6-羟多巴胺纹状体内注射制作大鼠帕金森病模型的研究

目的 为拓宽6-OHDA损毁多巴胺能神经元所制备大鼠帕金森病模型的应用范围,采用多位点纹状体内注入6-OHDA的途径来制备模型。方法 研究用SD大鼠,两个针道内四点定位注射,每点注射3μg／μ16-OHDA3μl。结果 术后两周出现缓慢旋转,4周旋转行为达到7转／分并保持稳定;形态学染色可见损毁1周后注射侧黑质酪氨酸羟化酶免疫组化阳性细胞减少20％,2周后减少38％,3～4周减少70％以上,6周后损伤趋缓。高效液相-电化学法活体检测纹状体内多巴胺的代谢产物3、4-二羟基苯乙酸(DOPAC)和高香草酸(HVA),发现注射侧和非注射侧相比含量分别下降98．33％和96．05％;组织匀浆检测损毁侧黑质多巴胺含量下降了73％以上,3、4-二羟基苯乙酸(DOPAC)含量下降60％。结论 纹状体内注射6-OHDA能够制备帕金森病大鼠模型。     

帕金森病模型大鼠脑内多巴胺与铁含量的关系

实验采用原子吸收分光光度法,快速周期伏安法,高效液相电化学检测等方法,研究以6－羟基多巴（6－OHDA）制备的帕金森病（PD）模型大鼠黑质内铁含量的变化。铁对多巴胺（DA）能神经元的直接毒性作用以及铁离子螯合剂甲磺酸去铁胺的神经保护作用。结果发现：（1）PD大鼠损毁侧黑质内铁含量为非标准PD大鼠的3倍左右;（2）PD大鼠损毁侧纹状体内铁含量无明显改变;（3）单纯注射6－OHDA的大鼠其损毁侧纹状体（CPu)DA的释放量和含量均明显降低;（4）侧脑室预先注射甲磺酸去铁胺,再重复上述实验,损毁侧CPu DA释放量和含量均无明显改变;（5）单侧黑质内注射40ug FeCl3后,大鼠损毁侧CPu内DA释放量和含量显著降低。上述结果提示,6－OHDA可导致CPu DA释放量及含量减少,此过程有铁的参与。由于铁可导致DA神经元死亡,因此铁含量的增加可能是DA含量减少的原因之一,甲磺酸去铁胺具有保护DA神经元的作用。     

6-OHDA制备的帕金森病大鼠黑质神经元的超微结构改变

目的观察6-羟多巴胺(6-OHDA)单侧注射制备的帕金森病(PD)大鼠多巴胺(DA)能神经元的超微结构改变。方法单侧微量注射6-OHDA制备PD大鼠模型,用免疫荧光组织化学方法观察正常侧与6-OHDA注射侧黑质酪氨酸羟化酶(TH)阳性神经细胞及神经纤维的变化;并利用免疫电镜技术观察大鼠正常侧与注射侧黑质致密部DA能神经元的超微结构。结果免疫荧光法显示注射侧黑质致密部TH阳性细胞数和网状部TH阳性纤维面积与正常侧的百分比平均值分别为21.83%,23.19%。免疫电镜显示:TH免疫反应阳性产物表达于PD大鼠正常侧DA能神经元的高尔基复合体质膜面及胞质内,电子密度较高,注射侧很少见或几乎未见,且注射侧线粒体嵴有不同程度的溶解,呈空泡样变或髓样变,粗面内质网脱颗粒。结论6-OHDA可引起DA能神经元发生凋亡的超微结构改变。     

脑室注射6-羟多巴胺对黄鼠冬眠入眠的影响

采用脑室内注射化学去交感药物6-羟多巴胺(6-OHDA)的方法,观察了人为地降低脑内去甲肾上腺素(NE)系统活动对达乌尔黄鼠入眠的影响。结果表明:脑室注射100-200μg6-OHDA使脑内NE含量减少50％以上,明显促进黄鼠入眠,平均入眠诱导期比自然冬眠动物明显缩短,整个冬眠季内冬眠时间延长,冬眠黄鼠仍具有正常的入眠觉醒周期。这些结果提示脑内NE系统活动水平降低是触发动物入眠的重要因素之一。     

大鼠灰翼区微量注射6-羟多巴胺对迷走-迷走抑胃反射的影响

本文用乌拉坦麻醉的大鼠,观察了灰翼区微量注射6-羟多巴胺(6-OHDA)对迷走-迷走抑胃反射的影响。实验动物分三组:空白对照组、溶媒组和6-OHDA 组。实验结果表明,在溶媒组刺激迷走神经中枢端使胃电慢波的振幅和胃内压分别下降到刺激前对照值的36.87±22.07%和32.52±25.41%,与空白对照组相比无显著的差异(P>0.05)。但是,在6-OHDA组,刺激迷走神经中枢端对胃电和胃运动的抑制效应明显减弱,慢波的振幅与胃内压分别下降到刺激前对照值的67.48±13.21%和50.88±21.40%,同溶媒组相比有非常显著的差异(P<0.01)。结果提示,延髓灰翼区的儿茶酚胺能神经参与迷走-迷走抑胃反射的中枢机制。     

脑内移植治疗帕金森病的研究概述

帕金森病(Parkinson’s disease,PD)是一种与年龄有关的神经系统退行性疾病,主要是由于中脑黑质多巴胺能神经元的变性和消失而造成纹状体内多巴胺水平下降。其行为上特征性表现为运动减少,肌肉僵直、震颤。除了运动感觉功能障碍外,病人还可能伴有心理障碍及认知能力的下降。20世纪初,人们开始     

不同剂量鱼藤酮对拟帕金森病模型大鼠行为学及纹状体多巴胺含量的影响

目的观察不同剂量鱼藤酮皮下注射对大鼠行为学及脑纹状体多巴胺含量的影响,探讨鱼藤酮拟帕金森病大鼠模型的适宜造模条件。方法Lewis大鼠分别给予鱼藤酮不同剂量（1．0、1．5和2．0ms／ks／d）皮下注射共28d。应用旷场实验和斜板实验分别测定大鼠的运动功能和协调性,高效液相色谱一电化学法检测大鼠纹状体内多巴胺含量。结果模型组大鼠存活率随鱼藤酮注射剂量的升高呈逐渐下降趋势。鱼藤酮2．0mg／kg组大鼠体重最先明显降低,随着注射时间的延长,各模型组大鼠均出现显著的体重降低。旷场实验结果显示,各剂量鱼藤酮组大鼠跨格次数和站立次数均明显下降。斜板实验结果显示,鱼藤酮1．5mg／kg组大鼠在斜板的停留角度较对照组显著减小。鱼藤酮I．5ms／kg组大鼠脑纹状体内多巴胺含量显著降低。结论鱼藤酮1．5ms／kg皮下注射28d,大鼠出现明显的运动功能障碍和脑内多巴胺含量减少,而死亡率相对较低,因此是建立帕金森病大鼠模型的适宜剂量。     

6-羟基多巴胺损毁大鼠内侧前脑束早期黑质-纹状体系统的铁水平与多巴胺能神经元退变的关系

应用快速周期伏安法(fast cyclic vohammetry,FCV)、原子吸收分光光度法及免疫组织化学方法,观察了6-羟基多巴胺(6-hydroxydopamine,6-OHDA)单侧损毁大鼠内侧前脑束(medial forebrain bundle,MFB)早期黑质(sub-stantia nigra,sN)铁水平与多巴胺(dopamine,DA)神经元损伤的变化,以及纹状体(striatum,Str)的DA释放。结果如下：6-OHDA单侧损毁大鼠MFB1d和3d后,SN的酪氨酸羟化酶(tyrosine hydroxylase,TH)阳性细胞分别下降了45％和66％;与正常鼠和未损毁侧相比,损毁侧SN的铁染色增强,铁浓度增加,而Str的DA释放量不变;6-OHDA损毁后1d与3d组相比,损毁侧的铁染色、铁浓度及DA释放量差别无显著性。上述结果表明,6-OHDA单侧损毁大鼠MFB的早期阶段,SN的DA能神经元数目中等程度减少时,铁染色及铁浓度即有增加,由于DA能神经系统有强大的代偿功能,使得Str的DA释放量仍趋于正常。     

选择性5-HT1A受体拮抗剂WAY-100635增加6-羟多巴胺损毁大鼠杏仁基底外侧核的神经活动

本文采用玻璃微电极细胞外记录法,观察正常大鼠和6-羟多巴胺(6-hydroxydopamine,6-OHDA)损毁黑质致密部大鼠杏仁基底外侧核(basolateral nucleus,BL)神经元电活动的变化,以及体循环给予选择性5-HT1A受体拮抗剂WAY-100635对神经元电活动的影响.结果显示,正常大鼠BL投射神经元和中间神经元的放电频率分别足(O.39±0.04)Hz和(0.83±0.16)Hz,6-OHDA损毁大鼠BL投射神经元和中间神经元的放电频率分别足(0.32±0.04)Hz和(0.53±0.12)Hz,与正常大鼠相比无显著差异.在正常大鼠,所有投射神经元呈现爆发式放电;94%的中间神经元为爆发式放电,6%为不规则放电.在6.OHDA损毁大鼠,85%的投射神经元呈现爆发式放电,15%为不规则放电;86%的中间神经元为爆发式放电,14%为不规则放电,与正常大鼠相比无显著差别.静脉给予0.1 mg/kg体重的WAY-100635不改变正常大鼠和6-OHDA损毁人鼠BL投射神经元和中间神经元的放电频率.然而,0.5 mg/kg体重的WAY-100635却显著降低正常大鼠BL投射神经元的平均放电频率(P<0.01),明显增加6-OHDA损毁大鼠BL投射神经元的平均放电频率(P<0.004).高剂量WAY-100635不影响正常大鼠和6-OHDA损毁大鼠BL中间神经元的平均放电频率.结果表明,黑质多巴胺能损毁后内在和外在的传入调节BL神经元的活动,在正常大鼠和6-OHDA损毁大鼠5-HT1A 受体调节投射神经元的活动,并且在6-OHDA损毁大鼠WAY-100635诱发投射神经元平均放电频率增加.结果提示,5-HT1A 受体在帕金森病情感性症状的产生中起重要作用.     

海马内注射6—OHDA对大鼠条件回避行为习得和保持的影响

本文研究了在大鼠两侧背海马内注射6-OHDA后,儿茶酚胺能纤维和多巴胺能纤维末梢在穿梭箱条件性回避行为习得和保持中的作用。结果表明:(1)6-OHDA使海马内多巴胺含量明显减少,特别是去甲肾上腺素含量的减少更为显著。(2)条件性回避行为的习得受到损害。(3)海马内注射加压素易化条件性回避行为的效应被阻断。     

Brain Catecholamine Alterations and Pathological Features with Aging in Parkinson Disease Model Rat Induced by Japanese Encephalitis Virus

We analyzed two disease model groups with rats infected by Japanese encephalitis virus (JEV), a 90-day group and a 180-day group after JEV infection. The time measured by the modified pole test showed that motor activities in these two groups were slower than those of age-matched control groups. Striatal dopamine (DA) levels were significantly decreased in all JEV-infected rats. Norepinephrine concentration in brain regions in the 180-day group was significantly decreased in the medulla oblongata and hypothalamus as compared with the control and 90-day group. Tyrosine hydroxylase-positive neurons were significantly decreased in both JEV-infected rat groups. These results suggest that DA decrease and pathological changes in JEV-infected model rats persist for a long time, at least up to 180 days, and this model will be useful for the evaluation of new anti-parkinsonian agents.     

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson's Disease

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson''s disease (PD) has proved to be invaluable in advancing our understanding of the mechanisms underlying parkinsonian symptoms, since it recapitulates the changes in basal ganglia circuitry and pharmacology observed in parkinsonian patients^1-4. However, the precise cellular and molecular changes occurring at cortico-striatal synapses of the output pathways within the striatum, which is the major input region of the basal ganglia remain elusive, and this is believed to be site where pathological abnormalities underlying parkinsonian symptoms arise^3,5.In PD, understanding the mechanisms underlying changes in basal ganglia circuitry following degeneration of the nigro-striatal pathway has been greatly advanced by the development of bacterial artificial chromosome (BAC) mice over-expressing green fluorescent proteins driven by promoters specific for the two striatal output pathways (direct pathway: eGFP-D1; indirect pathway: eGFP-D2 and eGFP-A2a)⁸, allowing them to be studied in isolation. For example, recent studies have suggested that there are pathological changes in synaptic plasticity in parkinsonian mice^9,10. However, these studies utilised juvenile mice and acute models of parkinsonism. It is unclear whether the changes described in adult rats with stable 6-OHDA lesions also occur in these models. Other groups have attempted to generate a stable unilaterally-lesioned 6-OHDA adult mouse model of PD by lesioning the medial forebrain bundle (MFB), unfortunately, the mortality rate in this study was extremely high, with only 14% surviving the surgery for 21 days or longer¹¹. More recent studies have generated intra-nigral lesions with both a low mortality rate >80% loss of dopaminergic neurons, however expression of L-DOPA induced dyskinesia^11,12,13,14 was variable in these studies. Another well established mouse model of PD is the MPTP-lesioned mouse¹⁵. Whilst this model has proven useful in the assessment of potential neuroprotective agents¹⁶, it is less suitable for understanding mechanisms underlying symptoms of PD, as this model often fails to induce motor deficits, and shows a wide variability in the extent of lesion^{17, 18}.Here we have developed a stable unilateral 6-OHDA-lesioned mouse model of PD by direct administration of 6-OHDA into the MFB, which consistently causes >95% loss of striatal dopamine (as measured by HPLC), as well as producing the behavioural imbalances observed in the well characterised unilateral 6-OHDA-lesioned rat model of PD. This newly developed mouse model of PD will prove a valuable tool in understanding the mechanisms underlying generation of parkinsonian symptoms.     

Melatonin reverses neurochemical alterations induced by 6-OHDA in rat striatum

The present work showed that the intrastriatal injection of 6-OHDA significantly decreases DA, DOPAC and HVA levels in that rat brain structure. Although there is also a decrease in 5-HT levels no changes were observed in 5-HIAA levels as compared to controls. On the other hand, melatonin (2, 5, 10 and 25 mg/kg. i.p., daily for 7 days) treatment starting 1 h after 6-OHDA lesions, partially reverses the decreases caused by 6-OHDA lesions on these neurotransmitter levels, and contents were brought to approximately 50% of that observed in the contralateral sides of controls or of melatonin treated group. Melatonin was more efficient at the doses of 5 and 10 mg/kg, i.p., and effects were similar between the lowest and highest doses characteristic of a bell-shaped type of response. The apomorphine-induced rotational behavior (3 mg/kg, i.p.) was blocked by 60, 89, 78 and 47% after the doses of 2, 5, 10 and 25 mg/kg, i.p., respectively. Similarly, in this case the doses of 5 and 10 mg/kg were also more efficient. Melatonin (5 mg/kg) produced an upregulation of D1 receptors associated with a decrease in Kd value. While no change was observed in maximum density of D2 receptors, the Kd value was also decreased.     

Time Dependent Effects of 6-OHDA Lesions on Iron Level and Neuronal Loss in Rat Nigrostriatal System

The early changes in iron level and neuronal loss in rat nigrostriatal system were investigated using 6-hydroxydopamine (6-OHDA) unilaterally lesioned rats. The results showed that: 1, 3, 5, 7, and 14 days of postlesion, there was a progressive reduction in the density of the tyrosine hydroxylase immunoreactive (TH-ir) cells in the lesioned substantia nigra (SN). Iron level increased in the lesioned SN from 1–14 days following 6-OHDA lesions, but there were no differences in iron level among them. Only on 14 days of postlesion, did the DA release decrease in striatum (Str) of the lesioned side, while there were no changes in other groups. These results implied that the increased iron level in SN occured when there was a moderate reduction of DA neurons. However, the DA release in Str was unchanged until TH-ir cells were highly reduced due to the immense compensatory mechanism of the DA system.     

大鼠实验性帕金森病的基因治疗

将稳定转染了大鼠酪氨酶羟化酶（Ｔｙｒｏｓｉｎｅｈｙｄｒｏｘｙｌａｓｅ,ＴＨ）基因的大鼠成肌细胞移植于帕金森病大鼠模型的纹状体,进行基因治疗研究。ＲＴ－ＰＣＲ和免疫组织化学检测都证明转基因细胞可在纹状体内存活并表达ＴＨ,动物的不对称旋转行为明显改善,而且疗效可维持半年以上。     

大鼠体外循环模型的建立

目的建立大鼠体外循环实验模型。方法雄性成年SD大鼠10只(450～550g),麻醉后给予气管切开,呼吸机辅助通气,股动脉置管接监护仪实时监测动脉血压并按时采集动脉血标本,股静脉置管用于持续补液和静脉血样采集。经右颈静脉置多孔引流管至右心房行中心静脉引流,血液经特制微型膜肺氧合后,由蠕动泵经右颈动脉实施灌注。本模型采用林格氏液和贺斯进行无血预充,总量为16ml,晶胶比为1∶1,灌注流量100～150ml(kg·min),转流时间60min。结果实验中膜肺氧合满意,血流动力学稳定,所有动物都成功脱机。结论利用大鼠可以建立简单、廉价的体外循环动物模型。本模型适用于研究与体外循环相关的全身炎症反应和多脏器功能损伤的病理生理机制及其防治策略。     

实验性X综合征大鼠模型的建立

目的　建立一种典型的X综合症动物模型。方法　雄性SD大鼠施行两肾一夹术后普通饲料喂养 4周 ,诱发肾性高血压 ,继以高果糖饲料喂养 4周 ,诱导建立X综合症模型。结果　术后 4周 ,大鼠仅出现收缩压升高 ,血糖、血脂未见明显改变。高果糖饲料喂养 4周后 ,大鼠出现高血糖、高胰岛素血症、胰岛素抵抗、高血压和高脂血症。结论　肾性高血压形成后高果糖饮食 1个月 ,可诱导SD大鼠出现典型的X综合症 ,为研究胰岛素抵抗及其伴随的心血管疾病提供了一种理想的动物模型。     

帕金森病抑郁的相关因素分析

目的:探讨抑郁在帕金森病中(Parkinson’s disease,PD)的发生率及其影响因素。方法:对确诊的PD患者,采用汉密尔顿抑郁量表(Hamilt depression scale,HAMD)、简易精神状态检查量表(Mini-Mental State Examination,MMSE)及Webster功能评分量表进行评定,分析抑郁的发生情况和相关影响因素。结果:PD伴发抑郁者32例,抑郁的发生率为49.2%,病程、文化程度、Webster评分、MMSE评分与帕金森抑郁的发生均有统计学意义(P<0.05),年龄、性别、婚姻状况、经济情况与帕金森抑郁的发生均无统计学意义(P>0.05)。回归分析发现病程和病情严重程度是PD患者抑郁的危险因素。结论:PD患者有较高的抑郁发生率,抑郁的发生可能是社会心理、神经生物学多种因素作用的结果。