遗传性视网膜变性rd小鼠及其感光细胞凋亡研究

目的 研究遗传性视网膜变性rd小鼠感光细胞层的发育变化及细胞凋亡。方法 对出生后5d到40d的rd小鼠及对照小鼠视网膜感光细胞层进行光镜及超微结构观察、TUNEL法检测及形态计量学分析。结果 与同龄对照鼠相比,rd小鼠出生后第10d视网膜开始变性,尔后1周内感光细胞迅速减少,第18d时只残留一层视椎细胞。rd小鼠出生后第10d感光细胞层开始出现TUNEL染色阳性细胞,第14d及16d达到高峰。电镜下变性高峰期rd小鼠视网膜感光细胞层可见大量浓缩核、染色质边聚及凋亡小体。结论 rd小鼠视网膜感光细胞在发育过程中变性,并通过凋亡的方式死亡。     

视网膜感光细胞退变过程中内层神经元结构和功能的变化

Retinitis pigmentosa (RP) is a retina disease which leads to a progressive photoreceptors degeneration. Although the inner neurons that mainly include bipolar cells and ganglion cells of retina remain relative stable during the photoreceptors degeneration, studies show that their morphologies and functions are changed during this process. Stu-dies indicate that rod bipolar cells (RBCs) and cone bipolar cells (CBCs) undergo dendritic retraction and remodelling in retinal degenerated mice, and they show changes in expression of the glutamate receptors as well as responses to glutamate and light stimuli. In many animal models of RP, significant decreases in receptive field size, contrast sensitivity, and threshold sensitivity, as well as an increasing spontaneous spiking in retinal ganglion cells (RGCs) were detected. As these modifications may impact the choice and success of intervention strategies for treating retinal degeneration, it is important to understand the characteristics of any functional deficits resulting from photoreceptors degeneration.     

新生犊牛视网膜节细胞的形态分布

目的:研究比较新生犊牛视网膜节细胞(RGCs)的数量、大小和密度的形态学变化。方法:采用Nissl染色、荧光染料DiI逆行标记技术和计算机图像处理方法。结果:RGCs在视神经乳头下方形成一个沿鼻颞侧轴方向伸展的高密度区(P1,2608/mm2;P21,2174/mm2),即视条纹。由视条纹至周边部细胞密度递减,颞侧周边部最低(P1,mm2;P21,217/mm2);细胞大小则呈递增的变化,这种变化趋势在颞侧最明显。由P1到P21,RGCs总数和平均细胞密度均递减而细胞大小递增。结论:RGCs大小由视条纹至周边部递增而细胞密度递减,并且由P1到P21,RGCs总数和细胞大小递增而细胞密度递减。     

兔视网膜LANT-6免疫反应节细胞的研究

用免疫组织化学ＡＢＣ法和间接免疫荧光双标法，研究ＬＡＮＴ－６免疫反应神经元在成年兔视网膜的定位与分布。结果表明，ＬＡＮＴ－６免疫阳性细胞体仅位于节细胞层，呈圆形或椭圆形，直径约为１０～４０μｍ，胞体可发生１个或多个树突伸向内网层，在神经纤维层偶见免疫视神经纤维。ＬＡＮＴ－６免疫细胞分布于视网膜各个区，细胞密度以视纹最高，其平均密度为３７８．３±（３０）／ｍｍ２，从视纹向边缘区密度变少，中央区、外围区和边缘区的平均密度分别为２０２±（１０）／ｍｍ２、１１４．３±（１５）／ｍｍ２和２１．６±（７）／ｍｍ２。双标实验的结果表明，兔视网膜的ＬＡＮＴ－６免疫神经元是视网膜节细胞。上述结果提示ＬＡＮＴ－６可能对视网膜节细胞与视觉中枢之间的神经传递起重要作用。     

移植周围神经对大鼠受损视网膜节细胞的影响

用成年雄性Ｗｉｓｔａｒ大鼠切断一侧视神经后，随即将自体坐骨神经段、神神经段或挤压后的坐骨神经段植入眼球玻璃体内，植入神经段长２ｍｍ。术后动物分别存活２、４或８周，将动物麻醉处死后，取同侧视网膜制备切片，Ｎｉｓｓｌ染色，观察视网膜节细胞的变化，并用计算机图像分析仪测量ＲＧＣ胞体截面积。结果表明，植入坐骨神经段各组ＲＧＣ大多数存活，并出现巨大的节细胞。存活２周时，ＲＧＣ胞体截面积均值比正常组大；４周时     

Nogo-A在小鼠胚胎新生视网膜节细胞及其轴突的表达

目的 研究小鼠胚胎阶段Nogo-A在视网膜节细胞(RGCs)及其轴突上的表达及时程变化. 方法 取不同发育阶段的小鼠胚胎,采用免疫荧光染色,以激光扫描共焦显微镜观察Nogo-A在视觉传导通路中的表达.并采用免疫双标染色确定视网膜中表达Nogo-A蛋白的细胞类型. 结果 在视网膜发育的早期阶段(E12),Nogo-A密集表达于具有放射状形态的细胞上,Nogo-A免疫阳性产物出现在胞质、胞膜以及轴突上.Nogo-A与Tuj-1双标染色显示,此阶段的视网膜中几乎所有RGCs及其轴突都表达有Nogo-A;在稍晚的发育阶段(E13),视网膜中表达Nogo-A的RGCs数量明显减少,且仅出现在节细胞层以外的室周带和睫状体边缘区.在视网膜的神经纤维层,大部分RGCs轴突不再表达Nogo-A,仅有少量视觉纤维为Nogo-A免疫阳性;RGCs的神经发生基本完成后(E15), 视网膜中几乎检测不到Nogo-A免疫阳性的细胞,但视网膜纤维层仍有少量表达Nogo-A的节细胞轴突.与之类似,视神经盘、视茎、视交叉和视束都观察到少量Nogo-A免疫阳性的轴突.值得注意的是,视束中表达Nogo-A的纤维集中位于表浅部位,而此处恰为新近到达轴突的通过部位. 结论 Nogo-A在视网膜RGCs以及轴突上表达的时程变化和位置特点提示,新生RGCs及其轴突表达Nogo-A,成熟后RGCs内Nogo-A的表达则下调.推测新生RGCs及其轴突中表达的Nogo-A可能与减少轴突分叉等细胞的内在功能有关.     

Response of X and Y cat retinal ganglion cells to moving stimuli

Summary Single optic tract fibers in the cat were classified as X or Y cells by a contrast reversal stimulus. A slit of light was then moved across the receptive field at velocities from 10–1000 ° /s. The preferred velocity was that velocity which elicited the strongest response from the cell. The maximum velocity was the highest velocity target to which a cell could respond. Y cells as a group both preferred and could follow faster targets than X cells, and were more broadly tuned to the preferred velocity.This study was supported by Public Health Service Grant EY 00701 and NEI Grant EY 00376     

远志皂苷元对氧化应激损伤的视网膜神经节细胞的保护作用

目的: 观察远志皂苷元(senegenin,Sen)对氧化应激损伤的视网膜神经节细胞(retinal ganglion cells,RGCs)的影响并初步探讨其作用机制。方法: 采用上丘荧光金逆行标记RGCs后,体外原代RGCs混合细胞培养,随机分为control组、H₂O₂组、Sen+H₂O₂和Sen组。检测带荧光金荧光细胞的活力。同上述分组处理视网膜,用Hoechst 33258 染色后观察视网膜细胞核形态的变化,Western blotting检测视网膜细胞cleaved caspase-3、细胞色素C及Bcl-2蛋白的表达。结果: 与control组比较,Sen浓度在10、20和40 μmol/L时, RGCs活力无明显变化(P>0.05),但Sen浓度达到80和160 μmol/L时,RGCs活力明显下降,差异显著(P<0.01)。25、50、100和200 μmol/L H₂O₂明显降低RGCs活力(P<0.05)。Sen浓度在10、20和40 μmol/L时,对50 μmol/L H₂O₂损伤的RGCs有较好的保护作用(P<0.05),其中40 μmol/L Sen保护作用最为明显。Hoechst 33258染色表明Sen可以减少H₂O₂引起的视网膜细胞凋亡。Western blotting结果表明Sen促进Bcl-2蛋白的表达,降低线粒体细胞色素C的释放,下调cleaved caspase-3的表达。 结论: Sen保护RGCs对抗氧化应激引起的损伤,其机制可能与其增强Bcl-2蛋白表达和减少氧化应激引起的细胞凋亡有关。     

Statistical dependence between neighboring retinal ganglion cells in goldfish

Summary Simultaneous recordings were made of the spontaneous discharge of neighboring retinal ganglion cells in goldfish. Crosscorrelation analysis indicates two types of correlation. Neighboring units with like receptive field organization tend to discharge together while units having complementary receptive field organization tend not to discharge together.     

Morphological classification of retinal ganglion cells in adult Xenopus laevis

Summary Retrograde transport of horseradish peroxidase (HRP) was used to characterise the soma and dendritic arborization of retinal ganglion cells in adultXenopus laevis toad. HRP was administered to the cut end of the optic nerve and the morphological characteristics of HRP-filled ganglion cells were analysed in retinal wholemount preparations using computer assisted morphometry. Ganglion cells were classified according to their soma size, dendritic branching pattern, dendritic field and the number of shaft dendrites. Ganglion cells were divided into 3 major classes on the basis of soma sizes and extent of dendritic field: large (soma size, mean 258.04 μm²±52.03 SD; dendritic field size 0.104 mm²±0.23), medium size (126.7 μm²±37.01; 0.041 mm²±0.013) and small (87.3 μm²±22.69; 0.0061 mm²±0.0035). A more detailed analysis allowed 12 morphologically distinct subgroups to be identified (Types I–XII). Quantitative studies showed that large cells comprise about 1%, medium size about 8–9% and the small cells over 90% of total ganglion cell population. The number of large and medium size ganglion cells corresponded well with the number of myelinated optic fibres and the number of small neurons with the number of unmyelinated optic fibres in the optic nerve. Large ganglion cells were correlated with Class 4 and 5, medium size ganglion cells with Class 3 and small ganglion cells with Class 1 and 2 functionally characterized ganglion cells in the frog retina (Maturana et al. 1960). The retinal distribution of large ganglion cells appear to suggest certain similarities to mammalian alpha type ganglion cells.     

原花青素对微波诱导视网膜神经节细胞凋亡的拮抗作用

目的： 探讨不同剂量的原花青素(procyanidin，PC)对微波致视网膜神经节细胞（retinal ganglion cells，RGCs）损伤的拮抗作用。方法: 体外培养RGCs，将细胞随机分为6组，即不同PC剂量(0 μg/L 、2 μg/L、20 μg/L和40 μg/L)4个实验组及维生素C 40 μg/L组和对照组，分别给予频率为2 450 MHz、功率密度为30 mW/cm2的微波连续辐照1 h，对照组不给予辐照。光镜下观察辐照前后细胞形态学的改变，台盼蓝染色检测细胞存活率，应用流式细胞仪和DNA末端标记法(TdT-mediated-dUTP nick end labeling，TUNEL)定量检测细胞早期凋亡。结果: 微波辐照后，细胞即有聚集现象，部分细胞突起消失。辐照后0 h，20 μg/L和40 μg/L PC组较0 μg/L PC组的细胞凋亡率有所降低(P<0.05)；辐照后6 h和12 h，3个PC剂量组较0 μg/L PC组的细胞凋亡率都有降低(P<0.05)；辐照后18 h，只有40 μg/L PC组较0 μg/L PC组的细胞凋亡率显著降低(P<0.01)。结论: 微波可诱导RGCs细胞凋亡，PC对此损伤具有一定的拮抗作用。     

Response of cat retinal ganglion cells to motion of visual texture

Summary Responses of retinal ganglion cells to motion of large fields of visual texture were recorded in the lightly anaesthetised, immobilized cat. Brisk sustained and brisk transient, on- or off-centre, units gave a modulated response to texture motion. The pattern of temporal modulation of the response was dependent upon the particular configuration (sample) of texture crossing the receptive field. The magnitude of the response depended on the size of the receptive field centre. For all units, whether sustained or transient the magnitude of response to a textured field of fixed angular subtense declined as centre-diameter increased from 0.9 deg. For brisk units the response magnitude levelled off for centre sizes smaller than 0.9 deg. Responses to texture were confined spatially to the region of the receptive field, and the overall characteristics of this response were due to interactions between the centre and surround mechanisms of the receptive field. In brisk transient units, no evoked response was evident when texture motion was confined to regions well away from the receptive field of the unit, i.e. no periphery or shift effect could be demonstrated. The results support previous suggestions that the differential sensitivity to texture motion evident in cortical neurones must be due to intra-cortical processing.Supported by Project Grant G978/558/N from the Medical Research Council     

Orientation bias of cat retinal ganglion cells: a reassessment

Summary For cat retinal ganglion cells whose receptive field centres were distributed in specified sections of the left visual field, the deviations of the major axis from the radial, horizontal, and circumferential directions were determined. The percentage of cells with deviations within ± 20° from the radial, horizontal, and circumferential directions were, respectively, 33%, 68%, 16%. In addition, comparison between values of deviation from the horizontal direction for cells located at eccentricities of 10° and 20° from the area centralis showed a statistically significant trend: the bias for the horizontal increased with eccentricity.     

Classification of cat retinal ganglion cells into X- and Y-cells with a contrast reversal stimulus

Summary A contrast reversal (alternating phase) stimulus was used to study the responses of 150 retinal ganglion cells from 15 adult cats. Because the majority of the cells did not show perfect linear spatial summation, a ratio of the firing rates at two time periods was used to express the degree of nonlinearity. Y-cells showed a high degree of nonlinearity, and their mean null ratio was significantly lower than that of X-cells. With the stimulus at the null position, X-cells had an unmodulated discharge rate which was significantly higher than maintained activity, while the firing rate of Y-cells was lower than maintained activity. With the stimulus placed at an eccentric position in the receptive field, X-cells responded in a sustained manner, while Y-cells respond transiently. Because of these observations, we conclude that X-cells correspond to the sustained cells, while Y-cells correspond to the transient cells.