Increased acid responsiveness in vagal sensory neurons in a guinea pig model of eosinophilic esophagitis

Abstract

Eosinophilic esophagitis (EoE) is characterized with eosinophils and mast cells predominated allergic inflammation in the esophagus and present with esophageal dysfunctions such as dysphagia, food impaction, and heartburn. However, the underlying mechanism of esophageal dysfunctions is unclear. This study aims to determine whether neurons in the vagal sensory ganglia are modulated in a guinea pig model of EoE. Animals were actively sensitized by ovalbumin (OVA) and then challenged with aerosol OVA inhalation for 2 wk. This results in a mild esophagitis with increases in mast cells and eosinophils in the esophageal wall. Vagal nodose and jugular neurons were disassociated, and their responses to acid, capsaicin, and transient receptor potential vanilloid type 1 (TRPV1) antagonist AMG-9810 were studied by calcium imaging and whole cell patch-clamp recording. Compared with naïve animals, antigen challenge significantly increased acid responsiveness in both nodose and jugular neurons. Their responses to capsaicin were also increased after antigen challenge. AMG-9810, at a concentration that blocked capsaicin-evoked calcium influx, abolished the increase in acid-induced activation in both nodose and jugular neurons. Vagotomy strongly attenuated those increased responses of nodose and jugular neurons to both acid and capsaicin induced by antigen challenge. These data for the first time demonstrated that prolonged antigen challenge significantly increases acid responsiveness in vagal nodose and jugular ganglia neurons. This sensitization effect is mediated largely through TRPV1 and initiated at sensory nerve endings in the peripheral tissues. Allergen-induced enhancement of responsiveness to noxious stimulation by acid in sensory nerve may contribute to the development of esophageal dysfunctions such as heartburn in EoE.

Keywords: antigen; transient receptor potential vanilloid type 1.

Fig. 1.

Ovalbumin (OVA) inhalation increases acid responsiveness in nodose neurons. A and B: fluorescence images of nodose neurons activated with acid (pH 6.4, 5.4, and 4.4) and KCl (50 mM) from naïve and OVA inhalation (OVA) guinea pigs. C: representative traces of nodose neurons from naïve (left) and OVA inhalation (right) guinea pigs that were activated by acid (pH 6.4, 5.4, and 4.4) and KCl (50 mM). D: summary of percentages of acid-responsive nodose neurons in all KCl-responsive nodose neurons from naïve (N = 6) and OVA inhalation (N = 4) guinea pigs. **P < 0.01 and ***P < 0.001 are the levels of significance for naïve vs. OVA inhalation (2-tailed unpaired t-test).

Fig. 2.

OVA inhalation increases acid responsiveness in jugular neurons. A and B: fluorescence images of jugular neurons activated with acid (pH 6.4, 5.4, and 4.4) and KCl (50 mM) from naïve guinea pigs and OVA inhalation guinea pigs. C: representative traces from jugular neurons from naïve (left) and OVA inhalation (right) guinea pigs that were activated by acid (pH 6.4, 5.4, and 4.4) and KCl (50 mM). D: summary of percentages of acid-responsive jugular neurons in KCl-responsive jugular neurons in naïve (N = 6) and OVA inhalation (N = 4) guinea pigs. *P < 0.05 for naïve vs. OVA inhalation.

Fig. 3.

OVA inhalation enhanced capsaicin responses in nodose and jugular neurons. A: representative traces of nodose neurons from naïve (left) and OVA inhalation (right) guinea pigs. The calcium response elicited by capsaicin (100 nM) was significantly enhanced in the OVA inhalation group. B: summary of percentages of capsaicin-responsive nodose neurons related to KCl-responsive nodose neurons. C: representative traces of jugular neurons from naïve (left) and OVA inhalation (right) guinea pigs. The calcium response elicited by capsaicin (100 nM) was enhanced in the OVA inhalation group. D: summary of percentages of capsaicin-responsive jugular neurons in KCl-responsive jugular neurons in naïve (N = 6) and OVA inhalation (N = 4) guinea pigs. ***P < 0.001 for naïve vs. OVA.

Fig. 4.

Transient receptor potential vanilloid type 1 (TRPV1) antagonist AMG-9810 inhibited the acid-evoked currents in nodose and jugular neurons in a dose-dependent manner. A–D: AMG-9810 significantly inhibited acid (pH 4.4)-evoked current in both nodose and jugular neurons (***P < 0.001). E and F: dose-dependence curve of the inhibition effects of AMG-9810 on acid-induced currents in nodose and jugular neurons from naïve guinea pigs. ***P < 0.001 for pH 4.4 vs. pH 4.4 + AMG-9810.

Fig. 5.

TRPV1 antagonist AMG-9810 reverses the acid-induced calcium response in nodose and jugular neurons from both naïve and OVA inhalation guinea pigs. A and B: representative traces of nodose and jugular neurons from naïve (N = 4) guinea pigs. The repeatable calcium response elicited by acid (pH 4.4) was inhibited by AMG-9810 (AMG 1 μM + acid pH 4.4). C and D: representative traces of nodose and jugular neurons from OVA inhalation (N = 3) guinea pigs. The repeatable calcium response elicited by acid (pH 4.4) was inhibited by AMG-9810 (AMG 1 μM + acid pH 4.4). E and F: summary of calcium-imaging experiments as shown in A–D. ***P < 0.001 for pH 4.4 vs. pH 4.4 + AMG-9810.

Fig. 6.

Vagotomy attenuated OVA inhalation-induced enhancements of acid and capsaicin responses in nodose and jugular neurons. A and C: representative traces of acid (pH 4.4)- and capsaicin (100 nM)-responsive nodose and jugular neurons in the control side (with intact vagus, labeled as OVA) and vagotomy side (labeled as OVA + vag) from OVA inhalation guinea pigs. B and D: summary of calcium-imaging experiments. Compared with the control side (with intact vagus), both acid- and capsaicin-responsive nodose and jugular neurons were significantly decreased in the vagotomy side after OVA inhalation. *P < 0.05, **P < 0.01, and ***P < 0.001 for OVA vs. OVA + vagotomy.

Fig. 7.

OVA inhalation increased acid-evoked inward current in Dil-labeled esophageal nodose and jugular neurons. A and B: representative traces of acid-evoked currents in nodose and jugular neurons from naïve and OVA-challenged (OVA) guinea pigs (N = 2 in each group). C and D: acid-elicited currents in nodose and jugular neurons from naïve (both n = 6) and OVA-challenged (both n = 7) guinea pigs. *P < 0.05, **P < 0.01, and ***P < 0.001 for naïve and OVA.