We have identified the tracheal and laryngeal afferent nerves regulating cough in anaesthetized guinea-pigs. Cough was evoked
by electrical or mechanical stimulation of the tracheal or laryngeal ...mucosa, or by citric acid applied topically to the trachea
or larynx. By contrast, neither capsaicin nor bradykinin challenges to the trachea or larynx evoked cough. Bradykinin and
histamine administered intravenously also failed to evoke cough. Electrophysiological studies revealed that the majority of
capsaicin-sensitive afferent neurones (both Aδ- and C-fibres) innervating the rostral trachea and larynx have their cell bodies
in the jugular ganglia and project to the airways via the superior laryngeal nerves. Capsaicin-insensitive afferent neurones
with cell bodies in the nodose ganglia projected to the rostral trachea and larynx via the recurrent laryngeal nerves. Severing
the recurrent nerves abolished coughing evoked from the trachea and larynx whereas severing the superior laryngeal nerves
was without effect on coughing. The data indicate that the tracheal and laryngeal afferent neurones regulating cough are polymodal
Aδ-fibres that arise from the nodose ganglia. These afferent neurones are activated by punctate mechanical stimulation and
acid but are unresponsive to capsaicin, bradykinin, smooth muscle contraction, longitudinal or transverse stretching of the
airways, or distension. Comparing these physiological properties with those of intrapulmonary mechanoreceptors indicates that
the afferent neurones mediating cough are quite distinct from the well-defined rapidly and slowly adapting stretch receptors
innervating the airways and lungs. We propose that these airway afferent neurones represent a distinct subtype and that their
primary function is regulation of the cough reflex.
Respiratory viral infection can lead to activation of sensory afferent nerves as indicated by the consequential sore throat, sneezing, coughing, and reflex secretions. In addition to causing ...troubling symptoms, sensory nerve activation likely accelerates viral spreading. The mechanism how viruses activate sensory nerve terminals during infection is unknown. In this study, we investigate whether coronavirus spike protein activates sensory nerves terminating in the airways. We used isolated vagally‐innervated mouse trachea‐lung preparation for two‐photon microscopy and extracellular electrophysiological recordings. Using two‐photon Ca2+ imaging, we evaluated a total number of 786 vagal bronchopulmonary nerves in six experiments. Approximately 49% of the sensory fibers were activated by S1 protein (4 μg/mL intratracheally). Extracellular nerve recording showed the S1 protein evoked action potential discharge in sensory C‐fibers; of 39 airway C‐fibers (one fiber per mouse), 17 were activated. Additionally, Fura‐2 Ca2+ imaging was performed on neurons dissociated from vagal sensory ganglia (n = 254 from 22 mice). The result showed that 63% of neurons responded to S1 protein. SARS‐CoV‐2 S1 protein can lead to direct activation of sensory C‐fiber nerve terminals in the bronchopulmonary tract. Direct activation of C‐fibers may contribute to coronavirus symptoms, and amplify viral spreading in a population.
We investigated whether the S1 subunit of the β‐CoV spike protein could directly activate sensory C‐fibers. Limited presence of ACE2 or TLR4 in mouse vagal sensory C‐fiber neurons suggests that direct activation doesn't occur via ACE2 or TLR4 binding. Other potential pathways, such as galectin‐3 dependent or TRP channel, warrant further investigation. Regardless of the specific mechanism, these findings suggest coronaviruses may directly trigger respiratory C‐fiber nerve endings, potentially intensifying symptoms and aiding viral spread.
Itch, or pruritus, is an important clinical problem whose molecular basis has yet to be understood. Recent work has begun to identify genes that contribute to detecting itch at the molecular level. ...Here we show that Pirt, known to play a vital part in sensing pain through modulation of the transient receptor potential vanilloid 1 (TRPV1) channel, is also necessary for proper itch sensation. Pirt(-/-) mice exhibit deficits in cellular and behavioral responses to various itch-inducing compounds, or pruritogens. Pirt contributes to both histaminergic and nonhistaminergic itch and, crucially, is involved in forms of itch that are both TRPV1-dependent and -independent. Our findings demonstrate that the function of Pirt extends beyond nociception via TRPV1 regulation to its role as a critical component in several itch signaling pathways.
We addressed the mechanism by which antigen contracts trachea isolated from actively sensitized mice. Trachea were isolated
from mice (C57BL/6J) that had been actively sensitized to ovalbumin (OVA). ...OVA (10 μg ml â1 ) caused histamine release (â¼70% total tissue content), and smooth muscle contraction that was rapid in onset and short-lived
( t 1/2 < 1 min), reaching approximately 25% of the maximum tissue response. OVA contraction was mimicked by 5-HT, and responses
to both OVA and 5-HT were sensitive to 10 μ m -ketanserin (5-HT 2 receptor antagonist) and strongly inhibited by atropine (1 μ m ). Epithelial denudation had no effect on the OVA-induced contraction. Histological assessment revealed about five mast cells/tracheal
section the vast majority of which contained 5-HT. There were virtually no mast cells in the mast cell-deficient ( sash â/â) mouse trachea. OVA failed to elicit histamine release or contractile responses in trachea isolated from sensitized mast
cell-deficient ( sash â/â) mice. Intracellular recordings of the membrane potential of parasympathetic neurons in mouse tracheal ganglia revealed
a ketanserin-sensitive 5-HT-induced depolarization and similar depolarization in response to OVA challenge. These data support
the hypothesis that antigen-induced contraction of mouse trachea is epithelium-independent, and requires mast cell-derived
5-HT to activate 5-HT 2 receptors on parasympathetic cholinergic neurons. This leads to acetylcholine release from nerve terminals, and airway smooth
muscle contraction.
Using single-unit extracellular recording techniques, we have examined the role of the vanilloid receptor-1 (VR1 aka TRPV1) in bradykinin-induced activation of vagal afferent C-fiber receptive fields ...in guinea pig isolated airways. Of 17 airway C-fibers tested, 14 responded to bradykinin and capsaicin, 2 fibers responded to neither capsaicin nor bradykinin, and 1 fiber responded to capsaicin but not bradykinin. Thus, every bradykinin-responsive C-fiber was also responsive to capsaicin. Bradykinin (200 microl of 0.3 microM solution) evoked a burst of approximately 130 action potentials in C-fibers. In the presence of the TRPV1 antagonist capsazepine (10 microM), bradykinin evoked 83 +/- 9% (n = 6; P < 0.01) fewer action potentials. Similarly, the TRPV1 blocker, ruthenium red (10 microM), inhibited the number of bradykinin-evoked action potentials by 75 +/- 10% (n = 4; P < 0.05). In the presence of 5,8,11,14-eicosatetraynoic acid (10 microM), an inhibitor of lipoxygenase and cyclooxygenase enzymes, the number of bradykinin-induced action potentials was reduced by 76 +/- 10% (n = 6; P < 0.05). Similarly, a combination of the 12-lipoxygenase inhibitor, baicalein (10 microM) and the 5-lipoxygenase inhibitor ZD2138 6-3-fluoro-5-4-methoxy-3,4,5,6-tetrahydro-2H-pyran-4-yl)phenoxy-methyl-1-methyl-2-quinolone (10 microM) caused significant inhibition of bradykinin-induced responses. Our data suggest a role for lipoxygenase products in bradykinin B(2) receptor-induced activation of TRPV1 in the peripheral terminals of afferent C-fibers within guinea pig trachea.
Lung vagal sensory fibres are broadly categorized as C fibres (nociceptors) and A fibres (non-nociceptive; rapidly and slowly
adapting low-threshold stretch receptors). These afferent fibre types ...differ in degree of myelination, conduction velocity,
neuropeptide content, sensitivity to chemical and mechanical stimuli, as well as evoked reflex responses. Recent studies in
nociceptive fibres of the somatosensory system indicated that the tetrodotoxin-resistant (TTX-R) voltage-gated sodium channels
(VGSC) are preferentially expressed in the nociceptive fibres of the somatosensory system (dorsal root ganglia). Whereas TTX-R
sodium currents have been documented in lung vagal sensory nerves fibres, a rigorous comparison of their expression in nociceptive
versus non-nociceptive vagal sensory neurons has not been carried out. Using multiple approaches including patch clamp electrophysiology,
immunohistochemistry, and single-cell gene expression analysis in the guinea pig, we obtained data supporting the hypothesis
that the TTX-R sodium currents are similarly distributed between nodose ganglion A-fibres and C-fibres innervating the lung.
Moreover, mRNA and immunoreactivity for the TTX-R VGSC molecules Na V 1.8 and Na V 1.9 were present in nearly all neurons. We conclude that contrary to findings in the somatosensory neurons, TTX-R VGSCs are
not preferentially expressed in the nociceptive C-fibre population innervating the lungs.
The influence of NaV1.9 on inflammatory mediator‐induced activation of airway vagal nodose C‐fibres was evaluated by comparing responses in wild‐type versus NaV1.9‐/‐ mice. A single‐cell RT‐PCR ...analysis indicated that virtually all nodose C‐fibre neurons expressed NaV1.9 (SCN11A) mRNA. Using extracellular electrophysiological recordings in an isolated vagally innervated mouse trachea–lung preparation, it was noted that mediators acting via G protein‐coupled receptors (PAR2), or ionotropic receptors (P2×3) were 70–85% less effective in evoking action potential discharge in the absence of NaV1.9. However, there was no difference in action potential discharge between wild‐type and NaV1.9‐/‐ when the stimulus was a rapid punctate mechanical stimulus. An analysis of the passive and active properties of isolated nodose neurons revealed no difference between neurons from wild‐type and NaV1.9‐/‐ mice, with the exception of a modest difference in the duration of the afterhyperpolarization. There was also no difference in the amount of current required to evoke action potentials (rheobase) or the action potential voltage threshold. The inward current evoked by the chemical mediator by a P2×3 agonist was the same in wild‐type versus NaV1.9‐/‐ neurons. However, the current was sufficient to evoke action potential only in the wild‐type neurons. The data support the speculation that NaV1.9 could be an attractive therapeutic target for inflammatory airway disease by selectively inhibiting inflammatory mediator‐associated vagal C‐fibre activation.
Key points
Inflammatory mediators were much less effective in activating the terminals of vagal airway C‐fibres in mice lacking NaV1.9.
The active and passive properties of nodose neurons were the same between wild‐type neurons and NaV1.9‐/‐ neurons.
Nerves lacking NaV1.9 responded, normally, with action potential discharge to rapid punctate mechanical stimulation of the terminals or the rapid stimulation of the cell bodies with inward current injections.
NaV1.9 channels could be an attractive target to selectively inhibit vagal nociceptive C‐fibre activation evoked by inflammatory mediators without blocking the nerves’ responses to the potentially hazardous stimuli associated with aspiration.
figure legend We tested the hypothesis that NaV1.9 plays an important role in the excitability of airway vagal nodose C‐fibre. The data reveal that NaV1.9 is important in the activation of nodose C‐fibre terminals evoked by inflammatory mediators that act via G protein‐coupled receptors or ionotropic receptors but less critical in the activation by rapid punctate mechanical stimulation. The presence of NaV1.9 does not influence the membrane properties of the neurons, nor their excitability as assessed by rapid injections of depolarizing current. NaV1.9 provides an intriguing target for treating chronic coughing and the excessive reflex bronchospasm and secretions associated with inflammatory airway disease.
The influence of Na
1.9 on inflammatory mediator-induced activation of airway vagal nodose C-fibres was evaluated by comparing responses in wild-type versus Na
1.9-/- mice. A single-cell RT-PCR ...analysis indicated that virtually all nodose C-fibre neurons expressed Na
1.9 (SCN11A) mRNA. Using extracellular electrophysiological recordings in an isolated vagally innervated mouse trachea-lung preparation, it was noted that mediators acting via G protein-coupled receptors (PAR2), or ionotropic receptors (P2×3) were 70-85% less effective in evoking action potential discharge in the absence of Na
1.9. However, there was no difference in action potential discharge between wild-type and Na
1.9-/- when the stimulus was a rapid punctate mechanical stimulus. An analysis of the passive and active properties of isolated nodose neurons revealed no difference between neurons from wild-type and Na
1.9-/- mice, with the exception of a modest difference in the duration of the afterhyperpolarization. There was also no difference in the amount of current required to evoke action potentials (rheobase) or the action potential voltage threshold. The inward current evoked by the chemical mediator by a P2×3 agonist was the same in wild-type versus Na
1.9-/- neurons. However, the current was sufficient to evoke action potential only in the wild-type neurons. The data support the speculation that Na
1.9 could be an attractive therapeutic target for inflammatory airway disease by selectively inhibiting inflammatory mediator-associated vagal C-fibre activation. KEY POINTS: Inflammatory mediators were much less effective in activating the terminals of vagal airway C-fibres in mice lacking Na
1.9. The active and passive properties of nodose neurons were the same between wild-type neurons and Na
1.9-/- neurons. Nerves lacking Na
1.9 responded, normally, with action potential discharge to rapid punctate mechanical stimulation of the terminals or the rapid stimulation of the cell bodies with inward current injections. Na
1.9 channels could be an attractive target to selectively inhibit vagal nociceptive C-fibre activation evoked by inflammatory mediators without blocking the nerves' responses to the potentially hazardous stimuli associated with aspiration.
The proper function of airway vagal afferent nerves is essential for the dynamic regulation of breathing and initiation of adequate airway defensive reflexes. A better understanding of the mechanisms ...controlling their activation bears physiological and pathological significance. The voltage‐gated potassium (KV) channels exert powerful controls on the neuronal excitability. Yet their expression and functions in the bronchopulmonary vagal afferent neurons remain largely unexplored. In this study we characterized the expression profile and inhibitory function of α‐dendrotoxin (α‐DTX)‐sensitive D‐type K+ channels composed of members of KV1subfamily of α‐subunits in mouse bronchopulmonary nodose neurons using a combination of single‐neurons RT‐PCR, patch clamp recording, ex‐vivo extracellular recording and two‐photon microscopic Ca2+ imaging techniques. The results showed that the vast majority of retrogradely labeled lung‐specific nodose neurons expressed Kcna1, 2 and 6 coding for the α‐DTX‐sensitive KV1.1, 1.2 and 1.6 α‐subunits, respectively. The D‐type K+ current (IK.D), defined as the α‐DTX‐sensitive K+ current, was recorded in 14/15 bronchopulmonary nodose neurons. It started to be activated at ‐65.7±4.3 mV with 50% channel activation at ‐24 ± 14 mV. IK.D opened rapidly in response to depolarization (activation time constants <10 ms at voltage ≥‐15 mV), and displayed no inactivation during 600 ms at subthreshold/ threshold voltages and little or slow inactivation at more positive potentials (≥‐5 mV). Inhibition of IK.D with 50 nM α‐DTX depolarized mouse pulmonary nodose neurons, increased input resistance, lowered the minimal depolarizing current needed to evoke an action potential, and increased the number and frequency of action potential firing in response to depolarizing current injections. Application of a bolus of 100 nM α‐DTX to the afferent terminals in the mouse lungs via trachea led to overt activation in 49% of capsaicin‐sensitive nodose neurons and in 32% of capsaicin‐insensitive nodose neurons as detected by two‐photon microscope. Further study with extracellular recording revealed that the application of an α‐DTX bolus evoked action potential discharges in 8/19 (42%) nodose C‐fibers terminating in the mouse lungs with a peak firing frequency of 4.4 ± 3.9 Hz. These results indicate that both the soma and terminals of bronchopulmonary nodose afferent nerves express functional IK.D characterized by low‐threshold, fast activation, and lack of or slow inactivation. With these unique biophysical properties, IK.D channels act as an activation brake on the airway vagal afferent nerves by stabilizing the resting membrane potential and by counterbalancing the subthreshold depolarizing forces. Down‐regulation of IK.D, as occurs in many inflammatory diseases, may result in a heightened excitability of airway vagal afferent nerves causing exaggerated dyspnea, excessive mucous secretion, bronchoconstriction and chronic unproductive coughs associated with airway inflammation.
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