Nbn基因敲除小鼠小脑形态发育的变化

目的:通过观察神经特异性Nbn基因敲除(Nbn-CNS-del)和对照(Nbn-CNS-Ctr)小鼠小脑发育的形态学变化,探讨DNA损伤修复基因Nbn在小鼠小脑发育过程中的作用.方法:应用光镜连续切片和体视学方法,对出生后(P)7、 14、 21d的实验组和对照组小鼠小脑进行系统地形态学观察和定量分析.结果:Nbn-CNS-del小鼠小脑发育比Nbn-CNS-Ctr明显滞后,至P21小脑截面积约减少4倍、小脑小叶截面积约减少6倍;外颗粒细胞层截面积、体密度和小脑小叶的体密度差异有统计学意义.结论:DNA损伤修复基因Nbn是小鼠小脑发育的一个关键基因,明显影响小鼠小脑的发育与成熟.     

小鼠小脑皮质发育中的神经元凋亡

目的探讨小鼠小脑皮质发育中神经元凋亡的规律和机制。方法用激活型Caspase-3多抗免疫组织化学标记及Hoechst 33258染色液染色,检测从出生至成年小鼠小脑皮质中神经元的凋亡,用Western blotting方法对小脑组织中Caspase-3和Caspase-8的活化片段进行半定量测定。结果外颗粒层、普肯耶细胞层和内颗粒层凋亡细胞密度最高分别在出生后第8d(P8)、P5及P9,P20各层凋亡细胞密度都很低。Caspase-3活化片段的表达量在P5较高,P5以后逐渐降低,至P14消失;Caspase-8活化片段的表达量从P0到P10都较高,P10以后逐渐降低,至P30消失。结论P0至P14是小脑皮质神经元凋亡的重要时期,通过启动Caspase-8的活化进而激活效应性Caspase-3的细胞凋亡途径存在于小脑皮质的塑型成熟过程。     

人胚胎视皮质神经元发育的体现学分析

实验取自１４例产后死亡的胎儿,胎龄１２～４０周,对其视皮质神经元进行电镜观察及立体计量分析。结果表明：（１）视皮质神经元在发育早期（１２周胎龄）时,可依细胞大小、核及胞质中的细胞器鉴别神经元和神经胶质细胞,（２）视皮质神经元发育过程中,核经例逐渐减少,细胞核体密度逐渐减少,核仁与胞质的体密度呈上升趋热（３）线粒体、粗面内质网、核糖体的体密度呈明显上升趋势,滑面内质网、高尔基复合体及溶酶体的体密度也     

小鼠放射状胶质细胞和小脑皮质的发育

目的探讨小鼠小脑皮质发育过程中放射状胶质细胞的分化。方法应用免疫荧光及5-溴脱氧尿嘧啶核苷(BrdU)检测技术,标记小鼠胚胎8d至生后180d小脑(57例,分为19组,每组3只)的神经干细胞、放射状胶质细胞、普肯耶细胞及颗粒细胞。结果放射状胶质细胞于胚胎13d的神经上皮出现,尔后该细胞分化为各种神经元和贝格曼胶质细胞,并在小脑皮质层状结构的形成中起着重要作用。结论放射状胶质细胞来源于神经上皮细胞,是神经细胞和神经胶质细胞的前体细胞。在小脑皮质的发育过程中,放射状胶质细胞能分化为普肯耶细胞和颗粒细胞,并为神经细胞的迁移提供路径和支架。     

小鼠小脑皮质的组织发生

目的:探讨小鼠小脑皮质的组织发生过程。方法:应用光镜和电镜技术对胚胎和生后小脑皮质进行形态学观察,对各层厚度和细胞密度进行测量。结果:胚胎12 d(E12)小脑原基有室管膜层、套层和边缘层构成,约出生当日(P0)出现外颗粒层、分子层、Purkinje细胞层和内颗粒层。外颗粒层P6/7达最厚,至P20消失。P0至P30内颗粒层细胞逐步分化发育成熟,Purkinje细胞树突树逐渐形成,约P7时Purkinje细胞排列成单层。结论:E12至P0片层化小脑主要经历了细胞增殖、分化与迁移;P0至P30片层化结构逐渐发育成熟,外颗粒层消亡以细胞迁移和凋亡为主,其他各层细胞主要经历了分化发育与凋亡。     

Nbn基因神经特异性敲除生后小鼠海马发育的形态学变化

目的:观察Nbn基因神经特异性敲除 (Nbn-CNS-del) 小鼠生后海马的形态学变化,探讨Nbn基因在小鼠海马发育中的作用.方法:应用H-E染色、免疫组织化学技术结合体视学方法对生后(P) 7、 14、 21d的Nbn-CNS-del小鼠海马进行系统观察和定量分析,以非基因敲除 (Nbn-CNS-ctr) 仔鼠为对照组.结果:P7～21 d,与对照组相比,Nbn-CNS-del小鼠海马发育迟缓;CA区分子层、锥体细胞层及多形层的截面积均减小;NeuN及神经胶质纤维酸性蛋白(GFAP) 阳性细胞面数密度也均明显减少.结论:Nbn-CNS-del小鼠海马CA区发育迟缓,Nbn基因可能是海马发育中的重要基因.     

噪声对小鼠神经元形态参数的影响

我们用昆明种雌性小白鼠４９只，以自配标准饲料喂养至老，其中３３只为对照组，１６只为实验组。实验组从幼年时期开始，每天施加２小时嗓声。在性成熟期、老年期分别将小鼠处死，用体视学方法测出两类小鼠额叶神经元的九种形态参数，结果发现，实验组与对照组比较，在性成熟期细胞体密度（Ｖｖ１）、细胞核体密度（Ｖｖ２）、核仁体密度（Ｖｖ３）、细胞核比表面（Ｓｖ２）有显著性差异。本文简要讨论了这些参数的意义。     

CTNND2基因敲除对小鼠小脑发育及运动功能的影响

目的 探讨CTNND2基因敲除对小鼠小脑神经元发育和运动功能的影响及可能的机制.方法 小鼠分为2组,每组10只,均为7周龄:野生型(WT)C57BL/6J小鼠为对照组,CTNND2基因敲除纯合子(CTNND2-/-)小鼠为实验组,PCR检测CTNND2-/-小鼠的基因型.平衡木实验、悬挂实验和步态分析实验检测两组运动功...     

早期应激对小鼠纹状体神经元发育的影响

目的探讨早期应激对背侧纹状体多棘神经元发育的影响。方法通过改变新生小鼠(出生后第2~9天)的生长环境建立早期应激动物模型,采用原位杂交、Golgi染色和体视学分析方法,定量分析应激小鼠背内侧纹状体和背外侧纹状体内神经元胞体、树突和树突棘的改变。结果 9d龄C57BL/6J小鼠的纹状体神经元含丰富的树突分支和树突棘。持续7d的应激主要影响背外侧纹状体,表现为纹状体神经元的近胞体树突分支增多(应激组9.50±0.38,n=8;对照组6.50±0.23,n=6;P0.05),树突分支上含大量丝状伪足(每20μm树突节段:应激组8.15±0.51,n=8;对照组3.85±0.33,n=6;P0.05),但树突棘数量减少(每20μm树突节段:应激组12.05±0.91,n=8;对照组20.02±0.73,n=6;P0.05)。结论早期应激主要干扰背外侧纹状体神经元树突的发育,导致树突棘的成熟延缓。     

C57/BL6小鼠小脑发生发育及衰老的形态学变化

目的:观察C57/BL6小鼠小脑发生发育及衰老的形态学变化规律.方法:应用石蜡连续切片、H-E染色结合体视学方法对胚龄(E)10、12、14、16、18、20 d和生后(P)1、3、7、14、21、28 d的仔鼠及生后2、3、6、15个月(M)的C57/BL6小鼠小脑的形态学变化进行系统观察和定量分析.结果:E12 d小脑原基出现.E16 d小脑半球、蚓部及皮质的外颗粒层出现.E18 d分子层的原基出现,小脑表面出现沟回结构.P1 d皮质分为外颗粒层、分子层、浦肯野细胞层和内颗粒层4层.P7 d皮、髓质分界清楚.P21d外颗粒层消失,皮质分为分子层、浦肯野细胞层和颗粒层3层.体视学分析显示,3个月之前,小脑总体积、皮质和髓质的体积、皮质各层（分子层、浦肯野细胞层和内颗粒细胞层/颗粒细胞层）的体积均逐渐增加,以P3～14 d增长最快;3个月之后趋稳定.E18 d～P14 d,外颗粒层体积先增大后减小,P21 d完全消失.结论:E12 d～P21 d是小鼠小脑发生发育的关键时期,细胞经历了增殖、分化和迁移,外颗粒层可能参与分子层和内颗粒层的形成.     

生后不同发育阶段的小鼠下丘脑大细胞神经元的超微结构

搞目的：研究小鼠下丘脑大细胞神经元在生后各发育阶段超微结构的变化规律。方法：以昆明小鼠为实验动物，取生后不同阶段的小鼠下丘脑，按常规的超微结构制作方法制作电镜标本，1200EX透射电镜观察。结果：生后1d粗面内质内网(PER)短，散在分布，凋亡细胞多；生后5d，凋亡细胞明显减少，有髓神经纤维开始形成；生后20d，各种细胞器均已出现，RER变长，平行排列，多聚核糖体布于其间，Golgi体明显；生后30d，细胞器发达，分泌颗粒明显；生后60d，Golgi体发达，分泌颗粒增多增大。结论：生后30d，小鼠下丘脑大细胞神经元已发育成熟。     

No change in neuron numbers in the dentate nucleus of patients with schizophrenia estimated with a new stereological method--the smooth fractionator

The dentate nucleus is phylogenetically the most recent nucleus in the cerebellum. Owing to its connections to the thalamus and the prefrontal cortex it may be involved in the symptomathology in schizophrenia and other psychiatric illnesses. In this stereological study we implemented the smooth fractionator, which combines the unbiased principles of the optical fractionator with a new and more efficient sampling strategy to the dentate nucleus. The smooth fractionator represents the most efficient sampling strategy described so far in stereology, in terms of reducing the sampling variance and thus increasing the efficiency. It is the first application of the smooth fractionator to human brain tissue and presents estimations of total number of neurons in the dentate nuclei of eight patients with schizophrenia compared to eight control persons. The total number of neurons in the dentate nucleus was estimated to 3.36 x 10(6) in subjects with schizophrenia, which was not statistically significant different from 3.65 x 10(6) in control subjects (P = 0.63). The advantages and disadvantages of the smooth fractionator method are discussed and its precision in practical application is estimated.     

Cultivation of mouse cerebellar cells in serum free,hormonally defined media: Survival of neurons

A hormonally defined medium is described which facilitates the survival of small neurons in primary cultures of mouse cerebellum. The defined medium consists of bovine serum albumin, insulin, transferrin, selenium, thyroxine, and the protease inhibitor aprotinin. About 95% of all cells were identified as neurons using tetanus toxin as a marker in an immunocytochemical assay. They survive for more than 4 weeks, showing a tendency to grow in cell clusters with a dense network of processes. The remaining cells (approximately 5%) were identified as astrocytes by their expression of vimentin and GFA protein and the lack of expression of fibronectin.     

Monoclonal antibody against SSEA-1 is specific for a subpopulation of astrocytes in mouse cerebellum

Monoclonal antibody to SSEA-1 reacts specifically in the mouse cerebellum with the cell surface of a subclass of estrocytes, but not of neurons, oligodendrocytes, fibroblasts and fibroblast-like cells, or macrophages. The antigen is weakly detectable on cultured cells by indirect immunofluorescence at embryonic day 13 after one day of maintenance in vitro, and strongly detectable in glial fibriallary acidic protein- and vimentin-positive astrocytes throughout later embryonic and early postnatal ages. In histological sections SSEA-1 is found at embryonic day 14 in the ventricular zones of the cerebellar anlage and at early postnatal ages in prospective while matter tracts and external granular and molecular layers. It continues to be detectable in these regions until the fourth postnatal week, when its expression becomes restricted to weakly positive, dot-like structures in the molecular layer.     

小鼠肾小体的生长曲线

目的:探讨小鼠肾脏发生发育过程中肾小体体积的增长规律。方法：光镜下应用体视学方法对生前和生后小鼠肾脏中各发育阶段的肾小体体积进行测量。结果：胚龄14d时，逗号小体和S小体出现，生后7d消失。胚龄16d时，Ⅲ、Ⅳ期肾小体出现，其体积从胚龄16d到生后40d大约增大70倍。结论：小鼠肾小体于胚龄14d发生，从生后7d到生后40d体积增长迅速。     

Design-based stereological study of the guinea-pig (Cavia porcellus) cerebellum

Guinea pigs have proved useful as experimental animal models in studying cerebellar anatomical and structural alterations in human neurological disease; however, they are also currently acquiring increasing veterinary interest as companion animals. The morphometric features of the normal cerebellum in guinea pigs have not been previously investigated using stereology. The objective of the present work was to establish normal volumetric and quantitative stereological parameters for cerebellar tissues in guinea pigs, by means of unbiased design-based stereology. Cerebellar total volume, gray and white matter volume fractions, molecular and granular layers volume fractions, cerebellar surface area, Purkinje cellular and nuclear volumes, and the Purkinje cell total count were stereologically estimated. For this purpose, cerebellar hemispheres from six adult male guinea pigs were employed. Isotropic, uniform random sections were obtained by applying the orientator method, and subsequently processed for light microscopy. The cerebellar total volume, the white and grey matter volume fractions, and the molecular and granular layer volumes were estimated using the Cavalieri's principle and the point counting system. The cerebellar surface area was estimated through the use of test lines; Purkinje cellular and nuclear volumes were analysed using the nucleator technique, whereas the Purkinje cell total count was obtained by means of the optical disector technique. The mean ± standard deviation total volume of a guinea-pig cerebellar hemisphere was 0.11 ± 0.01 cm³. The mean volumetric proportions occupied by the gray and white matters were, respectively, 78.0 ± 2.6% and 22.0 ± 2.6%, whereas their mean absolute volumes were found to be 0.21 ± 0.02 cm³ and 0.059 ± 0.006 cm³. The volumes of the molecular and granular layers were estimated at 112.4 ± 20.6 mm³ and 104.4 ± 7.3 mm³, whereas their mean thicknesses were calculated to be 0.184 ± 0.020 mm and 0.17 ± 0.02 mm. The molecular and granular layers accounted for 40.7 ± 3.9% and 37.4 ± 1.8% of total cerebellar volume respectively. The surface area of the cerebellum measured 611.4 ± 96.8 mm². Purkinje cells with a cellular volume of 3210.1 µm³ and with a nuclear volume of 470.9 µm³ had a higher incidence of occurrence. The mean total number of Purkinje cells for a cerebellar hemisphere was calculated to be 253,090 ± 34,754. The morphometric data emerging from the present study provide a set of reference data which might prove valuable as basic anatomical contribution for practical applications in veterinary neurology.     

小鼠小脑片层化过程中神经元迁移及reelin的可能调节作用

目的:观察小鼠小脑皮质片层化过程细胞迁移,探讨片层化、细胞迁移和reelin蛋白之间的关系。方法:不同年龄的小鼠172只,应用免疫荧光、Nissl和HE染色法对小脑形态结构及片层化、细胞迁移情况进行观察。结果:(1)小脑片层化:E15-E16的后脑主要由神经上皮构成,P0时小脑皮质基本形成了外颗粒层、分子层、浦肯野细胞层和颗粒层,P5时浦肯野细胞层形成2~3层细胞,分子层仍有大量细胞。P20时基本形成了小脑的典型三层结构。(2)细胞迁移过程:浦肯野细胞大约在E18时开始迁移,至P7时基本完成迁移,胞体形成均一的1~2层细胞结构。同时,小年龄时NeuN阳性细胞主要见于内颗粒层和外颗粒层,P7后阳性细胞只定位在内颗粒层,迁移基本停止。(3)野生鼠(WT)和reeler小鼠比较:约P14时,WT小鼠小脑形成分子层、浦肯野细胞层及颗粒层。但是在reeler小鼠,小脑分叶不良,浦肯野细胞未迁移至目的地,排列紊乱;内颗粒层(GL)细胞分布松散、不均一。结论:在小鼠小脑的片层化形成过程中,Reelin蛋白可能参与神经细胞增殖和迁移的调控过程。     

The midbrain dopaminergic system: anatomy and genetic variation in dopamine neuron number of inbred mouse strains

The mesotelencephalic dopamine system is genetically variable and affects motor behavior, motivation, and learning. Here we examine the genetic variation of mesencephalic DA neuron number in a quasi-congenic RQI mouse strain and its background partner and in a recombinant inbred strain with different levels of mesencephalic tyrosine hydroxylase activity (TH/MES). We used B6.Cb4i5-6/Vad, C57BL/6By, and CXBI, which are known to express high, intermediate, and low levels of TH/MES, respectively. Unbiased stereological sampling with optical disector counting methods were employed to estimate the number of TH-positive neurons in the A8-A9-A10 cell groups. Morphometric studies on the mesencephalic dopamine cell groups indicated that male mice of the B6.Cb4i5-6/Vad strain were endowed with a significantly lower number of TH-positive cells than CXBI mice. In all strains studied, the right retrorubral field (A8 area) had a higher number of dopamine neurons compared to the left A8 area. The results suggest an inverse relationship between TH/MES and number of dopamine neurons in the A9-A10 cell groups and significant lateral asymmetry in the A8 cell group. A detailed anatomical atlas of the mesencephalic A8-A9-A10 dopaminergic cell groups in the mouse is also presented to facilitate the assignment of TH-positive neurons to specific cell groups.