Phenotypic diversity of interstitial cells of Cajal and colonic motor functions

Cajal 间质细胞的表型多样性和结肠运动功能

• 批准号: 10359808
• 负责人: Sal Baker
• 金额: $ 39.65万
• 依托单位: UNIVERSITY OF NEVADA RENO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10359808
• 关键词: Address; Affect; Animal Model; Animals; Area; Behavior; Cells; Colon; Colonic Diseases; Confocal Microscopy; Constipation; Coupled; Data; Defect; Distal; Electrophysiology (science); Enteral; Evaluation; Event; Family; Ganglia; Gap Junctions; Gastrointestinal Motility; Gastrointestinal tract structure; Giant Cells; Hormonal; Human; Image; In Situ; Interstitial Cell of Cajal; Intramuscular; Investigation; Knowledge; Learning; Lesion; Light; Mechanics; Mediating; Membrane Potentials; Migrating Myoelectric Complex; Modeling; Molecular; Morphology; Motor; Motor Activity; Motor Neurons; Movement; Mus; Muscle; Nerve; Neurotransmitters; Organ; Outcome; Pacemakers; Pathology; Pattern; Peristalsis; Phase; Phenotype; Physiological; Physiology; Population; Pre-Clinical Model; Rectum; Regulation; Reporter; Reporting; Research; Role; Signal Transduction; Small Intestines; Smooth Muscle Myocytes; Stomach; Stretching; Submucosa; Surface; Techniques; Testing; Text; Therapeutic Agents; Time; Transgenic Organisms; Treatment Efficacy; antagonist; cell injury; cell motility; design; experience; gastrointestinal; hormone regulation; improved; in situ imaging; inhibitor; insight; interstitial cell; motility disorder; mutant; neuroregulation; neurotransmission; optogenetics; receptor; relating to nervous system; response; sensor; tool

Summary Colonic motility is the product of myogenic, neurogenic and endocrinogenic factors that regulate the excitability of smooth muscle cells (SMCs). The traditional concept of ‘myogenic’ has been revised to include regulatory inputs from interstitial cells (interstitial cells of Cajal (ICC) and PDGFRa+ cells). These cells are electrically coupled to SMCs, forming the SIP syncytium. While much has been learned about ICC in other organs of the GI tract, very little is known about the 4 types of ICC in colonic muscles. We developed reporter strains and mice with exclusive expression of optogenetic Ca2+ sensors in ICC. Using these mice it is possible to isolate specific types of ICC for physiological and molecular studies and to image Ca2+ transients in ICC in situ. Our preliminary data show that all ICC in the colon employ brief Ca2+ entry and release events (i.e. Ca2+ transients) to activate Ca2+-activated Cl- channels (CaCC), encoded by Ano1, in ICC. Activation of CaCC initiates inward current, and this has a depolarizing or excitatory impact on the SIP syncytium. Ca2+ transients occur spontaneously in colonic ICC, but they are also regulated by neural and hormonal inputs. Our overarching hypothesis is that Ca2+ transients in ICC are the ‘myogenic’ mechanism that establishes patterning of contractions in colonic muscles. Neural and hormonal modulation of ICC Ca2+ transients establish organ level mixing and propulsive motility. We will address the following questions to investigate this hypothesis: 1. What specific Ca2+ handling behaviors are manifest in the 4 classes of colonic ICC? 2. Is the behavior of ICC affected by neural and hormonal inputs and during colonic motility behaviors such as colonic migrating motor complexes (CMMC)? 3. What is the relationship between Ca2+ transients in ICC and the electrical and mechanical events that generate colonic motility? Ca2+ transients in ICC of GCaMP6f-Kit mice will be imaged in situ using confocal microscopy while recording movements and intracellular electrical activity. Preliminary data show that basal electrical and contractile patterning in colonic muscles are disrupted by ANO1 channel antagonists. These data demonstrate the key importance of ICC in colonic motor activity, because ICC express ANO1 exclusively in GI muscles. Colonic dysmotilities have been associated with reduced populations of ICC. Therefore, we will utilize animal models with reduced ICC to explore how deficiencies in these cells affect local propagating and mixing contractions and propulsive contractions, such as CMMC. Ca2+ transients will also be characterized in animals with reduced ICC to determine how loss of these cells affects spontaneous Ca2+ signaling and neural and hormonal regulation. This study will be the first comprehensive evaluation of the pacemaker activity of colonic ICC, and the first to show that ICC are responsible for developing basal motor patterning in colonic motility. Completion of the specific aims will provide understanding of why ICC loss has deleterious effects on colonic motility and provide new techniques for evaluating the heath/function of specific types of ICC and the efficacy of therapeutic agents designed to improve ICC function in colonic muscles.

摘要 结肠运动是肌源性、神经源性和内分泌因子调节兴奋性的产物。 平滑肌细胞(SMC)。“肌源性”的传统概念已被修订，以包括监管 从间质细胞(Cajal间质细胞(ICC)和PDGFRA+细胞)输入。这些电池是电的 与SMC偶联，形成SIP合胞体。虽然在国际刑事法院的其他机构中已经了解了很多关于国际刑事法院的情况 胃肠道，对结肠肌肉中的4种ICC类型知之甚少。我们开发了报道菌株和 在ICC中独占表达光遗传钙感受器的小鼠。利用这些小鼠，有可能分离出 用于生理和分子研究的特定类型的ICC，以及原位ICC中的钙瞬变图像。我们的 初步数据显示，结肠中的所有ICC都存在短暂的钙进入和释放事件(即钙瞬变)。 激活ICC中由Ano1编码的钙激活的氯通道(CACC)。CACC的激活向内启动 电流，这对SIP合胞体具有去极化或兴奋性影响。出现Ca2+瞬变 它们在结肠ICC中自发存在，但也受到神经和激素输入的调节。我们最重要的是 假说认为，ICC中的钙瞬变是建立心肌细胞构型的“生肌”机制。 结肠肌肉收缩。ICC钙瞬变的神经和激素调节建立器官水平 混合动力和推进动力。我们将回答以下问题来研究这一假设：1.什么 4类结肠ICC？2.ICC的行为受影响吗？ 通过神经和激素输入，以及在结肠运动行为中，如结肠移行运动复合体 (CMMC)？3.ICC内的钙瞬变与电和机械事件有什么关系 会产生结肠动力吗？GCaMP6f-Kit小鼠ICC中的钙瞬变将使用共聚焦进行原位成像 显微镜下记录运动和细胞内的电活动。初步数据显示，基本 结肠肌肉的电和收缩模式被ANO1通道拮抗剂破坏。这些数据 证明ICC在结肠运动活动中的关键重要性，因为ICC仅在GI中表达ANO1 肌肉。结肠动力障碍与ICC的减少有关。因此，我们将利用 用ICC降低的动物模型来探索这些细胞的缺陷如何影响局部繁殖和混合 宫缩和推进性收缩，如CMMC。钙离子瞬变也将在动物身上表现出来。 通过降低ICC来确定这些细胞的丢失如何影响自发的钙信号和神经和 荷尔蒙调节。这项研究将是首次全面评估结肠起搏器的活动。 ICC，并首次表明ICC在结肠运动中负责基础运动模式的形成。 具体目标的完成将有助于理解为什么ICC丢失对结肠有有害影响 并为评估特定类型ICC的健康/功能和疗效提供了新的技术 旨在改善结肠肌肉ICC功能的治疗药物。