CELL JUNCTIONS & CELL MEMBRANES IN THE LENS

我们建议扩大对连接复合体和细胞膜的研究 在脊椎动物的眼晶状体中。 此应用程序的长期目标是获得更好的 对结构特征、分布和功能的理解 各种类型的细胞连接在晶状体中的作用。细胞膜是 细胞连接的主要成分。据信，细胞 连接和细胞膜在维持正常状态中起着重要作用 建筑生理学和晶状体的透明度。我们延长了我们的 研究目标是深入了解细胞膜的作用 与有用大分子的内吞作用和囊泡运输有关 对着镜头。 研究计划的具体目标如下：(1)直接 晶状体天然缝隙连接外显子的结构分析与比较 使用快速冷冻技术的上皮和纤维细胞；(2)研究 缝隙连接与肌动蛋白束和肌动蛋白相关的关系 组件，使用细胞松弛素-D处理和改进的固定 含有戊二醛、赖氨酸和单宁酸；(3)免疫细胞化学 缝隙连接蛋白(MP70和/或MP26)、肌动蛋白和肌动蛋白的定位 (4)超微结构和免疫细胞化学分析 鸡晶状体形态发生过程中黏附连接的形成； 该菌发生、结构的形态和免疫细胞化学研究 晶状体中桥粒和相关细胞骨架成分的性质 不同种类的上皮；(6)超微结构和免疫细胞化学 与之相关的独特膜结构域(类笼状蛋白)的研究 纤维细胞中球窝连接的形成；(7)可视化 包膜囊泡和非包膜囊泡的结构和分布 不同物种的晶状体，使用改进的固定和蛋白质示踪剂 方法：(8)研究生长因子和其他因素对生长发育的影响。 晶状体内液相和受体介导的内吞作用； 免疫细胞化学方法检测某些生长发育过程中可能存在的损伤 以其特异性抗体为探针的内吞作用中的因素。 所使用的方法学包括(1)使用 本实验室改进的固定方法；(2)快速冷冻 在没有事先化学固定的液氦温度下，以及在 结合自由取代、冷冻断裂和深度蚀刻 技术；(3)常规冷冻断裂透射电子显微镜；(4)免疫荧光 和免疫金标记法，以及(5)蛋白质示踪剂和细胞化学 方法：研究方法。

We propose to extend the studies of junctional complexes and cell membranes in the ocular lens of vertebrates. The long-term objectives of this application are to gain a better understanding in structural characteristics, distributions and functional roles of various types of cell junctions in the lens. Cell membranes are the major component of al cell junctions. It is believed that cell junctions and cell membranes play important roles in maintaining normal architecture physiology, and transparency of the lens. We extend our research goals to gain some insights into the roles of cell membranes involved in endocytosis and vesicular transport of useful macromolecules into the lens. The specific aims of the research plan are as follows: (1) Direct structural analysis and comparison of native gap-junction connexons in lens epithelium and fiber cells, using rapid-freezing techniques; (2) Study association of gap junctions with actin bundles and actin-associated components, using cytochalasin-D treatments, and an improved fixation containing glutaraldehyde, lysine and tannic acid; (3) Immunocytochemical localization of gap junction proteins (MP70 and/or MP26), actin and actin- bundling proteins; (4) Ultrastructural and immunocytochemical analyses of formation of adherens junctions during chick lens morphogenesis; (5) Morphological and immunocytochemical studies of the occurrence, structure and nature of desmosomes and associated cytoskeletal components in lens epithelium of various species; (6) Ultrastructural and immunocytochemical investigations of unique membrane domains (clathrin-like) associated with formation of ball-and-socket junctions in fiber cells; (7) Visualization of structure and distribution of coated vesicles and non-coated vesicles in the lens of various species, using improved fixation and protein tracer methods; (8) Study the effects of growth factors and other factors on fluid-phase and receptor-mediated endocytosis in the lens; (9) Immunocytochemical determination of possible involvements of some growth factors in endocytosis using their specific antibodies as probes. Methodology used incudes (1) transmission electron microscopy using the improved fixation method developed in this laboratory; (2) rapid-freezing in liquid helium temperature without prior chemical fixation, and in conjunction with free-substitution, freeze-fracture and deep-etching techniques; (3) conventional freeze-fracture TEM; (4) immunofluorescence and immunogold labeling techniques, and (5) protein tracer and cytochemical methods.