_1788848102 20260908061502000 Rovedar Daryoushbabazadeh@gmail.com Rovedar Journal of Lab Animal Research J Lab Anim. Res 2980-9703 04 30 2026 5 2 Richard Mavunganidze https://orcid.org/0000-0003-0711-533X Vinny Naidoo https://orcid.org/0000-0003-2740-5983 Introduction: Limited space often presents a significant challenge in laboratory animal housing systems, as it restricts the animals' capacity to exhibit species-specific behaviors due to enclosures and structural designs that differ from their natural habitats. The present study aimed to develop a complex caging system (semi-natural cages) equipped with different enrichment items to assess the welfare of rats housed in these cages by observing home-cage behaviors, measuring fecal glucocorticoid metabolites, body weight, and certain organ weights.Materials and methods: Twenty-four, twelve-week-old female Sprague-Dawley rats with a mean weight of 239 ± 19 grams were randomly allocated to three semi-natural and three standard cages (four rats in each cage) and studied weekly for six weeks. Behavioral data were collected from four rats housed in each of the two cage types for five weeks via data-stamped video footage, which was randomly scored using scanning and focal analyses. Fecal glucocorticoid metabolite (FCM) concentrations were measured for six weeks using the Invitrogen progesterone competitive ELISA Kit, adapted for measuring 5a-pregnan-3b,11b,21-triol-20-one competitive enzyme immunoassay. Rats were weighed weekly, and the weights of the brain, thymus, spleen, and adrenal glands were measured at the end of the study.Results: Rats in semi-natural enriched cages were more active than in standard cages. Significantly higher counts of enrichment-directed (U = 617) and non-intake (U = 1,908.5) behaviors were recorded in semi-natural cages compared to the standard cages. The counts of social interaction behaviors were significantly higher in standard cages (U = 2,255) than in semi-natural cages. No significant differences in body weight and organ weights were observed among the rats in the two cage types. Average FCM concentrations indicated periodic fluctuations and an overall upward trend over time in both housing systems. 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