Impulse for animal welfare outside the experiment

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

Housing of female mice in a new environment and its influence on post-surgical behaviour and recovery

The transportation of mice into a new clean cage after surgery is a standard procedure but might have detrimental effects during the critical post-surgical recovery phase. To analyse the effect of post-surgical housing, female C57BL/6J mice housed in their familiar home cage or in a new environment after minor surgery ± analgesia, anaesthesia only or no treatment were monitored using non-invasive methods during the immediate postsurgical period to assess pain and general impairment. Behavioural investigations and burrowing test revealed no significant differences between housing conditions in untreated mice. While no appearance or posture abnormalities were observed post-experiment, home cage behaviours were affected distinctly. Behavioural rhythmicity was disrupted, and behaviours related to well-being, such as burrowing performance, were less compared to untreated mice. Burrowing latency ranged from an intermediate level following anaesthesia only and surgery with analgesia, to pronounced prolongation after surgery without analgesia in animals housed in their home cage, while after all experimental treatments burrowing latency in animals in new cages was prolonged dramatically. General activity and climbing behaviour in treatment groups housed in new cages tend to be higher compared to animals in familiar cages, leading to significant interactions between housing and treatment conditions (p = 0.006; p = 0.014). These behavioural differences in animals housed in a new environment compared to animals housed in their familiar environment might be interpreted as signs of reduced well-being, agitation and restlessness in the new cages and may hint that animals cope better with surgical stress when housed in their familiar environment. The post-surgical transport to a new and clean cage might therefore be an additional stressor after an exhausting event and may affect recovery

Implantation of Radiotelemetry Transmitters Yielding Data on ECG, Heart Rate, Core Body Temperature and Activity in Free-moving Laboratory Mice

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a manageable size and relatively easy to house. These factors, together with the availability of a wealth of spontaneous and experimentally induced mutants, make laboratory mice ideally suited to a wide variety of research areas

Impact of inhalation anaesthesia, surgery and analgesic treatment on home cage behaviour in laboratory mice

Anaesthesia and analgesia are used frequently in laboratory routine to ensure animal welfare and good scientific outcomes in experiments that may elicit pain or require immobilisation of the animal. However, there is concern regarding the effect of these procedures on animal behaviour in subsequent experiments. Our study determined the impact of short inhalation anaesthesia (sevoflurane, 15 min, 4.9%) and minor surgery (one-sided sham embryo transfer in females, one-sided sham vasectomy in males) with or without pain treatment (carprofen, 5 mg/kg, bid) on spontaneous species-specific home cage behaviours in inbred mice. Analysis of 18-h continuous video recordings showed clear post-procedural changes in spontaneous home cage behaviours, with changes of a moderate level after anaesthesia being marked after surgery. Self-grooming, resting and locomotion were the most important behaviours for group separation. Analysis of the temporal distribution of behavioural changes revealed that resting behaviour was altered contradictory to its circadian rhythm as it was decreased in the light phase and increased in the dark phase. Also, locomotion was decreased in the dark phase at 12 to 18 h after surgery and anaesthesia. In contrast, self-grooming was increased independently of circadian rhythm, being increased for up to 18 h after surgery and anaesthesia. Following surgery, there was no significant difference in duration of behaviours between animals that were treated with carprofen or left without pain relief. In conclusion, it can be assumed that the changes observed in home cage behaviours hint at reduced animal well-being. However, pain or the efficacy of post-operative pain treatment could not be discriminated reliably from the impact of the surgical procedure including inhalation anaesthesia by observing animals’ home cage behaviour. However for the interpretation of behavioural research data, the distinct impact of anaesthesia, surgery, pain treatment and other experimental procedures has to be considered. Our results highlight the requirement for knowledge of species-specific circadian rhythms of behaviours as well as the importance of determining the appropriate time of day for behavioural and welfare assessment

Administration of Tramadol or Buprenorphine via the drinking water for post-operative analgesia in a mouse-osteotomy model

Adequate analgesia is essential whenever pain might occur in animal experiments. Unfortunately, the selection of suitable analgesics for mice in bone-linked models is limited. Here, we evaluated two analgesics - Tramadol [0.1 mg/ml (Tlow) vs. 1 mg/ml (Thigh)] and Buprenorphine (Bup; 0.009 mg/ml) - after a pre-surgical injection of Buprenorphine, in a mouse-osteotomy model. The aim of this study was to verify the efficacy of these opioids in alleviating pain-related behaviors, to provide evidence for adequate dosages and to examine potential side effects. High concentrations of Tramadol affected water intake, drinking frequency, food intake and body weight negatively in the first 2-3 days post-osteotomy, while home cage activity was comparable between all groups. General wellbeing parameters were strongly influenced by anesthesia and analgesics. Model-specific pain parameters did not indicate more effective pain relief at high concentrations of Tramadol. In addition, ex vivo high-resolution micro computed tomography (µCT) analysis and histology analyzing bone healing outcomes showed no differences between analgesic groups with respect to newly formed mineralized bone, cartilage and vessels. Our results show that high concentrations of Tramadol do not improve pain relief compared to low dosage Tramadol and Buprenorphine, but rather negatively affect animal wellbeing

Burrowing Behavior as an Indicator of Post-Laparotomy Pain in Mice

Detection of persistent pain of a mild-to-moderate degree in laboratory mice is difficult because mice do not show unambiguous symptoms of pain or suffering using standard methods of short-term observational or clinical monitoring. This study investigated the potential use of burrowing performance – a spontaneous and highly motivated behavior – as a measure of post-operative pain in laboratory mice. The influence of minor surgery on burrowing was investigated in adult C57BL/6J mice of both genders in a modified rodent burrowing test (displacement of food pellets from a pellet-filled tube) within the animal's home cage. Almost all (98%) healthy mice burrowed (mean latency 1.3 h, SEM 0.5 h). After surgery without pain treatment, latency of burrowing was significantly prolonged (mean Δ latency 10 h). Analgesic treatment using the anti-inflammatory drug carprofen (5 mg/kg bodyweight) decreased latency of burrowing after surgery (mean Δ latency 5.5 h) to the level found in mice that had been anesthetized (mean Δ latency 5.4 h) or had received anesthesia and analgesia (mean Δ latency 4.6 h). Analgesia during surgery was associated with a significantly earlier onset of burrowing compared to surgery without pain treatment. A distinct gradation in burrowing performance was found ranging from the undisturbed pre-operative status to the intermediate level following anesthesia/analgesia and surgery with analgesia, to the pronounced prolongation of latency to burrow after surgery without pain relief. In conclusion, post-surgical impairment of general condition, probably mainly attributable to pain, can be conveniently assessed in laboratory mice on the basis of the burrowing test

Improving animal welfare using continuous nalbuphine infusion in a long-term rat model of sepsis

Abstract Background Sepsis research relies on animal models to investigate the mechanisms of the dysregulated host response to infection. Animal welfare concerns request the use of potent analgesics for the Refinement of existing sepsis models, according to the 3Rs principle. Nevertheless, adequate analgesia is often missing, partly because the effects of analgesics in this particular condition are unknown. We evaluated the use of nalbuphine, an opioid with kappa agonistic and mu antagonistic effects, in rats with and without experimental sepsis. Methods Male Wistar rats were anesthetized with isoflurane and instrumented with a venous line for drug administration. Arterial cannulation allowed for blood pressure measurements and blood sampling in short-term experiments of non-septic animals. Nalbuphine (or placebo) was administered intravenously at a dose of 1 mg/kg/h. Long-term (48 h) experiments in awake septic animals included repetitive clinical scoring with the Rat Grimace Scale and continuous heart rate monitoring by telemetry. Sepsis was induced by intraperitoneal injection of faecal slurry. Nalbuphine plasma levels were measured by liquid chromatography—high resolution mass spectrometry. Results In anesthetized healthy animals, nalbuphine led to a significant reduction of respiratory rate, heart rate, and mean arterial pressure during short-term experiments. In awake septic animals, a continuous nalbuphine infusion did not affect heart rate but significantly improved the values of the Rat Grimace Scale. Nalbuphine plasma concentrations remained stable between 4 and 24 h of continuous infusion in septic rats. Conclusions In anaesthetised rats, nalbuphine depresses respiratory rate, heart rate, and blood pressure. In awake animals, nalbuphine analgesia improves animal welfare during sepsis

Grimace scale, burrowing, and nest building for the assessment of post-surgical pain in mice and rats-A systematic review

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 behavioral parameters. However, our findings also point to relevant knowledge gaps concerning the post-surgical application in different strains, age levels, and following different surgical procedures. Future efforts are also necessary to directly compare the sensitivity and robustness of different readout parameters applied for the assessment of nest building and burrowing activities

Measuring Endogenous Corticosterone in Laboratory Mice - a Mapping Review, Meta-Analysis, and Open Source Database

Evaluating stress in laboratory animals is a key principle in animal welfare. Measuring corticosterone is a common method to assess stress in laboratory mice. There are, however, numerous methods to measure glucocorticoids with differences in sample matrix (e.g., plasma, urine) and quantification techniques (e.g., enzyme immunoassay or radioimmunoassay). Here, the authors present a mapping review and a searchable database, giving a complete overview of all studies measuring endogenous corticosterone in mice up to February 2018. For each study, information was recorded regarding mouse strain and sex; corticosterone sample matrix and quantification technique; and whether the study covered the research theme animal welfare, neuroscience, stress, inflammation, or pain (the themes of specific interest in our consortium). Using all database entries for the year 2012, an exploratory meta-regression was performed to determine the effect of predictors on basal corticosterone concentrations. Seventy-five studies were included using the predictors sex, time-since-lights-on, sample matrix, quantification technique, age of the mice, and type of control. Sex, time-since-lights-on, and type of control significantly affected basal corticosterone concentrations. The resulting database can be used, inter alia, for preventing unnecessary duplication of experiments, identifying knowledge gaps, and standardizing or heterogenizing methodologies. These results will help plan more efficient and valid experiments in the future and can answer new questions in silico using meta-analyses

Anesthesia and analgesia for experimental craniotomy in mice and rats: a systematic scoping review comparing the years 2009 and 2019

Experimental craniotomies are a common surgical procedure in neuroscience. Because inadequate analgesia appears to be a problem in animal-based research, we conducted this review and collected information on management of craniotomy-associated pain in laboratory mice and rats. A comprehensive search and screening resulted in the identification of 2235 studies, published in 2009 and 2019, describing craniotomy in mice and/or rats. While key features were extracted from all studies, detailed information was extracted from a random subset of 100 studies/year. Reporting of perioperative analgesia increased from 2009 to 2019. However, the majority of studies from both years did not report pharmacologic pain management. Moreover, reporting of multimodal treatments remained at a low level, and monotherapeutic approaches were more common. Among drug groups, reporting of pre- and postoperative administration of non-steroidal anti-inflammatory drugs, opioids, and local anesthetics in 2019 exceeded that of 2009. In summary, these results suggest that inadequate analgesia and oligoanalgesia are persistent issues associated with experimental intracranial surgery. This underscores the need for intensified training of those working with laboratory rodents subjected to craniotomies.Systematic review registrationhttps://osf.io/7d4qe