The gut microbiota, a complex community of microorganisms in the gastrointestinal tract, significantly impacts brain function through the gut-brain axis. Numerous studies have linked gut microbiota dysbiosis to neurological and neuropsychiatric disorders such as Alzheimer's disease, Parkinson's disease, and depression. Lifestyle factors, particularly exercise, influence gut microbiota composition and have demonstrable benefits on brain health. Exercise promotes adult hippocampal neurogenesis (AHN), a process crucial for learning, memory, and mood regulation. AHN is primarily located in the hippocampus and contributes to cognitive tasks requiring spatial and contextual memory, as well as anxiety-related behaviors. While exercise enhances AHN and improves cognitive function, the interaction between exercise, gut microbiota, and AHN remains poorly understood. This study aimed to determine if exercise could ameliorate the negative consequences of gut microbiota disruption (induced by an antibiotic cocktail) on AHN, cognitive function, and mood-related behaviors in adult male rats.

Extensive research demonstrates a strong bidirectional relationship between the gut microbiota and the brain. Alterations in gut microbiota composition are associated with various neurological and psychiatric diseases. Numerous studies have shown that exercise positively influences gut microbial composition, increasing the abundance of beneficial bacteria and their metabolites, such as short-chain fatty acids (SCFAs). Conversely, sedentary lifestyles are linked to cognitive impairment, while physical activity shows consistent benefits for brain health, increasing the levels of neurotrophic factors like brain-derived neurotrophic factor (BDNF). The hippocampus, a brain region crucial for learning, memory, and mood regulation, is highly responsive to exercise. Exercise is a robust enhancer of adult hippocampal neurogenesis (AHN), a process involving the generation of new neurons throughout life that is vital for several cognitive functions and mood regulation. While the gut microbiome is known to influence cognitive behaviors, neuroplasticity, and AHN, the interplay between exercise and gut microbiota on AHN remains an open question. This study directly addresses this gap by investigating the ability of exercise to counter the effects of gut microbiota disruption on AHN and associated functions.

Adult male Sprague-Dawley rats (9 weeks old) were randomly assigned to four groups: sedentary control (Sed), sedentary with antibiotics (Sed+ABX), voluntary running (Ex), and voluntary running with antibiotics (Ex+ABX). Antibiotics were administered in drinking water for the duration of the study. After two weeks of antibiotic treatment, rats in the Ex and Ex+ABX groups were given access to running wheels. Rats received BrdU injections (150 mg/kg/day for five days) to label newly generated neurons. Behavioral testing (modified spontaneous location recognition task (MSLRT), Y-maze, novelty-suppressed feeding test (NSFT), elevated plus maze (EPM), and forced swim test (FST)) began after three weeks of exercise. At week 10, rats were euthanized, and tissues (brain, colon, caecum, muscle) were collected for immunohistochemical analysis, mRNA expression analysis, and metabolomic profiling. Caecal content and hippocampal tissue underwent mass spectrometry-based untargeted metabolomics. Data analysis included two-way ANOVA (for normally distributed data) and Kruskal-Wallis test (for non-parametric data), followed by post-hoc tests. Metabolomics data were analyzed using FDR-adjusted p-values. Spearman correlations assessed relationships between metabolites and behavioral, biochemical, and immunohistochemical data.

Neither exercise nor antibiotics affected body weight or running distance. Exercise significantly increased PGC1α expression in skeletal muscle and decreased plasma corticosterone levels. Antibiotics induced a low-grade peripheral inflammation, increasing colonic TNFα mRNA expression and plasma TNFα levels in sedentary rats and IL-6 plasma levels in exercised rats. Metabolomic analysis revealed a substantial shift in the caecal metabolome due to antibiotics, with 527 metabolites differentially regulated between Sed+ABX and Sed rats. Exercise had minimal impact on the caecal metabolome. Antibiotics, but not exercise, impaired performance in the large separation paradigm of the MSLRT (pattern separation), while antibiotics and the exercise x antibiotics interaction affected performance in the small separation paradigm (pattern separation). Exercise reversed the antibiotics-induced impairment in the small separation paradigm. Antibiotics increased anxiety-like behavior (increased latency to approach the center in NSFT, reduced time spent in open arms in EPM), which was attenuated by exercise. Exercise increased swimming behavior in FST in rats treated with antibiotics. Antibiotics decreased AHN (BrdU+/NeuN+ cells) in the whole and dorsal hippocampus, and exercise reversed this effect in the dorsal hippocampus. Exercise increased hippocampal and plasma BDNF levels.

This study demonstrates that long-term antibiotic-induced gut microbiota disruption leads to peripheral and central inflammatory changes, cognitive deficits (specifically in pattern separation), and reduced AHN, which are partially reversed by voluntary exercise. The results confirm the gut microbiota's role in mediating cognitive and emotional behaviors. The lack of substantial changes in the hippocampal metabolome despite drastic caecal metabolomic shifts suggests that the hippocampus may be somewhat protected from peripheral changes. Exercise's ability to counteract the negative effects of microbiota disruption may be partially independent of direct hippocampal metabolomic effects, possibly involving other gut-brain communication pathways and factors like circulating hormones and metabolites. The identified correlation between a specific caecal metabolite (ethyl 2-(4-oxo-4,5-dihydro-1,3-thiazol-2-yl)acetate) and various biological and behavioral measures suggests a potential link between gut metabolism, neurogenesis, and behavior, although further investigation is warranted.

This study provides strong evidence that long-term antibiotic-induced gut dysbiosis negatively impacts AHN and related behaviors in adult male rats. Notably, voluntary exercise effectively mitigates these negative consequences, highlighting exercise as a potential therapeutic strategy to counteract antibiotic-induced cognitive and emotional impairments. Future research should explore the specific mechanisms underlying these interactions, including the role of specific gut metabolites and communication pathways between the gut and brain, and investigate these effects in female rodents to account for potential sex differences.

This study was conducted on male rats only, and results may not fully generalize to females. The MSLRT, while sensitive to neurogenesis, might not fully capture exercise's effects on AHN. Further research is needed to explore the specific metabolites and gut-brain communication pathways involved in the observed exercise-mediated mitigation of antibiotic-induced effects. The potential off-target actions of the antibiotics cannot be entirely excluded.