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Analysis of Nicotinic Acetylcholine Receptor (nAChR) Gene Expression in Zebrafish (Danio rerio) by In Situ Hybridization and PCR

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Nicotinic Acetylcholine Receptor Technologies

Part of the book series: Neuromethods ((NM,volume 117))

Abstract

Zebrafish (Danio rerio) have been established as an ideal model animal to study neural development, with several advantages over mice, rats, humans, or in vitro cell-based work. Zebrafish embryos develop externally and can either be genetically (using casper or albino strains) or pharmacologically (PTU) manipulated to be transparent, which persists throughout adulthood. Optical transparency allows the localization of gene expression in whole animals with relative ease. Many strains are available, including transgenic fish expressing a number of fluorescent markers in cell-specific manners. A large number of fish can be raised inexpensively, and all stages are free swimming and can be exposed to drugs for high-throughput screening. A number of behavioral assays used in mice, such as those used to test conditioned place preference (CPP), locomotor function, and anxiety, are available with zebrafish.

Zebrafish express a family of muscle and neural nicotinic acetylcholine receptor (nAChR) genes. Our laboratory has cloned α2a, α3, α4, α6, α7, β2, β3a, and β4 neuronal cDNAs, although genomic analysis indicates others exist as well. Several zebrafish nAChR subtypes have been expressed in Xenopus oocytes and we have shown that some zebrafish nAChR subtypes share similar pharmacological properties with those of mice, rat, and human nAChRs.

Given the many advantages of the zebrafish to study nAChRs, it is important to analyze the expression of nAChRs in developing zebrafish, as well as their expression in pharmacologically manipulated animals or in disease models. We describe how we have used in situ hybridization, polymerase chain reaction (PCR), and rapid amplification of cDNA ends (RACE) cloning identify and study zebrafish neuronal nAChR expression.

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Acknowledgements

We would like to thank Dr. Christine Beattie, the late Dr. Paul Henion along with the members of the Henion and Beattie labs at The Ohio State University for their support over the years. We would also like to thank Dr. Jeff Zirger, a former member of the lab, for his early work identifying nAChRs in zebrafish.

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Authors and Affiliations

  1. Department of Biology, High Point University, High Point, NC, 27268, USA

    Kristin M. Ackerman

  2. Department of Neuroscience, Wexner Medical Center, The Ohio State University College of Medicine, 333 West Tenth Avenue, 4198 Graves Hall, Columbus, OH, 43210, USA

    R. Thomas Boyd

Authors
  1. Kristin M. Ackerman
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  2. R. Thomas Boyd
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Corresponding author

Correspondence to R. Thomas Boyd .

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Editors and Affiliations

  1. University of Virginia, Charlottesville, Virginia, USA

    Ming D. Li

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Ackerman, K.M., Boyd, R.T. (2016). Analysis of Nicotinic Acetylcholine Receptor (nAChR) Gene Expression in Zebrafish (Danio rerio) by In Situ Hybridization and PCR. In: Li, M. (eds) Nicotinic Acetylcholine Receptor Technologies. Neuromethods, vol 117. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3768-4_1

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  • DOI: https://doi.org/10.1007/978-1-4939-3768-4_1

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3766-0

  • Online ISBN: 978-1-4939-3768-4

  • eBook Packages: Springer Protocols

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