Automated Screening System for Modifiers of Cardiac Output in Zebrafish

描述(申请人提供)：斑马鱼，Danio rerio，已被广泛接受为人类疾病的模型。已经进行了小分子筛选，以确定正常心血管过程的修饰物和心血管疾病的修饰物。正在开发的高通量系统旨在测量心率，但不涉及心血管功能的其他关键参数，如血流速度和分布。我们建议开发一种高通量系统，该系统将同时：1)测量心率；2)生成主要血管的流动图；以及3)测量这些血管的血流速度。这个基于显微镜的系统将利用带有多普勒的光谱域光学相干断层扫描，在几秒钟内生成斑马鱼胚胎和幼虫的三维结构图像，并测量血液流量。在第一阶段，我们将开发一个面包板系统，该系统将能够从动物的背侧和腹侧视图收集图像。将收集脑部和尾部的血管系统的图像。在面包板系统中，将对动物进行定位，并手动调整扫描角度。在第二阶段，将开发一个完整的自动筛选系统，其中包括：1)用于快速定位动物以进行成像并沿其轴向长度调整扫描角度的软件算法；2)自动图像采集；以及3)自动图像处理。第二阶段测试将包括一个试点复合筛查，以确定心输出量的调节剂。 这项研究将开发能够在高通量筛选活性药物化合物中使用人类疾病的斑马鱼模型的技术。特别是，这类筛查有望加速发现治疗心血管疾病的新药。

DESCRIPTION (provided by applicant): The zebrafish, Danio rerio, has become widely accepted as a model for human disease. Small molecule screens have been performed to identify both modifiers of normal cardiovascular processes and modifiers of cardiovascular disease. High-throughput systems under development aim to measure heart rate but do not address other critical parameters of cardiovascular function such as blood velocity and distribution. We propose to develop a high-throughput system that would simultaneously: 1) measure heart rate; 2) generate a flow map of the major blood vessels; and 3) measure blood velocity in those vessels. The microscope-based system will utilize spectral domain optical coherence tomography with Doppler to generate, within a matter of seconds, a three dimensional structural image of zebrafish embryos and larvae and measure blood flow. In Phase I we will develop a breadboard system that will enable collection of images from both the dorsal and ventral views of the animals. Images of the vasculature both of the brain and of the tail will be collected. In the breadboard system the animals will be located and the scan angle will be adjusted manually. In Phase II a complete, automated screening system will be developed that includes: 1) software algorithms to rapidly locate the animals for imaging and adjust the scan angle along their axial length; 2) automated image collection; and 3) automated image processing. Phase II testing will include a pilot compound screen to identify modifiers of cardiac output. This research will develop technologies that will enable the use of zebrafish models of human disease in high-throughput screens for active pharmaceutical compounds. In particular, such screens are expected to accelerate discovery of new pharmaceuticals to treat cardiovascular diseases.