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When introducing environmental enrichment in laboratory animals, positive and/or negative effects on behavioural and physiological parameters should be evaluated. This three-step randomised controlled trial in male C57BL/6 mice investigated the effect of supplementing the environment with one or more hemp ropes. In part 1, the effect of a hemp rope on aggressive and social behaviour, stress and anxiety levels was assessed by social interaction test, elevated plus maze behaviour, and faecal corticosterone metabolites, respectively (n = 224 mice). In part 2, the effect of 1, 2 or 7 hemp ropes on aggressive behaviour in mice subjected to routine handling was evaluated by assessing the number of wounded companion animals and wounds per animal (n = 224). In part 3, climbing activity in the rope and amount of material shredded from the rope was assessed (n = 56). Mice housed with one hemp rope engaged in social behaviour for longer time than mice housed without a hemp rope, while no difference was detected in stress and anxiety levels. No difference was seen in the number of wounded animals or wounds per animal when adding 1, 2 or 7 hemp ropes to the existing environment in mice undergoing minimal human handling. The mice continuously shredded and climbed the rope, even when provided with a new rope, although time spent climbing decreased slightly over time. Thus, a hemp rope can be used as additional environmental enrichment amongst male C57BL/6 mice.

Nest building and burrowing are highly motivated natural behaviors in rodents, and changes in these behaviors can serve as welfare assessment tools. In this study, we investigated: 1) the limits of agreement between 2 observers for a refined scoring method for nest-building behavior; 2) the effect of repeated exposure to 15 min of isoflurane on nest-building behavior; 3) the effect of 24 h of grid-floor housing, repeated exposure to 15 min isoflurane, and daily intraperitoneal injection of 0.2 mL 0.9% isotonic saline for 3 d on burrowing behavior; and 4) the effect of exposure to grid-floor housing, isoflurane, and intraperitoneal injections on fecal corticosterone metabolites, body weight, fur status, and sucrose preference in mice. SPF C57BL/6NTac female mice (n = 27) were included in the study and were assessed first for burrowing behavior, followed by 2 wk of rest and then for nesting behavior. The refined scoring method for nest-building activity had good inter observer agreement. According to this method, a single exposure to anesthesia with isoflurane led to a decrease in nest-building activity and sucrose preference; a second exposure to anesthesia with isoflurane had no effect on nest building. Neither grid-floor housing nor repeated exposure to isoflurane anesthesia had any effect on burrowing behavior in mice. In contrast, intraperitoneal injections increased burrowing behavior. In conclusion, a refined scoring method for nest-building activity test that we developed for this study proved to be objective and sensitive to the effect of an initial exposure to anesthesia with isoflurane.