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Introduction: Sheep are frequently used in translational surgical orthopedic studies. Naturally, a good pain management is mandatory for animal welfare, although it is also important with regard to data quality. However, methods for adequate severity assessment, especially considering pain, are rather rare regarding large animal models. Therefore, in the present study, accompanying a surgical pilot study, telemetry and the Sheep Grimace Scale (SGS) were used in addition to clinical scoring for severity assessment after surgical interventions in sheep. Methods: Telemetric devices were implanted in a first surgery subcutaneously into four German black-headed mutton ewes (4-5 years, 77-115 kg). After 3-4 weeks of recovery, sheep underwent tendon ablation of the left M. infraspinatus. Clinical scoring and video recordings for SGS analysis were performed after both surgeries, and the heart rate (HR) and general activity were monitored by telemetry. Results: Immediately after surgery, clinical score and HR were slightly increased, and activity was decreased in individual sheep after both surgeries. The SGS mildly elevated directly after transmitter implantation but increased to higher levels after tendon ablation immediately after surgery and on the following day. Conclusion: In summary, SGS- and telemetry-derived data were suitable to detect postoperative pain in sheep with the potential to improve individual pain recognition and postoperative management, which consequently contributes to refinement.

Good science in translational research requires good animal welfare according to the principles of 3Rs. In many countries, determining animal welfare is a mandatory legal requirement, implying a categorization of animal suffering, traditionally dominated by subjective scorings. However, how such methods can be objectified and refined to compare impairments between animals, subgroups, and animal models remained unclear. Therefore, we developed the RELative Severity Assessment (RELSA) procedure to establish an evidence-based method based on quantitative outcome measures such as body weight, burrowing behavior, heart rate, heart rate variability, temperature, and activity to obtain a relative metric for severity comparisons. The RELSA procedure provided the necessary framework to get severity gradings in TM-implanted mice, yielding four distinct RELSA thresholds L1<0.27, L2<0.59, L3<0.79, and L4<3.45. We show further that severity patterns in the contributing variables are time and model-specific and use this information to obtain contextualized between animal-model and subgroup comparisons with the severity of sepsis > surgery > restraint stress > colitis. The bootstrapped 95% confidence intervals reliably show that RELSA estimates are conditionally invariant against missing information but precise in ranking the quantitative severity information to the moderate context of the transmitter-implantation model. In conclusion, we propose the RELSA as a validated tool for an objective, computational approach to comparative and quantitative severity assessment and grading. The RELSA procedure will fundamentally improve animal welfare, data quality, and reproducibility. It is also the first step toward translational risk assessment in biomedical research.

Several studies suggested an informative value of behavioral and grimace scale parameters for the detection of pain. However, the robustness and reliability of the parameters as well as the current extent of implementation are still largely unknown. In this study, we aimed to systematically analyze the current evidence-base of grimace scale, burrowing, and nest building for the assessment of post-surgical pain in mice and rats. The following platforms were searched for relevant articles: PubMed, Embase via Ovid, and Web of Science. Only full peer-reviewed studies that describe the grimace scale, burrowing, and/or nest building as pain parameters in the post-surgical phase in mice and/or rats were included. Information about the study design, animal characteristics, intervention characteristics, and outcome measures was extracted from identified publications. In total, 74 papers were included in this review. The majority of studies have been conducted in young adult C57BL/6J mice and Sprague Dawley and Wistar rats. While there is an apparent lack of information about young animals, some studies that analyzed the grimace scale in aged rats were identified. The majority of studies focused on laparotomy-associated pain. Only limited information is available about other types of surgical interventions. While an impact of surgery and an influence of analgesia were rather consistently reported in studies focusing on grimace scales, the number of studies that assessed respective effects was rather low for nest building and burrowing. Moreover, controversial findings were evident for the impact of analgesics on post-surgical nest building activity. Regarding analgesia, a monotherapeutic approach was identified in the vast majority of studies with non-steroidal anti-inflammatory (NSAID) drugs and opioids being most commonly used. In conclusion, most evidence exists for grimace scales, which were more frequently used to assess post-surgical pain in rodents than the other behav

Voluntary wheel running (VWR) behaviour is a sensitive indicator of disturbed wellbeing and used for the assessment of individual experimental severity levels in laboratory mice. However, monitoring individual VWR performance usually requires single housing, which itself might have a negative effect on wellbeing. In consideration of the 3Rs principle, VWR behaviour was evaluated under group-housing conditions. To test the applicability for severity assessment, this readout was evaluated in a dextran sodium sulphate (DSS) induced colitis model. For continuous monitoring, an automated system with integrated radio-frequency identification technology was used, enabling detection of individual VWR. After a 14-day adaptation period mice demonstrated a stable running performance. Analysis during DSS treatment in combination with repeated facial vein phlebotomy and faecal sampling procedure resulted in significantly reduced VWR behaviour during the course of colitis and increased VWR during disease recovery. Mice submitted to phlebotomy and faecal sampling but no DSS treatment showed less reduced VWR but a longer-lasting recovery. Application of a cluster model discriminating individual severity levels based on VWR and body weight data revealed the highest severity level in most of the DSS-treated mice on day 7, but a considerable number of control mice also showed elevated severity levels due to sampling procedures alone. In summary, VWR sensitively indicated the course of DSS colitis severity and the impact of sample collection. Therefore, monitoring of VWR is a suitable method for the detection of disturbed wellbeing due to DSS colitis and sampling procedure in group-housed female laboratory mice., Federal Ministry of Economics and Energy, "Zentrales Innovationsprogramm Mittelstand (ZIM), Deutsche Forschungsgemeinschaft https://doi.org/https://dx.doi.org/10.13039/501100001659, Peer Reviewed

The Mouse Grimace Scale (MGS) is an established method for estimating pain in mice during animal studies. Recently, an improved and standardized MGS set-up and an algorithm for automated and blinded output of images for MGS evaluation were introduced. The present study evaluated the application of this standardized set-up and the robustness of the associated algorithm at four facilities in different locations and as part of varied experimental projects. Experiments using the MGS performed at four facilities (F1–F4) were included in the study; 200 pictures per facility (100 pictures each rated as positive and negative by the algorithm) were evaluated by three raters for image quality and reliability of the algorithm. In three of the four facilities, sufficient image quality and consistency were demonstrated. Intraclass correlation coefficient, calculated to demonstrate the correlation among raters at the three facilities (F1–F3), showed excellent correlation. The specificity and sensitivity of the results obtained by different raters and the algorithm were analysed using Fisher's exact test ( p < 0.05). The analysis indicated a sensitivity of 77% and a specificity of 64%. The results of our study showed that the algorithm demonstrated robust performance at facilities in different locations in accordance with the strict application of our MGS setup.

Animal welfare is a growing societal concern and the well-being of animals used for experimental purposes is under particular scrutiny. The vast majority of laboratory animals are mice living in small cages that do not offer very much variety. Moreover, the experimental procedure often takes very little time compared to the time these animals have been bred to the desired age or are being held available for animal experimentation. However, for the assessment of animal welfare, the time spent waiting for an experiment or the time spent after finishing an experiment has also to be taken into account. In addition to experimental animals, many additional animals (e.g. for breeding and maintenance of genetic lines, surplus animals) are related to animal experimentation and usually face similar living conditions. Therefore, in terms of improving the overall welfare of laboratory animals, there is not only a need for refinement of experimental conditions but especially for improving living conditions outside the experiment. The improvement of animal welfare thus depends to a large extent on the housing and maintenance conditions of all animals related to experimentation. Given the current state of animal welfare research there is indeed a great potential for improving the overall welfare of laboratory animals.