Ting Ruan

Sensitization by Pulmonary Reactive Oxygen Species of Rat Vagal Lung C-Fibers: The Roles of the TRPV1, TRPA1, and P2X Receptors

Sensitization of vagal lung C-fibers (VLCFs) induced by mediators contributes to the pathogenesis of airway hypersensitivity, which is characterized by exaggerated sensory and reflex responses to stimulants. Reactive oxygen species (ROS) are mediators produced during airway inflammation. However, the role of ROS in VLCF-mediated airway hypersensitivity has remained elusive. Here, we report that inhalation of aerosolized 0.05% H2O2 for 90 s potentiated apneic responses to intravenous capsaicin (a TRPV1 receptor agonist), α,β-methylene-ATP (a P2X receptor agonist), and phenylbiguanide (a 5-HT3 receptor agonist) in anesthetized rats. The apneic responses to these three stimulants were abolished by vagatomy or by perivagal capsaicin treatment, a procedure that blocks the neural conduction of VLCFs. The potentiating effect of H2O2 on the apneic responses to these VLCF stimulants was prevented by catalase (an enzyme that degrades H2O2) and by dimethylthiourea (a hydroxyl radical scavenger). The potentiating effect of H2O2 on the apneic responses to capsaicin was attenuated by HC-030031 (a TRPA1 receptor antagonist) and by iso-pyridoxalphosphate-6-azophenyl-2′,5′-disulphonate (a P2X receptor antagonist). The potentiating effect of H2O2 on the apneic responses to α,β-methylene-ATP was reduced by capsazepine (a TRPV1 receptor antagonist), and by HC-030031. The potentiating effect of H2O2 on the apneic responses to phenylbiguanide was totally abolished when all three antagonists were combined. Consistently, our electrophysiological studies revealed that airway delivery of aerosolized 0.05% H2O2 for 90 s potentiated the VLCF responses to intravenous capsaicin, α,β-methylene-ATP, and phenylbiguanide. The potentiating effect of H2O2 on the VLCF responses to phenylbiguanide was totally prevented when all antagonists were combined. Inhalation of 0.05% H2O2 indeed increased the level of ROS in the lungs. These results suggest that 1) increased lung ROS sensitizes VLCFs, which leads to exaggerated reflex responses in rats and 2) the TRPV1, TRPA1, and P2X receptors are all involved in the development of this airway hypersensitivity.

Mediator mechanisms involved in TRPV1, TRPA1 and P2X receptor-mediated sensory transduction of pulmonary ROS by vagal lung C-fibers in rats.

We investigated the mediator mechanisms involved in the sensory transduction of pulmonary reactive oxygen species (ROS) by vagal lung C-fibers in anesthetized rats. Airway challenge of aerosolized H₂O₂ (0.4%) stimulated these afferent fibers. The H₂O₂-induced responses were reduced by a cyclooxygenase inhibitor or ATP scavengers and also attenuated by an antagonist of TRPV1, TRPA1 or P2X receptors. The suppressive effect of the cyclooxygenase inhibitor was not affected by a combined treatment with the TRPV1 or TRPA1 antagonist, but was amplified by a combined treatment with the P2X antagonists. The suppressive effect of ATP scavengers was not affected by a combined treatment with the P2X antagonist, but was amplified by a combined treatment with the TRPV1 or TRPA1 antagonist. Thus, the actions of cyclooxygenase metabolites are mediated through the functioning of the TRPV1 and TRPA1 receptors, whereas the action of ATP is mediated through the functioning of P2X receptors.

A synergistic effect of simultaneous TRPA1 and TRPV1 activations on vagal pulmonary C-fiber afferents

Transient receptor potential ankyrin type 1 (TRPA1) and vanilloid type 1 (TRPV1) receptors are coexpressed in vagal pulmonary C-fiber sensory nerves. Because both these receptors are sensitive to a number of endogenous inflammatory mediators, it is conceivable that they can be activated simultaneously during airway inflammation. This study aimed to determine whether there is an interaction between these two polymodal transducers upon simultaneous activation, and how it modulates the activity of vagal pulmonary C-fiber sensory nerves. In anesthetized, spontaneously breathing rats, the reflex-mediated apneic response to intravenous injection of a combined dose of allyl isothiocyanate (AITC, a TRPA1 activator) and capsaicin (Cap, a TRPV1 activator) was ∼202% greater than the mathematical sum of the responses to AITC and Cap when they were administered individually. Similar results were also observed in anesthetized mice. In addition, the synergistic effect was clearly demonstrated when the afferent activity of single vagal pulmonary C-fiber afferents were recorded in anesthetized, artificially ventilated rats; C-fiber responses to AITC, Cap and AITC + Cap (in combination) were 0.6 ± 0.1, 0.8 ± 0.1, and 4.8 ± 0.6 impulses/s (n = 24), respectively. This synergism was absent when either AITC or Cap was replaced by other chemical activators of pulmonary C-fiber afferents. The pronounced potentiating effect was further demonstrated in isolated vagal pulmonary sensory neurons using the Ca(2+) imaging technique. In summary, this study showed a distinct positive interaction between TRPA1 and TRPV1 when they were activated simultaneously in pulmonary C-fiber sensory nerves.

Menthol suppresses laryngeal C-fiber hypersensitivity to cigarette smoke in a rat model of gastroesophageal reflux disease: the role of TRPM8

Patients with gastroesophageal reflux disease (GERD) display enhanced laryngeal reflex reactivity to stimuli that may be due to sensitization of the laryngeal C-fibers by acid and pepsin. Menthol, a ligand of transient receptor potential melastatin-8 (TRPM8), relieves throat irritation. However, the possibility that GERD induces laryngeal C-fiber hypersensitivity to cigarette smoke (CS) and that menthol suppresses this event has not been investigated. We delivered CS into functionally isolated larynxes of 160 anesthetized rats. Laryngeal pH 5-pepsin treatment, but not pH 5-denatured pepsin, augmented the apneic response to CS, which was blocked by denervation or perineural capsaicin treatment (a procedure that blocks the conduction of C fibers) of the superior laryngeal nerves. This augmented apnea was partially attenuated by capsazepine [an transient receptor potential vanilloid 1 (TRPV1) antagonist], SB-366791 (a TRPV1 antagonist), and HC030031 [a transient receptor potential ankyrin 1 (TRPA1) antagonist] and was completely prevented by a combination of TRPV1 and TRPA1 antagonists. Local application of menthol significantly suppressed the augmented apnea and this effect was reversed by pretreatment with AMTB (a TRPM8 antagonist). Our electrophysiological studies consistently revealed that laryngeal pH 5-pepsin treatment increased the sensitivity of laryngeal C-fibers to CS. Likewise, menthol suppressed this laryngeal C-fiber hypersensitivity and its effect could be reversed by pretreatment with AMTB. Our results suggest that laryngeal pH 5-pepsin treatment increases sensitivity to CS of both TRPV1 and TRPA1, which are presumably located at the terminals of laryngeal C-fibers. This sensory sensitization leads to enhanced laryngeal reflex reactivity and augmentation of the laryngeal C-fiber responses to CS, which can be suppressed by menthol acting via TRPM8.

Effects of simvastatin on pulmonary C-fiber sensitivity in rats with monocrotaline-induced pulmonary hypertension.

BACKGROUND The possible mechanisms of simvastatin attenuating pulmonary hypertension (PH) have been widely investigated in pulmonary vascular and hemodynamic systems, but few studies have examined the difference in respiratory response mediated by pulmonary C fibers (PCF) in animal models of PH. We hypothesized that PCF sensitivity would differ from normal in monocrotaline-induced pulmonary hypertension (MCT-PH) rats and the effects of simvastatin treatment would involve not only the pulmonary circulatory system, but also PCF sensitivity. METHODS The PCF sensitivity was investigated by measuring the apneic durations evoked by 3 chemical stimulants: capsaicin; α,β-methylene-adenosine triphosphate; and phenylbiguanide. The effects of simvastatin on PCF sensitivity were evaluated in the MCT-PH rat model. RESULTS The sensitivity of PCF was increased significantly after monocrotaline (MCT) application for 21 days. Bilateral vagatomy and high-dose perivagal capsaicin (250 μg/ml) treatment both blocked the PCF hypersensitivity induced by MCT. Three days of simvastatin (5 mg/kg) treatment significantly reduced the hypersensitive status of PCF. In MCT-PH rats, reactive oxygen species (ROS) production was significantly elevated in both blood and bronchoalveolar lavage, but both showed a significantly decrease after simvastatin treatment. These potential benefits of simvastatin were all abolished by co-application of tin protoporphyrin-IX (SnPP), a specific heme oxygenase-1 (HO-1) inhibitor. CONCLUSION Simvastatin treatment in MCT-PH rats not only attenuated pulmonary hypertension, but also desensitized PCF hypersensitivity and decreased the production of ROS. These cholesterol-independent effects were mainly through the HO-1 pathway and may all contribute to the therapeutic effects of PH treatment.

N-arachidonyl dopamine sensitizes rat capsaicin-sensitive lung vagal afferents via activation of TRPV1 receptors.

We investigated the effect of N-arachidonyl dopamine (NADA), an endogenous agonist of both transient receptor potential vanilloid 1 (TRPV1) and cannabinoid CB1 receptors, on the sensitivity of rat capsaicin-sensitive lung vagal afferent (CSLVA) fibers. In artificially ventilated rats, an intravenous infusion of NADA (400 microg/kg/ml, 0.5 ml/min for 2 min) mildly elevated the baseline CSLVA fiber activity, whereas it markedly potentiated CSLVA fiber responses to a right atrial injection of capsaicin or adenosine, and to lung inflation. The potentiating effect on CSLVA fiber sensitivity to an adenosine injection or lung inflation was blocked by capsazepine pretreatment (a TRPV1 receptor antagonist), but was unaffected by AM251 pretreatment (a CB1 receptor antagonist). In spontaneously breathing rats, a NADA infusion similarly potentiated the CSLVA fiber-mediated apneic response evoked by an adenosine injection, and this potentiating effect was also prevented by capsazepine pretreatment. We concluded that NADA at the dose tested non-specifically increases CSLVA fiber sensitivity to chemical and mechanical stimulation via activation of TRPV1 receptors.