J Neurogastroenterol Motil 2018; 24(4): 643-655  https://doi.org/10.5056/jnm18040
Role of High-affinity Choline Transporter 1 in Colonic Hypermotility in a Rat Model of Irritable Bowel Syndrome
Meng-juan Lin1,2 and Bao-ping Yu1,2*
1Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei, China; and 2Key Laboratory of Hubei Province for Digestive System Diseases, Wuhan, Hubei, China
Correspondence to: Bao-ping Yu, MD, PhD
Department of Gastroenterology, Renmin Hospital of Wuhan University, No.238 Jiefang Rd, Wuhan, Hubei 430060, China
Tel: +86-27-68759391, Fax: +86-27-68759391, E-mail: yubp62@163.com
Received: March 3, 2018; Revised: July 13, 2018; Accepted: July 31, 2018; Published online: October 1, 2018.
© The Korean Society of Neurogastroenterology and Motility. All rights reserved.

cc This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Background/Aims
Irritable bowel syndrome (IBS) is a common disease characterized by intestinal dysmotility, the mechanism of which remains elusive. We aim to determine whether the high-affinity choline transporter 1 (CHT1), a determinant of cholinergic signaling capacity, modulates intestinal motility associated with stress-induced IBS.
Methods
A rat IBS model was established using chronic water avoidance stress (WAS). Colonic pathological alterations were evaluated histologically and intestinal motility was assessed by intestinal transit time and fecal water content (FWC). Visceral sensitivity was determined by visceromotor response to colorectal distension. RT-PCR, western blotting, and immunostaining were performed to identify colonic CHT1 expression. Contractility of colonic muscle strips was measured using isometric transducers. enzyme-linked immunosorbent assay was used to measure acetylcholine (ACh). We examined the effects of MKC-231, a choline uptake enhancer, on colonic motility.
Results
After 10 days of WAS, intestinal transit time was decreased and fecal water content increased. Visceromotor response magnitude in WAS rats in response to colorectal distension was significantly enhanced. Protein and mRNA CHT1 levels in the colon were markedly elevated after WAS. The density of CHT1-positive intramuscular interstitial cells of Cajal and myenteric plexus neurons in WAS rats was higher than in controls. Ammonium pyrrolidine dithiocarbamate partly reversed CHT1 upregulation and alleviated colonic hypermotility in WAS rats. Pharmacological enhancement of CHT1 activity by MKC-231 enhanced colonic motility in control rats via upregulation of CHT1 and elevation of ACh production.
Conclusion
Upregulation of CHT1 in intramuscular interstitial cells of Cajal and myenteric plexus neurons is implicated in chronic stress-induced colonic hypermotility by modulation of ACh synthesis via nuclear factor-kappa B signaling.
Keywords: Choline tranporter; Gastrointestinal motility; Interstitial cells of Cajal; Irritable bowel syndrome; Myenteric plexus
Figures
Fig. 1. Immunostaining of high-affinity choline transporter 1 (CHT1) in the colon. (A) Double-immunostaining for CHT1 and transmembrane
member 16A (TMEM16A) in longitudinal colonic sections from control and water avoidance stress (WAS) rats (×400) revealing that CHT1 in the circular muscle of both naive and WAS rats is exclusively expressed in TMEM16A-immunoreactive (IR) cells and the number of CHT1-IR cells in the muscular layer of WAS rats is significantly increased compared to that of control rats. (B) Double-immunofluorescence on whole-mount preparations of colonic myenteric plexus (MP) (×400) illustrating that TMEM16A is robustly expressed in colonic MP and that all the CHT1-IR cells are co-labelled with TMEM16A. The percentage of CHT1-IR cells in the MP is substantially elevated after 10 days of WAS. Different layers were marked out. White arrows indicate CHT1-positive cells that are mainly fusiform in shape and found within circular muscle, exhibiting the morphological characteristics of interstitial cells of Cajal. White arrowheads indicate CHT1-positive cells that are predominantly round in shape and located between circular and longitudinal muscles, showing the hallmark of MP neurons. ML, mucosal layer; SML, submucosal layer; CML, circular muscle layer; LML, longitudinal muscle layer.



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