Article

Taming anxiety in laboratory mice

Mammalian Behaviour and Evolution Group, Institute of Integrative Biology, University of Liverpool, Leahurst Campus, Neston, UK.
Nature Methods (Impact Factor: 32.07). 10/2010; 7(10):825-6. DOI: 10.1038/nmeth.1500
Source: PubMed

ABSTRACT

Routine laboratory animal handling has profound effects on their anxiety and stress responses, but little is known about the impact of handling method. We found that picking up mice by the tail induced aversion and high anxiety, whereas use of tunnels or open hand led to voluntary approach, low anxiety and acceptance of physical restraint. Using the latter methods, one can minimize a widespread source of anxiety in laboratory mice.

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    • "Only two mice were notched per group for welfare reasons and in previous work we found no effect of ear notching[18]. For all procedures listed below, mice were handled using either a tunnel if EH or paper cup if SH, from their home cage, in order to minimize the effect of handling[4]. Due to a few instances of malocclusion, severe barbering, and accidental death, our final sample size was 216 mice arranged in the following groups: 8 SH and 9 EH C57BL/6 trios; 9 SH and 9 EH DBA/2 trios; 10 SH and 10 EH BALB/c trios; 9 SH and 8 EH mixed-strain trios. "
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    ABSTRACT: Background Inefficient experimental designs are common in animal-based biomedical research, wasting resources and potentially leading to unreplicable results. Here we illustrate the intrinsic statistical power of split-plot designs, wherein three or more sub-units (e.g. individual subjects) differing in a variable of interest (e.g. genotype) share an experimental unit (e.g. a cage or litter) to which a treatment is applied (e.g. a drug, diet, or cage manipulation). We also empirically validate one example of such a design, mixing different mouse strains -- C57BL/6, DBA/2, and BALB/c -- within cages varying in degree of enrichment. As well as boosting statistical power, no other manipulations are needed for individual identification if co-housed strains are differentially pigmented, so also sparing mice from stressful marking procedures. Methods The validation involved housing 240 females from weaning to 5 months of age in single- or mixed- strain trios, in cages allocated to enriched or standard treatments. Mice were screened for a range of 26 commonly-measured behavioural, physiological and haematological variables. Results Living in mixed-strain trios did not compromise mouse welfare (assessed via corticosterone metabolite output, stereotypic behaviour, signs of aggression, and other variables). It also did not alter the direction or magnitude of any strain- or enrichment-typical difference across the 26 measured variables, or increase variance in the data: indeed variance was significantly decreased by mixed- strain housing. Furthermore, using Monte Carlo simulations to quantify the statistical power benefits of this approach over a conventional design demonstrated that for our effect sizes, the split- plot design would require significantly fewer mice (under half in most cases) to achieve a power of 80 %. Conclusions Mixed-strain housing allows several strains to be tested at once, and potentially refines traditional marking practices for research mice. Furthermore, it dramatically illustrates the enhanced statistical power of split-plot designs, allowing many fewer animals to be used. More powerful designs can also increase the chances of replicable findings, and increase the ability of small-scale studies to yield significant results. Using mixed-strain housing for female C57BL/6, DBA/2 and BALB/c mice is therefore an effective, efficient way to promote both refinement and the reduction of animal-use in research.
    Full-text · Article · Jan 2016 · BMC Medical Research Methodology
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    • "Finally, we show that the cup/massage method reduces corticosterone responses to a novel EPM exposure, suggesting that this protocol is sufficient to reduce the HPA axis response to a stressor. The cup handling advocated by Hurst and West [21] and expanded on by Gouveia and West [16] demonstrated anxiety-reducing effects of the cup method. "
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    ABSTRACT: Studies focused on end-points that are confounded by stress are best performed under minimally stressful conditions. The objective of this study was to demonstrate the impact of handling designed to reduce animal stress on measurements of glucose tolerance. A cohort of mice (CD1.C57BL/6) naïve to any specific handling was subjected to either a previously described "cup" handling method, or a "tail-picked" method in which the animals were picked up by the tail (as is common for metabolic studies). Following training, an elevated plus maze (EPM) test was performed followed by measurement of blood glucose and plasma corticosterone. A second cohort (CD1.C57BL/6) was rendered obese by exposure to a high fat diet, handled with either the tail-picked or cup method and subjected to an intraperitoneal glucose tolerance test. A third cohort of C57BL/6 mice was exposed to a cup regimen that included a component of massage and was subjected to tests of anxiety-like behavior, glucose homeostasis, and corticosterone secretion. We found that the cup mice showed reduced anxiety-like behaviors in the EPM coupled with a reduction in blood glucose levels compared to mice handled by the tail-picked method. Additionally, cup mice on the high fat diet exhibited improved glucose tolerance compared to tail-picked controls. Finally, we found that the cup/massage group showed lower glucose levels following an overnight fast, and decreased anxiety-like behaviors associated with lower stress-induced plasma corticosterone concentration compared to tail-picked controls. These data demonstrate that application of handling methods that reduce anxiety-like behaviors in mice mitigates the confounding contribution of stress to interpretation of metabolic endpoints (such as glucose tolerance). Copyright © 2015. Published by Elsevier Inc.
    Full-text · Article · Jun 2015 · Physiology & Behavior
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    • "For example, stress can result in neophobia (Ennaceur et al., 2009), and so even the relatively small amount of stress that may be induced through handling (which may be considerable in these one trial a day tasks as animals are repeatedly taken in and out of the apparatus) may be sufficient to drive behaviour away from the novel stimulus, thereby masking true recognition abilities. Recent evidence supports this view and suggests that particular animal handling procedures can induce aversion and anxiety which can subsequently influence performance in behavioural tasks (Hurst and West, 2010). In this study, mice demonstrated greater anxiety in an elevated plus maze through reduced entry to the arms without protective walls when they were commonly handled with more anxiety-provoking methods such as being picked up by the tail. "
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    Full-text · Article · Apr 2015 · Neuroscience & Biobehavioral Reviews
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