Activation of TRPV1 and TRPM8 Channels in the Larynx and Associated Laryngopharyngeal Regions Facilitates the Swallowing Reflex

Abstract

The larynx and associated laryngopharyngeal regions are innervated by the superior laryngeal nerve (SLN) and are highly reflexogenic. Transient receptor potential (TRP) channels have recently been detected in SLN innervated regions; however, their involvement in the swallowing reflex has not been fully elucidated. Here, we explore the contribution of two TRP channels, TRPV1 and TRPM8, located in SLN-innervated regions to the swallowing reflex. Immunohistochemistry identified TRPV1 and TRPM8 on cell bodies of SLN afferents located in the nodose-petrosal-jugular ganglionic complex. The majority of TRPV1 and TRPM8 immunoreactivity was located on unmyelinated neurons. Topical application of different concentrations of TRPV1 and TRPM8 agonists modulated SLN activity. Application of the agonists evoked a significantly greater number of swallowing reflexes compared with the number evoked by distilled water. The interval between the reflexes evoked by the agonists was shorter than that produced by distilled water. Prior topical application of respective TRPV1 or TRPM8 antagonists significantly reduced the number of agonist-evoked reflexes. The findings suggest that the activation of TRPV1 and TRPM8 channels present in the swallowing-related regions can facilitate the evoking of swallowing reflex. Targeting the TRP channels could be a potential therapeutic strategy for the management of dysphagia.

Keywords: Dysphagia; Superior laryngeal nerve; Swallowing reflex; TRPM8; TRPV1.

Figure 1

(A) Photomicrographs of TRPV1 localization in the NPJc. Black-filled arrows indicate examples of cells positive for FG, TRPV1 and NF-200. White arrowheads indicate examples of cells positive for both FG and TRPV1, but negative for NF-200. (B) Number of TRPV1-positive cells in nodose (NG), petrosal (PG), and jugular (JG) ganglia. (C) Percentage of TRPV1-positive cells in nodose (NG), petrosal (PG), and jugular (JG) ganglia. FG(+): Cells stained with FG. FG(+)/TRPV1(+): FG-stained cells immune-positive for TRPV1. FG(+)/TRPV1(−): FG-stained cells immune-negative for TRPV1. FG(+)/NF(+): FG-stained cells immune-positive for NF-200. FG(+)/TRPV1(+)/NF(+): FG-stained cells immune-positive for TRPV1 and NF-200. FG(+)/TRPV1(+)/NF(−): FG-stained cells immune-positive for TRPV1 but not NF-200. NPJc: nodose-petrosal-jugular ganglionic complex. FG: fluoro-gold. NF-200: neurofilament-200. IR: immunoreactive. Scale bars represent 100 μm.

Figure 2

(A) Photomicrographs of TRPM8 localization in the NPJc. Black-filled arrows indicate examples of cells positive for FG, TRPM8, and NF-200. White arrowheads indicate examples of cells positive for both FG and TRPM8, but negative for NF-200. (B) Number of TRPM8-positive cells in nodose (NG), petrosal (PG), and jugular (JG) ganglia. (C) Percentage of TRPM8-positive cells in nodose (NG), petrosal (PG), and jugular (JG) ganglia. FG(+): Cells stained with FG. FG(+)/TRPM8(+): FG-stained cells immuno-positive for TRPM8. FG(+)/TRPM8 (−): FG-stained cells immuno-negative for TRPM8. FG(+)/NF(+): FG-stained cells immuno-positive for NF-200. FG(+)/TRPM8(+)/NF(+): FG-stained cells immuno-positive for TRPM8 and NF-200. FG(+)/TRPM8(+)/NF(−): FG-stained cells immuno-positive for TRPM8 but not NF-200. NPJc: nodose-petrosal-jugular ganglionic complex. FG: fluoro-gold. NF-200: neurofilament-200. IR: immunoreactive. Scale bars represent 100 μm.

Figure 3

(A) The response of the superior laryngeal nerve (SLN) to capsaicin and other stimulating solutions. Arrowheads indicate onset of delivery of the stimulating solutions. (B) SLN responses to different concentrations of capsaicin. (C) Comparison of SLN responses to capsaicin, saline, vehicle, and distilled water. The y-axis shows the area of the integrated SLN response calculated for every 2 s from the onset of infusion of stimulating solutions. Arb. Unit: arbitrary unit. S: seconds.

Figure 4

(A) The response of the superior laryngeal nerve (SLN) to menthol and other stimulating solutions. Arrowheads indicate onset of delivery of the stimulating solutions. (B) SLN responses to different concentrations of menthol. (C) Comparison of SLN responses to menthol, saline, vehicle, and distilled water. The y-axis shows the area of the integrated SLN response calculated for every 2 s from the onset of infusion of stimulating solutions. Arb. Unit: arbitrary unit. S: seconds.

Figure 5

Effect of prior application of TRPV1 and TRPM8 antagonists (AMG 9810 and AMTB as TRPV1 and TRPM8 antagonists, respectively) on respective agonist-evoked superior laryngeal nerve (SLN) responses. (A) SLN response following application of TRPV1 antagonist. (B) SLN response following application of TRPM8 antagonist. The y-axis shows the area of the integrated SLN response calculated for every 2 s from the onset of infusion of stimulating solutions. Arb. Unit: arbitrary unit. S: seconds.

Figure 6

(A) Swallowing reflexes evoked by application of capsaicin and other stimulating solutions. Arrowheads indicate onset of delivery of the stimulating solutions. (B) Swallowing reflexes evoked by capsaicin following application of TRPV1 antagonist. (C) Comparison of the number of swallowing reflexes evoked by distilled water and capsaicin. (D) Comparison of the interval between swallowing reflexes evoked by distilled water and capsaicin. Cap: capsaicin.

Figure 7

(A) Swallowing reflexes evoked by application of menthol and other stimulating solutions. Arrowheads indicate onset of delivery of the stimulating solutions. (B) Swallowing reflexes evoked by menthol following application of TRPM8 antagonist. (C) Comparison of the number of swallowing reflexes evoked by distilled water and menthol. (D) Comparison of the interval between swallowing reflexes evoked by distilled water and menthol. Men: Menthol.

Figure 8

Swallowing reflexes evoked by application of distilled water, capsaicin, and menthol before SLN transection (A), following unilateral (B) and following bilateral SLN transection (C). Arrowheads indicate onset of delivery of the stimulating solutions. (D) Comparison of the number of swallowing reflexes before and after unilateral SLN transection evoked by distilled water, menthol, and capsaicin. Men: Menthol, Cap: Capsaicin, Uni. SLN T.: Unilateral transection of the superior laryngeal nerve.