改良CCI 大鼠痛行为学检测与损伤区自发放电活动再观察

目的:应用改良CCI模型研究外周神经损伤后痛觉过敏和自发放电各自特征及相互关系.方法:雄性SD大鼠,随机分为CCI组和Sham组,分别于术前1天和术后1、4、7、9、11、14天测定机械刺激缩足反射阈值和热缩腿反射潜伏期,同时选取术侧机械刺激缩腿反射阈值低于4g或者术侧和健侧热缩腿反射潜伏期差异大于2s的CCI模型大鼠观察术后4-14天损伤区自发放电活动.结果:神经纤维损伤后,机械痛敏和热痛敏随时间表现为逐渐增强,同时在损伤区观察到三类放电模式:整数倍放电阵发放电和周期放电.结论:术后大鼠机械痛阈和热痛阈逐渐降低,机械痛敏的产生和损伤区自发放电活动关系密切,不同的放电模式可能代表不同的传入信息.     

川芎嗪对慢性压迫性损伤大鼠神经病理痛行为学的影响

目的探讨川芎嗪对慢性压迫性损伤（CCI）大鼠行为学的影响。方法建立大鼠坐骨神经CCI神经病理痛模型,取40只雄性大鼠随机分成4组,Ⅰ组为空白对照组,Ⅱ组为假手术组,Ⅲ组为CCI＋川芎嗪治疗组,Ⅲ组在术后第1天开始腹腔注射100 mg/kg川芎嗪注射液,Ⅳ组为CCI手术组。分别于术前（0 d）及术后1、3、5、7、91、1、14 d以von Frey细丝法和热辐射法测定机械缩足反射阈值（mechanical withdrawal threshold,MWT）和热缩足反射潜伏期（thermal withdrawal latency,TWL）,观察CCI大鼠神经病理痛的行为学变化。结果术后14 d,Ⅳ组和I、Ⅱ、Ⅲ组相比较,大鼠后爪的机械和热痛敏阈值明显降低（P〈0.01）;I、Ⅱ、Ⅲ组之间相比,大鼠后爪的机械和热痛敏阈值差异没有显著性（P〉0.05）。结论川芎嗪可以缓解CCI大鼠的慢性神经病理痛行为学表现。     

人参皂苷Rg2对慢性坐骨神经损伤大鼠痛觉敏化和抑郁状态的影响*

目的:观察人参皂苷Rg₂对慢性坐骨神经损伤大鼠痛觉敏化、抑郁状态的影响。方法: 将50只 SD 大鼠随机分为 5组(n=10): 空白对照组(Normal+生理盐水腹腔注射)、假手术组(手术但不结扎+生理盐水腹腔注射) 、坐骨神经慢性压迫损伤(CCI)组(CCI +生理盐水腹腔注射) 、人参皂苷Rg₂低剂量组(CCI+ Rg₂ 5 mg/kg腹腔注射)、人参皂苷Rg₂高剂量组(CCI+ Rg₂ 10 mg/kg 腹腔注射)。CCI模型建立后,药物通过注射器进行腹腔内注射 5 ml/kg,每天1次,连续14 d。分别在术前1 d和术后 1、3、5、7、10、14 d测定大鼠的机械性缩足反射阈值(MWT)和热缩足潜伏期(TWL);术前1 d和术后第14日时检测明暗箱实验和强迫游泳试验。 结果:与假手术组比较,CCI组术后14 d机械痛阈值和热痛潜伏期明显缩短(P＜0.01),明箱内停留时间明显缩短(P＜0.01),穿梭次数明显减少(P＜0.01),游泳潜伏期明显延长(P＜0.01)。与CCI组比较,人参皂苷Rg₂组术后14 d机械痛阈和热痛潜伏期明显增加(P＜0.01),大鼠在明箱内时间明显延长(P＜0.01),穿梭次数明显增多(P＜0.01),且游泳潜伏期明显缩短(P＜0.01)。结论:人参皂苷Rg₂能抑制 CCI 大鼠的机械痛敏和热痛敏,同时改善其抑郁状态。     

实验性高血糖对大鼠病理性痛反应的影响

目的:研究实验性高血糖大鼠血糖变化对病理性痛反应的影响.方法:用链脲佐菌素制造高血糖大鼠模型,并以生理盐水处理大鼠做为对照.在大鼠后足注入蜜蜂毒,观察其自发痛、持续性热刺激缩足反应潜伏期和机械刺激缩足反应阈值的变化.结果:在高血糖状态下,左足底注射蜜蜂毒引起大鼠持续性自发缩足反射的次数增加;对于病理性痛反应,与基础水平比较,热刺激和机械刺激缩足反应阙值均降低(P＜0.05),但是实验组大鼠和生理盐水组大鼠热刺激的反应性比较无差异(P＞0.05),实验组大鼠机械刺激缩足反应阈值较生理盐水组大鼠降低(P＜0.05).结论:高血糖大鼠皮下注射蜜蜂毒诱导外周组织局部炎症后,可引起大鼠的持续性自发缩足反射增强和热及机械痛敏.     

外源性的电磁干预对神经病理性疼痛大鼠的镇痛效果研究

目的:探讨外源性的电磁干预方法对神经病理性疼痛大鼠的镇痛效果。方法:将30只成熟的雄性SD大鼠随机等分成3组:空白对照组(Control),坐骨神经慢性压迫损伤(CCI)组以及坐骨神经慢性压迫损伤协同电磁刺激组(CCI+EMF)。CCI组和CCI+EMF组的20只大鼠建立坐骨神经慢性压迫损伤模型,CCI+EMF组大鼠行外源性的全身性电磁刺激干预(脉冲波形,频率15 Hz,强度30 Gs),每天刺激6小时。在CCI模型构建的第0、3、6、9、12及15天对大鼠测试和比较足底机械痛阈值、足底热痛阈值、运动功能评分和神经传导速率。结果:CCI组大鼠的足底机械痛阈值、足底热痛阈值及感觉神经传导速率从CCI手术后的第3天即出现显著性降低,其6、9、12、15天足底机械痛阈值、足底热痛阈值及感觉神经传导速率均显著低于Control组(P0.01),而运动功能评分均显著高于Control组(P0.05)。CCI+EMF组大鼠的足底机械痛阈值、足底热痛阈值及感觉神经传导速率在第9、12、15天显著高于CCI组大鼠(P0.05),而运动功能评分均显著高于CCI l组。结论:外源性的电磁刺激对于神经病理性疼痛大鼠具有良好的镇痛效果,有望成为一种临床治疗神经病理性疼痛的新的物理治疗手段。     

普瑞巴林对神经病理性痛大鼠行为学的影响

目的:探讨普瑞巴林对神经病理性痛大鼠行为学的影响.方法:建立大鼠神经病理性痛模型(CCI模型),取40只雄性Sprague-Dawley大鼠随机分成4组,Ⅰ组为空白对照组,Ⅱ组为假手术组,Ⅲ组为CCI+普瑞巴林治疗组,Ⅲ组在术后第1夭开始灌胃给予3 mg/kg普瑞巴林,Ⅳ组为CCI手术组.分别于术前0 d及术后1 d、3 d、5 d、7 d、9 d、11 d、14d以热辐射法测定热缩足反射潜伏期(Paw withdrawal thermal latency, PWTL),观察神经病理性痛大鼠行为学变化.结果:术后14 d,Ⅳ组和Ⅰ、Ⅱ、Ⅲ组相比较,大鼠后爪的热痛敏阈值明显降低(P<0.01);Ⅰ、Ⅱ、Ⅲ组之间相比,大鼠后爪的热痛敏阚值差异没有显著性(P>0.05).结论:普瑞巴林可以缓解神经病理性痛大鼠的慢性神经病理痛行为学表现.     

脊髓水平给予胍丁胺对鞘内吗啡镇痛的影响

目的：观察脊髓水平给予胍丁胺对鞘内吗啡镇痛作用的影响。方法：30只sD大鼠随机分为3组（n=10）：C1组：鞘内注射生理盐水（10出）;M1组：鞘内注射吗啡（15μg／10μl）;AMI组：鞘内同时给予吗啡15μg＋胍丁胺12．5μg／10出。所有大鼠均于鞘内给药后5min于跖部皮下注射蜜蜂毒50m（0．2mg）致痛,观察并纪录1h内大鼠的自发缩足反射次数。另30只SD大鼠分组为C2,M2和AM2（n=10）,每组给药分别同前,用来测定机械痛阈和热刺激阈值。结果：与C1组比较,M1组1h内大鼠的自发缩足反射次数显著减少,提示鞘内注射吗啡对蜜蜂毒诱致的自发痛具有显著性抑制作用（P〈0．05）;鞘内同时给予吗啡和胍丁胺（AM1组）,大鼠自发缩足反射次数进一步减少,与M1组比较具有显著性意义（P〈0．05）。与对照组C2组比较,M2组大鼠的热刺激闽值和机械性痛闽明显提高;AM2组热刺激潜伏期显著延长,机械刺激阈值显著提高;AM2组与M2组比较热刺激潜伏期和机械刺激阈值有统计学差异（P〈0．05）。结论：鞘内胍丁胺和吗啡联合用药可显著增强吗啡对蜜蜂毒诱致自发痛的抑制作用,具有加强效应。     

脑室注射白介素-1β引起的热痛敏作用

实验在SD大鼠上应用脑室微量注射和辐射热测痛的方法,研究了脑内微量注射白介素－1β对大鼠痛阈的影响。实验大鼠分为给药组和对照组,在给药组大鼠脑室注射不同剂量的白介素－1β（5、50和500pg/kg),对照组大鼠脑室注射配药液。白介素－1受体拮抗剂（IL－1ra,50ng/kg)在脑室注射白介素－1β前20min给予。实验以大鼠对光热刺激引起的缩爪反射潜伏期为痛阈指标。结果表明,脑室注射白介素－1β可显著缩短大鼠对光热刺激的缩爪反射潜伏期,并具有剂量依赖性关系。脑室给予500pg/kg的白介素－1β 20min后,大鼠对光热刺激的缩爪反射潜伏期显著缩短,40min时达峰值,然后逐渐恢复。该作用可被白介素－1β受体拮抗剂阻断。结果提示脑中白介素－1β可通过作用于白介素－1受体引起热痛敏作用。     

不同频率电针刺激对神经痛模型大鼠脊髓背角P物质的影响

目的:观察P物质(Substance P,SP)在慢性坐骨神经压迫损伤(chronic constriction injury,CCI)模型脊髓中表达的变化,探讨电针镇痛的机制是否与脊髓背角中SP表达的变化有关。方法:选择32只雄性、体重180-200 g的SD大鼠,并将其随机均分为4组(n=8)。空白组(Con组)为正常痛阈值大鼠;假电针组(CCI+A组)在损伤的坐骨神经旁置入电针,但无电流刺激;2 Hz组和100Hz组分别给予相应频率电流刺激30 min。在实验开始前和术后1、4、7、14、20、22天记录大鼠的热缩足反射潜伏期(Paw Withdrawal Latency,PWL)和机械刺激缩足反射阈值(Paw Withdrawal Threshold,PWT)。免疫组化方法检测脊髓背角SP的表达。结果:术后20天,电针治疗后,100 Hz组和2 Hz组PWT分别为(7.33±1.42)g和(7.80±1.42)g,均显著高于假电针组(2.60±1.46)g,差异有统计学意义(P0.05)。100 Hz组在术后20天后和2 Hz组在术后14天后PWL值均显著高于假电针组,差异有统计学意义(P0.05)。免疫组化显示:2 Hz组和100 Hz组大鼠脊髓背角中P物质阳性细胞显著低于假电针组(P0.05)。结论:坐骨神经旁电针刺激能够显著减轻CCI模型大鼠热痛觉及机械痛觉过敏,其机制可能与抑制脊髓背角SP的表达有关。     

HO/CO在甲醛炎性痛大鼠脊髓痛传导及痛觉过敏产生中的作用

目的:探讨血红素氧合酶/一氧化碳(HO/CO)在甲醛诱导的大鼠自发痛和痛觉过敏形成中的作用。方法:采用鞘内注射的方法,在甲醛炎性痛大鼠和正常大鼠分别给予HO抑制剂Znpp和HO激动剂Hemin;采用加权积分法对痛反应进行评分以代表痛反应程度;采用观察热辐射缩足潜伏期和机械刺激缩足反射阈值表示热和机械性痛觉过敏的程度。结果:Znpp各剂量组与单纯甲醛组相比,大鼠痛反应评分明显降低,且Znpp剂量越大,对大鼠痛反应的抑制作用越明显;与单纯甲醛组相比,Znpp各剂量组大鼠注射足热辐射缩足潜伏期和机械刺激缩足反射阈值均无明显变化,而非注射足热辐射缩足潜伏期和机械刺激缩足反射阈值均明显升高,且Znpp的剂量越大,这种改变越明显。正常大鼠鞘内注射HO的激动剂Hemin后,双侧足热辐射缩足潜伏期和机械刺激缩足反射阈值均明显降低。结论:鞘内给予HO抑制剂可明显抑制甲醛诱导的自发痛反应及热和机械性痛觉过敏程度;正常大鼠鞘内给予HO激动剂可诱发热和机械性痛觉过敏的产生,提示HO/CO系统参与脊髓伤害性信息的传导和痛觉过敏的形成过程。     

姜黄素对糖尿病大鼠疼痛过敏的影响

目的：神经病理性痛是糖尿病最常见的并发症之一,本课题旨在探讨姜黄素对糖尿病大鼠痛觉过敏的影响及其分子机制。方法：30只雄性SD大鼠随机分为对照组、糖尿病组和姜黄素治疗纽,模型纽和姜黄素治疗组利用腹腔注射链脲佐菌素（Streptozotocin,STZ）制备大鼠糖尿病模型,定期检测大鼠血糖、饮食、体重等变化,治疗组于STZ注射2wk后定期灌服姜黄素,分别在2wk和4wk后检测各组大鼠热痛敏和机械痛敏反应,在第4wk利用ELISA分别检测各组大鼠脊髓背角TNF-α表达变化。结果：STZ注射组大鼠2周后出现血糖〉14mol／L,并且该模型具有高血糖、体重增长缓慢、多饮多食多尿的特点,符合Ⅰ型糖尿病特征,痛行为测试结果显示糖尿病大鼠出现痛觉过敏,经过给予姜黄素灌服治疗后,痛觉过敏有所减轻,ELISA分析结果表明糖尿病大鼠脊髓背角TNF-α表达升高,经过姜黄素治疗后TNF-α表达有所下降。结论：成功制备STZ-型糖尿病大鼠模型,经过姜黄素治疗可以减轻糖尿病引起的疼痛过敏,姜黄素对糖尿病疼痛的治疗作用可能是通过降低大鼠脊髓背角TNF-α表达实现的。     

靶向毁损触液核对大鼠痛行为及脊髓背角5-HT和c-Fos表达的影响

目的：观察缺失触液核（CSF-contacting nucleus）对大鼠痛行为及脊髓背角痛相关物质5-羟色胺（5-HT）和c—Fas表达的影响,为触液核参与疼痛调制及机制提供实验依据。方法：成年雄性SD大鼠随机分为正常组（Control）,假手术组（Sham）,霍乱毒素亚单位B与辣根过氧化酶复合物（CB—HRP）组和毁损触液核组（Damage）。以机械缩足阈值（MWT）和热缩足潜伏期（耶儿）测定大鼠痛行为。免疫荧光法检测脊髓背角5-HT和c—Fos表达,并进行痛行为阈值与物质变化趋势的相关分析。结果：与Control、Sham和CB—HRP组相比,Damage组大鼠MWT和TWL明显降低（P〈0．05）。免疫荧光结果显示,正常大鼠触液核神经元高表达5-HT;Damage组大鼠触液核神经元数量随毁损天数延续逐渐减少,且在给予毁损剂CB—SAP第10天完全消失。与此同时脊髓背角5-HT和c—Fos表达量日趋增加,且与痛行为阈值变化趋势成负相关。结论：CB—SAP能科学可靠靶向毁损触液核,缺失触液核可致大鼠痛行为阈值减低,而脊髓背角5-HT和c—Fos表达量增加。本研究提示触液核参与了疼痛调制,且5-HT和c—Fos在此调制中发挥了重要作用。     

MiR‐101 promotes pain hypersensitivity in rats with chronic constriction injury via the MKP‐1 mediated MAPK pathway

This study was performed to characterize the effect of microRNA‐101 (miR‐101) on the pain hypersensitivity in CCI rat models with the involvement of mitogen‐activated protein kinase phosphatase 1 (MKP‐1) in spinal cord microglial cells. The mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) in the developed CCI models were determined to assess the hypersensitivity of rats to mechanical stimulation and thermal pain. To assess inflammation, the levels of interleukin (IL)‐1β, IL‐6 and tumour necrosis factor‐α (TNF‐α) in the spinal dorsal horns of CCI rats and lipopolysaccharide (LPS)‐activated microglial cells were examined. miR‐101 and MKP‐1 gain‐ and loss‐of‐function experiments were conducted in in vivo and in vitro settings to examine the roles of miR‐101 and MKP‐1 in CCI hypersensitivity and inflammation. The results showed that miR‐101 was highly expressed in the spinal dorsal horn and microglial cells of CCI rat models. Furthermore, overexpression of miR‐101 promoted the pain hypersensitivity in CCI rat models by reducing MWT and TWL. The overexpression of miR‐101 also promoted inflammation in LPS‐exposed microglial cells, as indicated by increased levels of IL‐1β, IL‐6 and TNF‐α. MiR‐101 was shown to target MKP‐1, inhibiting its expression. Moreover, miR‐101 promoted pain hypersensitivity in CCI rat models by inhibiting MKP‐1 expression and activating the mitogen‐activated protein kinase (MAPK) signalling pathway. Taken together, miR‐101 could potentially promote hypersensitivity and inflammatory response of microglial cells and aggravate neuropathic pain in CCI rat models by inhibiting MKP‐1 in the MAPK signalling pathway.     

Vatalanib decrease the positive interaction of VEGF receptor-2 and P2X(2/3) receptor in chronic constriction injury rats

Neuropathic pain can arise from a lesion affecting the peripheral nervous system. Selective P2X(3) and P2X(2/3) receptors' antagonists effectively reduce neuropathic pain. VEGF inhibitors are effective for pain relief. The present study investigated the effects of Vatalanib (VEGF receptor-2 (VEGFR-2) inhibitor) on the neuropathic pain to address the interaction of VEGFR-2 and P2X(2/3) receptor in dorsal root ganglia of chronic constriction injury (CCI) rats. Neuropathic pain symptoms following CCI are similar to most peripheral lesions as assessed by the Neuropathic Pain Symptom Inventory. Sprague-Dawley rats were randomly divided into sham group, CCI group and CCI rats treated with Vatalanib group. Mechanical withdrawal threshold and thermal withdrawal latency were measured. Co-expression of VEGFR-2 and P2X(2) or P2X(3) in L4-6 dorsal root ganglia (DRG) was detected by double-label immunofluorescence. The modulation effect of VEGF on P2X(2/3) receptor agonist-activated currents in freshly isolated DRG neurons of rats both of sham and CCI rats was recorded by whole-cell patch-clamp technique. The mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) in CCI group were lower than those in sham group (p<0.05). MWT and TWL in CCI rats treated with Vatalanib group were increased compared with those in CCI group (p<0.05). VEGFR-2 and P2X(2) or P2X(3) receptors were co-expressed in the cytoplasm and surface membranes of DRG. The co-expression of VEGFR-2 and P2X(2) or P2X(3) receptor in CCI group exhibited more intense staining than those in sham group and CCI rats treated with Vatalanib group, respectively. VEGF enhanced the amplitude of ATP and α,β-meATP -activated currents of both sham and CCI rats. Increment effects of VEGF on ATP and α,β-meATP -activated currents in CCI rats were higher than those in sham rats. Both ATP (100 μM) and α,β-meATP (10 μM)- activated currents enhanced by VEGF ( 1nM) were significantly blocked by Vatalanib (1 μM, an inhibitor of VEGF receptors). The stain values of VEGFR-2, P2X(2) and P2X(3) protein expression in L4/5 DRG of CCI treated with Vatalanib group were significantly decreased compared with those in CCI group (p<0.01). Vatalanib can alleviate chronic neuropathic pain by decreasing the activation of VEGF on VEGFR-2 and the positive interaction between the up-regulated VEGFR-2 and P2X(2/3) receptors in the neuropathic pain signaling.     

RvD1对大鼠2型糖尿病神经病理性痛的作用及机制研究

目的：采用2型糖尿病神经病理性痛大鼠,探讨其脊髓背角小胶质细胞极化情况以及消退素D1（RvD1）缓解大鼠2型糖尿病神经病理性痛的机制。方法：雄性SD大鼠高糖高脂饲养,腹腔注射链脲佐菌素（STZ）,制备大鼠2型糖尿病神经病理性痛模型。将2型糖尿病神经病理性痛大鼠随机分为3组（n=36）：2型糖尿病神经病理性痛组（D组）、2型糖尿病神经病理性痛注射RvD1组（R组）和溶剂对照组（S组）。R、S组分别于注射STZ 14 d后蛛网膜下腔置管,3 d后R、S组分别给予RvD1 10μl（10 ng/μl）和100%乙醇10μl,每天1次,连续14 d,D组不做任何处理。另取36只正常大鼠为正常对照组（N组）,普通饲料喂养。鞘内给药后第1、3、7、14天时测定机械缩足阈值（MWT）和热缩足潜伏期（TWL）,各组随机取9只大鼠处死,取L4-6脊髓膨大,采用Western blot法检测小胶质细胞M1、M2型极化标记物,即诱导型一氧化氮合酶（iNOS）、精氨酸酶1（Arg1）的表达。结果：与N组比较,D、S组第1、3、7、14天时MWT降低、TWL缩短,脊髓背角Arg1表达减少,iNOS表达增多（P < 0.05）;与D组比较,R组第7、14天时MWT升高、TWL延长,脊髓背角Arg1表达增多,iNOS表达减少（P < 0.05）;D组与S组各指标比较差异无统计学意义。结论：RvD1促进小胶质细胞M2型极化并缓解大鼠2型糖尿病神经病理性痛。     

Analgesic effect of electroacupuncture on chronic neuropathic pain mediated by P2X3 receptors in rat dorsal root ganglion neurons

Adenosine 5'-triphosphate disodium (ATP) gated P2X receptors, especially the subtype P2X(3), play a key role in transmission of pain signals in neuropathic pain, ATP has been documented to play a significant role in the progression of pain signals, suggesting that control of these pathways through electroacupuncture (EA) is potentially an effective treatment for chronic neuropathic pain. EA has been accepted to effectively manage chronic pain by applying the stimulating current to acupoints through acupuncture needles. To determine the significance of EA on neuropathic pain mediated by P2X(3) receptors in the dorsal root ganglion (DRG) neurons, mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were recorded, and the expression of P2X(3) receptors in the DRG neurons was assessed by immunohistochemistry (IHC) and in situ hybridization (ISH). In addition, the currents which were evoked in DRG neurons isolated from rats following chronic constriction injury (CCI) by the P2X(3) receptors agonists i.e. ATP and α,β-methylen-ATP (α,β-meATP) were examined through the experimental use of whole cell patch clamp recording. The present study demonstrates that EA treatment can increase the MWT and TWL values and decrease the expression of P2X(3) receptors in DRG neurons in CCI rats. Simultaneously, EA treatment attenuates the ATP and α,β-meATP evoked currents. EA may be expected to induce an apparent induce analgesic effect by decreasing expression and inhibiting P2X(3) receptors in DRG neurons of CCI rats. There is a similar effect on analgesic effect between rats with contralateral EA and those with ipsilateral EA.