Skip to main content
SpringerLink
Log in

Using the Elevated Plus Maze as a Bioassay to Assess the Effects of Naturally Occurring and Exogenously Administered Compounds to Influence Anxiety-Related Behaviors of Mice

  • Protocol
  • First Online:
Mood and Anxiety Related Phenotypes in Mice

Part of the book series: Neuromethods ((NM,volume 42))

Abstract

To assess a construct, such as anxiety, and to determine potential neurobiological underpinnings of this construct, it is often necessary to utilize an animal model. For example, the elevated plus maze is a widely used behavioral assay that has been validated to assess the anxiety-related behavior of rodents. There is great value in using a whole systems approach to assess the many potential targets for a complex disorder, such as anxiety. The elevated plus maze produces reliable results, can be fully automated, is easy and economical to use, and valid results are obtained in a single 5-minute testing session. Briefly, mice are placed at the intersection of the four arms of the maze (two open, two closed), facing an open arm. The number of entries and time spent in each arm is recorded. An increase in the open-arm time is an index of anti-anxiety behavior of mice. Importantly, the patterns of effects that we have observed investigating the role of steroids for effects on anxiety-related behavior has been replicable across cohorts, experiments, species, laboratories, and other anxiety models. Thus, the elevated plus maze is an indispensable tool for investigating the neurobiological substrates of anxiety disorders, using murine models.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Walf AA, Frye CA. The use of the elevated plus maze as an assay of anxiety-related behavior in rodents. Nat Protoc. 2007;2:322–8.

    Article  PubMed  CAS  Google Scholar 

  2. Belzung C, Griebel G. Measuring normal and pathological anxiety-like behaviour in mice: a review. Behav Brain Res. 2001;125:141–9.

    Article  PubMed  CAS  Google Scholar 

  3. Carobrez AP, Bertoglio LJ. Ethological and temporal analyses of anxiety-like behavior: the elevated plus-maze model 20 years on. Neurosci Biobehav Rev. 2005;29:1193–205.

    Article  PubMed  CAS  Google Scholar 

  4. Dawson GR, Tricklebank MD. Use of the elevated plus maze in the search for novel anxiolytic agents. Trends Pharmacol Sci. 1995;16:33–6.

    Article  PubMed  CAS  Google Scholar 

  5. File SE. Factors controlling measures of anxiety and responses to novelty in the mouse. Behav Brain Res. 2001;125:151–7.

    Article  PubMed  CAS  Google Scholar 

  6. Hogg S. A review of the validity and variability of the elevated plus-maze as an animal model of anxiety. Pharmacol Biochem Behav. 1996;54:21–30.

    Article  PubMed  CAS  Google Scholar 

  7. Kulkarni SK, Sharma AC. Elevated plus-maze: a novel psychobehavioral tool to measure anxiety in rodents. Methods Find Exp Clin Pharmacol. 1991;13:573–7.

    PubMed  CAS  Google Scholar 

  8. Rodgers RJ, Dalvi A. Anxiety, defence and the elevated plus-maze. Neurosci Biobehav Rev. 1997;21:801–810.

    Article  PubMed  CAS  Google Scholar 

  9. Wahlsten D, Metten P, Phillips TJ, Boehm SL 2nd, Burkhart-Kasch S, Dorow J, Doerksen S, Downing C, Fogarty J, Rodd-Henricks K, Hen R, McKinnon CS, Merrill CM, Nolte C, Schalomon M, Schlumbohm JP, Sibert JR, Wenger CD, Dudek BC, Crabbe JC. Different data from different labs: lessons from studies of gene-environment interaction. J Neurobiol. 2003;54:283–311.

    Article  PubMed  Google Scholar 

  10. Wall PM, Messier C. Methodological and conceptual issues in the use of the elevated plus-maze as a psychological measurement instrument of animal anxiety-like behavior. Neurosci Biobehav Rev. 2001;25:275–86.

    Article  PubMed  CAS  Google Scholar 

  11. Pellow S, Chopin P, File SE, Briley M. Validation of open:closed arm entries in an elevated plus-maze as a measure of anxiety in the rat. J Neurosci Methods. 1985;14:149–67.

    Article  PubMed  CAS  Google Scholar 

  12. Lister RG. The use of a plus-maze to measure anxiety in the mouse. Psychopharmacology. 1987;92:180–5.

    PubMed  CAS  Google Scholar 

  13. File SE, Mabbutt PS, Hitchcott PK. Characterisation of the phenomenon of “one-trial tolerance” to the anxiolytic effect of chlordiazepoxide in the elevated plus-maze. Psychopharmacology. 1990;102:98–101.

    Article  PubMed  CAS  Google Scholar 

  14. Almeida SS, Garcia RA, de Oliveira LM. Effects of early protein malnutrition and repeated testing upon locomotor and exploratory behaviors in the elevated plus-maze. Physiol Behav. 1993;54:749–52.

    Article  PubMed  CAS  Google Scholar 

  15. Bertoglio LJ, Carobrez AP. Previous maze experience required to increase open arms avoidance in rats submitted to the elevated plus-maze model of anxiety. Behav Brain Res. 2000;108:197–203.

    Article  PubMed  CAS  Google Scholar 

  16. Bertoglio LJ, Carobrez AP. Anxiolytic effects of ethanol and phenobarbital are abolished in test-experienced rats submitted to the elevated plus maze. Pharmacol Biochem Behav. 2002;73:963–9.

    Article  PubMed  CAS  Google Scholar 

  17. Bertoglio LJ, Carobrez AP. Behavioral profile of rats submitted to session 1-session 2 in the elevated plus-maze during diurnal/nocturnal phases and under different illumination conditions. Behav Brain Res. 2002;132:135–43.

    Article  PubMed  Google Scholar 

  18. Fernandes C, File SE. The influence of open arm ledges and maze experience in the elevated plus-maze. Pharmacol Biochem Behav. 1996; 54:31–40.

    Article  PubMed  CAS  Google Scholar 

  19. Lee C, Rodgers RJ. Antinociceptive effects of elevated plus-maze exposure: influence of opiate receptor manipulations. Psychopharmacology. 1990;102:507–13.

    Article  PubMed  CAS  Google Scholar 

  20. Treit D, Menard J, Royan C. Anxiogenic stimuli in the elevated plus-maze. Pharmacol Biochem Behav. 1993;44:463–9.

    Article  PubMed  CAS  Google Scholar 

  21. Espejo EF. Effects of weekly or daily exposure to the elevated plus-maze in male mice. Behav Brain Res 1991;87:233–238.

    Article  Google Scholar 

  22. Adamec R, Strasser K, Blundell J, Burton P, McKay DW. Protein synthesis and the mechanisms of lasting change in anxiety induced by severe stress. Behav Brain Res 2006;167:270–86.

    Article  PubMed  CAS  Google Scholar 

  23. Adamec R, Blundell J, Burton P. Anxiolytic effects of kindling role of anatomical location of the kindling electrode in response to kindling of the right basolateral amygdala. Brain Res. 2004;1024:44–5.

    Article  PubMed  CAS  Google Scholar 

  24. Frye CA, Petralia SM, Rhodes ME. Estrous cycle and sex differences in performance on anxiety tasks coincide with increases in hippocampal progesterone and 3α,5α-THP. Pharmacol Biochem Behav. 2000;67:587–596.

    Article  PubMed  CAS  Google Scholar 

  25. Frye CA, Walf AA, Rhodes ME, Harney JP. Progesterone enhances motor, anxiolytic, analgesic, and antidepressive behavior of wild-type mice, but not those deficient in type 1 5 alpha-reductase. Brain Res. 2004;1004:116–24.

    Article  PubMed  CAS  Google Scholar 

  26. Frye CA, Koonce CJ, Edinger KL, Osborne DM, Walf AA. Androgens with activity at estrogen receptor beta have anxiolytic and cognitive-enhancing effects in male rats and mice. Horm Behav. 2008;54:726–734.

    Google Scholar 

  27. Walf AA, Frye CA. Estradiol’s effects to reduce anxiety and depressive behavior may be mediated by estradiol dose and restraint stress. Neuropsychopharmacology. 2005;30:1288–301.

    Article  PubMed  CAS  Google Scholar 

  28. Walf AA, Frye CA. Estradiol or diarylpropionitrile decrease anxiety-like behavior of wildtype, but not estrogen receptor β knockout, mice. Behav Neurosci. 2008;122:974–81.

    Article  PubMed  CAS  Google Scholar 

  29. Hendrie CA, Eilam D, Weiss SM. Effects of diazepam and buspirone on the behaviour of wild voles (Microtus socialis) in two models of anxiety. Pharmacol. Biochem Behav. 1997;58:573–6.

    Article  PubMed  CAS  Google Scholar 

  30. Holmes A, Parmigiani S, Ferrari PF, Palanza P, Rodgers RJ. Behavioral profile of wild mice in the elevated plus-maze test for anxiety. Physiol Behav. 2000;71:509–16.

    Article  PubMed  CAS  Google Scholar 

  31. Rex A, Marsden CA, Fink H. Effect of diazepam on cortical 5-HT release and behaviour in the guinea-pig on exposure to the elevated plus maze. Psychopharmacology. 1993;110:490–6.

    Article  PubMed  CAS  Google Scholar 

  32. Stowe JR, Liu Y, Curtis JT, Freeman ME, Wang Z. Species differences in anxiety-related responses in male prairie and meadow voles: the effects of social isolation. Physiol Behav. 2005;86:369–78.

    Article  PubMed  CAS  Google Scholar 

  33. Rodgers RJ, Cole JC. Influence of social isolation, gender, strain, and prior novelty on plus-maze behaviour in mice. Physiol Behav. 1993;54:729–36.

    Article  PubMed  CAS  Google Scholar 

  34. Montgomery KC. The relation between fear induced by novel stimulation and exploratory behavior. J Comp Physiol Psychol. 1958;48:254–260.

    Article  Google Scholar 

  35. Handley SL, Mithani S. Effects of α-adrenoreceptor agonists and antagonists in a maze-exploration model of ‘fear’-motivated behaviour. Naunyn-Schmeideberg’s Arch Pharmacol. 1984;327:1–5.

    Article  CAS  Google Scholar 

  36. Barnett SA. The Rat- A Study in Behavior. University Chicago Press. (1975).

    Google Scholar 

  37. Frye CA, Sumida K, Dudek BC, Harney JP, Lydon JP, O'Malley BW, Pfaff DW, Rhodes ME. Progesterone's effects to reduce anxiety behavior of aged mice do not require actions via intracellular progestin receptors. Psychopharmacology. 2006;186:312–22.

    Article  PubMed  CAS  Google Scholar 

  38. Augustsson H, Dahlborn K, Meyerson BJ. Exploration and risk assessment in female wild house mice (Mus musculus musculus) and two laboratory strains. Physiol Behav. 2005;84:265–77.

    Article  PubMed  CAS  Google Scholar 

  39. Carola V, D’Olimpio F, Brunamonti E, Mangia F, Renzi P. Evaluation of the elevated plus-maze and open-field tests for the assessment of anxiety-related behaviour in inbred mice. Behav Brain Res. 2002;134:49–57.

    Article  PubMed  Google Scholar 

  40. Ramos A, Berton O, Mormede P, Chaouloff F. A multiple-test study of anxiety-related behaviours in six inbred rat strains. Behav Brain Res. 1997;85:57–69.

    Article  PubMed  CAS  Google Scholar 

  41. Trullas R, Skolnick P. Differences in fear motivated behaviors among inbred mouse strains. Psychopharmacology. 1993;111:323–31.

    Article  PubMed  CAS  Google Scholar 

  42. Voikar V, Koks S, Vasar E, Rauvala H. Strain and gender differences in the behavior of mouse lines commonly used in transgenic studies. Physiol Behav. 2001;72:271–81.

    Article  PubMed  CAS  Google Scholar 

  43. Frick KM, Burlingame LA, Arters JA, Berger-Sweeney J. Reference memory, anxiety and estrous cyclicity in C57BL/6NIA mice are affected by age and sex. Neuroscience. 2000;95:293–307.

    Article  PubMed  CAS  Google Scholar 

  44. Frye CA, Edinger K, Sumida K. Androgen administration to aged male mice increases anti-anxiety behavior and enhances cognitive performance. Neuropsychopharmacology. 2008;33:1049–61

    Article  PubMed  CAS  Google Scholar 

  45. Seeman MV. Psychopathology in women and men: focus on female hormones. Am J Psychiatry. 1997;154:1641–7.

    PubMed  CAS  Google Scholar 

  46. Bothe GW, Bolivar VJ, Vedder MJ, Geistfeld JG. Behavioral differences among fourteen inbred mouse strains commonly used as disease models. Comp Med. 2005;55:326–34.

    PubMed  CAS  Google Scholar 

  47. Bothe GW, Bolivar VJ, Vedder MJ, Geistfeld JG. Genetic and behavioral differences among five inbred mouse strains commonly used in the production of transgenic and knockout mice. Genes Brain Behav. 2004;3:149–57.

    Article  PubMed  CAS  Google Scholar 

  48. Hunt C, Hambly C. Faecal corticosterone concentrations indicate that separately housed male mice are not more stressed than group housed males. Physiol Behav. 2006;87:519–26.

    Article  PubMed  CAS  Google Scholar 

  49. Zhu SW, Yee BK, Nyffeler M, Winblad B, Feldon J, Mohammed AH. Influence of differential housing on emotional behaviour and neurotrophin levels in mice. Behav Brain Res. 2006;169:10–20.

    Article  PubMed  CAS  Google Scholar 

  50. Andrade MM, Tome MF, Santiago ES, Lucia-Santos A, de Andrade, TG. Longitudinal study of daily variation of rats' behavior in the elevated plus-maze. Physiol Behav. 2003;78:125–33.

    Article  PubMed  CAS  Google Scholar 

  51. Jones N, King SM. Influence of circadian phase and test illumination on pre-clinical models of anxiety. Physiol Behav. 2001;72:99–106.

    Article  PubMed  CAS  Google Scholar 

  52. Crawley JN, Chen T, Puri A, Washburn R, Sullivan TL, Hill JM, Young NB, Nadler JJ, Moy SS, Young LJ, Caldwell HK, Young WS. Social approach behaviors in oxytocin knockout mice: comparison of two independent lines tested in different laboratory environments. Neuropeptides. 2007;41:145–63.

    Article  PubMed  CAS  Google Scholar 

  53. Mechiel Korte S, De Boer SF. A robust animal model of state anxiety: fear-potentiated behaviour in the elevated plus-maze. Eur J Pharmacol. 2003;463:163–75.

    Article  PubMed  CAS  Google Scholar 

  54. Adamec RE, Shallow T. Lasting effects on rodent anxiety of a single exposure to a cat. Physiol Behav. 1993;54:101–9.

    Article  PubMed  CAS  Google Scholar 

  55. Andrews N, File SE. Handling history of rats modifies behavioural effects of drugs in the elevated plus-maze test of anxiety. Eur J Pharmacol. 1993;235:109–12.

    Article  PubMed  CAS  Google Scholar 

  56. Brett RR, Pratt JA. Chronic handling modifies the anxiolytic effect of diazepam in the elevated plus-maze. Eur J Pharmacol. 1990;178:135–8.

    Article  PubMed  CAS  Google Scholar 

  57. File SE, Andrews N, Wu PY, Zharkovsky A, Zangrossi, H Jr. Modification of chlordiazepoxide's behavioural and neurochemical effects by handling and plus-maze experience. Eur J Pharmacol 1992;218:9–14.

    Article  PubMed  CAS  Google Scholar 

  58. Lapin IP. Only controls: effect of handling, sham injection, and intraperitoneal injection of saline on behavior of mice in an elevated plus-maze. J Pharmacol Toxicol Methods. 1995;34:73–7.

    Article  PubMed  CAS  Google Scholar 

  59. Padovan CM, Guimaraes FS. Restraint-induced hypoactivity in an elevated plus-maze. Braz J Med Biol Res. 2000;33:79–83.

    Article  PubMed  CAS  Google Scholar 

  60. Schmitt U, Hiemke C. Strain differences in open-field and elevated plus-maze behavior of rats without and with pretest handling. Pharmacol Biochem Behav. 1998;59:807–11.

    Article  PubMed  CAS  Google Scholar 

  61. Steenbergen HL, Heinsbroek RP, Van Hest A, Van de Poll NE. Sex-dependent effects of inescapable shock administration on shuttlebox-escape performance and elevated plus-maze behavior. Physiol Behav. 1990;48:571–6.

    Article  PubMed  CAS  Google Scholar 

  62. Mong JA, Pfaff DW. Hormonal and genetic influences underlying arousal as it drives sex and aggression in animal and human brains. Neurobiol Aging. 2003;24:S83–8.

    Article  PubMed  CAS  Google Scholar 

  63. Morgan MA, Pfaff DW. Estrogen's effects on activity, anxiety, and fear in two mouse strains. Behav Brain Res. 2002;132:85–93.

    Article  PubMed  CAS  Google Scholar 

  64. Morgan MA, Pfaff DW. Effects of estrogen on activity and fear-related behaviors in mice. Horm Behav. 2001;40:472–82.

    Article  PubMed  CAS  Google Scholar 

  65. Walf AA, Frye CA. Estradiol decreases anxiety behavior and enhances inhibitory avoidance and gestational stress produces opposite effects. Stress. 2007;10:251–60.

    Article  PubMed  CAS  Google Scholar 

  66. File SE, Wardill AG. Validity of head-dipping as a measure of exploration in a modified hole-board. Psychopharmacologia. 1975;44:53–9.

    Article  PubMed  CAS  Google Scholar 

  67. File SE, Wardill AG. The reliability of the hole-board apparatus. Psychopharmacologia. 1975;44:47–51.

    Article  PubMed  CAS  Google Scholar 

  68. Cao BJ, Rodgers, RJ. Dopamine D4 receptor and anxiety: behavioural profiles of clozapine, L-745,870 and L-741,742 in the mouse plus-maze. Eur J Pharmacol. 1997;335:117–125.

    Article  PubMed  CAS  Google Scholar 

  69. Lamberty Y, Gower AJ. Arm width and brightness modulation of spontaneous behaviour of two strains of mice tested in the elevated plus-maze. Physiol Behav. 1996;59:439–44.

    Article  PubMed  CAS  Google Scholar 

  70. Holmes A, Rodgers RJ. Influence of spatial and temporal manipulations on the anxiolytic efficacy of chlordiazepoxide in mice previously exposed to the elevated plus-maze. Neurosci Biobehav Revs. 1999;23:971–980.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The research described in this paper from our laboratory was supported in part by grants from the National Institute of Mental Health (MH06769801), National Science Foundation (IBN03-16083), and U.S. Army Department of Defense (BC051001). Technical assistance provided by Fabiola Estrada, Carolyn Koonce, and Danielle Osborne is greatly appreciated.

Author information

Authors and Affiliations

  1. Department of Psychology Research, The University at Albany, SUNY, Albany, NY, USA

    Alicia A Walf

  2. Departments of Psychology, Biological Sciences, and The Centers for Neuroscience and Life Sciences Research, The University at Albany, SUNY, Albany, NY, USA

    Cheryl A Frye

Authors
  1. Alicia A Walf
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Cheryl A Frye
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. School of Medicine, University of Maryland, W. Baltimore Street 685, Baltimore, 21201, U.S.A.

    Todd D. Gould

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Walf, A.A., Frye, C.A. (2009). Using the Elevated Plus Maze as a Bioassay to Assess the Effects of Naturally Occurring and Exogenously Administered Compounds to Influence Anxiety-Related Behaviors of Mice. In: Gould, T. (eds) Mood and Anxiety Related Phenotypes in Mice. Neuromethods, vol 42. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-303-9_12

Download citation

  • DOI: https://doi.org/10.1007/978-1-60761-303-9_12

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60761-302-2

  • Online ISBN: 978-1-60761-303-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation