人体小脑神经元发育的定量观察

目的：研究人体小脑神经元的发育过程。方法：应用体视学方法,对18例不同时期人体小脑组织Golgi染色后进行观察,观测小脑皮质分层出现的时间,观测并计算神经元的数密度、体密度和表面积密度。结果：6月龄时,小脑皮质出现较明显的分子层、蒲肯野细胞层和颗粒层;星形细胞、篮状细胞、蒲肯野细胞、颗粒细胞和高尔基细胞的的数密度随月龄／年龄的增长而减少,体密度和表面积密度随月龄／年龄的增长而增加,但这些减小和增大是不等速的,6-8月龄变化最明显。结论：人体小脑神经元的发育呈现快慢交替、不均速发展,6～8月是小脑神经元发育的重要时期。     

猫小脑皮质GABA能神经元年龄相关性变化

为探讨青年猫和老年猫小脑皮质GABA能神经元及其表达的年龄相关性变化,利用Nissl染色显示小脑皮质结构及神经元,免疫组织化学ABC法标记GABA免疫阳性神经元。光镜下观察,采集图像,并利用图像分析软件对分子层、蒲肯野细胞层和颗粒层神经元及GABA免疫阳性神经元及其灰度值进行分析统计。结果显示,GABA免疫阳性神经元、阳性纤维及终末在青年猫和老年猫小脑皮质各层均有分布。与青年猫相比,老年猫分子层、蒲肯野细胞层神经元和GABA免疫阳性神经元密度及其GABA免疫阳性反应强度均显著下降(P<0.01),颗粒层神经元密度和GABA免疫阳性强度也显著下降(P<0.01),但其GABA免疫阳性神经元密度无显著变化(P>0.05);蒲肯野细胞的胞体萎缩,阳性树突分枝减少。因此认为,衰老过程中猫小脑皮质GABA能神经元的丢失和GABA表达的下降,可能是老年个体运动协调、精确调速和运动学习等能力下降的重要原因之一。     

多巴胺受体1在皖西白鹅小脑皮质发育中的表达

采用普通染色及免疫组化SABC染色法研究皖西白鹅小脑皮质的发育和多巴胺受体1(DRD1)阳性细胞在其发育中的表达.结果表明,小脑皮质在胚龄13 d(E13)由外向内分为外颗粒层(EGL)、浦肯野细胞层(PCL)和内颗粒层(IGL),E19由外向内分为EGL、分子层(ML)、PCL和IGL.随发育天数的增加,EGL的厚度和细胞层次呈先升后降的变化趋势,细胞密度逐渐下降;ML厚度逐渐增大,在E24到E28时增值最大;浦肯野细胞(PC)在E13、E19、E24和E28时随胚龄增大逐渐增大,在E28后趋于稳定,细胞密度随着发育天数的增加逐渐下降,在小脑皮质发育中还发现有一部分PC呈多层排列,且细胞层次逐渐变少;IGL厚度呈先升后降的变化趋势,细胞密度呈上升趋势.外颗粒层和内颗粒层在E13、E19、E24和E28时有DRD1阳性细胞表达,分子层在E24、E28、日龄7 d(P7)和15d(P15)有阳性细胞表达,PC在所检测的6个时段均有阳性表达.研究表明,小脑皮质的发育主要与细胞增殖、迁移和凋亡有关,外颗粒层的逐渐消失是以细胞迁移和凋亡为主,多层PC逐渐退化成单层是与细胞凋亡和正常突触联系的建立有关;DRD1在皖西白鹅小脑皮质发育中对外颗粒层细胞和PC起着重要作用.     

幼龄红嘴相思鸟小脑皮层组织学结构

为了探讨不同日龄幼龄红嘴相思鸟（Leiothrix lutea）小脑皮质的组织学结构变化,本文分别以1、5、9日龄幼龄红嘴相思鸟为研究对象,通过H.E.和甲苯胺蓝法进行染色,光镜下观察红嘴相思鸟小脑冠状切面的显微结构。结果显示,1日龄时小脑皮层由外颗粒层（EGL）、浦肯野细胞层（PCL）和内颗粒层（IGL）3层构成,外颗粒层较厚且清晰,而浦肯野细胞层和内颗粒层界限不清楚;5日龄和9日龄时小脑皮质均可见外颗粒层、分子层（ML）、浦肯野细胞层和内颗粒层4层结构。对不同日龄红嘴相思鸟小脑皮质各层厚度进行单因素方差分析显示,随日龄增长,小脑皮质、分子层和内颗粒层厚度极显著增厚（P < 0.01）,浦肯野细胞体积也极显著增加（P < 0.01）;外颗粒层厚度变化不明显（P < 0.01）,呈现先增厚后变薄的趋势,与皮质厚度的比例逐渐减小。研究表明,幼龄红嘴相思鸟在出生后,随日龄增长,小脑皮质层逐渐发育成熟。内颗粒层与外颗粒层的相对变化规律表明内颗粒层细胞是由外颗粒层细胞迁移而来。     

幼龄红嘴相思鸟小脑皮质组织学结构

为了探讨不同日龄幼龄红嘴相思鸟(Leiothrix lutea)小脑皮质的组织学结构变化,分别以1、5、9日龄红嘴相思鸟为研究对象,通过H.E和甲苯胺蓝法进行染色,光镜下观察红嘴相思鸟小脑冠状切面的显微结构。结果显示,1日龄时,小脑皮层由外颗粒层(EGL)、浦肯野细胞层(PCL)和内颗粒层(IGL)3层构成,外颗粒层较厚且清晰,而浦肯野细胞层和内颗粒层界限不清楚;5日龄和9日龄时,小脑皮质均可见外颗粒层、分子层(ML)、浦肯野细胞层和内颗粒层4层结构。对3个日龄红嘴相思鸟小脑皮质各层厚度进行单因素方差分析,随日龄增长,小脑皮质、分子层和内颗粒层厚度极显著增厚(P0.01),浦肯野细胞体积也极显著增加(P0.01);外颗粒层厚度变化不明显(P0.05),呈现先增厚后变薄的趋势,与皮质厚度的比例逐渐减小。研究表明,幼龄红嘴相思鸟在出生后,随日龄增长,小脑皮质层逐渐发育成熟。内颗粒层与外颗粒层的相对变化规律表明,内颗粒层细胞是由外颗粒层迁移而来的。     

NF200在猫小脑中的分布及其老年性变化

目的探讨NF200（神经丝蛋白-200）在猫小脑中的分布及其老年性变化,以及导致相关变化的原因与在小脑功能衰老中的意义。方法利用Nissl染色显示小脑内部结构和神经元,免疫组织化学ABC法标记NF200免疫阳性（NF200-IR）结构。光镜下观察,采集图像,并利用Image-Pro Express5.1图像分析软件对小脑中NF200免疫阳性神经元以及各层免疫阳性反应灰度值进行分析统计。结果NF200免疫阳性神经元主要分布于蒲肯野细胞层和分子层底部,阳性纤维及终末在青年猫和老年猫小脑各层均有分布。与青年猫相比,老年猫蒲肯野细胞层神经元密度及其各层NF200免疫阳性反应强度均显著下降（P〈0.01）;蒲肯野细胞的胞体萎缩,阳性树突分枝明显减少。结论衰老过程中猫小脑NF200含量的下降,导致细胞结构紊乱,轴浆运输能力下降,可能是老年个体小脑功能衰退的重要原因之一。     

人胎视皮质皮质下层NPY－IR神经元的发育

本文用免疫组化方法研究了１６周至足月人胎视皮质皮质下层ＮＰＹ－ＩＲ神经元的发育。各胎龄视皮质ＳＰ层内均有ＮＰＹ－ＩＲ神经元分布。从１６周至２６周,ＮＰＹ－ＩＲ神经元密度逐渐增高并于２６周达高峰;３２周以后阳性神经元密度随胎龄增长而下降。人胎视皮质ＳＰ层ＮＰＹ－ＩＲ神经元形态也随胎龄而变化;２０周以前,ＮＰＹ－ＩＲ神经元大多是胞体较小,突起短而少的未分化神经元、ＳＰ层内ＮＰＹ－ＩＲ纤维少。２０周以后,ＮＰＹ－ＩＲ神经元胞体增大,突起增多、变长;多极和双极、双簇神经元随胎龄增长而增多;ＳＰ层内的ＮＰＹ－ＩＲ纤维大量增加,部分纤维穿入皮质板。３２周以后,多极ＮＰＹ－ＩＲ神经元逐渐减少,双极双簇神经元所占比例相对增高。ＮＰＹ免疫组化结合ＮＡＤＰＨ－ｄ组化显示人胎视皮质ＳＰ层大多数ＮＰＹ－ＩＲ神经元同时呈ＮＯＳ阳性。本研究观察到人胎视皮质ＳＰ层内ＮＰＹ－ＩＲ神经元发育可分为发生、成熟和退化三个阶段。     

猫小脑皮质Glu/GABA表达的老年性变化

衰老导致小脑的生理功能下降,但其神经机制仍然不清楚。为此,利用免疫组织化学方法标记猫小脑皮质内谷氨酸(Glutamate,Glu)和γ-氨基丁酸(γ-Aminobutiric acid,GABA)免疫反应阳性(Glu-IR和GABA-IR)结构,探讨青年猫和老年猫小脑皮质Glu/GABA表达的老年性变化及其可能影响。并利用Image-Pro Express图像分析软件对小脑皮质各层Glu和GABA免疫反应阳性细胞密度及其灰度值进行测量。结果显示:与青年猫相比,老年猫小脑皮质内的Glu免疫反应阳性浦肯野细胞密度、颗粒层Glu免疫反应阳性细胞密度及其两者的免疫阳性反应灰度值均显著下降(P<0.01)(免疫反应强度与平均灰度值成反比);老年猫分子层、浦肯野细胞层GABA免疫反应阳性神经元密度及其免疫反应强度均显著下降(P<0.01);颗粒层GABA免疫反应阳性神经元密度无显著变化(P>0.05),但神经元免疫反应强度显著减弱(P<0.01)。研究结果提示,衰老过程中猫小脑皮质出现神经元Glu的表达增强、GABA的表达减少等,可能是小脑神经元丢失和精确调控能力下降等的重要原因之一。     

生长休止蛋白7在大鼠小脑皮质发育过程中的动态表达

目的研究生长休止蛋白7(Gas7)在大鼠小脑皮质不同发育时期的动态表达。方法采用逆转录聚合酶链反应(RT-PCR)方法检测Gas7mRNA在大鼠小脑皮质不同发育时期的表达;免疫组织化学方法观察Gas7蛋白在大鼠小脑皮质不同发育时期的表达和分布。结果 RT-PCR结果:Gas7mRNA在大鼠小脑皮质发育时期的表达呈现先增强后减弱的趋势,高峰出现在生后第21d(P21)。免疫组化实验结果:在胚胎第18.5d(E18.5)和E20.5仅Purkinje细胞层有Gas7免疫阳性产物分布;出生当天(P0)外颗粒层出现Gas7阳性神经纤维,Purkinje细胞层出现形态不规则的Gas7免疫阳性细胞;P7外颗粒层和Purkinje细胞层免疫反应增强,内颗粒层出现一些散在的Gas7强阳性细胞,胞体较小,突起清晰可见;P14小脑皮质4层均有Gas7阳性表达;P21小脑皮质3层Gas7免疫阳性反应较P14增强(P0.01);Adult(2月龄)较P21免疫反应减弱(P0.01)。结论 Gas7在大鼠小脑皮质发育过程中的动态表达呈现出时空特异性,提示Gas7基因在大鼠小脑皮质发育过程中可能起着重要的调控作用。     

短暂性缺血对小脑皮质影响的组织化学研究

为了探讨全脑短暂性缺血对小脑皮质蒲肯野细胞的影响 ,实验用组织化学方法对家兔全脑缺血 5分钟 (B组 )、 10分钟 (C组 )及缺血再灌 (D、 E组 )后蒲肯野细胞的酶组织化学变化进行了观察。结果显示 ,缺血 10分钟及缺血再灌后蒲肯野细胞的 SDH、Mg2 + - ATP活性及 PAS反应均降低 (P<0 .0 5 ) ,L DH增高 (P<0 .0 1)。结果提示家兔全脑缺血 10分钟和缺血再灌可损害小脑皮质蒲肯野细胞的能量代谢酶的活性     

Age-related changes of structures in cerebellar cortex of cat

We studied the structures of the cerebellar cortex of young adult and old cats for age-related changes, which were statistically analysed. Nissl staining was used to visualize the cortical neurons. The immunohistochemical method was used to display glial fibrillary acidic protein (GFAP)-immunoreactive (IR) astrocytes and neurofilament-immunoreactive (NF-IR) neurons. Under the microscope, the thickness of the cerebellar cortex was measured; and the density of neurons in all the layers as well as that of GFAP-IR cells in the granular layer was analysed. Compared with young adult cats, the thickness of the molecular layer and total cerebellar cortex was significantly decreased in old cats, and that of the granular layer increased. The density of neurons in each layer was significantly lower in old cats than in young adult ones. Astrocytes in old cats were significantly denser than in young adult ones, and accompanied by evident hypertrophy of the cell bodies and enhanced immunoreaction of GFAP substance. Purkinje cells (PCs) in old cats showed much fewer NF-IR dendrites than those in young adults. The above findings indicate a loss of neurons and decrease in the number of dendrites of the PCs in the aged cerebellar cortex, which might underlie the functional decline of afferent efficacy and information integration in the senescent cerebellum. An age-dependent enhancement of activity of the astrocytes may exert a protective effect on neurons in the aged cerebellum     

Distribution and morphology of ghrelin-immunopositive cells in the cerebellum of the African ostrich

Ghrelin, the endogenous ligand for the growth hormone secretagogue receptor, has been found in the cerebellum of many vertebrates and in the gastrointestinal tract of African ostrich chicks, but little is known about its distribution in the cerebellum of the African ostrich. In the present study, the distribution and morphological characteristics of ghrelin-producing cells in the cerebellum of the African ostrich were investigated using immunohistochemistry. The results indicate that the cerebellum is divided into two sections: the outer cerebellar cortex and the inner medulla of cerebellum. The cerebellar cortex comprises a molecular layer, a Purkinje cell layer and a granular layer; ghrelin-immunopositive (ghrelin-ip) cells were localized throughout the entire cerebellum, but sparsely in the medulla. The greatest number of ghrelin-ip cells was found in the stratum granulosum, and the density decreased gradually from the molecular layer to the Purkinje cell layer in the cerebellar cortex. The ghrelin-ip cells were fusiform or irregular polygons and their cytoplasm was stained intensely. These results clearly demonstrate the presence of ghrelin-ip cells in the cerebellum of the African ostrich. It is speculated that ghrelin may have a physiological function in the cerebellum.     

A single peripheral injection of basic fibroblast growth factor (bFGF) stimulates granule cell production and increases cerebellar growth in newborn rats

The control of neuronal number is critical for coordinating innervation and target organ requirements. Although basic fibroblast growth factor (bFGF) is known to regulate neuron number in the developing embryonic cortex, its potential role during postnatal brain development remains undefined. To address this issue, the cerebellum, a site of postnatal neurogenesis, was used. Previously, we found that a single peripheral injection of bFGF in newborn rats elicited mitosis of neuronal precursors in the external germinal layer (EGL) 8 h after administration. We now define the sustained effects of bFGF treatment on postnatal granule cell production and cerebellar growth. Seventy-two h after a single injection of bFGF (20 ng/g) in newborn rats, the fraction of BrdU-labeled cells in the EGL increased by 46% without altering apoptotic cell number, consistent with enhanced precursor proliferation. Moreover, bFGF increased mitotically labeled cells by 100% and total cell density by 33% in the internal granular layer (IGL), the final destination of the EGL precursors. Because cerebellar volume also increased by 22%, bFGF-induced proliferation enhanced generation of total IGL neurons and increased cerebellar growth. These morphometric measures were corroborated independently by using DNA quantitation: cerebellar DNA content increased 16% after bFGF injection, consistent with increased neuron number. Furthermore, using DNA quantitation as an index, increased total cerebellar cell number elicited by bFGF injection persisted beyond the neurogenetic period, until P35. We conclude that a single postnatal injection of bFGF increases granule neuron number and enhances cerebellar growth following mitotic stimulation.     

Relocalization of a microtubule-anchoring protein,ninein, from the centrosome to dendrites during differentiation of mouse neurons

Microtubules in typical cells form radial arrays with their plus-ends pointing toward the cell periphery. In contrast, microtubules in dendrites of neurons are free from centrosomes and have a unique arrangement in which about half have a polarity with a minus-end distal orientation. Mechanisms for generation and maintenance of the microtubule arrangement in dendrites are not well understood. Here, we examined dendritic localization of a centrosomal protein, ninein, which has microtubule-anchoring and stabilizing functions. Immunohistochemical analysis of developing mouse cerebral and cerebellar cortices showed that ninein is localized at the centrosome in undifferentiated neural precursors. In contrast, ninein was barely detected in migrating neurons, such as those in the intermediate layer of the cerebral cortex and the internal granular layer of the cerebellar cortex. High expression was observed in thick dendrite-bearing neurons such as pyramidal neurons of the cerebral cortex and Purkinje neurons in the cerebellar cortex. Ninein was not detected at the centrosome of these cells, but was diffusely present in cell soma and dendrites. In cultured cortical neurons, ninein formed granular structures in soma and dendrites, being not associated with γ-tubulin. About 60% of these structures showed resistance to detergent and association with microtubules. Our observations suggest that the minus-ends of microtubules may be anchored and stabilized by centrosomal proteins localized in dendrites.     

猫运动皮层神经元和S100、GFAP阳性细胞的年龄相关性变化

比较了青、老年猫运动皮层神经元与S100、GFAP免疫阳性胶质细胞的形态学变化,并探讨其与衰老过程中运动功能衰退的关系。采用Nissl染色显示青、老年猫运动皮层分层结构和神经元。免疫组织化学方法(SABC法)显示青、老年猫运动皮层S100免疫反应阳性(S100-immunoreactive,S100-IR)细胞及胶质纤维酸性蛋白免疫反应阳性(GFAP-immunoreactive,GFAP-IR)细胞。在Olympus显微镜下,用Moitcam5000数码成像与分析系统计数运动皮层各层神经元、S100-IR细胞及GFAP-IR细胞的数量,并随机抽样测量S100-IR、GFAP-IR细胞的胞体直径。与青年猫相比,老年猫运动皮层Ⅴ、Ⅵ层神经元密度显著下降(P<0.01),老年猫运动皮层中S100-IR和GFAP-IR细胞密度与胞体直径均显著增加(P<0.01),且细胞的免疫阳性反应较强。研究结果表明,猫运动皮层的神经元密度在衰老过程中Ⅴ、Ⅵ层神经元密度显著下降,有可能会降低老年个体运动皮层对运动的调控能力;随着衰老、运动皮层的星形胶质细胞出现明显的反应性活化与增生,这对维持大脑运动皮层神经元的活性和神经元之间的通讯联系,从而延缓老年性运动功能衰退具有重要意义。     

小熊猫小脑皮层的组织结构及RT-97、KGF和Bax蛋白在小脑皮层的表达

为了解小熊猫(Ailurus fulgens)小脑皮层的结构特征,观察神经丝蛋白抗体RT-97、角质细胞生长因子(KGF)及Bax蛋白在小脑皮层中的表达,利用组织学方法和免疫组织化学方法观察了小熊猫小脑皮层的显微结构,检测了RT-97、KGF和Bax蛋白的表达.结果表明,小脑皮层从外向内依次可分为分子层、Purkinje细胞层、颗粒层3层.RT-97在小熊猫小脑皮层Purkinje细胞层、颗粒层中神经细胞的轴突、分子层中颗粒细胞的轴突及小脑髓质中有阳性表达;KGF在小脑皮层分子层、Purkinje细胞层和颗粒细胞层及髓质中均有阳性表达;Bax蛋白在小脑皮层分子层、Purkinje细胞层和颗粒细胞层中有阳性表达.RT-97、KGF和Bax蛋白在小脑皮层神经结构的构筑中可能发挥着不同的功能.