Temperature-dependent differences in mouse gut motility are mediated by stress

The use of mice as models for human disease and biology is widespread, yet concerns exist regarding the translatability of research findings from mice to humans and the reproducibility of results across different institutions. The National Institutes of Health has highlighted the need to investigate the root causes of these issues, including the influence of environmental factors. One such factor gaining attention is the housing temperature of laboratory mice. Conventionally, mice are housed at approximately 22°C, a temperature not based on evidence but rather on human comfort. A growing body of research suggests that 22°C induces chronic cold stress in mice, leading to increased metabolism, heart rate, blood pressure, altered immune function, and increased tumor growth. These effects are alleviated at thermoneutrality (29-33°C for mice), where metabolic rate is minimal and constant. Studies have shown that mice housed at thermoneutrality (often 30°C) better model humans in various contexts than those housed at 22°C. Despite this, the impact of ambient temperature on mouse gastrointestinal physiology, particularly gut motility, remains largely unexplored. This study aims to address this gap, investigating the link between housing temperature, stress, and gut motility in mice.

Previous research on the effects of ambient temperature on gut motility in non-model organisms has yielded mixed results, with some studies showing faster gastric emptying in cold temperatures and others showing slower gastrointestinal motility in cold temperatures (except in the esophagus and crop). The potential link between environmental temperature and gut motility in mice has been hinted at by studies showing that cold exposure causes corticosterone production (the end hormone of the HPA axis stress response) in rodents, and that stress, via CRH administration or restraint, induces changes in rat gut motility. However, no studies had directly linked the effects of different temperatures on gut motility. This study bridges this knowledge gap by directly examining the relationship between temperature, stress (via the HPA axis), and gut motility in mice.

The study used C57BL/6 and 129x1/SvJ mice raised at either 22°C or 30°C. Gut motility was assessed using several methods: whole gut transit time (WGTT) assay (carmine red dye), gastric emptying (FITC-dextran), small intestinal transit (FITC-dextran and activated charcoal), and colon transit (glass bead expulsion). Gut microbiota composition was analyzed using shotgun metagenomic sequencing of fecal samples. To investigate the role of the gut microbiota, mice were treated with an antibiotic cocktail for two weeks before WGTT assays. Stress hormone levels (corticosterone, ACTH, and CRH) were measured using ELISAs and in situ hybridization. The effects of CRH were investigated through pharmacological intervention (astressin) and genetic deletion of Crh. Ex vivo colonic motility assays were conducted to determine whether intrinsic or extrinsic factors were driving the observed motility differences. Food intake and body weight were also monitored to rule out nutritional influences on the motility phenotype. Statistical analyses included t-tests and Wilcoxon rank-sum tests, as appropriate.

The study found that 22°C mice had WGTT nearly twice as fast as 30°C mice, mainly due to a threefold increase in colon transit speed. While the gut microbiota differed between 22°C and 30°C mice, antibiotic treatment did not abolish the temperature-dependent differences in WGTT, suggesting that microbiota is not the primary driver. 22°C mice showed significantly elevated plasma corticosterone and Crh expression compared to 30°C mice. Pharmacological (astressin) and genetic (Crh-/-) depletion of CRH slowed gut motility in 22°C mice but not in 30°C mice. Ex vivo colonic motility assays indicated that temperature-dependent differences are likely mediated by extrinsic factors, not intrinsic factors within the colon itself. No significant differences in food intake were observed between 22°C and 30°C mice. Acclimation of 30°C mice to 22°C resulted in decreased WGTT, and vice-versa, demonstrating the plasticity of gut motility in response to temperature changes.

These findings demonstrate that housing temperature significantly impacts mouse gut motility. The observed differences are primarily driven by the HPA axis stress response, which is more pronounced at 22°C. This suggests that cold stress, rather than gut microbiota composition or food intake, is the main factor influencing gut motility. The study's results have implications for translational research, highlighting the potential for environmental factors to introduce variability and hinder the reproducibility of mouse model studies. Using thermoneutral housing (30°C) may lead to more consistent and translatable results by minimizing stress-induced alterations in gut motility. Furthermore, the study's findings could influence the design of studies investigating gastrointestinal disorders and the impacts of stress on the gut-brain axis.

This research reveals a previously unknown link between ambient temperature, HPA axis stress, and gut motility in mice. Colder housing temperatures (22°C) induce a significant increase in gut motility primarily through the activation of the HPA axis stress response. These results underscore the importance of considering environmental factors, particularly temperature, in mouse model research to enhance the reproducibility and translatability of findings. Further research should explore the specific signaling pathways involved and examine the broader implications of temperature-induced stress on various physiological processes.

The study did not directly measure the impact of stress induction on gut motility, but relied on the correlation between cold stress and HPA axis activation. A brief exposure of 30°C mice to room temperature during procedures could have introduced an uncontrolled stressor. The influence of metabolic changes associated with thermoneutral housing on gut motility was not directly investigated. The role of water intake differences was also not explored. Although antibiotic treatment successfully reduced fecal microbial load, the possibility of residual uncultured taxa impacting WGTT remains. Future research should address these limitations to further solidify the conclusions drawn.