Action potential (AP) conduction depends on voltage-gated sodium channels, of which there are nine subtypes. The vagus nerve, comprising sensory afferent fibers and efferent parasympathetic fibers, provides autonomic regulation of visceral organs, but the voltage-gated sodium channels (Na(V)1) subtypes involved in its AP conduction are poorly defined. We studied the A- and C-waves of electrically stimulated compound action potentials (CAPs) of the mouse and rat vagus nerves with and without Na(V)1 inhibitor administration: tetrodotoxin (TTX), PF-05089771 (mouse Na(V)1.7), ProTX-II (Na(V)1.7), ICA-121341 (Na(V)1.1, Na(V)1.3, and Na(V)1.6), LSN-3049227 (Na(V)1.2, Na(V)1.6, and Na(V)1.7), and A-803467 (Na(V)1.8). We show that TTX-sensitive Na(V)1 channels are essential for all vagal AP conduction. PF-05089771 but not ICA-121341 inhibited the mouse A-wave, which was abolished by LSN-3049227, suggesting roles for Na(V)1.7 and Na(V)1.2. The mouse C-wave was abolished by LSN-3049227 and a combination of PF-05089771 and ICA-121341, suggesting roles for Na(V)1.7 and Na(V)1.6. The rat A-wave was inhibited by ProTX-II, ICA-121341, and a combination of these inhibitors but only abolished by LSN-3049227, suggesting roles for Na(V)1.7, Na(V)1.6, and Na(V)1.2. The rat C-wave was abolished by LSN-3049227 and a combination of ProTX-II and ICA-121341, suggesting roles for Na(V)1.7 and Na(V)1.6. A-803467 also inhibited the mouse and rat CAP suggesting a cooperative role for the TTX-resistant Na(V)1.8. Overall, our data demonstrate that multiple Na(V)1 subtypes contribute to vagal CAPs, with Na(V)1.7 and Na(V)1.8 playing predominant roles and Na(V)1.6 and Na(V)1.2 contributing to a different extent based on nerve fiber type and species. Inhibition of these Na(V)1 may impact autonomic regulation of visceral organs.NEW & NOTEWORTHY Distinct Na(V)1 channels are involved in action potential (AP) initiation and conduction from afferent terminals within specific organs. Here, we have identified the Na(V)1 necessary for AP conduction in the entire murine and rat vagus nerve. We show TTX-sensitive channels are essential for all AP conduction, predominantly Na(V)1.7 with Na(V)1.2 and Na(V)1.6 playing lesser roles depending on the species and fiber type. In addition, we show that Na(V)1.8 is also essential for most axonal AP conduction.
Publications
2023
2022
2021
OBJECTIVE: Transient receptor potential ankyrin 1 (TRPA1) is an excitatory ion channel expressed on a subset of sensory neurons. TRPA1 is activated by a host of noxious stimuli including pollutants, irritants, oxidative stress and inflammation, and is thought to play an important role in nociception and pain perception. TRPA1 is therefore a therapeutic target for diseases with nociceptive sensory signaling components. TRPA1 orthologs have been shown to have differential sensitivity to certain ligands. Cinnamaldehyde has previously been shown to activate sensory neurons via the selective gating of TRPA1. Here, we tested the sensitivity of cinnamaldehyde-evoked responses in mouse and guinea pig sensory neurons to the pore blocker ruthenium red (RuR). RESULTS: Cinnamaldehyde, the canonical TRPA1-selective agonist, caused robust calcium fluxes in trigeminal neurons dissociated from both mice and guinea pigs. RuR effectively inhibited cinnamaldehyde-evoked responses in mouse neurons at 30 nM, with complete block seen with 3 μM. In contrast, responses in guinea pig neurons were only partially inhibited by 3 μM RuR. We conclude that RuR has a decreased affinity for guinea pig TRPA1 compared to mouse TRPA1. This study provides further evidence of differences in ligand affinity for TRPA1 in animal models relevant for drug development.
Action potentials depend on voltage-gated sodium channels (Na(V)1s), which have nine α subtypes. Na(V)1 inhibition is a target for pathologies involving excitable cells such as pain. However, because Na(V)1 subtypes are widely expressed, inhibitors may inhibit regulatory sensory systems. Here, we investigated specific Na(V)1s and their inhibition in mouse esophageal mechanoreceptors-non-nociceptive vagal sensory afferents that are stimulated by low threshold mechanical distension, which regulate esophageal motility. Using single fiber electrophysiology, we found mechanoreceptor responses to esophageal distension were abolished by tetrodotoxin. Single-cell RT-PCR revealed that esophageal-labeled TRPV1-negative vagal neurons expressed multiple tetrodotoxin-sensitive Na(V)1s: Na(V)1.7 (almost all neurons) and Na(V)1.1, Na(V)1.2, and Na(V)1.6 (in ∼50% of neurons). Inhibition of Na(V)1.7, using PF-05089771, had a small inhibitory effect on mechanoreceptor responses to distension. Inhibition of Na(V)1.1 and Na(V)1.6, using ICA-121341, had a similar small inhibitory effect. The combination of PF-05089771 and ICA-121341 inhibited but did not eliminate mechanoreceptor responses. Inhibition of Na(V)1.2, Na(V)1.6, and Na(V)1.7 using LSN-3049227 inhibited but did not eliminate mechanoreceptor responses. Thus, all four tetrodotoxin-sensitive Na(V)1s contribute to action potential initiation from esophageal mechanoreceptors terminals. This is different to those Na(V)1s necessary for vagal action potential conduction, as demonstrated using GCaMP6s imaging of esophageal vagal neurons during electrical stimulation. Tetrodotoxin-sensitive conduction was abolished in many esophageal neurons by PF-05089771 alone, indicating a critical role of Na(V)1.7. In summary, multiple Na(V)1 subtypes contribute to electrical signaling in esophageal mechanoreceptors. Thus, inhibition of individual Na(V)1s would likely have minimal effect on afferent regulation of esophageal motility.