Characterising a working behavioural model to chronically stress zebrafish subjects and optimising brain RNA extraction in preparation for RNA-Seq analysis.

• Main Author: Siddharth Janarthanan.
• Other Authors: School of Biological Sciences
• Format: Final Year Project
• Language: English
• Published: 2013
• Subjects: DRNTU::Science::Biological sciences
• Online Access: http://hdl.handle.net/10356/53811
• Institution: Nanyang Technological University

The zebrafish has been established to be a good model to study the effects of fear and stress on the brain. It has been documented that events such as exposure to nicotine, netting, restraining in small spaces or exposure to air induce stress in fish. In addition, alarm signal molecules released from skin of injured fish (Schreckstoff) naturally elicit innate fear behaviour. This project attempted to characterise a working behavioural paradigm to induce chronic stress in fish. Two methods were used to induce stress – a variable, unpredictable chronic stress protocol in the form of netting, nicotine exposure, small volume restraining and Hypoxanthine-3-N-Oxide exposure and a predictable chronic stress protocol by Schreckstoff exposure. The stress levels were quantified by physiological measurements of HPI axis (Hypothalamus-Pitutary-Interrenal) activation and semi-quantitative genetic analyses of brain tissue RNA samples. A behavioural analysis by video tracking was also performed on a pair of stressed and control fish to serve as an example. Both stress models resulted in elevated cortisol levels and apparent increase in expression of stress-related gene Egr 1 as observed by semi-quantitative analysis by PCR. This project serves as a preparatory setup for further genetic and epigenetic comparative analysis of whole brain transcriptome of stressed fish as compared to control fish by RNA-seq.