Steroids, providing information regarding several biological patterns including stress and sexual behavior, have been investigated in different matrices in laboratory mice. Data regarding hair quantification, indicative of longer timespans when compared to blood and saliva, are lacking. The aim of the work was to analyze the hormonal hair profile of laboratory male mice and to investigate potential relationships with age and housing, as a potential tool for welfare assessment. Fifty-six adult male C57BL/6J and C57BL/6OlaHsd substrain mice were included in the study, housed in pairs or groups. Testosterone (T) and dehydroepiandrosterone (DHEA) were quantified by radioimmunoassay, corticosterone (CORT) by ELISA. Mean hormone levels were 6.42 pg/mg for T, 23.16 pg/mg for DHEA and 502.1 pg/mg for CORT. Age influenced all hormones by significantly increasing T and DHEA levels and decreasing CORT; only DHEA, significantly higher in grouped mice, was influenced by housing conditions. The influence of age indicates the need for accurate age-related reference intervals, while the higher levels of DHEA in grouped animals suggests that such housing practice may be beneficial for social interactions. In conclusion, it seems that hair hormones quantification may be a good tool for welfare assessment in laboratory mice and may help in refining husbandry.

Here we list species-specific recommendations for housing, care and management of cephalopod molluscs employed for research purposes with the aim of contributing to the standardization of minimum requirements for establishments, care and accommodation of these animals in compliance with the principles stated in Directive 2010/63/EU. Maximizing their psychophysical welfare was our priority. General recommendations on water surface area, water depth and tank shape here reported represent the outcome of the combined action of the analysis of the available literature and an expertise-based consensus reached – under the aegis of the COST Action FA1301 – among researchers working with the most commonly used cephalopod species in Europe. Information on water supply and quality, environmental conditions, stocking density, feeding and handling are also provided. Through this work we wish to set the stage for a more fertile ground of evidence-based approaches on cephalopod laboratory maintenance, thus facilitating standardization and replicability of research outcomes across laboratories, at the same time maximizing the welfare of these animals.

Article 23(2) of the European Union Directive 2010/63/EU, which regulates welfare provisions for animals used for scientific purposes, requires that staff involved in the care and use of animals for scientific purposes be adequately educated and trained before they undertake any such work. However, the nature and extent of such training is not stipulated in the Directive. To facilitate Member States in fulfilling their education and training obligations, the European Commission developed a common Education and Training Framework, which was endorsed by the Member States Competent Authorities. An Education & Training Platform for Laboratory Animal Science (ETPLAS) Working Group was recently established to develop further guidance to the Learning Outcomes in the Framework, with the objective to clarify the levels of knowledge and understanding required by trainees, and to provide the criteria by which these Learning Outcomes should be assessed. Using the Framework document as a starting point, assessment criteria for the Learning Outcomes of the modules required for Function A persons (carrying out procedures on animals) for rats, mice and zebrafish were created with sufficient detail to enable trainees, providers and assessors to appreciate the level of knowledge, understanding and skills required to pass each module. Adoption and utilization of this document by training providers and accrediting or approving bodies will harmonize introductory education and training for those involved in the care and use of animals for scientific purposes within the European Union, promote mutual recognition of training within and between Member States and therefore free movement of personnel.

Background and Aim: Environmental enrichment (EE) is a pivotal tool for creating ideal housing conditions that allow animals to fully display their ethograms. At the micro-environmental level, they should elicit cognitive and social responses and increase physical activities. Hair steroids may be among the best biomarkers to evaluate the effects of prolonged exposure to different enrichments because they are non-invasive and provide information regarding a longer period. This study aimed to compare the hair steroid profiles, specifically corticosterone, cortisol, and dehydroepiandrosterone (DHEA), of Wistar rats exposed to two different EE settings. Materials and Methods: Twenty (n = 20) outbred Wistar rats were enrolled in this study. First hair collection (T0) was performed 3 days before weaning (at 28 days of life), and then Wistar rats were randomly divided into two equal groups with a sex ratio of 1:1: standard EE group, provided with one rat tunnel, and extra EE group, provided with an additional tunnel hanging from the top of the cage. Environmental conditions were 20°C–24°C, 45%–65% relative humidity, and a 12:12 dark/light cycle, with water and pelleted diet ad libitum. The rats were housed in ventilated cages with poplar bedding and nesting material. Hair was sampled again after 3 months (T1). Steroids were quantified using radioimmunoassay or enzyme-linked immunosorbent assay after methanol extraction. Results: Cortisol was not quantifiable, while corticosterone and DHEA were. After 3 months of exposure (T1), no differences were noted between the experimental groups. On data categorization per sex, females showed higher levels of all steroids than males. In males, the extra EE group had higher corticosterone levels. Conclusion: This study showed that corticosterone and DHEA are quantifiable in rats’ hair, yet bigger datasets are needed to better understand the physiological levels of these hormones in such a matrix. Different enrichment settings induced differences between and within sex. Keywords: 3Rs, biomarker, environmental enrichment, Rattus norvegicus, refinement, steroid, welfare.

Since its publication, the 3Rs principle has provided a cornerstone for more ethical and humane biomedical and regulatory research. In Europe, the 3Rs principle has been incorporated into the European Directive 63/2010/EU, with the ultimate aim of fully replacing the procedures on live animals for scientific and educational purposes as soon as it is scientifically possible to do so. Thus, a critical shift in the discussion on animal use in biomedical and regulatory research is undergoing in Europe, a discussion where satisfying the “replacement” principle is becoming more and more defined as a scientific rather than ethical need. 3Rs Centres have been established in recent years across Europe. To date, Ireland has no 3Rs Centre, and the uptake of the 3Rs principle, and in particular of the “replacement” aspect, has been slow. In this Commentary, we present the Irish context of the use of animal models in biomedical and regulatory research, and urge for what, in the authors’ opinion, are the most critical actions that Ireland must undertake to align its biomedical (basic, applied and translational) research with the European 3Rs strategy.