Advancing understanding of anaesthesia and analgesia in the zebrafish

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The zebrafish (Danio rerio) is used widely as a model for understanding biological processes in development and various human diseases. Features of embryonic and larval zebrafish (embryo-larvae, <10 days post fertilisation, or dpf) that make them highly attractive as a research model include their optical transparency for visualising biological processes and ease of genetic manipulation. Consequently, each year millions of embryo-larval zebrafish are used in laboratories across the globe. Many of these animals undergo anaesthesia, however, very little information is available to guide appropriate anaesthetic choice, and even less information is available on analgesics (pain relievers). Our project will address these knowledge gaps and has 3 main aims. Aim 1) Will establish the concentrations of anaesthetics needed to induce sedation and euthanasia, and the concentrations of analgesics needed to provide pain relief. For this we will use functional brain imaging (via fluorescent dyes that reveal the brain's neural activity) to test the responsiveness of animals to stimulation whilst under treatment. Electrophysiology (direct recording of the brain's electrical activity) will be used to establish concentrations of anaesthesia required for euthanasia. We will also gather data to understand if traditional indicators of sedation in fish are appropriate (e.g. loss of balance), and measure compound uptake to support interpretation of compound exposure effect concentrations. Aim 2) Will determine if any of the anaesthetics inhibit movement, if any of the anaesthetics and analgesics are aversive, and if any of the analgesics reduce the effects of noxious stimulation (e.g. heat, CO2). For this we will assess behaviour such as the effect of treatment on locomotion, and the avoidance of areas within a test arena where an anaesthetic or analgesic has been deposited. In addition, we will measure the ability of analgesics to decrease the behavioural impact of noxious stimuli as an indicator of their pain relieving properties. Aim 3) Will provide a better understanding of the mechanisms of action of the chosen anaesthetics and analgesics, and of sedation and pain perception in the embryo-larval zebrafish. For this we will undertake functional brain imaging in genetically-modified animals in which we are able to visualise activity in specific types of neuron in the brain. This will allow us to understand which brain regions and cell types respond to treatment with each agent, and which regions/cell types are involved in the sedative process. We will also apply this approach to animals subjected to noxious stimulation to better understand pain perception in fish. This project will allow scientists to select the most scientifically and ethically appropriate anaesthetic and analgesic for use in their work and will provide a better understanding of how these agents work, as well as advancing knowledge on the processes of sedation and pain perception in fish. Beneficiaries will include veterinary and aquarium fish industries as these data will help refine practices such as how best to monitor anaesthesia in fish, and inform on treatments most appropriate for animal transportation. Government bodies such as the UK Home Office, who oversee the scientific use of animals, will benefit through the provision of high quality reference data to use in decision making on animal experimentation practices, and charities ,such as those associated with neurological disorders (e.g. epilepsy, Alzheimer's, Parkinson's), will be provided with further validation data on an alternative (and more cost effective) model species. The public and society in general will benefit from an improvement in animal welfare standards, a greater understanding of sedation and pain perception, and an increase in our understanding of the biology of a species increasingly being used in experimental, including biomedical, research.