Name: accumulation and distribution of fluorescent microplastics in the early life stages of zebrafish
Uploaded: 2021-07-04T13:00:00
Description: Watch this Scientific Journal Video about Accumulation and Distribution of Fluorescent Microplastics in the Early Life Stages of Zebrafish at JoVE.com

This work was funded by the National Natural Science Foundation of China (21777145, 22076170), and the Program for Changjiang Scholars and Innovative Research Team in University (IRT_17R97).

Adult zebrafish are originated from the China Zebrafish Resource Center (Wuhan, China). The experiments were conducted in compliance with the national guide &#34;Laboratory Animal Guideline for Ethical Review of Animal Welfare (GB/T35892-2018).
1. Embryo collection
<ol>
	<li>Maintain fish in 20 L glass tanks with recirculating charcoal-filtered tap water system (pH 7.0 &#177; 0.2) at a constant temperature (28 &#177; 0.5 &#176;C) on a photoperiod of 14:10 h light: dark.</li>
	<li>Feed fish t...

<ol>
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According to the guideline on the protection of animals used for scientific purposes, such as EU Directive 2010/63/EU, animal ethics permission is not mandatory for an experiment with early life-stages of zebrafish until the stage of being capable of independent feeding (5 days post fertilization)17. However, best welfare practice is important for optimizing the use of zebrafish, and, for example, the humane methods of anesthesia and euthanasia should be of concern. Ethyl 3-aminobenzoate methanesu...

Since first synthesized in the 1900s, plastics are widely used in various fields, resulting in rapid growth of global production1. In 2018, approximately 360 million tons of plastics were produced worldwide2. The plastics in the natural environment will degrade to fine particles due to chemical, physical or biological processes3. Generally, fine plastic particles &lt;5 mm in size are defined as microplastics4. Microplastics are also engineered for specific applications, such as microbeads from cosmetic products5. As near-permanent contaminants, microplastics are accumulated in the environment, and have attracted increasing attention from scientists, policymakers and the public1,6. Previous studies documented that microplastics could cause adverse effects in fish, such as gastrointestinal damage7, neurotoxicity8, endocrine disruption9, oxidative stress10 and DNA damage11. However, the toxicity of microplastics has not been fully revealed so far12,13.
Zebrafish embryos offer a lot of experimental advantages, including small size, external fertilization, optical transparency and large clutches, and is considered as an ideal model organism for in vivo studying the effects of pollutants on fish at early life stages. In addition, only limited amounts of test substances are needed for the evaluation of biological responses. Here, zebrafish embryos are exposed to different concentrations of microplastics (0.1, 1, 10 mg/L) for 5 days, and the bioaccumulation and distribution of microplastics in zebrafish embryos/larvae are evaluated. This result will advance our understanding about the toxicity of microplastics to fish, and the method described here can potentially be generalized to determine the accumulation and distribution of other types of fluorescent materials in the early life stages of zebrafish.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Fluorescent microscope</td>
			<td>Nikon, Japan</td>
			<td>Eclipse Ti-S</td>
			<td></td>
		</tr>
		<tr>
			<td>Green fluorescently labeled polystyrene beads</td>
			<td>Phosphorex, USA</td>
			<td>2103A</td>
			<td></td>
		</tr>
		<tr>
			<td>Tricaine</td>
			<td>Sigma-Aldrich, USA</td>
			<td>A5040</td>
			<td></td>
		</tr>
	</tbody>
</table>

accumulation and distribution of fluorescent microplastics in the early life stages of zebrafish

As a new type of environmental pollutant, microplastic has been widely found in the aquatic environment and poses a high threat to aquatic organisms. The bioaccumulation of microplastics plays a key role in their toxic effects; however, as a particulate, their bioaccumulations are different from many other pollutants. Described here is a feasible method to visually determine the accumulation and distribution of microplastics in zebrafish embryos or larvae using fluorescent microplastics. Embryos are exposed to different concentrations (0.1, 1, and 10 mg/L) of fluorescent microplastics with a diameter of 500 nm for 120 h. It is shown in the results that microplastics can bioaccumulate in zebrafish embryos/larvae in a concentration-dependent manner. Before hatching, strong fluorescence is found around the embryonic chorion; while in zebrafish larvae, the yolk sac, pericardium, and gastrointestinal tract are the main accumulated sites of microplastics. The results demonstrate the uptake and internalization of microplastics in zebrafish at early life stages, which will provide basis for better understanding the impact of microplastics on aquatic animals.

The distribution and accumulation of fluorescent microplastics are shown in Figure 1 and Table 1. No visible fluorescence is observed in the unexposed group (control). However, an accumulation of fluorescence is found surrounding the chorion after exposure to different concentrations of microplastics (24 hpf). Green fluorescence is also detected in larvae, and the fluorescence levels appear to increase in a concentration- and time-dependent manner. The yolk sac, pericardium,...

Watch this Scientific Journal Video about Accumulation and Distribution of Fluorescent Microplastics in the Early Life Stages of Zebrafish at JoVE.com

Accumulation and Distribution of Fluorescent Microplastics in the Early Life Stages of Zebrafish

Zebrafish embryos/larvae develop externally and are optically transparent. The bioaccumulation of microplastics in fish at early life stages is readily assessed with fluorescently labeled microbeads.

As a new type of environmental pollutant, microplastic has been widely found in the aquatic environment and poses a high threat to aquatic organisms. The ...

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