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Behavioral Test Questions

Should the animals be housed in groups or singly-housed before testing?

By September 3, 2019No Comments

The housing of laboratory animals either alone or in groups has been a subject of discussion for many years, with publications detailing the effects of these conditions on animal behavior in species ranging from mice[1] to rabbits.[2] In general, the consensus is that, when possible, group housing results in healthier, more emotionally well-adjusted animals that better reflect the conditions for which they are intended to model. However, here we will discuss several specific examples for consideration by researchers faced with the decision of how to house their animals prior to behavioral testing.


In their natural habitat, male mice live in small groups comprised of a single male, several females, and their offspring (until the juvenile age).[3] However, due to breeding concerns, amongst other confounding factors, this is not a viable approach for most researchers. Rather, scientists must consider the pros and cons of individual and group housing for male mice. Male mice consistently exhibit aggressive behaviors, which may cause non-alpha mice to experience stress and anxiety as well as, in extreme cases, injury. However, it has been found that the level of aggression in male mice is not uniform across strains. For example, while C57BL/6 mice are known to live together with little or no intra-male aggressive behavior,[4] others such as BALB/C mice are known to exhibit significantly more aggression between each other when group-housed.[5] These strain differences are of particular importance, however, on the whole, the incidence of stress induced by male-male aggression in laboratory mice may be outweighed by the detriment of single-housing.[6]

Social deprivation in male mice has been shown to increase anxiety-like behavior in such tests as the Open Field apparatus, Elevated Plus Maze, Forced Swim Test and Sucrose Preference, indicating significant psychological distress caused by a lack of social interaction.[7] These findings have also been correlated with reduced levels of brain-derived neurotrophic factor, a common biological marker of depression and chronic anxiety. Indeed, social isolation has been used as a model for inducing chronic anxiety[8] and schizophrenia.[9] Therefore, in the absence of observed aggressive behaviors, it is likely advantageous for researchers to prioritize group housing of male mice whenever possible. Surprisingly, when male mice are given the choice of group housing versus single housing, even subordinate mice choose to make nests closer to other males, suggesting that the perceived detriment of social isolation is outweighed by the potential for social domination by an alpha male.[5]

In addition to the consideration of strain in deciding whether or not to house male mice alone or in groups, researchers who perform in-house breeding must consider the aggressive behavioral outcomes of this practice. Specifically, it has been found that following breeding, male mice increase their territorial behavior and become increasingly aggressive towards male conspecifics, making reintroduction to group housing nearly impossible without significant danger to fellow mice.[10] Thus, once male mice have been separated for breeding initiation, group housing is no longer an option.


Similarly to laboratory mice, rats have been shown to prefer social interaction over social isolation.[11] Indeed, previous research using Sprague Dawley rats housed either in pairs, groups of four or groups of eight, found that the overall psychological welfare of rats was higher when they were housed in larger groups compared to in pairs.[12] Specifically, following seven weeks in one of these housing conditions, rats living in groups of four or eight were found to have a higher number of voluntary social interactions in a social-interaction test, and exhibited significantly less anxiety-like behavior in an Elevated Plus Maze compared with those housed in pairs.

Of note, a sexually-dimorphic effect of group-housing has been noted in at least one study of laboratory rats.[13] While group housing[14] and environmental enrichment[15] have been shown to have protective effects against chronic stress and its impact on the stress-regulating HPA axis in both male and female rats, a significant difference has been found with regards to housing conditions between sexes on the endocannabinoid system. While stressful conditions decrease the density of cannabinoid receptors in male rats, the inverse is true in females. This is consistent with findings that the stress hormone corticosterone is increased in males following group housing, while it is decreased in females in the same conditions.[16] These sexually-dimorphic effects of housing conditions are of particularly important consideration for researchers investigating pharmaceutical or other drug-associated effects, as changes in cannabinoid receptor density or corticosterone levels may further impact drug effects.

Other considerations

In addition to the naturalistic behavior of rodents in groups, there are additional considerations that must be addressed by researchers deciding whether to house their laboratory animals alone or in groups. For instance, following surgical procedures such as canula implantations, optogenetic fiber implantations, or other procedures which leave exposed wounds, single-housing is required for the adequate recuperation of individual animals. In some cases, this housing procedure may remain necessary if animals are found to exhibit behavior which may disturb the implanted apparatus, such as exposed lenses for cranial imaging. Nonetheless, recent advances in implantable technologies have allowed for the post-surgery group-housing of animals in some cases.[17]


On the whole, the emotional welfare and social well-being of rodents is higher in group-housed animals as compared to those who are socially isolated by individual housing. While certain exceptions to this have been found, such as in particularly aggressive mouse strains or in male mice following breeding practices, the overall benefits of group-housing are clear. Aside from considerations such as surgical procedure recovery and the potential for sexually-dimorphic regulation of certain neuropharmacological characteristics, single-housing of animals should be used only with a logical justification wherein the possible detriment outweighs the clear benefits of group-housing.


  1. Kappel, S., Hawkins, P., & Mendl, M. T. (2017). To Group or Not to Group? Good Practice for Housing Male Laboratory Mice. Animals : An Open Access Journal from MDPI, 7(12).
  2. The effects of group housing on the research use of the laboratory rabbit. (1993). Laboratory Animals, 27(4), 330–341.
  3. Zegeren, K. V. (1979). Variation in Aggressiveness and the Regulation of Numbers in House Mouse Populations. Netherlands Journal of Zoology, 30(4), 635–770.
  4. Bisazza, A. (1981). Social organization and territorial behaviour in three strains of mice. Bollettino Di Zoologia, 48(2), 157–167.
  5. Van Loo, P. L. P., Van de Weerd, H. A., Van Zutphen, L. F. M., & Baumans, V. (2004). Preference for social contact versus environmental enrichment in male laboratory mice. Laboratory Animals, 38(2), 178–188.
  6. Brain, P. (1975). What does individual housing mean to a mouse? Life Sciences, 16(2), 187–200
  7. Berry, A., Bellisario, V., Capoccia, S., Tirassa, P., Calza, A., Alleva, E., & Cirulli, F. (2012). Social deprivation stress is a triggering factor for the emergence of anxiety- and depression-like behaviours and leads to reduced brain BDNF levels in C57BL/6J mice. Psychoneuroendocrinology, 37(6), 762–772.
  8. Ieraci, A., Mallei, A., & Popoli, M. (2016). Social Isolation Stress Induces Anxious-Depressive-Like Behavior and Alterations of Neuroplasticity-Related Genes in Adult Male Mice. Neural Plasticity, 2016, 6212983.
  9. Pietropaolo, S., Singer, P., Feldon, J., & Yee, B. K. (2008). The postweaning social isolation in C57BL/6 mice: Preferential vulnerability in the male sex. Psychopharmacology, 197(4), 613–628.
  10. Hurst J. Making Sense of Scents: Reducing Aggression and Uncontrolled Variation in Laboratory Mice.National Centre for the Replacement, Refinement and Reduction of Animals in Research; London, UK: 2005. pp. 1–8.
  11. Patterson-Kane, E. G., Hunt, M., & Harper, D. (2002). Rats demand social contact. Animal Welfare, 11(3), 327-332.
  12. Spangenberg, E., Dahlborn, K., Essén-Gustavsson, B., & Cvek, K. (2009, May). Effects of physical activity and group size on animal welfare in laboratory rats.
  13. Dow-Edwards, D., Frank, A., Wade, D., Weedon, J., & Izenwasser, S. (2016). Sexually-dimorphic alterations in cannabinoid receptor density depend upon prenatal/early postnatal history. Neurotoxicology and Teratology, 58, 31–39.
  14. Fox, C., Merali, Z., & Harrison, C. (2006). Therapeutic and protective effect of environmental enrichment against psychogenic and neurogenic stress. Behavioural Brain Research, 175(1), 1–8.
  15. Belz, E. E., Kennell, J. S., Czambel, R. K., Rubin, R. T., & Rhodes, M. E. (2003). Environmental enrichment lowers stress-responsive hormones in singly housed male and female rats. Pharmacology, Biochemistry, and Behavior, 76(3–4), 481–486.
  16. Brown, K. J., & Grunberg, N. E. (1995). Effects of housing on male and female rats: Crowding stresses male but calm females. Physiology & Behavior, 58(6), 1085–1089.
  17. Jensen, V. F. H., Mølck, A.-M., Mårtensson, M., Strid, M. A., Chapman, M., Lykkesfeldt, J., & Bøgh, I. B. (2016). Assessment of implantable infusion pumps for continuous infusion of human insulin in rats: potential for group housing. Laboratory Animals, 51(3), 273–283.
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