Effect of Lonely Mouse Syndrome in a Drug Safety Study
Katharine Sewell, BVetMed, MRCVS
Data from a new study show that the isolation felt by single-housed mice may skew the results of clinical studies.
In many clinical studies, single caging (i.e., 1 animal per cage) is advised to avoid inadvertent drug transfer and to lower the number of animals used, because each cage counts as a unit for statistical analysis. A trial intended to evaluate the safety of a lipid-bound analgesic in mice raised important questions about the behavior of mice housed in single cages.
Although behavioral analyses in many species have shown isolation effects, these effects generally are seen only after several weeks of isolation. Because the present study would last for a maximum of 12 days, single caging was still recommended. The investigators analyzed the behavior of the mouse subjects in the trial to determine whether the cage effect was real.
Male and female BALB/cAnNCrl mice (6-8 weeks old weighing 20-22 g) were obtained and kept in groups of 3 or 4 for 10 days before the start of the study; the mice were housed separately throughout the study. All the mice were anesthetized, given a 4- to 5-mm skin incision with a 2 ´ 4—cm subcutaneous pocket, and closed with staples.
In the first trial, 8 males and 8 females were used to compare the effect of a single 5-fold dose of buprenorphine (16.25 mg/kg) following the procedure with no other medication over 4 days. Trial 2, which included a different group of 8 males and 8 females, compared three 16.25-mg/kg doses of buprenorphine at 4-day intervals (days 0, 4, and 8 post procedure) versus no medication over 12 days. Half of the mice in each group were euthanized at the midpoint of each trial (day 2 for trial 1 and day 6 for trial 2); the remainder were euthanized at the end of each trial. After euthanasia the mice were evaluated for body weight, hematologic parameters, clinical chemistries, and anatomic pathology.
Behavioral data were collected twice daily throughout each trial, on 4 days in trial 1 and 5 days in trial 2. Mouse behavior was analyzed by measuring respiration, nasal/skin appearance, fur appearance, motor activity, ocular activity (closed or open eyes), actions suggesting stress (i.e., aggressiveness), presence of tremors, and surgical-site erythema, edema, or infection (swelling, pus, and exudate).
The results of the 2 drug-free groups were compared to evaluate the effect of solitude on stress and mouse behaviour. Significantly more mice in the control group of trial 2 were observed with closed eyes than in trial 1. A gender difference was seen in motor signs, with more females than males observed with rapid, darting movements and more males than females seen to have a hunched posture. Additionally, the experiments also showed a significant time effect on ocular score and motor activity on mice without drug interventions, with both groups showing increasing signs of stress and lethargy over time (day 1 vs day 5).
These study results show that single housing can induce lonely mouse syndrome as early as day 3 of isolation. This affects not only the welfare of the animals used but also potentially the results of the studies. Further research is warranted in this area.
Dr Sewell received her BVetMed and MRCVS from the Royal Veterinary College in London in 2008. She worked in mixed animal practice before settling in Essex, UK, to focus on companion animals and exotics.