Cultured Peptidergic Neurons: Electrical, Secretory, Immunological and Morphological Correlates

培养的肽能神经元：电、分泌、免疫和形态学相关性

• 批准号: 8910432
• 负责人: Ian Cooke
• 金额: $ 20.86万
• 依托单位: University of Hawaii
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-01 至 1992-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8910432&HistoricalAwards=false
• 关键词: Cultured; Peptidergic; Neurons; Electrical; Secretory

Information is sought to answer questions about the relationship between the differing electrical characteristics of neurons that secrete different peptide hormones and their biochemical and morphological characteristics. The neurons will be studied intact within an acutely isolated preparation and isolated in cell culture. In culture, the neurons synthesizing different hormones, as indicated by their immunoreactivity to specific antisera, assume distinct morphologies under defined conditions. The cultured neurons having different form show distinct differences in their electrical characteristics. The material used for these studies is the X-organ - sinus gland of the tropical land crab, Cardisoma carnifex. Earlier work has demonstrated that this crustacean neurosecretory system is directly analogous to the major vertebrate neurosecretory system, the hypothalamic-neurohypophyseal system, and that at every level of analysis yet undertaken the physiological and cell biological mechanisms involved are directly comparable. Many of the observations obtainable with the crab material, however, would not be possible with vertebrate material. The crab neurosecretory system is easily isolated and consists exclusively of peptidergic neurons and supporting tissue. The terminals are unusually large (up to 30um), permitting recording with intracellular or patch electrodes. Antisera to several peptide- hormonal activities permit typing of neurons and terminals. Isolated X-organ neurons immediately commence regeneration of processes in unconditioned dishes and defined media. Conventional microelectrode techniques and dye injection will be used to characterize neurons in acutely isolated systems. Both conventional and patch-clamping techniques will be applied to the neurons in culture. The regional distribution in the membrane of channels giving rise to different ionic currents will be explored by testing the effects of axotomy and by localized recording. Regional sensitivity to putative neurotransmitter agents will be explored. As feasible, changes with time in culture or with changed morphology induced by altered culturing conditions will be detailed.

信息是寻求回答有关神经元分泌不同肽激素的不同电特性及其生化和形态特征之间的关系的问题。神经元将在急性分离制备和细胞培养中完整地进行研究。在培养中，合成不同激素的神经元，根据它们对特定抗血清的免疫反应，在规定的条件下呈现不同的形态。不同形态的神经元在电特性上表现出明显的差异。这些研究使用的材料是热带陆地蟹Cardisoma carnifex的x器官窦腺。早期的研究表明，这种甲壳类动物的神经分泌系统与主要的脊椎动物的神经分泌系统——下丘脑-神经垂体系统——有直接的相似之处，并且在每一层次的分析中，所涉及的生理和细胞生物学机制都是直接可比较的。然而，许多用螃蟹材料得到的观察结果，用脊椎动物材料是不可能得到的。蟹的神经分泌系统很容易分离，只由多肽能神经元和支持组织组成。终端异常大（可达30um），允许使用细胞内或贴片电极进行记录。几种肽激素活性的抗血清允许神经元和终末分型。分离的x器官神经元在无条件培养皿和确定的培养基中立即开始再生过程。传统的微电极技术和染料注射将用于表征急性分离系统中的神经元。常规技术和贴片夹紧技术都将应用于培养的神经元。我们将通过测试肛门切开术的效果和局部记录来探讨引起不同离子电流的通道在膜中的区域分布。将探讨对假定的神经递质药物的区域敏感性。如果可行，将详细说明随培养时间的变化或因改变培养条件而引起的形态变化。