Knowledge Base

Introduction

The Elevated Plus Maze Test

The elevated plus-maze was designed to provide measures of anxiety that were relatively uncontaminated by changes in overall motor activity, and has been extensively validated by Pellow et al. (1985) using behavioral, physiological, and pharmacological measures.

The open and closed arms are considered to evoke the same exploratory drive in the animals, therefore avoidance of the open arms is considered to be a result of the induction of higher levels of fear (Rodgers and Dalvi, 1997). It is thought that the aversion of animals to explore the open arms of the maze is caused by fear of open and/or elevated spaces.

The plus-maze is sensitive to the anxiolytic effects of neurotoxic lesions of serotonergic neurons and to the anxiogenic effects of drugs (Pellow and File, 1986), drug withdrawal (File and Andrews, 1991), and predator odor (Zangrossi and File, 1992). For a discussion of the additional, ethological measures that can be taken in this test, see Rodgers et al. (1995) and Fernandes and File (1996). Unlike the social interaction and light/dark tests, the elevated plus-maze does not rely on aversion to bright light, and it has been found repeatedly that behavior in the maze is independent of light level (e.g., Becker and Grecksch, 1996).

For a review of the protocol, see Walf and Frye (2007).

What to Measure

• Principal component analysis of the conventional plus-maze (File, 1991) has shown that the percentage of time spent on the open arms and the number of entries onto the open arms are the best measures of anxiety (these are increased by anxiolytic and decreased by anxiogenic treatments).
• The number of closed arm entries is the best measure of locomotor activity.
• There are marked strain differences in baseline scores and even between the scores of different batches of animals. If the scores are high it will be hard to detect an anxiolytic effect. If they are very low it will be difficult to detect an anxiogenic effect.
• The plus-maze was originally developed for male rats, but it can be used with female rats. However, while in male rats anxiety is the main factor measured, in females it is activity (Fernandes et al., 1999). The plus maze has also been validated for mice (Lister, 1987; Walf and Frye, 2007). For a discussion of the factors controlling measures of anxiety in the mouse, see File (2001).

Quantifying Head Dips

EthoVision XT can detect when the subject's nose is outside the open arms.

However, you can also score head dips manually. First define head dips in the Manual Scoring Settings. Next, during acquisition, press the assigned keyboard key when a head dip occurs.

The Sample Experiment

To see how a plus maze test is carried out in EthoVision XT, see also the sample experiment Elevated plus maze XT180 on the downloads section of the Noldus website (my.noldus.com). Download this file and save it on your computer. In EthoVision XT, choose File > Restore Backup and select the file. For more information, see the document Description of sample experiments of EthoVision XT.pdf.

References

Papers

• Becker, A. and Grecksch, G. (1996). Illumination has no effect on rats' behavior in the elevated plus maze. Physiol. Behav., 1175–1177.
• Fernandes, C. and File, S.E. (1996). The influence of open arm ledges and maze experience in the elevated plus-maze. Pharmacol. Biochem. Behav., 31–40.
• Fernandes, C., Gonzalez, M.I., Wilson, C.A. and File, S.E. (1999). Factor analysis reveals that female rat behaviour is characterised by activity, male rats are driven by sex and anxiety. Pharmacol. Biochem. Behav., 731–738.
• File, S.E. (1991). Animal models of anxiety. In Biological Psychiatry (G. Racagni, N. Brunello, and T. Fukuda, eds.), 596–599. Elsevier, New York.
• File, S.E. (2001). Factors controlling measures of anxiety and responses to novelty in the mouse. Behav. Brain Res., 151–157.
• File, S.E. and Andrews, N. (1991). Low but not high doses of buspirone reduce the anxiogenic effects of diazepam withdrawal. Psychopharmacology, 578–582.
• File, S.E., Lippa, A.S., Beer, B., Lippa, M.T. (2004). Animal Tests of Anxiety. Current Protocols in Neuroscience, Unit 8.3. DOI: 10.1002/0471142301.ns0803s26 (online).
• Lister, R.G. (1987). The use of a plus-maze to measure anxiety in the mouse. Psychopharmacology, 180–185.
• Pellow, S., Chopin, P., File, S.E., and Briley, M. (1985). Validation of open:closed arm entries in an elevated plus-maze as a measure of anxiety in the rat. J. Neurosci. Methods, (3), 149–167.
• Pellow, S. and File, S.E. (1986). Anxiolytic and anxiogenic drug effects in exploratory activity in an elevated plus-maze: A novel test of anxiety in the rat. Pharmacol. Biochem. Behav., 525–529.
• Rodgers, R.J., Cole, J.C., Aboualfa, K., and Stephenson, L.H. (1995). Ethopharmacological analysis of the effects of putative "anxiogenic" agents in the mouse elevated plus-maze. Pharmacol. Biochem. Behav., 805–813.
• Rodgers, R.J. and Dalvi, A. (1997). Anxiety, defense and the elevated plus-maze. Neurosci. Behav. Rev., 801–810.
• Violle, N., Balandras, F., Le Roux, Y., Desor, D., and Schroeder, H. (2009). Variations in illumination, closed wall transparency and/or extramaze space influence both baseline anxiety and response to diazepam in the rat elevated plus-maze. Behav. Brain Res., 35–42.
• Walf, A.A. and Frye, C.A. (2007). The use of the elevated plus maze as an assay of anxiety-related behavior in rodents. Nature Protocols, 322–328. doi:10.1038/nprot.2007.44. Web: www.nature.com/nprot/journal/v2/n2/pdf/nprot.2007.44.pdf
• Zangrossi, H. and File, S.E. (1992). Behavioral consequences in animal tests of anxiety and exploration of exposure to cat odor. Brain Res. Bull., 381–388.

Videos

• https://www.youtube.com/watch?v=4rRsxf1U6-w
• http://www.dailymotion.com/video/x2e25p_4tgndox16_animals

Source: EthoVision XT 18 - Application Manual, Noldus Information Technology