FIGURE SUMMARY
Title

A suitable anaesthetic protocol for metamorphic zebrafish

Authors
Owen, J.P., Kelsh, R.N.
Source
Full text @ PLoS One

Standard length (SL) as described by Parichy <italic>et al</italic> [<xref rid='pone.0246504.ref027' ref-type='bibr'>27</xref>].

SL is the distance from the snout to the caudal peduncle. In pre-flexion larvae that do not have a caudal penduncle, SL is the distance from the snout to the posterior tip of the notochord.

Metamorphic fish become sensitive to MS-222 around 10mm SL and later become de-sensitised around 24mm SL.

(A), (C), (E) Percentage of fish that recovered by length for concentrations of 0.008%, 0.009% and 0.01% MS-222 respectively. (B), (D), (F) Percentage of fish that were not MUA for concentrations of 0.008%, 0.009% and 0.01% MS-222 respectively. Black lines represent the 90% confidence interval for each value, calculated using the sample size (see materials and methods).

Protocols 1, 4, 5 and 6 vary in the time taken to induce, lose touch responsiveness, regain movement, regain equilibrium as well as respiratory rate.

Boxplots of the time taken to (A) induce anaesthesia, (B) lose touch responsiveness, (C) regain movement post anaesthesia and (D) regain equalibrium post anaesthesia for each protocol 1–6 as defined in Table 1. (E) Boxplot of the respiratory rate (breaths per minute—bpm) at 5 minutes for each protocol. Stars indicate significant difference of subsequent repeats as determined using the Mann-Whitney U-test. ns = not significant, ‘*’ indicates p<0.05, ‘**’ indicates p<0.01, ‘***’ indicates p < 0.001 and ‘****’ indicates p<0.0001.

Growth of the fish is not significantly impacted by repeated dosing using any of the 4 protocols.

Barchart with 95% confidence intervals for age vs length (SL (mm)) of fish anaesthetised using protocols 1,4,5 and 6 respectively.

Protocols 1, 4, 5 and 6 vary in the average time taken to induce, lose touch responsiveness, regain movement, regain equilibrium as well as respiratory rate when repeatedly anaesthetised every 4 days.

Barplot of the mean (A) induction time, (B) time taken to lose touch responsivity, (C) heartbeats per minute at 5:00, (D) visible breaths per minute at 5:00, (E) time taken to regain movement and (F) time taken to fully recover against repeat number for each protocol 1,4,5 and 6. Mean taken over all repeats. P/L corresponds to Propofol/Lidocaine, PE corresponds to phenoxyethanol, CP corresponds to Cotton Pad maintenance.

The time taken to induce, lose touch responsiveness, regain movement, regain equilibrium as well as respiratory rate varies within an acceptable range for fish repeatedly anaesthetised using protocol 4 every 2 days.

Boxplot of (A) induction time, (B) time taken to lose touch responsiveness, (C) time taken to regain movement and (D) time taken to recover. Stars indicate significant difference of subsequent repeats as determined using the Mann-Whitney U test. ns = not significant, ‘*’ indicates p<0.05, ‘**’ indicates p<0.01, ‘***’ indicates p < 0.001 and ‘****’ indicates p<0.0001.

Acknowledgments
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