A Multi-Site Assessment of Anesthetic Overdose, Hypothermic Shock, and Electrical Stunning as Methods of Euthanasia for Zebrafish (Danio rerio) Embryos and Larvae.

Euthanasia in zebrafish (Danio rerio) younger than 5 days post fertilization (dpf) is poorly described in the literature, and standardized protocols are lacking, most likely because larvae not capable of independent feeding are often not protected under national legislations. We assessed the euthanasia efficacy in laboratories in different countries of a one hour anesthetic overdose immersion with buffered lidocaine hydrochloride (1 g/L, with or without 50 mL/L of ethanol), buffered tricaine (1 g/L), clove oil (0.1%), benzocaine (1 g/L), or 2-phenoxyethanol (3 mL/L), as well as the efficacy of hypothermic shock (one hour immersion) and electrical stunning (for one minute), on zebrafish at <12 h post fertilization (hpf), 24 hpf, and 4 dpf. Based on the survival/recovery rates 24 h after treatment, the most effective methods were clove oil, lidocaine with ethanol, and electrical stunning. For 4 dpf larvae, signs of aversion during treatment demonstrated that all anesthetics, except lidocaine, induced aversive behavior. Therefore, the most suited euthanasic treatment was lidocaine hydrochloride 1 g/L, buffered with 2 g/L of sodium bicarbonate and mixed with 50 mL/L of ethanol, which euthanized both embryos and larvae in an efficient and stress-free manner. Electrical stunning also euthanized embryos and larvae efficiently and without signs of aversion; this method needs further assessment in other laboratories to draw firm conclusions.

Anaesthesia of laboratory, aquaculture and ornamental fish: Proceedings of the first LASA-FVS Symposium.

Following on from the Annual Fish Veterinary Society Conference, this symposium was organised with the Laboratory Animal Science Association and brought together experts from ornamental (pond and aquarium) fish practice, aquaculture and aquatic-research facilities to discuss good practice of anaesthesia. This proceedings paper gives an overview of relevant experiences involving a range of immersion drugs including tricaine, benzocaine and isoeugenol, as well as a summary of the main topics of discussion. While fish anaesthesia is commonplace, administration methods, drugs and monitoring procedures may often be regarded as antiquated when compared with mammalian practice. These limitations notwithstanding, individual fish will benefit from good anaesthetic monitoring. Although the most common anaesthetic drugs may be perceived as equally efficacious and therefore interchangeable for different settings, challenges are different for the anaesthesia of grouped fish, when determining species-dependent anaesthetic dosing in a multi-species tank, or adapting to farming requirements, nationally licensed products, costs and withdrawal periods. The fish anaesthetic arsenal fails to address premedication, analgesia and issues of averseness. The two latter factors should be part of the evaluation of anaesthetic protocols; therefore, instructions for the analgesic provision of lidocaine to fin clipped zebrafish are proposed. Euthanasia practices could sometimes be refined too. Alternative physical methods such as electrical stunning are options to be considered.