High-fat diet impact on the gut microbiota, serotonergic systems within the dorsal raphe nuclei, and anxiety-like behavior

Anxiety and obesity are two conditions that affects the population in a consistent manner. In experimental rats’ model of long-term induced-obesity by HFD intake, anxiety-like behavioral responses have been recently reinforced, whereas a short period-intake, independently of obesity condition, was also associated to this condition. Moreover, HFD intake has a pivotal role in modulating microbiome, thus participating to gut dysbiosis which is an important condition associated to a range of detrimental outcomes, such as intestinal permeability, inflammation, immune reactivity, impaired cognitive function, depression-, and anxiety-like responses, among others. Dysbiotic gut is known to impact the gut-brain-axis (GBA) mostly through endocrine, immune, metabolic, and neural pathways. This condition may influence neural networks including serotonergic system within raphe brain nuclei, limiting in this way gene expression of tph2, htr1a and slc6a4 thus impairing release, storage, signalization, re-uptake, and synthesis of central serotonin (5-HT). In this study, we conducted a 9-week diet protocol to induce obesity and gut microbiome shift in rats, followed by the investigation of anxiety-related defensive behavioral responses using the elevated-plus maze (EPM), light/dark box (LDB), and open-field arena (OF). We also investigated the microbiome through analysis of alpha- and beta-diversity, relative abundance, principal components, correlations, analysis of composition (ANCOM), Firmicutes and Bacteroidetes ratio (F/B), and supervised machine learning. Further, we investigated the differential mRNA expression of limiting 5-HT serotonergic genes – tph2, htr1a, and slc6a4, in the pontine brainstem and raphe nuclei. Here we demonstrated that nine weeks of HFD intake induced obesity, increased fat pad tissues and weight gain, lead to anxiety-related defensive behavioral responses, shifted the microbiome in terms of alpha- and beta-diversity, and impacted the relative abundance. Further, we found that diet was capable to change microbiome from middle-adolescence to late-adolescence, to adult in rats. Within control diet (CD) rats we found increased in Prevotella sp., Prevotella copri, and Prevotella stecorea, and Lactobacillus helveticus, while in HFD we found increases in the species such as Blautia producta, Clostridium spp. and Clostridium celatum. In terms of serotonergic analysis, we found some species to be correlated with increased in gene expression in the caudal aspect of the dorsomedial raphe nucleus (cDRD), which in turns is associated with stress- and anxiety-related behavioral responses. We found that tph2, htr1a and slc6a4 gene expression was overall and subset increased in the pontine brainstem and raphe nuclei – reinforcing the role of HFD and our model for studying microbiome, serotonergic system, and behavioral responses in rats. These data are consistent with the hypothesis that HFD induces obesity and microbial shifts, thus impacting serotonergic-limiting gene expression, and as consequence, arising anxiety-related behavioral responses to stressful stimuli due to enhanced stress reactivity in rats.