γ-氨基丁酸能神经元在新生大鼠缺氧性痫性发作中的作用

目的 通过观察新生大鼠早期发育过程中及缺氧性痫性发作后海马组织病理改变以及原癌基因c-fos蛋白、谷氨酸脱羧酶(glutamate decarboxylase,GAD)的变化,探讨γ-氨基丁酸(γ-amino butylic acid,GABA)能神经元在缺氧性痫性发作中的作用及可能的影响机制.方法 采用出生后10d的SD大鼠建立改良Jensen缺氧诱导痫性发作模型,分为痫性发作后1d、3d、7d、14d 4组,并选取相应时间点正常大鼠为对照组,采用尼氏染色方法检测海马组织的组织病理变化,免疫组织化学法检测各组海马c-fos蛋白灰度值以及GAD阳性神经元数量的改变.结果 尼氏染色结果显示,各缺氧性痫性发作组海马区形态结构正常,细胞排列略稀疏,但未见明显的细胞丢失.免疫组化结果显示,与对照组比较,c-fos蛋白灰度值在痫性发作后7d,在缺氧性痫性发作组海马CA2、CA3和DG区明显地降低(P ＜0.05);GAD阳性细胞数在痫性发作后7～ 14d,缺氧性痫性发作组海马CA1、CA3和DG区明显地减少(P＜0.05).结论 缺氧性痫性发作后14d内并没有造成大鼠海马区及时或迟发性细胞丢失,但c-fos表达在大鼠海马区有迟发性增高;缺氧性痫性发作后海马GABA神经元数量的减少可能是新生大鼠缺氧诱导痫性发作后癫痫易感性升高的原因之一.     

大鼠癫痫持续状态后海马组织各区脑红蛋白表达的实验研究

目的观察大鼠癫痫持续状态(Status epilepticus,SE)后海马组织脑红蛋白(Neuroglobin,NGB)表达动态变化,探讨NGB在癫痫发作中的作用。方法健康成年雄性SpragueDawley大鼠40只,随机分为对照组(n=5)、癫痫模型实验组(n=35);实验组再依据观察时间分为:0、1、3、12、24 h和10、30 d。应用锂-匹罗卡品(20~127 mg/kg)建立大鼠SE模型,观察大鼠致痫期间行为学变化;采用尼氏(Nissl)染色检测海马组织神经元损伤情况;SABC免疫组化法检测海马组织NGB表达水平。结果 SE后,海马组织各区均出现不同程度神经元细胞损伤坏死,随着发作时程进展,CA1、CA3区存活神经元呈近直线下降趋势。其中CA1区(12、24 h,10、30d)、CA3区(0、12、24 h,10、30 d)和(DG区12、24 h,10、30 d)神经元存活数较对照组明显减少(P0.05)。大鼠SE后,海马各区NGB表达水平均上调,CA1、DG区NGB表达均于SE后24 h达顶峰后轻度下降,但仍持续高于对照组,CA3区NGB表达呈持续升高趋势。其中CA1区(24 h,10、30 d)、CA3区(24 h,10、30 d)和DG区(12、24 h,10、30 d)NGB表达水平均较对照组显著升高(P0.05)。另外,海马CA1和CA3区神经元存活数与NGB表达水平呈正相关(R=0.206,P=0.015;R=0.306,P=0.011)。结论大鼠SE后海马各区NGB表达上调,且与CA1、CA3区神经元存活数呈正相关,提示NGB表达上调可能是癫痫发作所致缺血缺氧损害的一种代偿保护机制,参与癫痫相关神经元损害的保护。     

雌激素和姜黄素对海人酸杏仁核点燃大鼠行为学、脑电图及海马神经元损伤的影响

目的了解雌激素和姜黄素对海人酸(kainic acid,KA)杏仁核点燃大鼠癫痫发作的影响。方法给去势的雌性大鼠添加雌激素治疗,添加姜黄素治疗,或添加雌激素和姜黄素治疗,比较各组大鼠致痫后癫痫发作的行为学、脑电图和海马神经元损伤的变化。结果给雌激素治疗的大鼠重型发作(Racine 4/5级)评分最高,而雌激素加姜黄素治疗组评分最低(P<0.05)。脑电图的变化与行为学的改变基本一致。致痫后大鼠注射KA侧海马CA3区、CA4区可见到明显的细胞损伤,而该侧海马CA1区、齿状回区(DG)及对侧海马CA3区、CA1区及DG区神经元损害不明显。雌激素组大鼠双侧海马CA3区均出现加重的神经元损害,姜黄素组及雌激素加姜黄素组大鼠海马注射对侧CA3区存活神经元较雌激素组明显增加(P<0.01)。结论高水平的雌激素可以加重癫痫的发作,给姜黄素治疗可以减轻大鼠海马CA3区神经元损害。     

红藻氨酸致痫大鼠海马神经元凋亡及其与caspase-3关系的研究

目的　研究大鼠癫痫发作后海马神经元凋亡及其与天冬氨酸特异性半胱氨酸蛋白酶 -3 (cysteinylasparate-specific proteinase,caspase-3 )表达的关系。方法　采用红藻氨酸 (kainic acid,KA)诱导大鼠癫痫模型 ,以原位末端标记 (TUNEL)及透射电镜检测癫痫发作后 6h及 1、3、7d海马神经元凋亡 ;半定量 RT-PCR及免疫组化法检测 caspase-3 m RNA及 caspase-3阳性表达。结果　KA致痫后 1 d,海马 CA1、CA3及 CA4区开始出现凋亡细胞 ,3 d时明显增多 ,7d时最多。 3个时间组相应区域间凋亡神经元数比较差异均有显著性 (P<0 .0 0 1 )。透射电镜观察可见典型的凋亡细胞形态学改变。 RT-PCR结果显示 ,KA致痫后 6h,海马组织 caspase-3 m RNA表达较对照组显著增高 (P <0 .0 5 ) ,1、3、7d caspase-3 m RNA仍持续高水平表达 (P <0 .0 5 )。免疫组化结果显示 ,KA致痫后 1 d,海马 CA1、CA3、CA4区开始出现 caspase-3阳性表达 ,3 d时阳性表达进一步增强 ,7d时表达最强。结论　凋亡参与 KA致痫大鼠癫痫发作后海马神经元迟发性死亡过程 ,caspase-3可能在癫痫后神经元凋亡过程中具重要的作用。     

低剂量伽玛刀照射对癫痫大鼠皮层及海马NMDA受体亚基表达的影响

目的观察低剂量伽玛刀照射对癫痫大鼠皮层和海马N-甲基-D-天氡氨酸(N-methyl-Daspartate,NMDA)受体亚基表达的影响。方法根据动物是否致痫及接受伽玛刀照射,将大鼠分为4组:对照组、伽玛刀组、药物致痫组、伽玛刀+药物组。腹腔连续注射戊四氮(pentylenetetrazole,PTZ)制备癫痫大鼠模型,以双侧额叶为照射靶区对大鼠进行低剂量伽玛刀照射,边缘剂量为15Gy。观察并记录各组大鼠伽玛刀照射前、后癫痫发作情况,并于伽玛刀照射后12周后留取脑组织,分别利用免疫组化及免疫蛋白印迹法对大鼠皮层及海马NMDA受体亚基NR1、NR2A和NR2B进行检测。结果对照组及伽玛刀组大鼠无痫性发作表现,与药物致痫组大鼠相比,伽玛刀+药物组大鼠经低剂量伽玛刀照射后12周,痫性发作明显减轻(P0.05)。与对照组相比,药物致痫组大鼠额叶皮层及海马CA1、CA3区NR1、NR2A和NR2B表达均明显增强(P0.05),阳性神经元数目及平均吸光度值均明显增加(P0.05);与药物致痫组比较,伽玛刀+药物组额叶皮层及海马CA1、CA3区NR1、NR2A和NR2B表达均明显降低(P0.05),阳性神经元数目及平均吸光度值明显减少(P0.05);伽玛刀组与对照组无明显差别(P0.05)。结论癫痫大鼠额叶皮层及海马NR1、NR2A及NR2B亚单位蛋白表达增强,低剂量伽玛刀照射可能引起癫痫大鼠皮层及海马NMDA受体亚基表达减少,这可能是低剂量伽玛刀抑制癫痫发作的分子机制之一。     

红藻氨酸致颞叶癫痫大鼠海马内Semaphorin3C、Semaphorin3F基因表达的研究

目的研究神经轴索导向分子Sem aphorin3C(Sem a3C),Sem aphorin3F(Sem a3F)mRNA对颞叶癫痫(TLE)大鼠海马神经轴索环路重建的调控作用。方法采用侧脑室内注射红藻氨酸(KA)制作TLE大鼠模型,用N issl染色及原位杂交的方法,分别检测致痫后1d、1w、2w、3w、4w大鼠海马的齿状回(DG),CA1区、CA3区神经细胞丢失程度以及Sem a3C、Sem a3F mRNA的表达。结果KA致痫后1d始出现神经元丢失,至4w神经元丢失明显增多。KA致痫后1w,Sem a3C、Sem a3F mRNA在海马的CA1区、Sem a3F mRNA在海马的CA3区表达明显下降,持续至3w(P<0.01),4w时恢复至正常(P>0.05);Sem a3C、Sem a3F mRNA在DG的表达,Sem a3C在CA3区的表达,实验组与对照组均无明显差别(P>0.05)。结论KA致痫后海马CA1区神经元下调Sem a3C、Sem a3F mRNA的表达,CA3区神经元下调Sem a3F mRNA的表达,可能促进TLE大鼠海马神经轴索环路重建。     

钙结合蛋白与颞叶癫痫大鼠海马神经元易损性的关系研究

目的 探讨微白蛋白(Parvalbumin,PV)、钙视网膜蛋白(Calretinin,CR)、钙结合蛋白-D28K(Calbindin-D28k,CB)与颞叶癫痫大鼠海马神经元易损性的关系及在颞叶癫痫的发生发展中的作用.方法 采用氯化锂-匹罗卡品颞叶癫痫大鼠模型,应用免疫组化动态观察海马神经元内PV、CR、CB的表达.结果 实验组大鼠PV阳性中间神经元的数量在CA3区无明显变化(P>0.05);在CA1区,呈进行性下降,到7d时达最低值(P<0.01);在齿状回及门区,先进行性下降,15d时达最低值(P<0.01),30d后开始上升,到60d几乎达正常水平(P>0.05).大鼠各区CR阳性中间神经元的数量较对照组均明显下降;在CA3区,急性期和慢性期下降最明显(P<0.01);在CA1区24h组的数量最少(P<0.01);在齿状回的门区15d组的数量最低(P<0.01).CB阳性中间神经元的数量在CA3区与对照组无明显变化(P>0.05);在CA1区,6h后开始减少(P<0.05),7d后达最低值(P<0.01),然后稍有回升,但仍明显低于对照组(P<0.05);门区CB阳性中间神经元的数量较对照组增加(P<0.05).结论 氯化锂-匹罗卡品颞叶癫痫动物模型中海马内含PV、CR、CB的GABA能中间神经元存在选择性易损性,CA1区含PV、CR、CB的GABA能中间神经元的急剧减少可能诱发了颞叶癫痫的急性发作;含PV、CB的GABA能中间神经元在齿状回的改变可能在颞叶癫痫自发性复发性发作的发生和发展中扮演了非常重要的角色.     

氯化锂-匹罗卡品致痫大鼠海马结构中CLC-2、CLC-3氯通道表达变化的研究

目的 研究ClC-2、ClC-3氯通道在氯化锂-匹罗卡品大鼠慢性癫痫模型中分布和表达的变化,探讨其在癫痫发作病理机制中的作用.方法 Wistar大鼠采用随机数字表法分成致痫组(60只)与对照组(20只),其中致痫组根据处死及处理时间又分为24h组、14d组与30d组,每组20只.致痫组复制氯化锂-匹罗卡品大鼠慢性癫痫模型,在发作后24h、14d、30d时,分别予以:(1)免疫组化染色,观察ClC-2、ClC-3氯通道蛋白在海马表达的分布情况及其致病后不同时点的吸光度(A)值的变化;(2)RT-PCR,观察ClC-2、ClC-3氯通道mRNA在致痫后不同时点的变化.结果 (1)与对照组比较,致痫后14d至30d,致痫组免疫反应阳性神经元数和A值明显减少和降低,差异有统计学意义(P<0.05);ClC-2 mRNA表达降低,差异有统计学意义(P<0.05).(2)与对照组比较,致痫组致痫后24 h,海马CA1、CA3及齿状回各层ClC-3免疫反应阳性神经元数和A值明显增加和升高,差异有统计学意义(P<0.05);ClC-3氯通道mRNA表达明显增加,差异有统计学意义(P<0.05).结论 癫痫慢性期的发作和ClC-2氯通道的减少有关.     

吡咯烷二硫代氨基甲酸对匹鲁卡品诱导癫痫大鼠海马神经元及小胶质细胞的影响

目的观察吡咯烷二硫代氨基甲酸(PDTC)对匹鲁卡品诱导癫痫大鼠海马CA1、CA3和DG各区神经元及小胶质细胞的影响。方法 30只大鼠随机分为正常对照组(n=6)、癫痫组(n=12)及PDTC干预组(n=12)。采用一次性腹腔注射匹鲁卡品(320 mg/kg)诱导大鼠癫痫发作;PDTC干预组分别于注射匹鲁卡品前24 h、20 min腹腔注射PDTC(100 mg/kg);正常对照组注射等量生理盐水。监测大鼠行为学改变;采用Fluoro-Jade C(FJC)染色法检测海马DG、CA1、CA3区退行性改变神经元;免疫组织化学方法检测各组大鼠海马各区小胶质细胞表达情况。结果癫痫组及PDTC干预组各有10只大鼠制模成功。正常对照组大鼠无癫痫发作;癫痫组大鼠平均跌倒次数[(32.30±4.37)次]显著多于PDTC干预组[(17.50±2.37)次](P0.05)。正常对照组大鼠海马各区未见FJC阳性细胞及少量Iba-1标记的小胶质细胞。与癫痫组比较,正常对照组DG、CA1、CA3锥体层Iba-1标记的小胶质细胞数显著减少(P0.01);PDTC干预组CA1、CA3区FJC阳性细胞数及锥体层Iba-1标记的小胶质细胞数明显减少(P0.05~0.01)。结论 PDTC可能通过抑制小胶质细胞活性,影响小胶质细胞介导的神经炎症反应,从而对癫痫持续状态所致脑损伤起到保护作用,改善癫痫发作的严重程度。     

海人酸致痫大鼠海马神经元凋亡研究

目的　研究大鼠癫痫发作后海马神经元凋亡的时空分布。方法　采用海人酸 (KA)诱导大鼠癫痫模型 ,以原位末端标记 (TUNEL)及透射电镜检测癫痫发作后 6h、1d、3d、7d海马神经元凋亡。结果　对照组及KA致痫后 6h组 ,海马区均未发现凋亡细胞。KA致痫后 1d ,海马CA1、CA3及CA4区开始出现凋亡细胞 ,3d时明显增多 ,7d时最多。KA致痫后 1d、3d、7d ,海马CA1锥体层线性长度1mm的TUNEL阳性细胞数分别为 (6 .6 0± 3.6 9)个、(13.5 7± 5 .17)个和 (2 5 .96± 4 .87)个 ;CA3区分别为 (6 .4 8± 2 .4 5 )个、(13.89± 2 .5 2 )个和 (2 8.80± 5 .39)个 ;CA4区分别为 (4 .6 0± 1.4 5 )个、(12 .2 0± 2 .0 4 )个和 (2 5 .2 0± 5 .83)个。 3个时间组相应区域凋亡神经元数比较均存在显著性差异(P <0 .0 0 1)。透射电镜观察可见典型的凋亡细胞形态学改变。结论　凋亡参与KA致痫大鼠癫痫发作后海马神经元迟发性死亡过程。     

Denervation study of synapses of organ of corti of old world monkeys

Fine structural characteristics of synapses in the spiral organ of Corti were examined, with reference to differences between inner and outer haircell systems, and to location of neurons of origin of efferent axons. Surgical interruption of crossed olivocochlear bundle, of vestibular nerve, of facial nerve, and excision of superior cervical ganglia were used to determine the pathways of efferent axons. Interruption of the vestibular nerve near the brainstem results in degeneration of all efferent terminals on outer hair cells. Mid-line lesions at, and caudal to, the facial colliculus result in degeneration of about half of these efferent terminals. Efferent synaptic bulbs to the inner hair-cell system are small, of the order of one micron, and form type 2 junctions with afferent dendrites. They tend to have more large dense-core vesicles (about 80 nm) than the large efferent terminals of the outer hair-cell system, and appear to be the terminals of axons in the habenula perforata, which exhibit varicosities laden with large dense core vesicles. The varicosities are unaffected by excision of the superior cervical ganglia. So far as our material can reveal, it appears that the varicosities in the habenula perforata do not survive vestibular root interruption, nor do the efferent processes in the internal spiral bundle or at the base of inner hair cells. Most interestingly, the afferent processes of the inner hair-cell system, as identified for example by their relation to pre-synaptic bodies in the inner hair cells, are subject to a trans-synaptic reaction after severance of the vestibular root. They undergo a dramatic cytological transformation, characterized by increase of volume, engorgement with microtubules, microfilaments, microvesicles of various sizes, and clusters of lysosomes. Thus, both the efferent and afferent terminals of the inner hair-cell system show marked cytological differences from the corresponding terminals of the outer hair cell system.     

Mechanism of action of tubocurarine on nicotinic cholinoreceptors of neurons of the sympathetic ganglia of rats

Tubocurarine (Tc) effect on membrane currents elicited by acetylcholine (ACh) was studied in isolated superior cervical ganglion neurons of rat using patch-clamp method in the whole-cell recording mode. The "use-dependent" block of ACh current by Tc was revealed in the experiments with ACh applications, indicating that Tc blocked the channels opened by ACh. Mean lifetime of Tc-open channel complex, tau, was found to be 9.8 +/- 0.5 s (n = 7) at -50 mV and 20-24 degrees C. tau exponentially increased with membrane hyperpolarization (e-fold change in tau corresponded to the membrane potential shift by 61 mV). Inhibition of the ACh-induced current by Tc (3-30 microM/1) was completely abolished by membrane depolarization to the level of 80-100 mV. Inhibition of ACh-induced current was augmented at increased ACh doses. It is concluded that the open channel block produced by Tc is likely to be the only mechanism for Tc action on nicotinic acetylcholine receptors in superior cervical ganglion neurons of rat.     

The effect of age of onset of PD on risk of dementia

Background Dementia occurs in the majority of patients with Parkinson’s disease (PD). Late onset of PD has been reported to be associated with a higher risk for dementia. However, age at onset (AAO) and age at baseline assessment are often correlated. The aim of this study was to explore whether AAO of PD symptoms is a risk factor for dementia independent of the general effect of age. Methods Two community-based studies of PD in New York (n = 281) and Rogaland county, Norway (n = 227) and two population-based groups of healthy elderly from New York (n = 180) and Odense, Denmark (n = 2414) were followed prospectively for 3–4 years and assessed for dementia according to DSM-IIIR. All PD and control cases underwent neurological examination and were followed with neurological and neuropsychological assessments. We used Cox proportional hazards regression based on three different time scales to explore the effect of AAO of PD on risk of dementia, adjusting for age at baseline and other demographic and clinical variables. Findings In both PD groups and in the pooled analyses, there was a significant effect of age at baseline assessment on the time to develop dementia, but there was no effect of AAO independent of age itself. Consistent with these results, there was no increased relative effect of age on the time to develop dementia in PD cases compared with controls. Interpretation This study shows that it is the general effect of age, rather than AAO that is associated with incident dementia in subjects with PD. Received in revised form: 22 December 2005     

Limits of deinstitutionalization--perspectives of relatives of psychiatric patients

After a hopeful beginning, the social process of the reintegration of those with severe mental illness has come to a standstill. I am led to wonder whether "the community" really wants to live together with people suffering from severe mental illness, and if so, how closely? As long as the medical treatment of mental illness provided by the general practitioners is fundamentally deficient, as they are not able to prescribe the necessary interventions--such as out-patient psychiatric nursing, and service providers in the out-patient sector are content with offering increasingly intensive forms of care for the less seriously ill at the cost of the Social Welfare System--the reintegration of those with serious mental illness remains an illusion--which is mainly to the benefit of providers of residential care in homes and hostels.