Sex Differences: To freeze or not to freeze

  1. Debra Bangasser  Is a corresponding author
  1. Temple University, United States

Much of our understanding of the neurobiological basis of psychiatric disorders, such as post-traumatic stress disorder and depression, is based on studies that involve rodents. However, although post-traumatic stress disorder and depression occur more frequently in women (Kessler et al., 2012), most preclinical studies in this area have been conducted on male rodents (Beery and Zucker, 2011). This means many of the tests that researchers use to measure fear, anxiety and depressive-like behavior have been validated in male rodents, but not in female rodents (Kokras and Dalla, 2014). Moreover, male and female rodents differ in terms of their size, strength and other characteristics, and these differences can influence how the animals behave. For example, female rats are more active than males, which may explain why some classic tests that define anxiety on the basis of inactivity and avoidance detect fewer behavioral changes in females than in males (Fernandes et al., 1999; Kokras and Dalla, 2014). There is, therefore, a clear need to include female rats in preclinical studies and to allow for the differences between males and females when measuring fear, anxiety and depressive-like behavior. Now, in eLife, Rebecca Shansky of Northeastern University and colleagues – including Tina Gruene as first author – report the results of experiments that found that male and female rats respond to a fearful experience in different ways (Gruene et al., 2015).

Assessing sex differences in fear responses is particularly critical because fear-conditioning tasks are widely used to study the neurobiological underpinnings of learning and remembering distressing or traumatic events. In the typical rodent fear-conditioning task, a tone that initially produces no behavioral response is paired with a slight electric shock to the foot that elicits a fear response. The tone is played so that it precedes and predicts the shock: this results in the rodent becoming ‘conditioned’ to the tone and displaying a fear response when the tone is played without the shock being applied. However, once the conditioned fear response has been learned it can be extinguished, or diminished, by playing the tone without applying the shock a number of times.

The challenge in these experiments is to find a way to measure the degree of fear. Most studies do this by measuring freezing, which is characterized by the rodents ceasing to move. Freezing is generally regarded as a valid measure of fear learning in male rodents. However, Gruene, Shansky and their colleagues – Katelyn Flick and Alexis Stefano of Northeastern, and Stephen Shea of Cold Spring Harbor Laboratories – found that instead of freezing, many female rats display a brief, high-velocity movement termed darting.

Before conditioning, the rats did not dart when they heard the tone. However, darting increased over time as the rats learned to associate the tone with a shock to the foot. Additionally, darting could be extinguished by playing the tone without applying the shock. Taken together, these findings indicate that darting, like freezing, reflects learned fear.

Gruene et al. found that female rats performed more darts per minute than males. However, not all females dart, and not all males freeze: in the experiments approximately 40% of the females engaged in darting behavior, but only about 10% of males did so. These findings reveal sex differences in the presentation and prevalence of darting behavior and illustrate that there is greater variability in the fear responses produced by female rats.

The finding that a higher proportion of female rats dart may explain why previous studies have reported less freezing in females (e.g., Maren et al., 1994; Pryce et al., 1999). Moreover, the Gruene et al. study underscores that darting behavior needs to be assessed in fear-conditioning studies in the future, particularly when females are included. Additional research should also determine the prevalence of conditioned darting in other species, such as mice, which are commonly used in fear-conditioning studies.

More broadly, when including female subjects in tests that were developed using male subjects, it is critical to observe and validate behaviors that are unique to or more commonly expressed by females. Considering these behaviors is timely because the National Institutes of Health in the US has instituted a new policy to promote the inclusion of female rodents in preclinical studies (Clayton and Collins, 2014). With more researchers studying female rodents, a failure to consider sex differences in the behavioral expression of fear, anxiety and depressive-like behavior can potentially lead to inaccurate interpretations of results, thereby hindering scientific progress. The work of Gruene et al. can serve as a model for how to validate common behavioral tests with female rodents. Finally, using behavior procedures that have been validated in both sexes will be critical to understanding the neurobiological processes that underlie post-traumatic stress disorder and depression in both men and women.

References

Article and author information

Author details

  1. Debra Bangasser

    Department of Psychology and Neuroscience Program, Temple University, Philadelphia, United States
    For correspondence
    debra.bangasser@temple.edu
    Competing interests
    The author declares that no competing interests exist.

Publication history

  1. Version of Record published:

Copyright

© 2015, Bangasser

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,622
    views
  • 183
    downloads
  • 3
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Debra Bangasser
(2015)
Sex Differences: To freeze or not to freeze
eLife 4:e13119.
https://doi.org/10.7554/eLife.13119
  1. Further reading

Further reading

  1. Male rats and female rats respond to stress in different ways.

    1. Chromosomes and Gene Expression
    2. Neuroscience
    Robyn D Moir, Emilio Merheb ... Ian M Willis
    Research Article

    Pathogenic variants in subunits of RNA polymerase (Pol) III cause a spectrum of Polr3-related neurodegenerative diseases including 4H leukodystrophy. Disease onset occurs from infancy to early adulthood and is associated with a variable range and severity of neurological and non-neurological features. The molecular basis of Polr3-related disease pathogenesis is unknown. We developed a postnatal whole-body mouse model expressing pathogenic Polr3a mutations to examine the molecular mechanisms by which reduced Pol III transcription results primarily in central nervous system phenotypes. Polr3a mutant mice exhibit behavioral deficits, cerebral pathology and exocrine pancreatic atrophy. Transcriptome and immunohistochemistry analyses of cerebra during disease progression show a reduction in most Pol III transcripts, induction of innate immune and integrated stress responses and cell-type-specific gene expression changes reflecting neuron and oligodendrocyte loss and microglial activation. Earlier in the disease when integrated stress and innate immune responses are minimally induced, mature tRNA sequencing revealed a global reduction in tRNA levels and an altered tRNA profile but no changes in other Pol III transcripts. Thus, changes in the size and/or composition of the tRNA pool have a causal role in disease initiation. Our findings reveal different tissue- and brain region-specific sensitivities to a defect in Pol III transcription.