The mouse light/dark box test

https://doi.org/10.1016/S0014-2999(03)01274-3Get rights and content

Abstract

The light/dark test is based on the innate aversion of rodents to brightly illuminated areas and on the spontaneous exploratory behaviour of rodents in response to mild stressors, that is, novel environment and light. The test apparatus consists of a small dark safe compartment (one third) and a large illuminated aversive compartment (two thirds). The test was developed with male mice. The strain, weight and age may be crucial factors. The extent to which an anxiolytic compound can facilitate exploratory activity depends on the baseline level in the control group. Differences between the type and severity of external stressors might account for the variable results reported by different laboratories. The light/dark test may be useful to predict anxiolytic-like or anxiogenic-like activity in mice. Transitions have been reported to be an index of activity-exploration because of habituation over time, and the time spent in each compartment to be a reflection of aversion. Classic anxiolytics (benzodiazepines) as well as the newer anxiolytic-like compounds (e.g. serotonergic drugs or drugs acting on neuropeptide receptors) can be detected using this paradigm. It has the advantages of being quick and easy to use, without requiring the prior training of animals.

Introduction

The light/dark test is based on the innate aversion of rodents to brightly illuminated areas and on the spontaneous exploratory behaviour of rodents in response to mild stressors, that is, novel environment and light (Crawley and Goodwin, 1980). A natural conflict situation occurs when an animal is exposed to an unfamiliar environment or novel objects. The conflict is between the tendency to explore and the initial tendency to avoid the unfamiliar (neophobia). The exploratory activity reflects the combined result of these tendencies in novel situations. Thus, in the light/dark test, drug-induced increase in behaviours in the white part of a two-compartment box, in which a large white compartment is illuminated and a small black compartment is darkened, is suggested as an index of anxiolytic activity. An increase in transitions without an increase in spontaneous locomotion is considered to reflect anxiolytic activity. It is interesting to note that this effect is only observed in certain strains of mice or with certain drugs. This model differs from other purported models of anxiety which are not equivalent in terms of elicited/induced emotional state File 1992, Njung'e and Handley 1991, Treit, 1990, De Vry et al., 1993. The goal of this paper is to review the main data obtained with the light/dark test and point out the inconsistent findings linked with the various modifications of the apparatus as well as the methodology.

Section snippets

Test apparatus

Although the light/dark test was based on the initial model described by Crawley and Goodwin (1980), many authors have used it with several structural modifications (Table 1).

Typical dimensions of the compartment are generally one third for the dark compartment and two thirds for the light compartment with an exterior size of 46×27×30 cm (l×b×h). Nevertheless, Costall et al. (1989) have differently distributed the compartments with two thirds for the dark compartment. The model is based on the

Animals

This test was developed with male mice. As inbred mice displayed substantial variability in spontaneous behaviour, the choice of strain may be a crucial parameter. Early studies with this paradigm concluded that highly active strains of mice show consistently larger percentage increases in exploratory behaviour after diazepam treatment. The C57Bl/6J and the SW-NIH strains seem to be the strains of choice for anti-anxiety testing in the light/dark test (Crawley and Davis, 1982).

The C57Bl/6J

Test conditions

The extent to which an anxiolytic compound can facilitate exploratory activity depends on the baseline level in the control group. There are a number of non-genetic non-pharmacological manipulations that modulate the general stress levels of animals, which, when performed before testing, have profound effects on behaviour. Deliberate or accidental manipulation of these influential factors can also dramatically alter the effects of drugs (Hoggs, 1996). Differences between the type and severity

Scoring of behaviour

Crawley and Goodwin (1980) described a model in which benzodiazepines produced a facilitation of exploratory behaviour between an illuminated open field and a dark enclosure. Mice placed in the white area (which they found aversive) would generally move around the periphery until they found an opening, at floor level, to enable access to the black compartment, and this usually occurred within 7–12 s. The essential feature was the measurement of increased transitions between the light and dark

Drugs

In the present section, the results were obtained after systemic administration of the drugs by the intraperitoneal or subcutaneous route Table 2, Table 3, Table 4, Table 5, Table 6, Table 7.

Conclusion

In conclusion, the light/dark test may be useful to predict the anxiolytic-like or anxiogenic-like activity of drug in mice. It has the advantages of being quick and easy to use, without the prior training of animals, food and water deprivation is unnecessary and natural stimuli are used. Transitions have been reported to be an index of activity-exploration because of habituation over time, and the time spent in each compartment to be a reflection of aversion (Belzung et al., 1987), but the

References (106)

  • M. Bourin et al.

    Cyamemazine as an anxiolytic drug on the elevated plus maze and light/dark paradigm in mice

    Behav. Brain Res.

    (2001)
  • C.H.K. Cheng et al.

    Actions of 5-hydroxytryptophan to inhibit and disinhibit mouse behaviour in the light/dark test

    Eur. J. Pharmacol.

    (1994)
  • B. Costall et al.

    Exploration of mice in a black and white box: validation as a model of anxiety

    Pharmacol. Biochem. Behav.

    (1989)
  • B. Costall et al.

    Sites of action of ondansetron to inhibit withdrawal from drugs of abuse

    Pharmacol. Biochem. Behav.

    (1990)
  • B. Costall et al.

    Anxiolytic effects of CCK-B antagonists

    Neuropeptides

    (1991)
  • B. Costall et al.

    The effect of the 5-HT3 receptor antagonist, RS 42358-197, in animal models of anxiety

    Eur. J. Pharmacol.

    (1993)
  • J.N. Crawley

    Neuropharmacologic specificity of a simple model for the behavioural actions of benzodiazepines

    Pharmacol. Biochem. Behav.

    (1981)
  • J.N. Crawley

    Exploratory behaviour models of anxiety in mice

    Neurosci. Biobehav. Rev.

    (1985)
  • J.N. Crawley et al.

    Base line exploratory activity predicts anxiolytics responsiveness to diazepam in five mouse strains

    Brain Res. Bull.

    (1982)
  • J.N. Crawley et al.

    Preliminary report of a simple animal behaviour for the anxiolytic effects of benzodiazepines

    Pharmacol. Biochem. Behav.

    (1980)
  • J.N. Crawley et al.

    Absence of intrinsic antagonist actions of benzodiazepine antagonist on an exploratory model of anxiety in the mouse

    Neuropharmacology

    (1984)
  • L. De Angelis et al.

    The anxiolytic like properties of two selective MAOIs, moclobemide and selegiline, in a standard and an enhanced light/dark aversion test

    Pharmacol. Biochem. Behav.

    (2000)
  • J. De Vry et al.

    Shock-induced ultrasonic vocalization in young adults rats: a model for testing putative anti-anxiety drugs

    Eur. J. Pharmacol.

    (1993)
  • B. Gao et al.

    Effect of acute administration of the 5-HT3 receptor antagonist, BRL 46470A, on the behavior of mice in a two compartment light–dark box and during social interactions in their home cage and an unfamiliar neutral cage

    Neuropharmacology

    (1992)
  • B. Gao et al.

    Effect of acute and subchronic administration of ritanserin on the social behaviour of mice

    Neuropharmacology

    (1993)
  • G. Griebel

    5-Hydroxytryptamine-interacting drugs in animal models of anxiety disorders: more then 30 years of research

    Pharmacol. Ther.

    (1995)
  • G. Griebel et al.

    Preclinical profile of the mixed 5-HT1A/5-HT2A receptor antagonist S21357

    Pharmacol. Biochem. Behav.

    (1996)
  • G. Griebel et al.

    A comparative study of the effects of selective and non-selective 5-HT2C receptor subtype antagonists in rat and mouse models of anxiety

    Neuropharmacology

    (1997)
  • S.L. Handley

    5-Hydroxytryptamine pathways in anxiety and its treatment

    Pharmacol. Ther.

    (1995)
  • S.L. Handley et al.

    Multiple serotonin mechanisms in animal models of anxiety: environmental, emotional and cognitive factors

    Behav. Brain Res.

    (1993)
  • M. Hascoët et al.

    A new approach to the light/dark procedure in mice

    Pharmacol. Biochem. Behav.

    (1998)
  • M. Hascoët et al.

    Influence of age on behavioural response in the light–dark paradigm

    Physiol. Behav.

    (1999)
  • M. Hascoët et al.

    Anxiolytic like effects of antidepressants after acute administration in a four plate test in mice

    Pharmacol. Biochem. Behav.

    (2000)
  • M. Hascoët et al.

    The influence of buspirone and its metabolite1-PP on the activity of paroxetine in the mouse light dark paradigm and four plates test

    Pharmacol. Biochem. Behav.

    (2000)
  • M. Imaizumi et al.

    The behavioural and biochemical effects of thioperamide, a histamine H3-receptor antagonist, in a light/dark test measuring anxiety in mice

    Life Sci.

    (1993)
  • T. Kilfoil et al.

    Effects of anxiolytic and anxiogenic drugs on exploratory activity in a simple model of anxiety in mice

    Neuropharmacology

    (1989)
  • S. Lightowler et al.

    Anxiolytic like effect of paroxetine in a rat social interaction test

    Pharmacol. Biochem. Behav.

    (1994)
  • R.G. Lister

    Ethologically-based models of anxiety disorders

    Pharmacol. Ther.

    (1990)
  • C. Lopez-Rubalcava et al.

    Interaction of GABA and serotonin in the anxiolytic action of diazepam and serotonergic anxiolytics

    Pharmacol. Biochem. Behav.

    (1992)
  • K. Njung'e et al.

    Evaluation of marble-burying behavior as a model of anxiety

    Pharmacol. Biochem. Behav.

    (1991)
  • B. Olivier et al.

    5-HT3 receptor antagonists and anxiety; a preclinical and clinical review

    Eur. Neuropsychopharmacol.

    (2000)
  • M.V. Pletnikov et al.

    Relationship between memory and fear: developmental and pharmacological studies

    Pharmacol. Biochem. Behav.

    (1996)
  • T. Shimada et al.

    The modified light/dark transition test in mice: evaluation of classic and putative anxiolytic and anxiogenic drugs

    Gen. Pharmacol.

    (1995)
  • C.B. Smith et al.

    Anxiolytic action of CGS 9896 on mouse explorator behaviour

    Eur. J. Pharmacol.

    (1986)
  • R. Stefanski et al.

    Serotonergic innervation of the hippocampus and nucleus accumbens septi and the anxiolytic-like action of the 5-HT3 receptor antagonists

    Neuropharmacology

    (1993)
  • D. Treit

    A comparison of anxiolytic and nonanxiolytic agents in the shock probe/burying test for anxiolytics

    Pharmacol. Biochem. Behav.

    (1990)
  • H.J.G.M. Van Megen et al.

    Cholecystokinin in anxiety

    Eur. Neuropsychopharmacol.

    (1996)
  • L. Arborelius et al.

    The role of corticotrophin-releasing factor in depression and anxiety

    J. Endocrinol.

    (1999)
  • J.E. Barret et al.

    5-HT receptors as targets for the development of novel anxiolytic drugs/models, mechanisms and future directions

    Psychopharmacology

    (1993)
  • J.M. Barry et al.

    A simple habituation test in the mouse

    Br. J. Pharmacol.

    (1987)
  • Cited by (997)

    View all citing articles on Scopus
    View full text