Pain: Why sex matters

The immune mechanisms underlying hypersensitivity to pain after nerve injury are different in male and female mice.
  1. Josette J Wlaschin
  2. Sangeetha Hareendran
  3. Claire E Le Pichon  Is a corresponding author
  1. Eunice Kennedy Shriver National Institute on Child Health and Human Development, National Institutes of Health, United States
  2. Department of Biology, Johns Hopkins University, United States

Pain cautions our bodies against harmful stimuli – such as a burning flame or the pointy end of a needle – and protects us when we are injured. These stimuli are detected by sensory neurons, which transmit signals to the spinal cord and brain. Damaging these neurons can lead to persistent and chronic pain, but the mechanisms underlying this are not fully understood.

One important player in controlling pain related to nerve damage is the immune system (Calvo et al., 2012; Scholz and Woolf, 2007). Previous work showed that injured sensory neurons release a protein called CSF1 (short for colony stimulating factor 1), which activates microglia, the main immune cell type in the brain and spinal cord. In this activated state, microglia proliferate, change their form and alter their behavior.

In 2016, a group of scientists discovered that male mice became hypersensitive to touch when their microglia were activated by nerve injury or by injecting CSF1 in to the space around the spinal cord (Guan et al., 2016). However, microglia have also been shown to be sexually dimorphic, playing different roles in disease and pain in males and females (Mogil, 2020). Now, in eLife, Allan Basbaum, Anna Molovsky and colleagues from the University of California, San Francisco – including Julia Kuhn and Ilia Vainchtein as co-first authors – report that microglia and another immune cell population respond differently to pain signals in male and female mice (Kuhn et al., 2021).

To investigate the mechanisms underlying hypersensitivity to touch, the team (which includes some of the researchers involved in the 2016 study) damaged the sciatic nerves of male and female mice lacking the gene for the CSF1 protein in their sensory neurons. Pain was assessed using the Von Frey assay, where mice are placed on an elevated grate and their paws are poked with different sized filaments (Decosterd and Woolf, 2000; Shields et al., 2003). Thick filaments will evoke a pain response that causes the mouse to flinch and withdraw its paw; whereas, thinner filaments only elicit this response when mice are hypersensitive to touch.

As shown previously, male mice deficient in CSF1 were not hypersensitive to touch after nerve injury. Female mice lacking CSF1, however, still withdrew their paws when poked with thinner filaments, suggesting that the mechanism underlying hypersensitivity in females is different to males. To confirm these findings, Kuhn et al. injected CSF1 near the spinal cord and assessed pain in the absence of nerve injury. As expected based on the previous results, the male mice became hypersensitive to touch, whereas the females did not (Figure 1). Further experiments examining the genes expressed by microglia after injection of CSF1 revealed that male mice upregulated different genes compared to females, including genes associated with disease, and the activation and recruitment of immune cells.

The differing effects of CSF1 injection on male and female mice.

When CSF1 is injected into wild-type mice, microglia in the spinal cord become activated (red cells) in male mice (top) but not females (middle). In females, regulatory T-cells (Tregs, blue circles) present in the membrane layers surrounding the spinal cord block CSF1 from activating microglia, which remain in the resting state (green cells); when regulatory T-cells are depleted (Treg KO; bottom), the microglia of female mice respond to CSF1 the same way as in males (bottom). During Von Frey pain assessment tests, female mice with depleted levels of regulatory T-cells and male mice exhibit the paw withdrawal response typical of hypersensitivity (top and bottom); however, female mice do not elicit a hypersensitive pain response. This indicates that regulatory T-cells suppress the activation of microglia and development of a pain response after CSF1 injection, but only in female mice.

Image credit: Figure created using

Other types of immune cells are known to influence how the central nervous system works under both normal and diseased conditions. To see if any of these might be involved in female pain sensation, Kuhn et al. examined which immune cells were present in the membrane layers surrounding the spinal cords of mice injected with CSF1. Females were found to have more regulatory T-cells, which are potent inflammation suppressors. Kuhn et al. wondered if having a greater number of regulatory T-cells counteracts the effects of CSF1, so they repeated the experiments in female mice in which regulatory T-cells had been depleted. This revealed that without regulatory T-cells, female mice also develop hypersensitivity after CSF1 injection, and their microglia express a more similar pattern of genes to the microglia of males (Figure 1).

This study demonstrates that the immune system plays different roles in the pain pathways of male and female mice after nerve injury. In male mice, microglia are the major immune cell type driving pain induced by CSF1 injection, while regulatory T-cells repress this pathway in females. This work highlights the need to include males and females in scientific research, and the importance of considering sex-specific approaches for pain management. It also opens up interesting questions for future investigation. For example, it is unclear how regulatory T-cells are recruited in females after CSF1 injection, and the mechanisms underlying pain hypersensitivity in female mice remain to be discovered.


Article and author information

Author details

  1. Josette J Wlaschin

    Josette J Wlaschin is in the Eunice Kennedy Shriver National Institute on Child Health and Human Development, National Institutes of Health, Bethesda, and the Department of Biology, Johns Hopkins University, Baltimore, United States

    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1532-3905
  2. Sangeetha Hareendran

    Sangeetha Hareendran is in the Eunice Kennedy Shriver National Institute on Child Health and Human Development, National Institutes of Health, Bethesda, United States

    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9070-8023
  3. Claire E Le Pichon

    Claire E Le Pichon is in the Eunice Kennedy Shriver National Institute on Child Health and Human Development, National Institutes of Health, Bethesda, United States

    For correspondence
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9274-3615

Publication history

  1. Version of Record published: December 2, 2021 (version 1)


© 2021, Wlaschin et al.

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.


  • 1,497
  • 131
  • 0

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. Josette J Wlaschin
  2. Sangeetha Hareendran
  3. Claire E Le Pichon
Pain: Why sex matters
eLife 10:e74935.
  1. Further reading

Further reading

    1. Neuroscience
    Noah J Steinberg, Zvi N Roth ... Elisha Merriam
    Research Article

    In the ‘double-drift’ illusion, local motion within a window moving in the periphery of the visual field alters the window’s perceived path. The illusion is strong even when the eyes track a target whose motion matches the window so that the stimulus remains stable on the retina. This implies that the illusion involves the integration of retinal signals with non-retinal eye-movement signals. To identify where in the brain this integration occurs, we measured BOLD fMRI responses in visual cortex while subjects experienced the double-drift illusion. We then used a combination of univariate and multivariate decoding analyses to identify (1) which brain areas were sensitive to the illusion and (2) whether these brain areas contained information about the illusory stimulus trajectory. We identified a number of cortical areas that responded more strongly during the illusion than a control condition that was matched for low-level stimulus properties. Only in area hMT+ was it possible to decode the illusory trajectory. We additionally performed a number of important controls that rule out possible low-level confounds. Concurrent eye tracking confirmed that subjects accurately tracked the moving target; we were unable to decode the illusion trajectory using eye position measurements recorded during fMRI scanning, ruling out explanations based on differences in oculomotor behavior. Our results provide evidence for a perceptual representation in human visual cortex that incorporates extraretinal information.

    1. Neuroscience
    Evan D Vickers, David A McCormick
    Tools and Resources

    The flow of neural activity across the neocortex during active sensory discrimination is constrained by task-specific cognitive demands, movements, and internal states. During behavior, the brain appears to sample from a broad repertoire of activation motifs. Understanding how these patterns of local and global activity are selected in relation to both spontaneous and task-dependent behavior requires in-depth study of densely sampled activity at single neuron resolution across large regions of cortex. In a significant advance toward this goal, we developed procedures to record mesoscale 2-photon Ca2+ imaging data from two novel in vivo preparations that, between them, allow for simultaneous access to nearly all 0f the mouse dorsal and lateral neocortex. As a proof of principle, we aligned neural activity with both behavioral primitives and high-level motifs to reveal the existence of large populations of neurons that coordinated their activity across cortical areas with spontaneous changes in movement and/or arousal. The methods we detail here facilitate the identification and exploration of widespread, spatially heterogeneous neural ensembles whose activity is related to diverse aspects of behavior.