偏头痛大鼠颈髓中间神经元的变化及其对运动神经元的影响

目的:探讨偏头痛大鼠模型颈髓1~2中间神经元的变化及其对运动神经元的影响。方法:雄性SD大鼠随机分为空白组、假手术组、致炎剂(inflammatory soup,IS)组(IS 3 d、6 d、10 d)、生理盐水组。采用IS刺激大鼠硬脑膜模型,应用Von Frey纤维丝测定大鼠眶周痛觉阈值,应用Western blot和免疫荧光法观察颈髓中间神经元、运动神经元的变化。结果:随着IS反复刺激,大鼠眶周痛阈逐渐下降;IS组大鼠运动神经元乙酰胆碱转移酶Ch AT和c-Fos表达量较对照组增多,平均吸光度值增加;谷氨酸能中间神经元表达增强,GABA能中间神经元表达减弱。结论:反复硬脑膜给予致炎剂能有效诱导大鼠反复发作偏头痛并出现痛觉超敏,颈髓前角运动神经元兴奋性增加,这可能与颈髓中间神经元"兴奋—抑制"稳态失调相关。     

头痛宁对偏头痛大鼠硬脑膜及三叉神经节IL-1β表达影响

目的:研究头痛宁对反复发作性偏头痛大鼠硬脑膜及三叉神经节IL-1β(interleukin-1beta)表达的影响。方法:选择健康成年雄性SD大鼠60只,初始质量180~220 g。先取20只随机分为4组,每组5只,分别为生理盐水7天组(sham)、致炎剂(inflammatory soup,IS)1天组(IS1)、致炎剂3天组(IS3)和致炎剂7天组(IS7),建立反复发作性偏头痛大鼠模型;再取25只随机分为5组,每组5只,分别为致炎剂7天对照组(IS7)、0.375 g/kg头痛宁干预组(T1)、0.75 g/kg头痛宁干预组(T2)、1.5 g/kg头痛宁干预组(T3)和3.0 g/kg头痛宁干预组(T4),取硬脑膜及三叉神经节进行IL-1β蛋白免疫印迹分析(western blot)及IL-1βm RNA实时荧光定量PCR检测。最后取15只,随机分为3组,每组5只,分别为生理盐水7天组(sham)、致炎剂7天组(IS7)和3.0 g/kg头痛宁干预组(T4),灌流固定后各组分别取大鼠硬脑膜及三叉神经节进行IL-1β蛋白免疫荧光检测。结果:(1)与生理盐水7天组比较,致炎剂7天组IL-1β的表达在硬脑膜(P<0.01)及三叉神经节(P<0.01)均明显升高;(2)与致炎剂7天组相较,在大鼠硬脑膜/三叉神经节,0.75 g/kg头痛宁干预组(P<0.01/P<0.01)、1.5 g/kg头痛宁干预组(P<0.01/P<0.05)和3.0 g/kg头痛宁干预组(P<0.01/P<0.01)IL-1β表达均明显降低,0.375 g/kg头痛宁干预组IL-1β表达下降不明显;(3)与致炎剂7天组相较,3.0 g/kg头痛宁干预组三叉神经节神经元细胞胞质内免疫荧光斑点数明显减少,大鼠硬脑膜未能成功染色;(4)与致炎剂7天组相较,3.0 g/kg头痛宁干预组在硬脑膜(P<0.01)及三叉神经节(P<0.05)IL-1βm RNA表达均减少。结论:头痛宁抑制硬脑膜及三叉神经节IL-1β基因转录,下调IL-1β的表达可能是其治疗偏头痛的机制之一。     

P2X4受体在慢性偏头痛大鼠三叉神经脊束尾核的表达及意义

目的:观察P2X4受体(P2X4 receptor,P2X4R)在慢性偏头痛(chronic migraine,CM)大鼠三叉神经脊束尾核(trigeminal nucleus caudalis,TNC)的表达变化,探讨其在CM中的作用。方法:SD雄性大鼠随机分为5组,对照组(n=12)、模型组(n=12)、模型+溶剂组(n=6)、模型+TNP-ATP30 nmol组(n=6)、模型+TNP-ATP 60 nmol组(n=6)。硬脑膜反复滴注"炎性汤"(inflammatory soup,IS)建立CM模型。von Frey test检测眶周及后足机械痛阈值,Western blot检测P2X4R及神经元激活标志蛋白c-Fos的表达,免疫荧光检测P2X4R定位情况。结果:荧光染色显示P2X4R主要表达于TNC小胶质细胞。与对照组相比,模型组大鼠眶周及后足痛阈值显著降低(P<0.01),P2X4R及c-Fos表达量显著上调(P<0.05)。给予P2X4R抑制剂TNP-ATP后,大鼠眶周及后足痛阈值显著升高(P<0.05),c-Fos表达量显著降低(P<0.05)。结论:TNC小胶质细胞P2X4R表达上调,可能通过影响神经元的激活从而参与CM发病过程。     

蛋白激酶C参与大鼠慢性偏头痛中枢敏化

目的:本文旨在研究蛋白激酶C(PKC)是否通过中枢敏化参与慢性偏头痛病理生理过程。方法:SD雄性大鼠随机分为7组,假手术组(n=11),模型组(n=12),模型+溶剂组(n=8),PKC抑制剂4μg组(n=11),PKC抑制剂8μg组(n=10),PKC激动剂100 ng组(n=10),PKC激动剂200 ng组(n=11)。"炎性汤"反复刺激硬脑膜,模拟硬脑膜痛觉感受器的反复激活,建立慢性偏头痛大鼠模型,侧脑室给予PKC抑制剂和PKC激动剂,使用von Frey test测定大鼠面部及后足的机械痛阈值,Western blot检测PKC、CGRP、c-Fos蛋白的表达变化。结果:"炎性汤"反复刺激硬脑膜后,大鼠面部及足底的机械痛阈值显著降低,而三叉神经脊束尾核部位的CGRP、c-Fos及PKC蛋白表达显著增加。使用CHE抑制PKC可以显著增高大鼠面部及足底痛阈值,显著降低CGRP、c-Fos的表达;而使用PMA激活PKC则降低痛阈值,显著增高CGRP、c-Fos的表达。结论:本研究表明PKC可以调节机械痛阈值以及CGRP、c-Fos的表达变化,提示PKC参与慢性偏头痛中枢敏化的病理生理过程。     

偏头痛大鼠硬脑膜及三叉神经节上TRPV1的表达变化

目的:观察TRPV1在偏头痛大鼠硬脑膜及三叉神经节上的表达变化。方法:取健康成年雄性SD大鼠60只,按随机数字法分为对照组(control组)、实验复方致炎剂(inflammation soup,IS)1 d组(IS 1 d组)、实验IS 3 d组(IS 3 d组)和实验IS 6 d组(IS 6 d组)。在雄性SD大鼠硬脑膜上埋置PE-10管,实验组分别给予1 d、3 d、6 d IS 20μl/d及对照组给予6天等量生理盐水。采用WesternBlot、免疫荧光法和Real-Time PCR法检测TRPV1在硬脑膜及三叉神经节上的表达情况。结果:对照组和实验组大鼠硬脑膜及三叉神经节上均可见TRPV1表达;与对照组相比,实验组硬脑膜、三叉神经节上TRPV1蛋白表达量、阳性细胞数及mRNA明显升高,以实验IS 6 d组最高(P<0.05)。结论:TRPV1在偏头痛大鼠硬脑膜及三叉神经节上的表达增加。TRPV1表达上调可能是偏头痛外周敏化的重要机制之一。     

PTEN下调表达对反复发作性偏头痛大鼠模型行为学的影响

目的:探讨下调第10号染色体同源丢失性磷酸酶张力蛋白基因(phosphatase and tensin homolog deleted on chromosome ten,PTEN)对反复发作性偏头痛大鼠模型行为学的影响。方法:SD大鼠分4组,对照组(Normal组,n=8),假手术组(Sham+Ad-RFP组,n=8),模型组(IS组,n=15),干预组(IS+Ad R-si PTEN组,n=15)。炎性汤(inflammation soup,IS)刺激大鼠硬脑膜,建立反复发作性偏头痛大鼠模型,干预组在建立模型前1周三叉神经核(trigeminal nucleus caudalis,TNC)注射Ad R-si PTEN干预,通过畏光、痛阈等方法对大鼠行为学评价。结果:滴注IS后大鼠出现抓耳挠头,过度理毛等症状;9次IS后大鼠的痛阈阈值下降到<2.0 g;12次IS后大鼠表现为恐光,但是下调PTEN能有效逆转上述行为学改变(P<0.05)。结论:下调PTEN基因能有效逆转反复发作性偏头痛大鼠模型的行为学改变,对反复发作性偏头痛具有保护作用。     

探讨偏头痛大鼠三叉神经节内可能调控CGRP表达的微小RNA

目的:探索偏头痛大鼠三叉神经节内异常表达的微小RNA(micro RNA,miRNA)以及可能参与调节降钙素基因相关肽(calcitonin gene-related peptide,CGRP)基因表达的miRNA。方法:本研究中共选取36只180~220 g的雄性SD大鼠,每个实验中的大鼠均随机分组。首先,用炎性复合物(inflammation soup,IS)刺激大鼠硬脑膜来进行偏头痛大鼠模型的制备,并监测术后以及IS刺激硬脑膜后大鼠眶周机械痛阈的变化;其次,通过三个计算机程序预测出可能作用于CGRP基因的miRNA;然后,用蛋白质印迹法获得致炎剂组(IS组)大鼠三叉神经节内CGRP的量的变化;再次,用miRNA芯片技术得到IS-2 d组大鼠三叉神经节内miRNA的表达谱差异,并用实时定量PCR法对特别挑选出的miRNA的表达差异进行验证;最后,结合上述实验结果获得可能靶向作用于CGRP基因的miRNA。结果:根据观察,术后第五天大鼠的眶周痛阈与术前无明显差异,而IS-2 d组大鼠在术后第7天和第8天注射IS后2 h,其大鼠眶周痛阈较术前以及对照组均明显降低;根据miRanda、Target Scan和PITA三个软件的预测结果,共207个miRNA被预测出可能参与CGRP基因的表达调控;另外,与sham-8 d组相比,IS-2 d组大鼠三叉神经节内CGRP表达量显著增加、且有158个异常表达的miRNA;最后,在根据芯片结果的miRNA异常表达倍数及预测出的与CGRP基因表达调节相关性挑选出的6个miRNA的定量PCR实验结果中,miR-30a-3p和miR-449a-5p的表达量均显著下调。结论:miR-30a-3p和miR-449a-5p有可能参与了偏头痛大鼠三叉神经节中CGRP基因的表达调控。     

季德胜蛇药抗炎镇痛作用和对脊髓c-Fos表达影响的实验研究

目的:研究季德胜蛇药在福尔马林实验中的抗炎镇痛作用和对脊髓c-Fos表达的影响。方法:实验一:雄性SD大鼠40只,随机分为生理盐水组(NS组)、福尔马林组(F组)、季德胜蛇药组+福尔马林(S组)、扶他林组+福尔马林(V组)和复方利多卡因乳膏组+福尔马林(L组)(n=8)。记录各组注射后1小时的伤害性反应时间,每天观察各组注射足的水肿情况并测定机械缩足阈值和热缩足潜伏期,至第10天测量体重并做后足病理切片。实验二:雄性SD大鼠18只,随机分为生理盐水组(NS1组)、福尔马林组(F1组)和季德胜蛇药组+福尔马林(S1组)(n=6)。采用免疫组化方法结合电子计算机图像分析技术研究脊髓背角c-Fos蛋白表达。结果:S组的第一、二相反应时间小于V组、L组(P<0.05)。S组机械缩足阈值降低程度较F组和L组低(P<0.01)。S组的水肿增长率小于F组(P<0.05)。S组的体重增长与NS组比较无统计学差异(P>0.05)。病理方面,S组的炎症反应最轻。S1组c-Fos样免疫阳性神经元数量少于F1组(P<0.01)。结论:季德胜蛇药在大鼠福尔马林实验中具有降低大鼠伤害性反应,抗炎,抑制痛觉过敏的作用。季德胜蛇药抗大鼠伤害性反应的机制可能与降低脊髓水平c-Fos蛋白表达有关。     

布洛芬对偏头痛大鼠模型三叉神经节神经元电压门控型钙通道的影响

目的:探讨布洛芬对偏头痛大鼠模型三叉神经节神经元高电压激活钙通道电流(HVA-ICa)的调控作用。方法:雄性SD大鼠24只随机分为3组:生理盐水(NS)组、致炎剂(IS)+降钙素基因相关肽(CGRP)(IS+CGRP)组和布洛芬组,各8只。采用新型IS构建偏头痛大鼠模型,全细胞式膜片钳技术记录急性分离的三叉神经节神经元钙电流(HVA-ICa)。结果:与NS组相比较,(IS+CGRP)组HVA-ICa峰值密度增高,激活电压向超级化方向移动(P0.05);与(IS+CGRP)组相比,布洛芬组钙电流峰电流降低(P0.05)。与NS组相比,(IS+CGRP)组半数激活电压向超级化方向移动(P0.05);与(IS+CGRP)组相比,布洛芬组半数激活电压向去级化方向移动(P0.05)。与NS组相比,(IS+CGRP)组半数失活电压向去级化方向移动(P0.05);与(IS+CGRP)组相比,布洛芬组半数失活电压向超级化方向移动(P0.05)。结论:布洛芬能够降低三叉神经节神经元HVA-ICa的幅度、促进HVA-ICa的失活。     

ASIC3在慢性偏头痛大鼠三叉神经脊束尾核的表达

目的:本文通过检测酸敏感离子通道3(acid sensing ion channel 3,ASIC3)在慢性偏头痛大鼠三叉神经脊束尾核(trigeminal nucleus caudalis,TNC)的表达变化,探讨其在慢性偏头痛发病中的作用及可能机制。方法:将SD雄性大鼠随机分为7组,假手术组(n=18),模型组(n=18),模型+溶剂组(n=5),模型+ASIC3抑制剂5μM组(n=5),模型+ASIC3抑制剂10μM组(n=5),模型+ASIC3激动剂100μM组(n=5),模型+ASIC3激动剂200μM组(n=5)。使用Von Frey纤维丝测定大鼠眶周及足底的机械痛阈值,实时定量聚合酶链反应(q RT-PCR)检测ASIC3 m RNA,蛋白质印迹法(Western blotting)检测ASIC3、CGRP蛋白表达,免疫荧光技术检测ASIC3和CGRP在TNC部位的表达和定位情况。结果:(1)硬脑膜反复给予"炎性汤"后,大鼠眶周及足底的机械痛阈值显著降低(P<0.01);(2)与Sham组相比,CM组大鼠ASIC3在TNC的表达明显升高(P<0.05);(3)给予ASIC3抑制剂后,大鼠眶周及足底机械痛阈值均显著增加(P<0.05),CGRP蛋白的表达也明显降低(P<0.05),而给予ASIC3激动剂后,大鼠眶周及足底机械痛阈值均显著降低(P<0.05),CGRP蛋白的表达明显增加(P<0.05);(4)免疫荧光检测显示在TNC部位,ASIC3主要表达于神经元,且与CGRP有少量共表达。结论:ASIC3在慢性偏头痛大鼠TNC部位表达上调,且通过调节机械痛阈值的变化及CGRP的表达参与了慢性偏头痛的发生发展过程。     

TIMBER IN AUSTRALIA IN WARTIME

Fire history since 1850 was reconstructed by analysing tall wet eucalypt forest stands in the Warra long-term ecological research (LTER) site in southern Tasmania. Major fires occurred in 1898, 1906, 1914 and 1934. Fifty-seven percent of the forest has remained unburnt since 1850. Sixty percent of the forest burnt since 1850 occurs as mixed age stands, while 40 percent is pure regrowth. Ring counts of stumps of three veteran eucalypt trees indicated that the mature and oldgrowth forests of the Warra site represent a number of pre-1850 fires. One extant oldgrowth eucalypt stand was estimated to be over 450 years old. The common occurrence of multi-aged wet eucalypt stands has silvicultural implications where a management objective is to emulate pre-existing stand structures. While the fire history reported here is limited mainly to the period since 1850, it provides a good basis for understanding the current stand structures of the tall wet eucalypt forests of the Warra site. Such an understanding is essential to interpret results from current and planned ecological studies associated with the LTER program at Warra.     

STAGGERS IN SHEEP IN PATAGONIA

After observations and experimental work both in the field and laboratory, the following conclusions seem justified. 1. Staggers is a non-infectious disorder affecting horses, cattle, and sheep. 2. The disease is characterized by weakness, muscular twitching, irregular movements of the head, stiffness of the limbs, and transient motor paralysis, accompanied with spastic spasms on excitement. There is also a derangement of vision and conjunctivitis. 3. The postmortem lesions are not characteristic. 4. We readily produced the disease by feeding susceptible sheep on a coarse tuft grass commonly known as coiron or pampa grass (Poa argentina). 5. The time required to produce definite symptoms by feeding the grass varied. Two animals developed typical staggers after two feedings; in another instance a period of 21 days of feeding was required. The average time for the production of unmistakable symptoms in our experiments was 10 days. 6. Many sheep recover from staggers spontaneously. A complete change of diet will usually effect a cure within 2 weeks. 7. Older .animals that have pastured for long periods on lands where the grass grows become tolerant and are rarely affected with staggers. 8. The grass is toxic to sheep at all seasons of the year. We fed late winter and early spring grass and grass in flower, and produced staggers in every instance. The young green grass is as toxic as any edible portion of the plant.     

BACTERIUM PNEUMOSINTES IN CLINICAL INFLUENZA IN NEW YORK CITY IN 1926

The presence of Bacterium pneumosintes has been demonstrated in nasopharyngeal washings from 2 patients in a sporadic outbreak of clinical influenza in New York City in March, 1926. 2 strains of bacteria morphologically similar to Bact. pneumosintes, but differing in certain cultural characters, and 2 other anaerobic filter-passing organisms were also isolated from the 9 patients examined. The blood serum of 16 among 17 persons convalescent from clinical influenza, and of 6 among 10 supposedly normal persons, agglutinated 1 or more strains of Bact. pneumosintes.     

RENAL TRANSPLANTATION IN DIABETES MELLITUS IN RATS

Immunoglobulins and complement are deposited in the glomerular mesangium of rats with progressive glomerulosclerosis secondary to chemically induced diabetes mellitus. Isotransplantation of a kidney from a rat diabetic for 6 mo into a normal recipient results within 2 mo in the disappearance of IgG, IgM, and β₁C from the mesangium and arrest or reversal of the light microscopic glomerular lesions. Kidneys isotransplanted from normal donors into diabetic rats developed mesangial matrix thickening and deposition of IgG, IgM) and β₁C in the mesangium. No glomerular changes occur upon transplantation of a normal kidney into a normal rat. These findings indicate that diabetic glomerular changes in the rat are reversible and are secondary to the diabetic state rather than to the inducing agent.