Lu Xin Yi, Wen Yu Xiang, Jiang Ni, Zhou Si Qi, Yang Tian, Shi Liang Liang, Guo Hui Min, Zhang Wei, Zhang Qi Peng, Zhang Ni Na
Department of Gastroenterology, Nanjing Drum Tower Hospital Clinical College of Nanjing University of Chinese Medicine, Nanjing, Jiangsu Province, China.
Department of Gastroenterology, Nanjing University Medical School Affiliated Nanjing Drum Tower Hospital, Nanjing, Jiangsu Province, China.
J Dig Dis. 2025 Jan-Feb;26(1-2):62-73. doi: 10.1111/1751-2980.13344. Epub 2025 Apr 14.
Designer receptors exclusively activated by designer drugs (DREADDs)-based chemogenetic tools are commonly used to activate or silence targeted neurons by the agonistic ligand deschloroclozapine (DCZ). This study aimed to establish a Gi-DREADD-based murine model of slow transit constipation (STC) and elucidate its pathophysiological mechanisms.
Adeno-associated virus (AAV) 9-hM4Di was injected into the intestinal wall of mice, and colonic motility was evaluated. The efficiency and immunogenicity of AAV9-hM4Di transduction in the enteric nervous system (ENS) were evaluated. Nitric oxide (NO), acetylcholine (ACh), and substance P (SP) in the colonic tissues and serum samples were analyzed. Calcium (Ca) imaging was performed to evaluate the responses of AAV9-hM4Di on enteric nerves.
AAV9-hM4Di-treated mice showed gastrointestinal motility dysfunction, including reduced fecal pellets and decreased fecal mass and water content. Electrophysiological recording of muscle contraction in the isolated colonic tissues from the chemogenetic mice showed decreased frequency and amplitude after DCZ treatment. The mice treated with AAV9-hM4Di showed the highest levels of transduction in the myenteric plexuses of the ENS. There were no differences in transduction in neuronal nitric oxide synthase (nNOS) and choline acetyltransferase (ChAT) neurons. Gi-DREADDs significantly downregulated ACh but not NO or SP expression in the distal colon in the chemogenetic mice. Ca transient in neurons of ENS in chemogenetic mice was strongly inhibited by DCZ.
It is feasible to apply the DREADDs-based chemogenetic tools to the ENS. Gi-DREADDs can selectively modulate the ENS, inducing STC without excitatory-neural bias, offering targeted neuromodulation for gastrointestinal motility disorders.
基于设计药物特异性激活的设计受体（DREADDs）的化学遗传学工具通常用于通过激动剂配体去氯氯氮平（DCZ）激活或沉默靶向神经元。本研究旨在建立一种基于Gi-DREADD的慢传输型便秘（STC）小鼠模型，并阐明其病理生理机制。
将腺相关病毒（AAV）9-hM4Di注射到小鼠肠壁，评估结肠运动。评估AAV9-hM4Di在肠神经系统（ENS）中的转导效率和免疫原性。分析结肠组织和血清样本中的一氧化氮（NO）、乙酰胆碱（ACh）和P物质（SP）。进行钙（Ca）成像以评估AAV9-hM4Di对肠神经的反应。
AAV9-hM4Di处理的小鼠表现出胃肠运动功能障碍，包括粪便颗粒减少、粪便质量和含水量降低。对化学遗传学小鼠分离的结肠组织中的肌肉收缩进行电生理记录显示，DCZ处理后频率和幅度降低。用AAV9-hM4Di处理的小鼠在ENS的肌间神经丛中表现出最高的转导水平。神经元型一氧化氮合酶（nNOS）和胆碱乙酰转移酶（ChAT）神经元中的转导没有差异。Gi-DREADDs显著下调化学遗传学小鼠远端结肠中ACh的表达，但不影响NO或SP的表达。DCZ强烈抑制化学遗传学小鼠ENS神经元中的钙瞬变。
将基于DREADDs的化学遗传学工具应用于ENS是可行的。Gi-DREADDs可以选择性地调节ENS，诱导STC而无兴奋性神经偏向，为胃肠运动障碍提供靶向神经调节。
