Establishment of a Mouse Model with Cough Hypersensitivity via Inhalation of Citric Acid

Summary

This protocol describes the development of a mouse model with cough hypersensitivity, which can serve as an ideal model for studying the mechanisms of chronic cough.

Abstract

Cough is one of the most common symptoms of many respiratory diseases. Chronic cough significantly impacts quality of life and imposes a considerable economic burden. Increased cough sensitivity is a pathophysiological hallmark of chronic cough. It has been observed that cough hypersensitivity is related to airway inflammation, remodeling of airway sensory nerves, and alterations in the central nervous system. However, the precise molecular mechanisms remain unclear and require further elucidation using suitable animal models. Previous studies have utilized guinea pigs as models for studying cough, but these models present several experimental limitations, including high costs, a lack of transgenic tools, and a scarcity of commercial reagents. In addition, guinea pigs typically exhibit poor environmental tolerance and high mortality when exposed to stimuli. In contrast, mice are smaller, easier to maintain, more cost-effective, and amenable to genetic manipulation, making them more suitable for mechanistic investigations. In this study, we established a mouse model with cough hypersensitivity via continuous inhalation of citric acid (CA). This model is straightforward to operate and yields reproducible results, making it a valuable tool for further studies on the mechanisms and potential novel treatments for chronic cough.

Introduction

Cough is a crucial defensive reflex that helps clear respiratory secretions or foreign materials from the airway. However, it is also one of the most common symptoms of many respiratory diseases, often prompting patients to seek medical attention¹. Chronic cough, defined as a persistent cough lasting more than 8 weeks in adults, significantly impacts the quality of life, causing issues such as incontinence, insomnia, reflux, and other unpleasant experiences, along with a substantial economic burden^2,3,4. It is widely believed that increased cough sensi....

Protocol

All animal experiment procedures were approved by the Laboratory Animal Ethics Committee of the First Affiliated Hospital of Guangzhou Medical University (20230656). Adult male specific-pathogen-free C57BL/6 mice, aged 8-10 weeks and weighing 20-25 g, were used in this study. The details of the reagents and equipment used are listed in the Table of Materials.

1. Chemical reagent preparation

1. Citric acid (CA) solution preparation
   1. For.......

Discussion

This study successfully established a mouse model with cough hypersensitivity through continuous inhalation of citric acid (CA). This model demonstrated a reliable increase in cough sensitivity for both spontaneous coughs and reflexive coughs elicited by citric acid and capsaicin. Citric acid and capsaicin are widely used to assess cough reflex sensitivity¹⁶.

Several critical steps in this protocol ensure.......

Materials

Name	Company	Catalog Number	Comments
0.9% normal saline	Biosharp	BL158A
Capsaicin	Cayman chemical	92350
Citric Acid	Sigma-Aldrich	C2404
Ethanol	Guangzhou chemical reagent factory	GSHB15-AR-0.5L
First-strand cDNA synthesis kit	TransGen Biotech	AT341
Methacholine	Sigma-Aldrich	A2251
Non-invasive whole-body plethysmography (WBP) system	DSI	601-1400-001
Pentobarbital sodium	Merk	P3761
PerfectStart Green qPCR SuperMix	TransGen Biotech	AQ601
Phosphate Buffered Saline (PBS)	Meilunbio	MA0015
Real-time quantitative PCR detecting system	Bio-rad	CFX  Connect
TRIzol reagent	Invitrogen	15596026CN
Tween-80	Solarbio	T8360-100
Ultrasonic nebulizer	Yuwell	402AI