An optogenetic platform for high-throughput interrogation of cellular basis of physiological function in zebrafish

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We will take advantage of recent technological breakthroughs that now enable cellular function to be directly linked to animal physiology and behaviour. Determining the outcome of cell activity in multicellular organisms traditionally has been hampered by an inability to directly manipulate cells of interest, particularly in living animals. Most methods of studying cell function such as pharmacological agents or electrical stimulation are flawed owing to widespread off-target effects. Therefore, our ability to understand how cell function relates to physiology and behaviour is limited using these techniques. These limitations can now be overcome through the use of proteins that respond to light of specific wavelengths. Illuminating cells expressing these light-sensitive proteins can instantly shape the activity of the cells, while leaving other cells unaffected. By genetically controlling which cells express the light-sensitive molecules, an approach termed optogenetics, we can directly influence cell activity. The approach is particularly powerful in species with life stages where animals are transparent, so that light is readily transmitted to living tissue, as is the case for larval zebrafish. Optogenetics studies in zebrafish where changes in physiology or behaviour can be directly attributed to light-induced changes in cell activity have led to important advances in our understanding of animal physiology. A multidisciplinary team of researchers at the University of Ottawa with expertise in studying cell function and animal physiology and behaviour in zebrafish will assemble an optogenetics system specialized for the study of zebrafish. This system will be comprised of two platforms. The first platform consists of a microscope and a light patterning device to enable the selective stimulation of targeted cells within specific regions of zebrafish larvae. The outcome of optogenetic stimulation will be monitored by a camera sufficiently fast and sensitive to detect even minute and rapid changes in cell signalling. This platform will allow us to study the role of cell activity on animal physiology at the microscopic level. A second platform will allow simultaneous observation and analysis of the behavioral consequences of optogenetic activation in up to 96 larval zebrafish at a time, dramatically increasing research output. Another component of this platform will enable the study of how optogenetic stimulation at larval stages shapes behaviour in adult stages. The proposed system will allow researchers to make fundamental insights into the roles of specific cell types on many important biological processes. Optogenetics is rapidly become a standard technique for physiological studies and our HQP will be provided with an invaluable opportunity for the cutting edge training needed for fields such as biology, biochemistry, biomedical engineering, ecology, and physiology.