A High-Content Larval Zebrafish Brain Imaging Method for Small Molecule Drug Discovery.

Harrison Liu,Zhiqiang Dong,Julie Gendre,Steven Chen,Han Mo,Qiang Li,Raffael Iovine,Michelle Arkin,Jeffrey Kim,Su Guo,Pauline Pascal,Kevin Huang,Bo Huang,Yaping Sun,Harold A Burgess

doi:10.1371/journal.pone.0164645

Harrison Liu, Zhiqiang Dong + Show 13 more

Open Access

https://doi.org/10.1371/journal.pone.0164645

Copy DOI

Abstract

Drug discovery in whole-organisms such as zebrafish is a promising approach for identifying biologically-relevant lead compounds. However, high content imaging of zebrafish at cellular resolution is challenging due to the difficulty in orienting larvae en masse such that the cell type of interest is in clear view. We report the development of the multi-pose imaging method, which uses 96-well round bottom plates combined with a standard liquid handler to repose the larvae within each well multiple times, such that an image in a specific orientation can be acquired. We have validated this method in a chemo-genetic zebrafish model of dopaminergic neuron degeneration. For this purpose, we have developed an analysis pipeline that identifies the larval brain in each image and then quantifies neuronal health in CellProfiler. Our method achieves a SSMD* score of 6.96 (robust Z’-factor of 0.56) and is suitable for screening libraries up to 105 compounds in size.

Highlights

The drug discovery process typically involves screening large libraries of compounds using in vitro or cell culture-based disease models. Using these simplified models allows for expedited assessment of compound binding and efficacy, but makes other drug parameters, such as in vivo efficacy and toxicity, absorption, distribution, metabolism, and excretion (ADME), difficult to assay [1]
Highthroughput screening at the whole-organism level can assay both compound effectiveness and ADME; it can be performed in the absence of a known target [3]
We discovered that using a liquid handler to quickly aspirate and replace 40uL of liquid in each well caused the anesthetized embryo within to tumble and assume a new pose

Summary

Introduction

The drug discovery process typically involves screening large libraries of compounds using in vitro or cell culture-based disease models. Using these simplified models allows for expedited assessment of compound binding and efficacy, but makes other drug parameters, such as in vivo efficacy and toxicity, absorption, distribution, metabolism, and excretion (ADME), difficult to assay [1]. This approach often results in final stage compounds that fail to show efficacy in whole-organism disease models or have unwanted toxicity [2].

Methods

Results

Conclusion