特异性敲除小鼠肠神经系统多巴胺D2受体促进肠道转运

doi:10.16352/j.issn.1001-6325.2026.06.0765

基础医学与临床 ›› 2026, Vol. 46 ›› Issue (6): 765-771.doi: 10.16352/j.issn.1001-6325.2026.06.0765

特异性敲除小鼠肠神经系统多巴胺D2受体促进肠道转运

汪国庆, 朱愔喆, 王益嘉, 冯小燕^*

首都医科大学基础医学院生理学与病理生理学系, 北京 100069

收稿日期:2025-08-29 修回日期:2025-11-26 出版日期:2026-06-05 发布日期:2026-05-27
通讯作者: ^* fengxy@ccmu.edu.cn
基金资助:
北京市自然科学基金(7222010);国家自然科学基金(32071126)

Specific knockout of dopamine D2 receptor in the enteric nervous system promotes intestinal transit in mice

WANG Guoqing, ZHU Yinzhe, WANG Yijia, FENG Xiaoyan^*

Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Capital Medical University, Beijing 100069, China

Received:2025-08-29 Revised:2025-11-26 Online:2026-06-05 Published:2026-05-27
Contact: ^* fengxy@ccmu.edu.cn

摘要/Abstract

摘要： 目的构建特异性敲除肠神经系统(ENS)多巴胺D2受体(D₂R)的小鼠模型,阐明其促进肠道转运的机制。方法利用Cre-LoxP系统构建特异性敲除ENS D₂R(D₂R^Nmu-CKO)小鼠及其对照组(D₂R^flox/flox)小鼠,免疫荧光染色、DAB染色、RT-qPCR和Western blot等检测D₂R和乙酰胆碱转移酶(ChAT)的表达以及ELISA技术检测乙酰胆碱(ACh)的含量;利用活体小动物X线成像技术检测小鼠肠道转运情况。结果免疫荧光染色证实野生型小鼠的十二指肠肌间神经丛(MP)中D₂R与神经元标志物(PGP 9.5)以及胆碱能神经元标志物(ChAT)3者共存,说明D₂R表达于小鼠十二指肠ENS的胆碱能神经元。鼠尾鉴定结果显示成功构建基因型为Nmu-Cre^+/+D₂R-flox^+/+或Nmu-Cre^+/-D₂R-flox^+/+的D₂R^Nmu-CKO小鼠以及基因型为Nmu-Cre^-/-D₂R-flox^+/+的对照组D₂R^flox/flox小鼠模型。与对照组D₂R^flox/flox小鼠相比,D₂R^Nmu-CKO小鼠十二指肠肌层D₂R mRNA(n=6,P＜0.001)以及蛋白表达(n=8,P＜0.01)水平显著降低,而ChAT mRNA(n=6,P＜0.001)以及蛋白表达(n=6,P＜0.05)水平明显升高,且ACh含量也显著升高(n=8,P＜0.01)。DAB染色进一步显示D₂R^Nmu-CKO小鼠十二指肠MP中ChAT阳性神经元数量明显升高(n=6,P＜0.01)。肠道转运实验表明D₂R^Nmu-CKO小鼠肠道转运速度显著加快,全肠转运时间显著缩短(n=5,P＜0.05)。结论本研究成功构建特异性敲除ENS D₂R小鼠模型,并阐明ENS D₂R缺失通过增加ACh的释放进而促进肠道转运,为特异性干预ENS D₂R治疗功能性消化不良、慢传输型便秘等肠动力障碍疾病的新策略提供理论依据。

关键词: 肠神经系统, 多巴胺D2受体, 乙酰胆碱, 肠道转运

Abstract: Objective To establish a mouse model with specific knockout of dopamine D2 receptor (D₂R) in the enteric nervous system (ENS) and elucidate its mechanism in promoting intestinal motility. Methods The Cre-LoxP system was employed to generate ENS-specific D₂R knockout mice (D₂R^Nmu-CKO) and control mice (D₂R^flox/flox). Expression levels of D₂R and choline acetyltransferase (ChAT) were detected using immunofluorescence staining, DAB staining, RT-qPCR, and Western blot. Acetylcholine (ACh) content was measured by ELISA. Intestinal transit was assessed in vivo using X-ray imaging in live small animals. Results Immunofluo- rescence staining confirmed the co-localization of D₂R with the neuronal marker PGP 9.5 as well as the cholinergic neuronal marker ChAT in the duodenal myenteric plexus (MP) of wild-type mice, indicating that D₂R is expressed in cholinergic neurons of the duodenal ENS. Genotyping confirmed successful establishment of D₂R^Nmu-CKO mice (genotypes: Nmu-Cre^+/+D₂R-flox^+/+ or Nmu-Cre^+/-D₂R-flox^+/+) and control D₂R^flox/flox mice (genotype: Nmu-Cre^-/- D₂R-flox^+/+). Compared with control mice, D₂R^Nmu-CKO mice exhibited significantly reduced mRNA(n=6, P＜0.001) and protein (n=8, P＜0.01) expression of D₂R in the duodenal muscle layer, while mRNA(n=6, P＜0.001) and protein (n=6, P＜0.05) expression of ChAT were markedly increased. The content of ACh was also significantly elevated (n=8, P＜0.01). DAB staining further revealed a significant increase in the number of ChAT-positive neurons in the duodenal MP of D₂R^Nmu-CKO mice (n=6, P＜0.01). Intestinal transit assays demonstrated accelerated intestinal transit and significantly shortened whole-gut transit time in D₂R^Nmu-CKO mice (n=5, P＜0.05). Conclusions This study successfully established an ENS-specific D₂R knockout mouse model and revealed that loss of ENS D₂R promotes intestinal transit by enhancing ACh release. These findings provide a theoretical basis for novel strategies targeting ENS D₂R to treat gastrointestinal motility disorders such as functional dyspepsia and slow-transit constipation.

Key words: enteric nervous system, dopamine D2 receptor, acetylcholine, intestinal transit

中图分类号:

R592

汪国庆, 朱愔喆, 王益嘉, 冯小燕. 特异性敲除小鼠肠神经系统多巴胺D2受体促进肠道转运[J]. 基础医学与临床, 2026, 46(6): 765-771.

WANG Guoqing, ZHU Yinzhe, WANG Yijia, FENG Xiaoyan. Specific knockout of dopamine D2 receptor in the enteric nervous system promotes intestinal transit in mice[J]. Basic & Clinical Medicine, 2026, 46(6): 765-771.

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特异性敲除小鼠肠神经系统多巴胺D2受体促进肠道转运

Specific knockout of dopamine D2 receptor in the enteric nervous system promotes intestinal transit in mice

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