Confocal laser scan system to image parallel processing of the brain neurons in behaving animals

共焦激光扫描系统对行为动物大脑神经元的并行处理进行成像

基本信息

批准号：
12358013
负责人：
ODA Yoichi
金额：
$ 22.83万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

We developed two confocal laser systems to image neuronal activities of behaving animals. The first system consists of a high-speed scanner built on an upright microscope. It can scan 500 frames per second but has a low signal-to-noise ratio to detect faint change in fluorescence signals. The second system is based on a conventional confocal laser system, but the scanning speed was improved to collect 10 full-frames to 500 line-frames every second. This confocal system has a higher signal-to-noise ratio to detect fluorescence change associated with synaptic inputs without averaging them. Using the system we observed the activities of reticulospinal neurons in the hindbrain of zebrafish larvae. Since the body and brain of larval zebrafish are transparent, the neuronal activities in vivo can be observed by calcium imaging of the neurons in the intact brain. Especially we succeeded in in vivo imaging of functional inhibitory networks on the Mauthner cell which is known to initiate fast escape response behavior of fish. It is the first report of imaging inhibitory synaptic action in intact animals. Finally we developed the optical system to observe neuronal activities of behaving animals. For this purpose, the high-speed video camera system was added to the laser-scan system to measure the fluorescence responses of neurons and behavioral responses of zebrafish simultaneously. The high-speed confocal laser scanning system associated with motion detector will help us to understand which and how central neurons contribute to controlling the simple behavior of animals.

我们开发了两种共焦激光系统来对行为动物的神经元活动进行成像。第一个系统由建立在正置显微镜上的高速扫描仪组成。它每秒可以扫描 500 帧，但信噪比较低，无法检测荧光信号的微弱变化。第二个系统基于传统的共焦激光系统，但扫描速度提高到每秒收集 10 个全帧到 500 个线帧。该共焦系统具有更高的信噪比，可以检测与突触输入相关的荧光变化，而无需对其进行平均。使用该系统，我们观察了斑马鱼幼虫后脑中网状脊髓神经元的活动。由于斑马鱼幼体的身体和大脑是透明的，因此可以通过完整大脑神经元的钙成像来观察体内的神经元活动。特别是我们成功地对毛特纳细胞的功能抑制网络进行了体内成像，已知毛特纳细胞可以引发鱼类的快速逃逸反应行为。这是对完整动物的抑制性突触作用进行成像的第一份报告。最后，我们开发了光学系统来观察行为动物的神经元活动。为此，在激光扫描系统中添加高速摄像机系统，以同时测量神经元的荧光反应和斑马鱼的行为反应。与运动探测器相关的高速共焦激光扫描系统将帮助我们了解哪些中枢神经元以及如何有助于控制动物的简单行为。