HTS assay for zebrafish embryonic heart function (RMI)

Massachusetts General Hospital, Boston MA

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Linked publications & trials

Abstract

[unreadable] DESCRIPTION (provided by applicant): Embryonic heart function is a complex biological process regulated by a host of cardiac proteins and, directly or indirectly, by proteins and pathways in other organ systems. If technology existed for automated determination of embryonic heart function in a high-throughput molecular screening (HTS) assay, then screening for small molecules (novel compounds and FDA approved drugs) that influence embryonic cardiac physiology would become possible. The small molecules identified in such screens would have important applications in cardiovascular physiology, toxicology, and drug discovery. An automated assay for zebrafish embryonic heart rate has been developed that utilizes a 96-well format and transgenic embryos (cmlc2::GFP) expressing GFP specifically in the embryonic heart. Briefly, an automated fluorescence microscope scans a 96-well plate, identifies GFP+ hearts, and acquires short digital videos. Raw data from image analysis are used, in conjunction with a Fast Fourier Transform function, to quickly and automatically determine heart rates. Heart rates are outputted in a simple format. Experiments in specific Aim I will determine the reproducibility, responsiveness, and selectivity of assay-response by screening the NINDS custom collection (approximately 1,000 bioactive molecules) available from the Harvard Institute of Chemistry and Cell Biology. Experiments in specific aim II will investigate the viability of incorporating indices of embryonic heart contractility into the assay so that two parameters, heart rate and contractility, can be assay simultaneously thereby yielding a more-complete measure of embryonic heart function. In its current incarnation, the assay has important applications in cardiovascular physiology, toxicology, and drug discovery. The breadth of applications would increase if contractility were incorporated. The long-term goal is to perform high throughput screening to identify small-molecule-modifiers of embryonic heart function for use as biological probes and as lead compounds in the treatment of human heart disease. [unreadable] [unreadable]