Brain signalling pathways involved in subclinical anxiety and depressed mood can be modulated via the gut brain axis (GBA), providing the potential for diet and dietary components to affect mood. We investigated behavioural, physiological and gut microbiome responses to the Lacticaseibacillus rhamnosus strain HN001 (LactoB HN001™), which has been shown to reduce postpartum anxiety and depression, and a milk fat globule membrane-enriched product, Lipid 70 (Surestart^TM MFGM Lipid 70), which has been implicated in memory in stress-susceptible Wistar Kyoto rats. We examined behaviour in the open field, elevated plus maze and novel object recognition tests in conjunction with the expression of host genes in neuro-signalling pathways, and we also assessed brain lipidomics. Treatment-induced alterations in the caecal microbiome and short-chain fatty acid (SCFA) profiles were also assessed. Neither ingredient induced behavioural changes or altered the brain lipidome (separately or when combined). However, with regard to brain gene expression, the L. rhamnosus HN001 + Lipid 70 combination produced a synergistic effect, reducing GABAA subunit expression in the amygdala (Gabre, Gat3, Gabrg1) and hippocampus (Gabrd). Treatment with L. rhamnosus HN001 alone altered expression of the metabotropic glutamate receptor (Grm4) in the amygdala but produced only minor changes in gut microbiota composition. In contrast, Lipid 70 alone did not alter brain gene expression but produced a significant shift in the gut microbiota profile. Under the conditions used, there was no observed effect on rat behaviour for the ingredient combination. However, the enhancement of brain gene expression by L. rhamnosus HN001 + Lipid 70 implicates synergistic actions on region-specific neural pathways associated with fear, anxiety, depression and memory. A significant shift in the gut microbiota profile also occurred that was mainly attributable to Lipid 70.

Although effects of stress-induced anxiety on the gastrointestinal tract and enteric nervous system (ENS) are well studied, how ENS dysfunction impacts behaviour is not well understood. We investigated whether ENS modulation alters anxiety-related behaviour in rats. We used loperamide, a potent μ-opioid receptor agonist that does not cross the blood–brain barrier, to manipulate ENS function and assess changes in behaviour, gut and brain gene expression, and microbiota profile. Sprague Dawley (male/female) rats were acutely dosed with loperamide (subcutaneous) or control solution, and their behavioural phenotype was examined using open field and elevated plus maze tests. Gene expression in the proximal colon, prefrontal cortex, hippocampus, and amygdala was assessed by RNA-seq and caecal microbiota composition determined by shotgun metagenome sequencing. In female rats, loperamide treatment decreased distance moved and frequency of supported rearing, indicating decreased exploratory behaviour and increased anxiety, which was associated with altered hippocampal gene expression. Loperamide altered proximal colon gene expression and microbiome composition in both male and female rats. Our results demonstrate the importance of the ENS for communication between gut and brain for normo-anxious states in female rats and implicate corticotropin-releasing hormone and gamma-aminobutyric acid gene signalling pathways in the hippocampus. This study also sheds light on sexually dimorphic communication between the gut and the brain. Microbiome and colonic gene expression changes likely reflect localised effects of loperamide related to gut dysmotility. These results suggest possible ENS pharmacological targets to alter gut to brain signalling for modulating mood.

Background: Social isolation (SI) is an established rat model of chronic stress. We applied this to the stress‐sensitive Wistar Kyoto (WKY) strain to explore brain‐to‐gut interactions associated with mood. Whether SI stress‐induced behavioral changes are sex‐specific or if they affect the microbiome in WKY is unknown. We hypothesized individually housed (IH) animals would be more anxious than pair‐housed (PH), with sex differences. Male and female rats were either IH or PH from 70 to 112 days old and behavior was assessed in modified open field (OFTmod), elevated plus maze (EPM), and novel object recognition (NOR) tests. Cecal content DNA was analyzed by shotgun metagenome sequencing.Results: IH rats, particularly females, spent more time in the center of the OFTmod where the semi‐novel feed was presented compared to PH group rats. There was a tendency for greater distance traveled, or potential hyperactivity, in IH female rats. Males stayed in the EPM closed arms more than females. No treatment difference occurred for recognition memory. SI altered cecal microbiome composition in females where housing was associated with seven differentially abundant taxa and 49 differentially abundant KEGG Level 3 ortholog/gene categories. Several relationships were noted between behavioral traits and relative abundance of microbiome taxa. There was a greater shift in female microbiome composition.Conclusions: In summary, behavioral responses to the housing treatment were minimal. IH animals, particularly females, spent more time in the center of an OFT that contained food; this may have been an indication of depression, as opposed to anxiety. Housing status had a differential impact on the microbiome for females compared to males. The associations between cecal microbiota and activity in the modified OFT suggest that dietary interventions that influence the relative abundance of Bifidobacteria, Alistipes, and Muribaculaceae should be explored.

Adolescent social isolation is a known risk factor for anxiety and depression related disorders, yet its effects on brain–gut communication and potential sex differences remain unclear. We hypothesised that isolation would heighten anxiety- and depression-related behaviors and consequently impair memory in both sexes. To investigate this we exposed male (M) and female (F) rats to four weeks of social isolation beginning at 3 weeks of age and assessed behavior, brain and gut gene expression and microbiota, in single- (S) or pair-housed (P) animals. Contrary to our hypothesis, the results showed higher novel object recognition memory in socially-isolated females (FS vs FP). No isolation-induced changes in anxiety-related behaviours were detected in either sex. Social isolation in females (FS vs FP) increased expression of hippocampal Grik5 (glutamate receptor/memory/learning), and decreased expression of prefrontal cortex genes: Mbp, Mobp, Plp1 (neuroplasticity), Cnp (neuroprotection) and Tph2 (serotonin synthesis). There was a trend toward lower microbial diversity in socially-isolated females (FS vs FP). Although no behaviour change was detected in isolated males, amygdala c-Fos (neuronal activity) and prefrontal cortex Gabbr1 (inhibitory) expression were decreased. Il6r, Tgfb1, Tlr9 (immune-related) were increased in the colon (MS vs MP). In both sexes, social isolation increased Tph1 expression in the colon (FS vs FP; MS vs MP). These findings indicate sex-specific responses to adolescent social isolation, with females showing enhanced novel object recognition memory performance alongside changes in genes linked to neuroplasticity and memory, while males showed altered brain and gut gene expression linked to brain neuro-activity and gut-immune function.