Roles of Nav1.9 in inflammatory Mediator‐induced Activation of Vagal C‐fibers Innervating Mouse Lungs

Abstract

We used NaV1.9 knockout (KO) mice to evaluate the role of NaV1.9 in chemical activation of vagal C‐fiber terminals in the mouse lungs. Single‐cell rt‐PCR of lung‐labeled neurons revealed that the vast majority of TRPV1 expressing vagal sensory neurons innervating the lungs express NaV1.9 mRNA. The mouse lungs were isolated with the vagus nerves and vagal ganglia intact. Action potentials (APs) were recorded using extracellular electrodes positioned near the relevant cell bodies in the vagal ganglion. The stimuli were applied to the receptive field via perfusion through the mouse trachea. The PAR1 agonist, TFLLR(10mM) evoked 270 ± 62 APs in wild type mice (n=23), but only 85 ± 38 action potentials in NaV1.9 ‐/‐ mice (P< 0.01, n=29). PAR1 is a GPCR stimulus, so we next evaluated a,b‐methylene ATP (1ml, 1µM) that stimulates nodose C‐fibers in the mouse lung via activation of the ionotropic P2X2/3 receptor. a,b‐Methylene ATP evoked 93 ± 18 APs in wildtype mice (n=33), compared to only 13 ± 4 APs the NaV1.9 ‐/‐ mice (P=0.0001, n= 35). The C‐fibers of both WT and NaV1.9 ‐/‐ were equally activated by rapid punctate mechanical stimulation of the receptive field with Von Frey fibers. At the patched clamped cell soma, there was no difference in active or passive electrophysiological properties of nodose neurons isolated from WT versus KO animals; rheobases were also the same with AP threshold averaging 0.34 ± 0.1 and 0.29 ± 0.06 mV/ms, respectively. However, bath‐applied ATP was more effective in evoking action potentials in neurons isolated from WT (40 % responding) vs. KO (0 % responding). The data reveal that NaV1.9 is important in the activation of nodose C‐fiber terminals evoked by inflammatory mediators that act via GPCRs or ionotropic receptors but less critical in the activation by rapid punctate mechanical or electrical stimulation. NaV1.9 provides an intriguing target for treating symptoms caused by inflammatory mediator‐induced C‐fiber activation in the airways, e.g. chronic coughing and excessive reflex bronchospasm and secretions associated with inflammatory airway disease.