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Open Access Evaluation of Rapid Cooling and Tricaine Methanesulfonate (MS222) as Methods of Euthanasia in Zebrafish (Danio rerio)

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Despite the progressively increasing use of zebrafish (Danio rerio) in research, the most humane method of euthanasia for these fish has not been determined. Contemporary guidance documents state that hypothermia is an unacceptable method of euthanasia. The goal of this study was to compare rapid cooling and tricaine methanesulfonate (MS222) for zebrafish euthanasia. Zebrafish (n = 46) were euthanized by immersion in either an ice-water (4 °C or less) bath or unbuffered MS222 solution (250 mg/L; 25 to 30 °C). Another cohort (n = 10) was exposed to buffered MS222 to determine whether the acidity of the water alone caused distress. The times from exposure until the animals became unable to swim, right themselves, and death were measured, and signs of distress were recorded. Fish then were placed in a 'recovery tank' of system water to verify that recovery did not occur. Tissues were examined histologically. The mean time for euthanasia was longer and the number of fish exhibiting signs of distress was greater for fish exposed to MS222 than those exposed to chilled water. In addition, 4 of the 23 fish exposed to MS222 regained consciousness in the recovery tank, whereas none of 23 fish exposed to chilled water recovered. No histopathologic changes or evidence of ice crystal formation were seen in either group. In light of the faster time to death and fewer signs of distress in zebrafish euthanized by rapid cooling, we advocate this method as a humane veterinary practice.

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Document Type: Research Article

Affiliations: University Laboratory Animal Resources, University of Pennsylvania, Philadelphia, Pennsylvania, USA

Publication date: November 1, 2009

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  • The Journal of the American Association for Laboratory Animal Science (JAALAS) serves as an official communication vehicle for the American Association for Laboratory Animal Science (AALAS). The journal includes a section of refereed articles and a section of AALAS association news. The mission of the refereed section of the journal is to disseminate high-quality, peer-reviewed information on animal biology, technology, facility operations, management, and compliance as relevant to the AALAS membership. JAALAS accepts research reports (data-based) or scholarly reports (literature-based), with the caveat that all articles, including solicited manuscripts, must include appropriate references and must undergo peer review.

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