节内可见大量中、小型神经元胞体和神经纤维表达VR1,一群独特的VR1阳性特小神经元胞体(直径8～11 μm)呈荧光强阳性.荧光双标显示背根节内许多VR1阳性神经无与CGRP共存或结合IB4,而CGRP/IB4双标神经元数量稀少.有41.1%±3.2%VR1阳性细胞呈CGRP阳性,有54.9%±3.8%VR1阳性神经元结合IB4.VR1强荧光特小神经元胞体未见CGRP阳性标记或结合IB4.VR1/CGRP/IB4三标神经元数量较少,只有1.5%±1.1% VR1阳性神经元同时呈CGRP阳性并结合IB4.结论:背根节内可能存在VR1/CGRP与VR1/IB4两种不同的与伤害性刺激相关的VR1阳性神经元亚群.     

坐骨神经慢性卡压损伤模型大鼠背根神经节纤维化改变

文题释义：组织纤维化：是结缔组织过度增生和细胞外基质沉积的结果,纤维化过程是一个异常的、不受控制的组织修复过程,其中组织损伤和自身免疫疾病是导致组织纤维化的主要因素,过度的组织纤维化最终导致组织结构和功能改变。 慢性神经卡压损伤：指周围神经在特定部位受到慢性卡压引起的相应神经功能障碍。 背景：既往的研究主要集中于周围神经慢性卡压损伤所导致的靶器官——骨骼肌萎缩及其纤维化发生的机制研究,关于周围神经慢性卡压损伤信号向上引起背根神经节功能改变的研究较少。 目的：观察大鼠坐骨神经慢性卡压损伤对背根神经节纤维化的影响。方法：按照Mackinnon设计的方法制备大鼠坐骨神经慢性卡压模型,术后3周,分别取大鼠坐骨神经卡压侧和对侧L_4-6背根神经节,采用RT-PCR、Western blot及免疫荧光检测大鼠卡压侧和对照侧背根神经节内转化生长因子β1、结缔组织生长因子及胶原蛋白Ⅰ的表达变化。结果与结论：①损伤3周后,相比于对照侧,背根神经节内转化生长因子β1、结缔组织生长因子及胶原蛋白Ⅰ的mRNA及蛋白表达均具有相同的升高趋势(P < 0.05);②损伤3周后,大鼠卡压侧和对照侧背根神经节内转化生长因子β1和结缔组织生长因子主要表达在背根神经元内和轴突内,而胶原蛋白Ⅰ主要表达在背根神经元和轴突的周围,形成包绕背根神经元和轴突的网状结构;③上述数据证实,大鼠坐骨神经慢性卡压损伤可以导致背根神经节纤维化的改变,并且背根神经节纤维化的改变可能与背根神经元内转化生长因子β1及结缔组织生长因子表达升高有关。ORCID: 0000-0003-1632-0837(黎琴文) 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

成年猕猴背根神经节nNOS免疫阳性神经元的分布

目的:观察正常成年猕猴背根神经节神经元型一氧化氮合酶(nNOS)免疫阳性神经元的分布。方法:ABC免疫细胞化学方法显示nNOS免疫阳性神经元,并用体视方法进行定量分析。结果:猕猴颈、胸、腰各段背根神经节nNOS免疫阳性神经元的分布相似,数量较多,阳性神经元的大小不等,多呈圆形或椭圆形;胞浆着色较深,胞核位于细胞中央,不着色,细胞被神经纤维束分隔成群。nNOS免疫阳性神经元以中型神经元为主,其次为小型神经元,其胞浆呈强阳性染色,细胞直径<50μm,大型神经元较少。颈、胸、腰各段背根神经节nNOS免疫阳性神经元的密度以及阳性细胞与总细胞数的比值均无明显差异。结论:猕猴背根神经节nNOS主要表达在中、小型神经元,提示NO可能主要参与痛觉等浅感觉的传导和调制。     

足底伤害性刺激条件下大鼠腰骶髓背根节内SP mRNA和α-CGRP mRNA的表达

将Formalin注入大鼠一侧后肢足底，分别于术后不同时间（0．5、1、2、4、8、12、24、48h和5d）将动物断头取材，用放射性同位素[α－(35)S]dATP标记的寡核苷酸探针进行原位杂交组织化学实验，检测大鼠腰骶髓（L5、L6和S1）背根节内SPmRNA和α－CGRPmRNA表达阳性细胞占整个背根节细胞的百分率。结果显示：注射后2h背根节内SPmRNA和α－CGRPmRNA表达阳性细胞百分率即明显增高；分别于12h（α－CGRPmRNA阳性细胞表达率增加了3．3％，P＜0．01）和24h（SPmRNA阳性细胞表达率增加了3．5％，P＜0．01）达到高峰，其中SPmRNA可持续到48h后，而α－CGRPmRNA则在12h后逐渐回落；5d时二者的表达阳性率均降至术前水平．提示SP和α－CGRP的mRNA的表达水平与伤害性刺激的施加有关，从而进一步证明了SP和CGRP在伤害性刺激信息的传递过程中具有重要作用．     

Sprouting of sensory neurons in dorsal root ganglia after transection of peripheral nerves

Morphological reaction of sensory neurons of dorsal root ganglia after peripheral nerve transection was investigated by a nerve tracing method using E. coli lacZ (beta-galactosidase) gene recombinant adenovirus. The sciatic nerve of the rat was transected and inoculated with the gene recombinant adenovirus from the cutting end of nerve fibers. The fixation was accomplished from one to six weeks after inoculation. A whole mount specimen was observed after the reaction in a X-galactocidase substrate. Newly formed sprouting processes of dorsal root ganglion (DRG) cells appeared, all of them sprouting from the primary segment of DRG cells. Developed branches were morphologically categorized in to two types: one was the "linear type" which showed diverged branches running straightly along the major axis of the DRG; the other was the "winding type" which exhibited a random running pattern to the original axons and wound and extended in all directions in dorsal root ganglia with many branches. Many of this type encircled other cell bodies and formed a ring-like structure. There was no difference in the size of cell bodies in either type or between the ring-like structure forming the cells and those cells encircled by them.     

DRASIC contributes to pH-gated currents in large dorsal root ganglion sensory neurons by forming heteromultimeric channels

For many years it has been observed that extracellular acid activates transient cation currents in large-diameter mechanosensory dorsal root ganglion (DRG) neurons. However, the molecular basis of these currents has not been known. Large DRG neurons express the dorsal root acid sensing ion channel (DRASIC), suggesting that DRASIC might contribute to H+-gated DRG currents. To test this, we examined whole cell currents in large DRG neurons from mice in which the DRASIC gene had been disrupted. We found that DRASIC null neurons retained H+-gated currents, indicating that DRASIC alone was not required for the currents. However, without DRASIC, the properties of the currents changed substantially as compared with wild-type neurons. In DRASIC -/- neurons, the rate of current desensitization in the continued presence of an acid stimulus slowed dramatically. H+-gated currents in DRASIC null neurons showed a decreased sensitivity to pH and an enhanced sensitivity to amiloride. The loss of DRASIC also altered but did not abolish the current potentiation generated by FMRF-related peptides. These data indicate that the DRASIC subunit makes an important contribution to H+-gated currents in large DRG sensory neurons. The results also suggest that related acid-activated DEG/ENaC channel subunits contribute with DRASIC to form heteromultimeric acid-activated channels.     

Immunohistochemical localization of adenosine deaminase in primary afferent neurons of the rat

Adenosine deaminase (ADA) was detected immunohistochemically in neuronal cell bodies of dorsal root ganglia (DRG) of the rat. ADA-immunoreactivity was confined exclusively to small type B ganglion neurons in cervical, thoracic and lumbar sensory ganglia; large type A neurons in sensory ganglia were devoid of immunostaining for ADA. It was consistently found that only a small proportion of type B neurons in DRG contain immunohistochemically detectable ADA. It is suggested that the expression of high ADA levels is a distinguishing feature of a subpopulation of type B DRG neurons and, further, that ADA in these neurons may reflect their utilization of purines (adenosine or adenine nucleotides) as transmitters or cotransmitters.     

Nerve injury increases an excitatory action of neuropeptide Y and Y2-agonists on dorsal root ganglion neurons

Damage to sensory nerves invokes the expression of neuropeptide Y in the cell bodies of sensory neurons in dorsal root ganglia. We therefore compared the action of this peptide on control dorsal root ganglia neurons with its action on neurons from animals in which the sciatic nerve had been cut. Neuropeptide Y (0.1-1.0 microM) increased the excitability of 24% of control neurons and its effect was stronger and more cells (56%) were affected after axotomy. Increased excitability was mediated via a Y2-receptor and resulted from attenuation of Ca2+-sensitive K+-conductance(s) secondary to suppression of N-type Ca2+ channel current. Y1-agonists potentiated L-type Ca2+ channel current in control neurons without altering excitability. This Y1-effect was attenuated whereas effects mediated via Y2-receptors were enhanced after axotomy. No evidence was found for involvement of Y4- or Y5-receptor subtypes in the actions of neuropeptide Y either on control or on axotomized dorsal root ganglion neurons. It is concluded that neuropeptide Y increases the excitability of sensory neurons by interacting with a Y2-receptor and thereby decreasing N-type Ca2+ channel current and Ca2+-sensitive K+-conductance(s). When peripheral nerves are damaged, dorsal root ganglion neurons start to express neuropeptide Y and its excitatory Y2-excitatory effects are enhanced. The peptide may therefore contribute to the generation of aberrant sensory activity and perhaps to the etiology of injury-induced neuropathic pain.     

Spontaneous discharge originates in the dorsal root ganglion at the onset of a painful peripheral neuropathy in the rat.

The activity of myelinated primary afferents was recorded from the dorsal roots 1-3 days after creation of a painful peripheral neuropathy in rats. The effects on spontaneous discharge of acute transections at various points along the injured sciatic nerve and the dorsal root were determined, as were the effects of K+ channel blockers applied topically to two putative sites of impulse origin: the injured region of the nerve and the dorsal root ganglion (DRG). Transections just proximal to the nerve injury and just distal to the DRG failed to halt the discharge, but spontaneous discharge disappeared when the transection was made just proximal to the DRG (i.e. between the DRG and recording electrode). K+ channel blockers (4-aminopyridine and gallamine triethiodide) applied to the DRG increased the frequency of spontaneous discharge or initiated activity from silent fibers. Applications of K+ channel blockers to the injured region of the nerve were without effect. Thus, the spontaneous discharge and the sensitivity to K+ channel blockade seen in A beta and A delta primary afferents at the time of the onset of the neuropathic pain syndrome appear to originate in the DRG.     

大鼠背根神经节分离神经元的延时整流的钾离子单通道

运用膜片钳技术对急性分离的大鼠背根神经节神经元细胞上的电压依赖性钾离子通道进行了研究.在细胞贴附式记录模式下,去极化可以激活一个电导为20pS的钾离子单通道电流,分析了其单通道的特性,提出了其动力学的模型.对于了解大鼠背根神经节神经元细胞的电活动机制具有重要的意义.     

Functional Na+/K+ pump in rat dorsal root ganglia neurons.

Steady-state Na+/K+ pump current (Ip) in isolated adult rat dorsal root ganglia neurons was studied to determine if the plasma membrane Na+/K+ pump activity is uniform across the population of dorsal root ganglia neurons. Cells were voltage-clamped at -40 mV and holding current (Ih) was recorded using whole-cell patch-clamp techniques under conditions that stimulate the Na+/K+ pump (60 mM intracellular Na+ and 5.4 mM extracellular K+). Ip was defined as the 1 mM ouabain-sensitive fraction of Ih. Data suggest the existence of three subpopulations of dorsal root ganglia neurons having mean steady-state Ip densities of 1.6+/-0.1, 3.8+/-0.2 and 7.5+/-0.4 pA/pF. Neurons with small Ip had an average soma perimeter of 95+/-3 microm, while neurons with medium and large Ip density had significantly larger soma sizes (131+/-8 and 141+/-7 microm, respectively). Neurons with a large Ip density had a significantly lower specific membrane resistance (Rm; mean 4.0+/-0.3 kohm x cm2) than neurons with medium or small Ip density (19+/-6 and 31+/-6 kohm x cm2, respectively). Regardless of these differences, in all groups of neurons Ip had a low sensitivity to ouabain (Ip half inhibition by ouabain was observed at 80-110 microM). These data suggest that the Na+/K+ pump site density and/or its activity is not uniform throughout the dorsal root ganglia neuron population; however, this non-uniformity does not appear to relate to the functional expression of the different alpha isoforms of the Na+/K+ pump. The major functional Na+/K+ pump in the dorsal root ganglia neuron plasma membrane appeared to be the low ouabain affinity (alpha1) isoform.     

Modulation of action potential firing by iberiotoxin and NS1619 in rat dorsal root ganglion neurons

The present study investigated the effects of iberiotoxin (IbTx), a peptide toxin blocker of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels and NS1619, a BK(Ca) channel opener, on action potential firing of small and medium size afferent neurons from L6 and S1 dorsal root ganglia of adult rats. Application of IbTx (100 nM) reduced whole-cell outward currents in 67% of small and medium size neurons. Analysis of action potential profile revealed that IbTx significantly prolonged the duration of action potential and increased firing frequency of afferent neurons. IbTx did not significantly alter the resting membrane potential, threshold for action potential activation and action potential amplitude. The benzimidazolone NS1619 (10 microM) increased opening activity of a Ca(2+)-dependent channel as assessed by single channel measurements. In contrast to IbTx, NS1619 reversibly suppressed action potential firing, attributable to increases in threshold for evoking action potential, reduction in action potential amplitude and increases in amplitude of afterhyperpolarization. The effect of NS1619 on neuronal firing was sensitive to IbTx, indicating the attenuation of neuronal firing by NS1619 was mediated by opening BK(Ca) channels. NS1619 also reduced neuronal hyperexcitability evoked by 4-aminopyridine (4-AP), a transient-inactivated K(+) channel (A-current) blocker, in an IbTx-sensitive manner.These results indicate that IbTx-sensitive BK(Ca) channels exist in both small and medium diameter dorsal root ganglion (DRG) neurons and play important roles in the repolarization of action potential and firing frequency. NS1619 modulates action potential firing and suppresses 4-AP-evoked hyperexcitability in DRG neurons, in part, by opening BK(Ca) channels. These results suggest that opening BK(Ca) channels might be sufficient to suppress hyperexcitability of afferent neurons as those evoked by stimulants or by disease states.     

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.