Exercise is More Effective at Altering Gut Microbial Composition and Producing Stable Changes in Lean Mass in Juvenile versus Adult Male F344 Rats

Abstract

The mammalian intestine harbors a complex microbial ecosystem that influences many aspects of host physiology. Exposure to specific microbes early in development affects host metabolism, immune function, and behavior across the lifespan. Just as the physiology of the developing organism undergoes a period of plasticity, the developing microbial ecosystem is characterized by instability and may also be more sensitive to change. Early life thus presents a window of opportunity for manipulations that produce adaptive changes in microbial composition. Recent insights have revealed that increasing physical activity can increase the abundance of beneficial microbial species. We therefore investigated whether six weeks of wheel running initiated in the juvenile period (postnatal day 24) would produce more robust and stable changes in microbial communities versus exercise initiated in adulthood (postnatal day 70) in male F344 rats. 16S rRNA gene sequencing was used to characterize the microbial composition of juvenile versus adult runners and their sedentary counterparts across multiple time points during exercise and following exercise cessation. Alpha diversity measures revealed that the microbial communities of young runners were less even and diverse, a community structure that reflects volatility and malleability. Juvenile onset exercise altered several phyla and, notably, increased Bacteroidetes and decreased Firmicutes, a configuration associated with leanness. At the genus level of taxonomy, exercise altered more genera in juveniles than in the adults and produced patterns associated with adaptive metabolic consequences. Given the potential of these changes to contribute to a lean phenotype, we examined body composition in juvenile versus adult runners. Interestingly, exercise produced persistent increases in lean body mass in juvenile but not adult runners. Taken together, these results indicate that the impact of exercise on gut microbiota composition as well as body composition may depend on the developmental stage during which exercise is initiated.

Fig 1. Running distance and body weight.

A) Weekly total running distance across six weeks of exercise, estimated per rat. Adults ran more in the first half of exercise, whereas juveniles ran more during the second half of exercise, although total distance summed across six weeks did not differ between age groups. B) Body weight across the duration of the experiment; adult runners weighed less than their sedentary counterparts during exercise, then returned to sedentary levels shortly following exercise cessation. Juvenile runners weighed more than their sedentary counterparts toward the end of exercise and continued to weigh more after exercise cessation. Data are represented as mean ± SEM; *p<0.05.

Fig 2. Early life exercise and age altered alpha diversity.

Measures of alpha diversity for adult and juvenile run and sed rats after three days (3d) and six weeks (6 wk) of wheel running, and 25 days following exercise cessation (25d post). A) Shannon entropy, an indicator of an even community structure, was significantly higher in the adults than juveniles. Juvenile runners displayed decreased Shannon entropy overall and at 6 wk. B) Species richness was significantly higher in the adults relative to juveniles. Runners had significantly fewer species overall than their sedentary counterparts, and juvenile runners had significantly fewer species than juvenile sedentary rats 3d following the start of exercise. Data are represented as mean ± SEM; *p<0.05.

Fig 3. Early life exercise and age altered beta diversity.

Principle coordinates analysis (PCoA) using unweighted UniFrac distances with an explicit time axis depicts clustering of microbial communities due to age after three days (3d) and six weeks (6 wk) of exercise and 25 days following exercise cessation (25d post). After 6 wk, a clear clustering of juvenile run versus juvenile sed samples is noticeable.

Fig 4. Effects of early life exercise at the phylum level.

The relative abundance of nine phyla for adult and juvenile run and sed rats following three days (3d) and six weeks (6 wk) of exercise, and 25 days following exercise cessation (25d post). Significant differences in phyla due to exercise were only observed in juvenile runners. Specifically, juvenile onset exercise increased relative abundance of Euryarchaeota and Bacteroidetes and decreased relative abundance of Firmicutes and Proteobacteria, overall as well as at 6 wk. *p<0.05.

Fig 5. Supervised learning analyses: early life exercise altered specific genera.

Classification accuracy generated from supervised learning depicted across each level of taxonomy. A) When samples were collapsed into age of running onset, running status, and running status and time point categories, the highest classification accuracy was observed for age of running onset. B) Next, the algorithm attempted to classify time of sample collection separately for each experimental group. The classification accuracy of predicting time point increased to 16.875 times better than random guessing for juvenile runners only, indicating that certain genera were significantly altered across time in juvenile runners.

Fig 6. Effects of age of exercise onset on body composition across the lifespan.

A) Weekly total running distance across time for the separate cohort used for chemical carcass analyses. Adult runners ran less toward the end of exercise, whereas running distance steadily increased for juvenile runners throughout the duration of exercise. Total distance summed across six weeks did not differ by age. B-D depict body weight, lean mass and fat mass, respectively, for juvenile and adult run and sed chemical carcass rats after six weeks of exercise (6 wk) or 25 days following exercise cessation (25d post). B) Body weight; adult runners weighed less than adult seds at 6 wk, however they returned to sedentary levels 25d post. In contrast, juvenile onset runners consistently weighed more than their sedentary counterparts at both time points. C) Lean mass; juvenile onset runners had sustained increases in lean mass at both time points. D) Fat mass; sustained decreases in fat mass were observed in adult onset runners only. Data are represented as mean ± SEM; *p<0.05.