Study suggests the gut microbiota may be one of the factors determining the physiological inter-sex differences in pain perception

By Tarun Sai LomteReviewed by Danielle Ellis, B.Sc.Jun 10 2022

In a recent study published in Brain, Behavior, and Immunity, researchers investigated whether the gut-brain axis could influence pain thresholds.

Background

Pain is an unpleasant sensory and emotional event, often due to tissue injury. Acute pain is usually short-lived after injury and typically resolves on its own. Contrastingly, failure to resolve the pain or continual nerve stimulation could lead to sensitization, impairment of central nervous system (CNS) mechanisms, and chronic pain. The physiological pain threshold is influenced by biological sex differences and other biopsychosocial factors.

It has been demonstrated that females generally exhibit greater sensitivity to pain than males. Differences in pain perception between adult males and females are profound for pressure and electrical stimuli. The peripheral nervous system (PNS) regulates the pain sensation pathway as it senses sensory variations and relays them to the brain. The CNS processes this information and allows pain perception. Lately, gut microbes have been observed to play critical roles in pain regulatory pathways.

About the study

The current study investigated the diversity of gut microbiota and the relationship of microbial taxa with electrical pain sensitivity in healthy adults (males and females). Participants were adult employees and students of the University Cork College (UCC) aged 18-to-35 years, with a body mass index (BMI) of 30 kg/m­² or lower and a regular menstrual cycle.

Excluded subjects were 1) users of psychotropic drugs and beta-blockers in the last three months, 2) recipients of hormonal treatment in the past three months, except those using hormonal contraceptives, 3) those with gastrointestinal (GI), neurologic diseases, or diabetes, and 4) those who took analgesics 24 hours before measurements. Participants were asked to abstain from strenuous physical activities or alcohol consumption and sleep regularly a day before the experiment.

Males paid a visit to the Department of Neurophysiology once, and females visited thrice across a menstrual cycle. Subjects were asked to provide saliva and stool samples at the test visits. Participants provided four saliva samples collected within an hour of awakening and stool samples collected within 24 hours before test visits. A trained clinician performed quantitative sensory testing; They assessed the pain sensation threshold (PST) to transcutaneous constant current stimulation. Pain tolerance threshold (PTT) and PST were recorded during the stimulation characterized by a manual increase of 5mA (up to 100mA).

Venous blood samples were obtained after the neurophysiology assessment. The soluble cluster of differentiation 12 (sCD14) and lipopolysaccharide-binding protein (LBP) levels were quantified. Further, interleukin (IL)-6, IL-1β, IL-8, interferon (IFN-γ), and tumor necrosis factor (TNF)-α levels were also measured. DNA was extracted from the fecal samples and subject to amplification of the V3-V4 variable region of 16s rRNA. Short-chain fatty acids (SCFAs) in the fecal samples were analyzed. Cortisol levels were measured in saliva samples.

Findings

Fifteen males and females were included in the study, eight women used oral contraceptives, and one used a Nuova ring. No significant differences were evident in PTT or PST between males and females, including those using hormonal contraceptives. When stratified based on contraceptive use, PTT-to-PST ratios were significantly lesser for contraceptive users and non-users during the early follicular (EF) and mid-luteal (ML) phases, respectively. Nonetheless, a significantly lower PTT/PST ratio was observed in all females across the menstrual cycle phases.

The authors determined the relative abundance of bacterial taxa and richness/α or β-diversity indices. There were no differences in α-diversity indices or β-diversity between males and females (irrespective of contraceptive use) and across the menstrual cycle phases. Notably, significant differences were observed for 10 and five taxa across the menstrual cycle of contraceptive users and females with normal menstruation, respectively.

They noted that contraceptive use was associated with an increased relative abundance of bacteria from the Erysipelatoclostridium genus in the late follicular (LF) phase, but not SCFAs. Plasma LBP levels in contraceptive users were higher than in males and non-users during LF and ML phases. There were no differences in sCD14 levels. Moreover, no differences were observed in plasma IFN-γ, IL-8, and TNF-α concentrations.

The team investigated the association between gut microbiota and SCFA levels, LBP, sCD14, pro-inflammatory cytokines, and electrical pain thresholds. Specifically, they focused on the relative abundance of bacterial genera correlated with other parameters in females and males. No significant correlations were observed for males. However, in the LF phase, PTT and PST positively correlated with bacteria from Megasphaera and Prevotella genera, respectively. A significant negative correlation was observed between the Eggerthella genus and sCD14 and between the Rothia genus and TNF-α.

In the ML phase, a positive correlation was observed between the Anaerofustis bacteria and the increase in salivary cortisol levels within 30 minutes of awakening. A negative correlation was found between IL-8 levels and bacteria from Lachnospiraceae UCG-005 in the ML phase. Interestingly, the SCFA data, PST, PTT, and some inflammatory markers correlated significantly within each dataset for females. Of note, PST correlated positively with IL-8 levels in females.

Conclusions

The findings revealed that healthy adult females have a lower PTT-to-PST ratio upon electrical stimulation than men. Further, cortisol awakening levels, somatic pain thresholds, and contraceptives were associated with some bacterial genera in females and not males. The relative abundance of Erysipelatoclostridium bacteria in contraceptive users might suggest that its use stimulated growth.

The results also indicated that hormonal contraceptive use might increase systemic levels of LBP through a hormone-dependent mechanism and not by the increased intestinal permeability. In summary, these data support that gut microbiota might determine the physiologic differences in pain perception between sexes, and further research is required to elucidate the underlying molecular mechanisms.