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Key points: The vagus nerve has been implicated in mediating behavioural effects of the gut microbiota on the central nervous system. This study examined whether the secretory products of commensal gut bacteria can modulate the excitability of vagal afferent neurons with cell bodies in nodose ganglia.Cysteine proteases from commensal bacteria increased the excitability of vagal afferent neurons via activation of protease‐activated receptor 2 and modulation of the voltage dependence of Na+ conductance activation.Lipopolysaccharide, a component of the cell wall of gram‐negative bacteria, increased the excitability of nodose ganglia neurons via TLR4‐dependent activation of nuclear factor kappa B.Our study identified potential mechanisms by which gut microbiota influences the activity of vagal afferent pathways, which may in turn impact on autonomic reflexes and behaviour. Behavioural studies have implicated vagal afferent neurons as an important component of the microbiota‐gut‐brain axis. However, the mechanisms underlying the ability of the gut microbiota to affect vagal afferent pathways are unclear. We examined the effect of supernatant from a community of 33 commensal gastrointestinal bacterial derived from a healthy human donor (microbial ecosystem therapeutics; MET‐1) on the excitability of mouse vagal afferent neurons. Perforated patch clamp electrophysiology was used to measure the excitability of dissociated nodose ganglion (NG) neurons. NG neuronal excitability was assayed by measuring the amount of current required to elicit an action potential, the rheobase. MET‐1 supernatant increased the excitability of NG neurons by hyperpolarizing the voltage dependence of activation of Na+ conductance. The increase in excitability elicited by MET‐1 supernatant was blocked by the cysteine protease inhibitor E‐64 (30 nm). The protease activated receptor‐2 (PAR2) antagonist (GB 83, 10 μm) also blocked the effect of MET‐1 supernatant on NG neurons. Supernatant from Lactobacillus paracasei 6MRS, a component of MET‐1, recapitulated the effect of MET‐1 supernatant on NG neurons. Lastly, we compared the effects of MET‐1 supernatant and lipopolysaccharide (LPS) from Escherichia coli 05:B5 on NG neuron excitability. LPS increased the excitability of NG neurons in a toll‐like receptor 4 (TLR4)‐dependent and PAR2‐independent manner, whereas the excitatory effects of MET‐1 supernatant were independent of TLR4 activation. Together, our findings suggest that cysteine proteases from commensal bacteria increase the excitability of vagal afferent neurons by the activation of PAR2. Key points: The vagus nerve has been implicated in mediating behavioural effects of the gut microbiota on the central nervous system. This study examined whether the secretory products of commensal gut bacteria can modulate the excitability of vagal afferent neurons with cell bodies in nodose ganglia.Cysteine proteases from commensal bacteria increased the excitability of vagal afferent neurons via activation of protease‐activated receptor 2 and modulation of the voltage dependence of Na+ conductance activation.Lipopolysaccharide, a component of the cell wall of gram‐negative bacteria, increased the excitability of nodose ganglia neurons via TLR4‐dependent activation of nuclear factor kappa B.Our study identified potential mechanisms by which gut microbiota influences the activity of vagal afferent pathways, which may in turn impact on autonomic reflexes and behaviour. [ABSTRACT FROM AUTHOR]

Peripheral pain signaling reflects a balance of pro and anti-nociceptive influences; the contribution by the gastrointestinal (GI) microbiota to this balance has received little attention. Disorders such as inflammatory bowel disease and irritable bowel syndrome are associated with exaggerated visceral nociceptive actions that may involve altered microbial signaling, particularly given the evidence for bacterial dysbiosis. Thus, we tested whether a community of commensal GI bacteria derived from a healthy human donor (microbial ecosystem therapeutics; MET-1) can affect the excitability of male mouse dorsal root ganglion (DRG) neurons. MET-1 reduced the excitability of DRG neurons by significantly increasing rheobase, decreasing responses to capsaicin (2μM) and reducing action potential discharge from colonic afferent nerves. The increase in rheobase was accompanied by an increase in the amplitude of voltage-gated K+ currents. A cocktail of bacterial protease inhibitors abrogated the effect of MET-1 effects on DRG neuron rheobase. A serine protease inhibitor but not inhibitors of cysteine proteases, acid proteases, metalloproteases, or aminopeptidases abolished the effects of MET-1. The serine protease cathepsin G recapitulated the effects of MET-1 on DRG neurons. Inhibition of protease-activated receptor (PAR)-4, but not PAR-2, blocked the effects of MET-1. Furthermore, Faecalibacterium prausnitzii recapitulated the effects of MET-1 on excitability of DRG neurons. We conclude that serine proteases derived from commensal bacteria can directly impact the excitability of DRG neurons, through PAR-4 activation. The ability of microbiota-neuronal interactions to modulate afferent signaling suggests that therapies that induce or correct microbial dysbiosis may impact visceral pain. [ABSTRACT FROM AUTHOR]