Optical study of secretion in mammalian nerve terminals

哺乳动物神经末梢分泌的光学研究

• 批准号: 8431349
• 负责人: BRIAN Matthew SALZBERG
• 金额: $ 48.01万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-24 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8431349
• 关键词: Alzheimer' s Disease; Amplifiers; Argipressin; Atomic Force Microscopy; Axon; Biological Process; Blood Circulation; Cell physiology; Cells; Clinic; Collection; Coupling; Detection; Development; Disease; Electron Microscopy; Equilibrium; Event; Exocytosis; Extracellular Space; Fiber; Fluorescence; Goals; Gold; Hormones; Hypertension; Hypothalamic structure; Immune response; Impairment; In Vitro; Infundibular Stalk; Laboratories; Lambert-Eaton Myasthenic Syndrome; Lead; Measurement; Mechanics; Membrane; Membrane Fusion; Mental Depression; Metabolic; Metabolism; Methods; Milk Ejection; Mitochondria; Moods; Mus; NADH; Names; Nerve; Nervous system structure; Neurologic; Neurons; Neuropeptides; Neurosecretion; Optical Methods; Optics; Oxytocin; Physiological; Pituitary Gland; Posterior Pituitary Gland; Preparation; Process; Proteins; Regulation; Reproduction; Resolution; Scanning Probe Microscopes; Schizophrenia; Secretory Vesicles; Sex Behavior; Signal Transduction; Staging; Synaptic Transmission; System; Techniques; Technology; Temperature; Testing; Time; Transgenic Mice; Variant; Vasopressins; Water; base; blood pressure regulation; cell motility; coral; detector; fluorophore; information processing; insight; light scattering; magnocellular; median eminence; millisecond; neurotransmitter release; novel; novel strategies; paraventricular nucleus; peptide hormone; presynaptic; promoter; red fluorescent protein; research study; response; social cognition; supraoptic nucleus; tool

DESCRIPTION (provided by applicant): Secretion is one of the most ubiquitous of cellular processes. Regulated exocytosis is central to such diverse biological functions as information processing, reproduction, motility, temperature regulation, metabolism, the immune response, and signal transduction, but the elucidation of its mechanism(s) remains a challenge to cell physiologists. The neurohypophysis is a classic in vitro preparation, uniquely relevant for studying evoked release of peptide hormones. Magnocellular neurons from the supraoptic and paraventricular nuclei in the hypothalamus project their axons as bundles of fibers through the median eminence and infundibular stalk to arborize extensively and terminate in the neurohypophysis, a quasi-pure collection of nerve terminals, where the neuropeptides oxytocin and vasopressin are released into the circulation. Despite remarkable progress in identifying components of the macromolecular machinery essential for their release, direct observation in real time of the intraterminal events that follow electrical excitation, but precede membrane fusion, has proven extremely difficult. To identify these steps we have developed an arsenal of biophysical techniques possessing time scales ranging from microseconds to minutes, with emphasis on the former. Three of them, in particular, constitute the core of each of our specific aims: 1) High Bandwidth Dynamic Atomic Force Microscopy (HBDAFM), to study the rapid, but very small (~1.0 ¿), mechanical events ("spike" and "dip") which accompany neuropeptide secretion in mammalian nerve terminals. 2) Real-time detection of Ca2+-transients, from whole terminals as well as from near- membrane regions, to identify intermediate steps between Ca2+-entry and exocytosis. 3) Millisecond time resolved optical detection of secretion per se. These technologies, already at different stages of development in our laboratory, will be combined in this proposal to capture, in a single experiment, sequential, albeit partially overlapping events, which are essential components of excitation-secretion coupling.

描述（由应用程序提供）：分泌是蜂窝过程中最普遍的一种。受调节的胞吐作用对于信息处理，繁殖，运动，温度调节，代谢，免疫响应和信号转导等多种生物学功能至关重要，但其机制的阐明仍然是对细胞生理学家的挑战。 Neuroholypophysy是一种经典的体外制剂，与研究诱发的意大利辣味激素的释放相关。下丘脑中的上型和旁腔核核核元的巨细胞神经元将其轴突作为纤维束作为纤维束作为纤维束，并通过中位数和漏斗性茎，以广泛的神经嗜知和终止神经嗜知分析，在神经素中终止，释放的Never-pure终端集合，释放了循环蛋白蛋白蛋白蛋白蛋白蛋白蛋白和VasopressIns resepressIns and VasopressIns和VasopressIns rate and vasopressIns at vasopressIns and VasopressIns。尽管在确定释放大分子机械的组件方面取得了显着的进展，但事实证明，在实时的及时兴奋（但之前的膜融合）之前，直接观察到了直接观察，但事实证明非常困难。为了确定这些步骤，我们开发了一种具有从微秒到几分钟的时间尺度的生物物理技术库，重点是前者。其中三个尤其构成了我们每个特定目的的核心：1）高带宽动态原子力显微镜（HBDAFM），以研究快速但很小的（〜1.0€），机械事件（“ Spike”和“ DIP”），可容纳哺乳动物的神经肽的分泌。 2）从整个末端以及近膜区域的CA2+过量剂的实时检测，以识别Ca2+ - 饮食和胞吐作用之间的中间步骤。 3）毫秒末解析了分泌本身的光学检测。这些技术已经在我们实验室的发展阶段已经不同，将在此提案中结合使用，以捕获单个实验，即顺序的，尽管部分重叠的事件，这是兴奋 - 分泌耦合的必不可少的组成部分。