Presynaptic Mechanisms at Ribbon-class Synapses

描述（由申请人提供）：这项工作的长期目标是了解耳朵、眼睛和其他在带状突触上使用分级传递的感觉系统中感觉受体细胞的突触传递机制。这种特殊形式的化学信号似乎是一种适应，允许传递有关感觉输入的微小变化的信息，这些信息在使用动作电位的传统突触传递过程中会丢失。拟议的实验将研究这些突触的生理学、解剖学和生物化学。耳朵中感觉受体（毛细胞）的研究将使用青蛙和斑马鱼作为模型物种。斑马鱼的工作还将研究视网膜中的光感受器和双极细胞，以及侧线器官中的毛细胞。所有这些声音都有带状神经突触。之所以选择这些物种进行研究，是因为这些细胞和分子工具可以回答突触生理学中的基本问题。在青蛙的例子中，也有大量的信息已经可以用来建立。在过去的10年中，几个实验室已经开发出技术，允许详细的电生理分析突触传递使用密封电压钳测量膜电容的微小变化，以观察突触传递在毫秒的时间尺度。这种生理学方法将与电子断层扫描和最近开发的膜示踪染料结合起来，用于研究突触囊泡周期带状突触的超微结构。我们的目标是测试几个关键的假设，关于功能的突触“丝带”，突出的解剖特征，这些突触被命名。该项目还将重点关注两种主要蛋白质（calretinin和parvalbumin 3），它们被认为通过捕获和运输钙离子远离突触，在这些细胞的突触传递中发挥核心作用。该项目将研究决定这些蛋白质结合钙的速度有多快，它们可以螯合多少钙，以及它们在细胞内扩散的速度有多快的重要生化特性。这些特性对于理解毛细胞中的突触传递至关重要，并且与这些和相关钙结合蛋白在中风和其他脑损伤期间保护钙超载的拟议功能具有更广泛的相关性。斑马鱼研究人员在过去十年中开发的遗传和分子工具将允许直接测试最近被确定为丝带主要成分的蛋白质（Ribeye）的功能。

DESCRIPTION (provided by applicant): The long-term objective of this work is to understand the mechanism of synaptic transmission in sensory receptor cells in the ear, eye and other sensory systems that use graded transmission at ribbon-class synapses. This specialized form of chemical signaling appears to be an adaptation to allow the transmission of information about small changes in sensory input that would be lost during conventional synaptic transmission that uses action potentials. The proposed experiments will investigate the physiology, anatomy and biochemistry of these synapses. The work on sensory receptors (hair cells) in the ear will use frogs and zebrafish as model species. The zebrafish work will also study photoreceptors and bipolar cells in the retina, and hair cells in the lateral line organs. All of these calls have ribbon-class synapses. These species were chosen for study because of the cellular and molecular tools that are available to answer fundamental questions in synaptic physiology. In the case of frogs, there is also a wealth of information already available upon which to build. During the past 10 years, several laboratories have developed techniques that allow detailed electrophysiological analysis of synaptic transmission using tight-seal voltage clamp to measure small changes in membrane capacitance to observe synaptic transmission on a millisecond time scale. This physiological method will be used in conjunction with electron tomography and recently developed membrane tracer dyes to study the ultra structure of the synaptic vesicle cycle ribbon synapses. The goal is to test several key hypotheses concerning the function of the synaptic "ribbon", the prominent anatomical feature for which these synapses are named. The project will also focus on two major proteins (calretinin and parvalbumin 3) that are believed to serve central roles in synaptic transmission in these cells by capturing and transporting calcium ions away from the synapses. The project will investigate the important biochemical properties that determine how fast these proteins bind calcium, how much calcium they can sequester, and how fast they can diffuse within the cell. These properties are central to understanding synaptic transmission in hair cells, and have a wider relevance to the proposed function of these and related calcium-binding proteins in protection from calcium overload during strokes and other brain injuries. The genetic and molecular tools developed by zebrafish researchers during the past decade will allow a direct test the function of a protein (Ribeye) that has recently been identified as a major component of the ribbon.