Activation of mu-opioid receptors in the gut influences anxiety related behaviour in female rats

Background: The enteric nervous system (ENS) is an important pathway through which the gut communicates with the brain. However, there is little information on how modulating ENS activity influences higher cognitive function. The objective of this experiment was to investigate if modulating the ENS affects anxiety related behaviour in rats. We used loperamide, a μ-opioid receptor agonist that does not cross the blood brain barrier, to manipulate ENS function and assess whether any behavioural effects are associated with changes in gut and brain gene expression, and microbiota profile.  

Methods: Male and female Sprague Dawley rats were treated with an acute dose of loperamide, or control solution and anxiety related behaviour assessed using the Open Field and Elevated-Plus maze tests. Gene expression in the caecum, prefrontal cortex, hippocampus, and amygdala was assessed by RNAseq, and caecal microbiota composition determined by shotgun metagenome sequencing. 

Results: In female rats, loperamide treatment significantly decreased total distance moved (P=0.04) and frequency of supported rearing (P=0.03) compared to controls, which indicated a decrease in exploratory behaviour and increased anxiety. However, loperamide did not significantly alter behaviour in male rats. Similarly, loperamide did not alter gene expression in any brain region in male rats. However, in female rats, loperamide significantly altered expression of 10 genes in the hippocampus (FDR<0.05). In contrast, loperamide altered caecum tissue gene expression and microbiome composition in both male and female rats (FDR<0.05). 

Conclusion: The ability of loperamide to modulate behaviour and brain gene expression indicates that gut to brain signaling involved μ-opioid receptors. Because loperamide slows motility, the changes in the microbiome are likely a consequence of this.