Systematic Assessment of Well-Being in Mice for Procedures Using General Anesthesia

Abstract

In keeping with the 3R Principle (Replacement, Reduction, Refinement) developed by Russel and Burch, scientific research should use alternatives to animal experimentation whenever possible. When there is no alternative to animal experimentation, the total number of laboratory animals used should be the minimum needed to obtain valuable data. Moreover, appropriate refinement measures should be applied to minimize pain, suffering, and distress accompanying the experimental procedure. The categories used to classify the degree of pain, suffering, and distress are non-recovery, mild, moderate, or severe (EU Directive 2010/63). To determine which categories apply in individual cases, it is crucial to use scientifically sound tools. The well-being-assessment protocol presented here is designed for procedures during which general anesthesia is used. The protocol focuses on home cage activity, the Mouse Grimace Scale, and luxury behaviors such as burrowing and nest building behavior as indicators of well-being. It also uses the free exploratory paradigm for trait anxiety-related behavior. Fecal corticosterone metabolites as indicators of acute stress are measured over the 24-h post-anesthetic period. The protocol provides scientifically solid information on the well-being of mice following general anesthesia. Due to its simplicity, the protocol can easily be adapted and integrated in a planned study. Although it does not provide a scale to classify distress in categories according to the EU Directive 2010/63, it can help researchers estimate the degree of severity of a procedure using scientifically sound data. It provides a way to improve the assessment of well-being in a scientific, animal-centered manner.

Toward evidence‐based severity assessment in rat models with repeated seizures: II. Chemical post–status epilepticus model

“…In this context, it is of relevance to identify sensitive and robust parameters that should be included in severity‐assessment schemes aiming to validate putative refinement measures and to classify new epilepsy models. Both nest‐building and burrowing have been discussed as nonessential behaviors that can serve as easy‐to‐use indicators of well‐being in laboratory rodents . In contrast to the persistent decrease of burrowing behavior throughout all experimental phases, an influence on nest‐building was only observed in the chronic phase in animals with preceding telemetric recordings.…”

Assessing Affective State in Laboratory Rodents to Promote Animal Welfare—What Is the Progress in Applied Refinement Research?

“…While burrowing activity in rats is mainly used to assess different pain types or analgesic efficacy (e.g., [67][68][69]), the use of this test in mice is more heterogeneous. Burrowing behavior is known to be reduced due to post-surgical pain (e.g., [70]), experimental procedures like anesthesia [71], or experimental housing conditions like grid housing [72]. The recovery of burrowing has been used as a confirmation of the beneficial effects of experimental refinements of surgical techniques (e.g., [73]) or improvement of post-surgical housing conditions [74,75].…”

Towards a fully automated surveillance of well-being status in laboratory mice using deep learning

“…A dosage of 80 mg/kg 104 ketamine and 16 mg/kg xylazine [32], warmed to body temperature, was administered 105 intraperitoneally at a volume of 10 µL/g body weight using 27 3 4 Gauge needles. Then the Images were generated as described previously in Hohlbaum et al [34]. All images were taken 139 in observation cages (22 × 29 × 39 cm) ( Fig 1) with three white walls to contrast the black mice 140 and one clear wall.…”

“…For MGS scoring, one image of high quality showing the mouse face from frontal or lateral 171 view was randomly selected per mouse, and point in time [29,30,34] and the face of the mouse 172 were manually cropped from the image so that, if possible, the body posture was not visible. MGS score.…”

“…As we could not predict the MGS score directly, we followed the approach of Tuttle 184 et al [36] and assigned one of two defined states, "pain" and "no pain", to all images in order to 185 train a binary classifier on the whole data set. To do so, time points of image acquisition were 186 either defined as "pain" or "no pain" based on the MGS scores obtained by humans and the 187 statistical analysis performed in our previous studies [29,30,34]: if MGS scores were 188 significantly higher when compared to untreated mice (Fig 2), all images of this point in time 189 were considered to display a "pain face" and were assigned to the label "pain" (Table 1).…”