ELEMENTAL AND STRUCTURAL ORGANIZATION OF NEURONS AND GLIA

这两部分的项目研究的组织和功能， 神经元和神经胶质中的特殊膜。第一部分旨在 表征突触活动期间的钙调节 小脑皮质纤维-浦肯野细胞突触和突触内 海马CA 3区锥体细胞。 新的冰冻切片技术， 结合扫描透射电子显微镜（STEM）， 已经允许直接研究结构和钙调节， 一种新的海马器官型培养物的冷冻制备。 结构分析结合电生理记录， 证明了CA 3苔藓纤维突触的良好组织， 这些文化。 我们还成功地获得了来自 这些突触，和初步的元素分析提供了 总钙浓度和分布的首次数据 在海马体培养中有完整的神经回路。 在第二部分， 特化髓鞘膜的表达、转运和组装 研究蛋白质。共聚焦显微镜先前已经显示， 雪旺细胞依赖微管进行细胞内转运， 髓鞘特异性蛋白的组装。 我们现在用微管 组装-拆卸实验表明，P0蛋白，髓鞘- 相关糖蛋白（MAG）和层粘连蛋白，其靶向于 离散的和不同的质膜域，被分选成单独的 载体囊泡，因为它们退出trans-Golgi网络。 以下 分选，某些载体囊泡沿着髓鞘运输 微管上的节间，即使微管似乎不 选择性地将这些囊泡靶向特定的膜位点。

This two-part project studies the organization and function of specialized membranes in neurons and glia. The first part aims to characterize calcium regulation during synaptic activity in parallel fiber- Purkinje cell synapses of the cerebellar cortex and in synapses of CA3 hippocampal pyramidal cells. New frozen sectioning techniques, in combination with scanning transmission electron microscopy (STEM), have permitted studies of structure and calcium regulation in directly frozen preparations of a new kind of organotypic culture of hippocampus. Structural analysis in conjunction with electrophysiology recordings has demonstrated excellent organization of the CA3 mossy fiber synapses in these cultures. We have also successfully obtained cryosections from these synapses, and preliminary elemental analysis has provided for the first time data on the concentrations and distributions of total calcium in a hippocampal cultures with intact circuitry. In part two, the expression, transport and assembly of specialized myelin membrane proteins are studied. Confocal light microscopy had previously shown that Schwann cells depend on microtubules for intracellular transport and assembly of myelin-specific proteins. We have now used microtubule assembly-disassembly experiments to show that the P0 protein, the myelin- associated glycoprotein (MAG), and laminin, which are targeted to discrete and different plasma membrane domains, are sorted into separate carrier vesicles as they exit the trans-Golgi network. Following sorting, certain carrier vesicles are transported along the myelin internode on microtubules, even though microtubules do not appear to selectively target these vesicles to specific membrane sites.