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. 2012;7(9):e44437.
doi: 10.1371/journal.pone.0044437. Epub 2012 Sep 7.

Evaluation of EMLA cream for preventing pain during tattooing of rabbits: changes in physiological, behavioural and facial expression responses

Affiliations

Evaluation of EMLA cream for preventing pain during tattooing of rabbits: changes in physiological, behavioural and facial expression responses

Stephanie C J Keating et al. PLoS One. 2012.

Abstract

Background: Ear tattooing is a routine procedure performed on laboratory, commercial and companion rabbits for the purpose of identification. Although this procedure is potentially painful, it is usually performed without the provision of analgesia, so compromising animal welfare. Furthermore, current means to assess pain in rabbits are poor and more reliable methods are required. The objectives of this study were to assess the physiological and behavioural effects of ear tattooing on rabbits, evaluate the analgesic efficacy of topical local anaesthetic cream application prior to this procedure, and to develop a scale to assess pain in rabbits based on changes in facial expression.

Methodology/principal findings: In a crossover study, eight New Zealand White rabbits each underwent four different treatments of actual or sham ear tattooing, with and without prior application of a topical local anaesthetic (lidocaine/prilocaine). Changes in immediate behaviour, heart rate, arterial blood pressure, serum corticosterone concentrations, facial expression and home pen behaviours were assessed. Changes in facial expression were examined to develop the Rabbit Grimace Scale in order to assess acute pain. Tattooing without EMLA cream resulted in significantly greater struggling behaviour and vocalisation, greater facial expression scores of pain, higher peak heart rate, as well as higher systolic and mean arterial blood pressure compared to all other treatments. Physiological and behavioural changes following tattooing with EMLA cream were similar to those in animals receiving sham tattoos with or without EMLA cream. Behavioural changes 1 hour post-treatment were minimal with no pain behaviours identifiable in any group. Serum corticosterone responses did not differ between sham and tattoo treatments.

Conclusions: Ear tattooing causes transient and potentially severe pain in rabbits, which is almost completely prevented by prior application of local anaesthetic cream. The Rabbit Grimace Scale developed appears to be a reliable and accurate way to assess acute pain in rabbits.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist. The commercial organisation Harlan UK donated the animals used in the study. This does not alter the authors’ adherence to all the PLOS ONE policies on sharing data and materials.

Figures

Figure 1
Figure 1. Change in mean heart rate from pre to post-procedure.
Change in mean heart rate (beats per minute ±1 SE) from pre to post-procedure observed in rabbits (y-axis) when undergoing the various treatments (x-axis); sham tattooing with and without prior application of EMLA and tattooing with and without prior application of EMLA (n = 8 per treatment) (P = 0.022, ★★P = 0.031, ★★★P = 0.004).
Figure 2
Figure 2. Change in mean systolic arterial pressure (SAP) from pre to post-procedure.
Change in mean systolic arterial pressure (mmHg ±1 SE) from pre to post-procedure observed in rabbits (y-axis) when undergoing the various treatments (x-axis); sham tattooing with and without prior application of EMLA and tattooing with and without prior application of EMLA (n = 8 per treatment) (P = 0.024, ★★P = 0.028, ★★★P = 0.003).
Figure 3
Figure 3. Change in mean arterial pressure (MAP) from pre to post-procedure.
Change in mean arterial pressure (mmHg ±1SE) from pre to post-procedure observed in rabbits (y-axis) when undergoing the various treatments (x-axis); sham tattooing with and without prior application of EMLA and tattooing with and without prior application of EMLA (n = 8 per treatment) (P = 0.05, ★★P = 0.035, ★★★P = 0.01).
Figure 4
Figure 4. Mean serum corticosterone concentration from pre to post-procedure.
Mean corticosterone concentration (ng/ml ±1SE) observed in rabbits (y-axis) at baseline and various post-procedure time points (x-axis) when undergoing the various treatments; sham tattooing with and without prior application of EMLA and tattooing with and without prior application of EMLA (n = 8 per treatment) (Comparison to baseline: P<0.05, ★★P<0.01, ★★★P<0.001, Comparison between treatments: +P = 0.05, ++P = 0.01).
Figure 5
Figure 5. Mean Rabbit Grimace Scale scores pre and intra procedures.
Mean RbtGS scores (±1SE) observed in rabbits (y-axis) at baseline and intra-procedure (x-axis) when undergoing the various treatments; sham tattooing with and without prior application of EMLA and tattooing with and without prior application of EMLA (n = 8 per treatment) (P = 0.018, ★★P = 0.002, ★★★P = 0.001, ★★★★P = 0.000).
Figure 6
Figure 6. Example images and RbtGs scores.
Example images and associated RbtGS scores of a rabbit in the pre-tattoo (a) and intra-tattoo (b) periods without prior application of EMLA and of a rabbit in the pre-sham tattoo (c) and intra-sham tattoo (d) periods without prior application of EMLA.
Figure 7
Figure 7. Rabbit Grimace Scale (RbtGS).
The Rabbit Grimace Scale with images and explanations for each of the 5 facial action units (FAU); orbital tightening, cheek flattening, nose shape, whisker position and ear position. Each FAU is scored according to whether it is not present (score of 0), moderately present (score of 1) and obliviously present (score of 2).

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