Identification of enteric nerve circuits controlling gut motility

控制肠道运动的肠神经回路的识别

基本信息

批准号：
10652992
负责人：
James J. Galligan
金额：
$ 34.45万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-17 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10652992
关键词：
Afferent Neurons Antibodies Anus Autonomic nervous system Biosensor Blood flow Cavia Choline O-Acetyltransferase Code Colon Cre driver Cre lox recombination system Data Dependovirus Diamond Dinucleoside Phosphates Electrodes Electrophysiology (science)Enteral Enteric Nervous System Enzymes Esthesia Ganglia Gastrointestinal Motility Gastrointestinal tract structure Genes Genetic Goals Individual Interneurons Intestines Ion Channel Knowledge Large Intestine Light LoxP-flanked allele Maps Measurement Measures Mediating Membrane Potentials Methods Microelectrodes Motor Motor Neurons Mus Muscle Myenteric Plexus Nerve Nerve Endings Nerve Fibers Nervous System control Neurons Neurotransmitters Nicotinic Acids Nitric Oxide Nitric Oxide Synthase Nucleotides Nutrient Optics Oral Pathway interactions Periodicals Phenotype Purines Reflex action Relaxation Rhodopsin Satellite Viruses Serotonin Side Signaling Molecule Site Small Intestines Smooth Muscle Stomach Synapses Synaptic Transmission Synaptic Vesicles Techniques Testing Tracer Varicosity Vesicle Water absorption cell motility cell type cholinergic effective therapy gastrointestinal motility disorder neurobiotin neurochemistry neuromuscular transmission neurophysiology neuroregulation neurotransmission novel therapeutics photoactivation promoter protein biomarkers selective expression

项目摘要

Project Summary The enteric nervous system (ENS) is a semi-autonomous division of the autonomic nervous system. The ENS controls gastrointestinal motor function, absorption and secretion of nutrients and water and gut sensation. The nerve circuits controlling these important functions are incompletely mapped. In specific aim 1, we will use an antibody against the vesicular nucleotide transporter (VNUT), a protein marker for purinergic nerves to identify these pathways. Purinergic neurotransmission is important in the ENS but there are no data describing the purinergic neurons or pathways in the ENS. These will be important new data that will broaden our knowledge of enteric synaptic connectivity. In specific aim 2 we will use cre-lox technology and adeno-associated virus (AAV9) transduction of myenteric neurons to express the light-activated ion channel, channel rhodopsin-2 (ChR2) in specific neuronal subtypes. This will allow selective activation of specific functional classes of neurons (interneurons, motorneurons, sensory neurons) to determine their specific synaptic connections and the neurotransmitters that these neurons release. These studies will be done in mice and guinea pigs. We will use cre driver mice where cre is driven by neuron-subtype specific promoter (choline acetyltransferase, purines, nitric oxide synthase, 5-HT, for example) crossed with mice containing the floxed gene encoding ChR2. We will measure excitatory and inhibitory junction potentials (EJPs, IJPs) using microelectrode electrophysiology techniques to determine the neurochemical phenotype of neurons synapsing with excitatory and inhibitory motorneurons supplying the longitudinal and circular muscle layers. In specific aim 3, we will also use the cre- lox approach to study synaptic connections in myenteric ganglia. We will also use electrochemical methods to measure local release of ATP and nitric oxide (NO) from myenteric neurons. We will optically stimulate individual ganglia and make intracellular recordings from myenteric neurons on the oral and anal sides of the site of stimulation. We will use microelectrodes filled with neurobiotin so the recorded neurons can be identified in subsequent immunohistochemical studies to identify the neurochemical phenotype of neurons receiving synaptic input from the optically stimulated neurons. These studies will identify synaptic connections responsible for periodic propulsive colonic contractions. Successful completion of these studies will identify synaptic connections between neurochemically identified subsets of myenteric neurons. These connections control coordinated contractions and relaxation of gut smooth muscle leading to propulsion of gut content. A more complete understanding of these pathways will aid in identifying deficits in neural control of gut motility and identification of new drug or genetic treatments of gut motility disorders.

项目概要肠神经系统（ENS）是自主神经系统的半自主部分。 ENS 控制胃肠道运动功能、营养物质和水的吸收和分泌以及肠道感觉。这控制这些重要功能的神经回路尚未完全绘制出来。在具体目标 1 中，我们将使用针对囊泡核苷酸转运蛋白 (VNUT) 的抗体，这是嘌呤能神经识别的蛋白质标记这些途径。嘌呤能神经传递在 ENS 中很重要，但没有数据描述 ENS 中的嘌呤能神经元或通路。这些将是重要的新数据，将扩大我们的知识肠突触连接。在具体目标 2 中，我们将使用 cre-lox 技术和腺相关病毒 (AAV9) 转导肌间神经元表达光激活离子通道、通道视紫红质-2 (ChR2) 存在于特定的神经元亚型中。这将允许选择性激活特定功能类别的神经元（中间神经元、运动神经元、感觉神经元）以确定它们的特定突触连接和这些神经元释放的神经递质。这些研究将在小鼠和豚鼠身上进行。我们将使用 cre 驱动小鼠，其中 cre 由神经元亚型特异性启动子（胆碱乙酰转移酶、嘌呤、硝酸氧化物合酶，例如 5-HT）与含有编码 ChR2 的 floxed 基因的小鼠杂交。我们将使用微电极电生理学测量兴奋性和抑制性连接电位（EJP、IJP）确定具有兴奋性和抑制性突触的神经元的神经化学表型的技术运动神经元供应纵向和环形肌肉层。在具体目标 3 中，我们还将使用 cre- lox 方法研究肌间神经节突触连接。我们还将使用电化学方法测量肌间神经元 ATP 和一氧化氮 (NO) 的局部释放。我们将对个体进行光学刺激神经节并从口腔侧和肛门侧的肌间神经元进行细胞内记录刺激。我们将使用充满神经生物素的微电极，以便可以识别记录的神经元随后的免疫组织化学研究以确定接受突触的神经元的神经化学表型来自光刺激神经元的输入。这些研究将确定负责的突触连接周期性推进性结肠收缩。成功完成这些研究将确定突触神经化学鉴定的肌间神经元亚群之间的联系。这些连接控制肠道平滑肌的协调收缩和松弛导致肠道内容物的推进。一个更多完全了解这些途径将有助于识别肠道蠕动的神经控制缺陷和肠道动力障碍的新药或基因治疗的鉴定。

项目成果

期刊论文数量（3）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

In Vitro Monitoring of Nitric Oxide Release in the Mouse Colon Using a Boron-Doped Diamond Microelectrode Modified with Platinum Nanoparticles and Nafion.

使用铂纳米粒子和 Nafion 修饰的掺硼金刚石微电极体外监测小鼠结肠中一氧化氮的释放。

DOI：
10.1021/acs.analchem.2c03731
发表时间：
2023
期刊：
Analytical chemistry
影响因子：
7.4
作者：
Henderson,Skye;Bhardwaj,Kirti;Perugachi,Victoria;Espinoza-Montero,Patricio;Galligan,JamesJ;Swain,GregM
通讯作者：
Swain,GregM

Effects of Striatal Amyloidosis on the Dopaminergic System and Behavior: A Comparative Study in Male and Female 5XFAD Mice.

纹状体淀粉样变性对多巴胺能系统和行为的影响：雄性和雌性 5XFAD 小鼠的比较研究。