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Assessment of distress associated with carbon dioxide euthanasia in laboratory rats Niel, Lee Erin


Carbon dioxide (CO₂) gas is the most widely used euthanasia agent for laboratory rodents. However, it has the potential to cause both pain and dyspnea [i.e. dyspnœa], an unpleasant sensation of breathless, while animals are still conscious. The aims of this dissertation were to determine whether gradual-fill CO₂ euthanasia causes distress in laboratory rats, and to examine potential sources of distress, including pain, dyspnea and novelty. The first study examined the behavioural responses of rats during gradual-fill CO₂ euthanasia. Rats showed increased exploratory behaviours and escape behaviours during CO₂ euthanasia, suggesting that this procedure does cause distress. The second and third studies used approach-avoidance testing to investigate aversion to CO₂ in rats, by examining their willingness to enter a test cage containing CO₂ for access to an attractive food reward. Rats were found to avoid CO₂ concentrations that are sufficient to cause unconsciousness. Specifically, when tested with static concentrations of CO₂ rats showed avoidance at 15% CO₂ and greater, and when tested with gradually increasing concentrations of CO₂ at flow rates ranging from 3 to 27% per minute, rats showed avoidance at 13 to 16% CO₂. This avoidance indicates that rats are at least moderately averse to CO₂ concentrations occurring during gradual-fill CO₂ euthanasia, and that forced exposure likely causes distress. Concentrations of CO₂ that were associated with behavioural responses and aversion were not consistent with previous data on pain due to CO₂. However, similar concentrations have been shown to cause dyspnea in humans. The final study examined the role of novelty in rats’ responses to CO₂, and found that novelty was not a major source of distress during gradual-fill CO₂ euthanasia. In summary, these studies suggest that gradual-fill CO₂ euthanasia causes distress in rats, and that this distress is likely due to dyspnea. Further research is necessary to examine the effects of CO₂ on other rodent species such as mice, and to identify alternative methods of euthanasia that cause unconsciousness without distress.

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