Studying Neurobehavioral Effects of Environmental Pollutants on Zebrafish Larvae

Summary

A detailed experimental protocol is presented in this paper for the evaluation of neurobehavioral toxicity of environmental pollutants using a zebrafish larvae model, including the exposure process and tests for neurobehavioral indicators.

Abstract

Recent years more and more environmental pollutants have been proved neurotoxic, especially at the early development stages of organisms. Zebrafish larvae are a preeminent model for the neurobehavioral study of environmental pollutants. Here, a detailed experimental protocol is provided for the evaluation of the neurotoxicity of environmental pollutants using zebrafish larvae, including the collection of the embryos, the exposure process, neurobehavioral indicators, the test process, and data analysis. Also, the culture environment, exposure process, and experimental conditions are discussed to ensure the success of the assay. The protocol has been used in the development of psychopathic drugs, research on environmental neurotoxic pollutants, and can be optimized to make corresponding studies or be helpful for mechanistic studies. The protocol demonstrates a clear operation process for studying neurobehavioral effects on zebrafish larvae and can reveal the effects of various neurotoxic substances or pollutants.

Introduction

In recent years more and more environmental pollutants have been proved neurotoxic^1,2,3,4. However, the assessment of neurotoxicity in vivo after exposure to environmental pollutants is not as easy as that of endocrine disruption or developmental toxicity. In addition, early exposure to pollutants, especially at environmentally relevant doses, has attracted increasing attention in toxicity studies^5,6,7,8.

Protocol

The protocol is in accordance with guidelines approved by the Animal Ethics Committee of Tongji University.

1. Zebrafish embryo collection

1. Put two pairs of healthy adult Tubingen zebrafish into the spawning box on the night before exposure, keeping the sex ratio at 1:1.
2. Remove the adult fish back to the system 30-60 min after daylight the next morning.
3. Remove the embryos out of the spawning box.
4. Rinse the embryos with system water.
5. Transfer t.......

Discussion

This work provides a detailed experimental protocol to evaluate the neurotoxicity of environmental pollutants using zebrafish larvae. Zebrafish go through the process from embryos to larvae during the exposure period, which means that good care of the embryos and larvae is essential. Anything that affects the development of the embryos and larvae can influence the final result. Here the culture environment, exposure process, and experimental conditions are discussed to ensure the success of the whole assay.

Materials

Name	Company	Catalog Number	Comments
48-well-microplate	Corning	3548	Embyros housing
6-well-microplate	Corning	3471	Embyros housing
BDE-47	AccuStandard	5436-43-1	Pollutant
DMSO	Sigma	67-68-5	Cosolvent
Microscope	Olympus	SZX 16	Observation instrument
Pipette	Eppendorf	3120000267	Transfer solution
Zebrabox	Viewpoint	ZebraBox	Behavior instrument
Zebrafish	Shanghai FishBio Co., Ltd.	Tubingen	Zebrafish supplier
ZebraLab	Viewpoint	ZebraLab	Behavior software