Conditioned Place Avoidance of Zebrafish (Danio rerio) to Three Chemicals Used for Euthanasia and Anaesthesia

Abstract

Zebrafish are becoming one of the most used vertebrates in developmental and biomedical research. Fish are commonly killed at the end of an experiment with an overdose of tricaine methanesulfonate (TMS, also known as MS-222), but to date little research has assessed if exposure to this or other agents qualifies as euthanasia (i.e. a “good death”). Alternative agents include metomidate hydrochloride and clove oil. We use a conditioned place avoidance paradigm to compare aversion to TMS, clove oil, and metomidate hydrochloride. Zebrafish (n = 51) were exposed to the different anaesthetics in the initially preferred side of a light/dark box. After exposure to TMS zebrafish spent less time in their previously preferred side; aversion was less pronounced following exposure to metomidate hydrochloride and clove oil. Nine of 17 fish exposed to TMS chose not to re-enter the previously preferred side, versus 2 of 18 and 3 of 16 refusals for metomidate hydrochloride and clove oil, respectively. We conclude that metomidate hydrochloride and clove oil are less aversive than TMS and that these agents be used as humane alternatives to TMS for killing zebrafish.

The efficacy and effect of repeated exposure to 2-phenoxyethanol, clove oil and tricaine methanesulphonate as anesthetic agents on juvenile Angelfish (Pterophyllum scalare)

“…Moreover, from a welfare perspective, it causes less aversion than MS-222 in D. rerio (Wong et al, 2014). For these reasons it should be considered as an anesthetic option for juvenile angelfish.…”

Effects of repeated anaesthesia on gill and general health of Atlantic salmon, Salmo salar

“…() reported that freshwater S. salar smolts also had no rise in plasma glucose after a single dose of metomidate (up to 10 mg l −1 ) or AQUI‐S (up to 100 mg l −1 active ingredient). However, behavioural (Pirhonen & Schreck, ; Readman et al ., ; Wong et al ., ) and blood glucose stress responses (Bourne, ; Davis & Griffin, ; Larter & Rees, ; Molinero & Gonzalez, ; Small & Chatakondi, ; Soivio et al ., ; Thomas & Robertson, ; Wagner et al ., ) have been reported in a variety of fish species exposed to a single dose of the same anaesthetics tested in this study. While the role of in‐tank anaesthesia in the reduction of stress has yet to be quantified, it seems likely this methodology plays an important role in minimizing any stress response.…”

Ethical considerations in fish research

“…() argue that in addition to assessing the ease of delivery and rate and stability of anaesthesia, researchers should also consider the way a fish responds to anaesthesia when initially exposed to the chemical. Danio rerio demonstrate a negative behavioural response when exposed to benzocaine and MS‐222 (Readman et al ., ) and appear to remember exposure to some anaesthetics (Wong et al ., ). Medaka Oryzias latipes (Temminck & Schlegel 1846) also show a negative behavioural response to these chemicals, whereas carp Cyprinus carpio L. 1758 and O. mykiss do not (Readman et al ., ).…”

“…Readman et al (2017) argue that in addition to assessing the ease of delivery and rate and stability of anaesthesia, researchers should also consider the way a fish responds to anaesthesia when initially exposed to the chemical. Danio rerio demonstrate a negative behavioural response when exposed to benzocaine and MS-222 (Readman et al, 2013) and appear to remember exposure to some anaesthetics (Wong et al, 2014) Mettam et al, 2011;Newby et al, 2006Newby et al, , 2009Nordgreen et al, 2009;Rizzo et al, 2017;Schroeder & Sneddon, 2017;Sneddon, 2003b;Taylor et al, 2017). Where not effective is stated these drugs did not prevent the changes in behaviour associated with treatment with a noxious stimulus.…”