Thirty-Second Net Stressor Task in Adult Zebrafish
成年斑马鱼30秒应激实验   

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Behavioral Neuroscience
Oct 2014

 

Abstract

Zebrafish have become a popular animal model for behavioral neuroscience (Gerlai, 2014). Recent studies have demonstrated that brief experimental handling prior to euthanizing animals can subsequently alter biological measures quantified post-mortem (e.g. cortisol levels) (Ramsay et al., 2009; Tran et al., 2014). Here we provide a detailed protocol for a simple 30-sec net stressor task for adult zebrafish that increases whole-body cortisol levels without altering the levels of whole-brain dopamine, 3, 4-dihydroxyphenylacetic acid, serotonin, and 5-hydroxyindoleacetic acid (Tran et al., 2014).

Materials and Reagents

  1. Adult zebrafish
  2. System water: Reverse osmosis purified water supplemented with 60 mg/L instant ocean sea salt
  3. 2.8 L zebrafish housing tank and lid (Aquaneering, catalog number: ZT280 )

Equipment

  1. Fish net (soft material)
  2. Timer
  3. Zebrafish housing (single sided, five shelf stand alone rack) (Aquaneering)

Procedure

  1. Adult zebrafish were individually isolated in 2.8 L housing tanks and placed on a stand alone rack system with filtration.
  2. Zebrafish were allowed to habituate to being in isolation for 7 days prior to the stressor (Tran et al., 2014).
  3. On the day of the stressor, the housing tank was moved to the testing room and zebrafish were allowed to acclimatize to the new environment for 1 h.
  4. Zebrafish were netted out of the tank using a fish net and suspended above the tank and fully exposed in air for 30 sec, with the lid covering the net.
  5. Following the 30 sec net stressor, zebrafish were placed back in their housing tanks.
  6. Zebrafish behavioral responses can be recorded and/or tissue samples can be collected after 15 min post-stressor,.

Representative data



Figure 1. Note that the fish is exposed to air in the net held in place by the tank lid on top

Notes

Zebrafish were isolated because the stressor and subsequent behavioral recordings are conducted on individual zebrafish. In the case of multiple days of testing, it is recommended that the stressor be applied at the same time of the day to minimize the effects of circadian rhythms on endogenous cortisol levels.

Acknowledgments

This protocol was adapted from our previous publication: Tran et al. (2014). This project was funded by an NSERC Discovery grant (#311637) issued to R. G.

References

  1. Gerlai, R. (2014). Fish in behavior research: unique tools with a great promise! J Neurosci Methods 234: 54-58.
  2. Ramsay, J. M., Feist, G. W., Varga, Z. M., Westerfield, M., Kent, M. L. and Schreck, C. B. (2009). Whole-body cortisol response of zebrafish to acute net handling stress. Aquaculture 297(1-4): 157-162.
  3. Tran, S., Chatterjee, D. and Gerlai, R. (2014). Acute net stressor increases whole-body cortisol levels without altering whole-brain monoamines in zebrafish. Behav Neurosci 128(5): 621-624.

简介

斑马鱼已成为行为神经科学的流行动物模型(Gerlai,2014)。 最近的研究已经证明,在安乐死动物之前的短暂的实验操作可以随后改变生后量化的生物学测量(例如皮质醇水平)(Ramsay等人,2009; em> et al。,2014)。 在这里我们提供一个简单的30秒净应激子任务的成年斑马鱼,增加全身皮质醇水平,而不改变全脑多巴胺,3,4-二羟基苯乙酸,血清素和5-羟基吲哚乙酸的水平的简单协议 Tran et al。,2014)。

材料和试剂

  1. 成人斑马鱼
  2. 系统水:补充有60mg/L即时海洋海水的反渗透净化水
  3. 2.8 L斑马鱼容器罐和盖(Aquaneering,目录号:ZT280)

设备

  1. 鱼网(软材料)
  2. 计时器
  3. 斑马鱼壳(单面,五架独立架)(水中)

程序

  1. 成年斑马鱼单独隔离在2.8升容纳罐中,并放置在单独的架系统中过滤
  2. 斑马鱼被允许习惯于在应激前隔离7天(Tran等人,2014)。
  3. 在紧张性刺激的当天,将容纳槽移动到测试室,使斑马鱼适应新环境1小时。
  4. 使用鱼网将斑马鱼从网箱中取出并悬挂在罐上,并在空气中完全暴露30秒,盖子覆盖网。
  5. 在30秒净应激后,斑马鱼被放回他们的容器中。
  6. 可以记录斑马鱼行为反应和/或在应激后15分钟后收集组织样品。

代表数据



图1.请注意,鱼被暴露在网上的空气中,通过顶部的罐盖固定到位

笔记

斑马鱼被隔离,因为应激者和随后的行为记录是对个体斑马鱼进行。 在多天测试的情况下,建议应激者在一天的同一时间应用,以最小化昼夜节律对内源皮质醇水平的影响。

致谢

此协议改编自我们先前的出版物: 等人(2014)。 该项目由颁发给R. G.的NSERC发现补助金(#311637)资助。

参考文献

  1. Gerlai,R。(2014)。 鱼行为研究:独特的工具,带来巨大的希望! J Neurosci 方法 234:54-58。
  2. Ramsay,J.M.,Feist,G.W.,Varga,Z.M.,Westerfield,M.,Kent,M.L.and Schreck,C.B。(2009)。 全身皮质醇对斑马鱼对急性净处理压力的反应。水产养殖 297(1-4):157-162。
  3. Tran,S.,Chatterjee,D。和Gerlai,R。(2014)。 急性净应激物增加全身皮质醇水平,而不改变斑马鱼中的全脑单胺。 Behav Neurosci 128(5):621-624。
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引用:Tran, S. and Gerlai, R. (2015). Thirty-Second Net Stressor Task in Adult Zebrafish. Bio-protocol 5(5): e1413. DOI: 10.21769/BioProtoc.1413.
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