SERT KO rats are a model of sex specific visceral pain

SERT KO 大鼠是性别特异性内脏疼痛模型

• 批准号: 8302494
• 负责人: James J. Galligan
• 金额: $ 21.86万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-15 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8302494
• 关键词: Acetylcholine; Action Potentials; Affect; Afferent Neurons; Animal Model; Cell physiology; Cells; Chemical Stimulation; Chemicals; Colon; Colorectal; Complex; Coupled; Data; Disease; Drug Delivery Systems; Enterochromaffin Cells; Estradiol; Estrogen Receptor 1; Estrogen Replacements; Estrogens; Estrous Cycle; Exhibits; Female; Functional disorder; GTP-Binding Proteins; Gender; Genes; Genetic Polymorphism; Human; Hypersensitivity; Immunohistochemistry; In Vitro; Ion Channel; Irritable Bowel Syndrome; Knock-out; Label; Lead; Measures; Mechanical Stimulation; Mechanics; Methods; Modeling; Molecular; Mucous Membrane; Neurons; Organic Cation Transporter; Ovariectomy; Pain; Patients; Pharmaceutical Preparations; Play; Population; Property; Publishing; Rattus; Resistance; Role; Safe Sex; Serotonin; Signal Transduction; Sodium Channel; Study models; Techniques; Testing; Tetrodotoxin; Time; Tissues; Up-Regulation; Visceral; Visceral Afferents; Visceral pain; Western Blotting; Whole-Cell Recordings; Woman; afferent nerve; antagonist G; cell motility; dopamine transporter; electrical property; extracellular; human female; immunocytochemistry; male; men; nerve supply; neuronal excitability; noradrenaline transporter; patch clamp; response; serotonin receptor; serotonin transporter; sex

DESCRIPTION (provided by applicant): This project will test the hypothesis that 5-hydroxytryptamine (5-HT, serotonin) and estrogen signaling interact to increase excitability of primary afferent neurons supplying the colon. Increased excitability is caused by changes in ion channel expression that result in visceral hypersensitivity in female rats. This hypothesis will be tested using male and female serotonin transporter (SERT) knockout (KO) rats, which we propose is a unique animal model of gender specific visceral hypersensitivity. The underlying pathophysiology of visceral pain is unclear and this is partly due to a lack of animal models where mechanistic and interventional studies can be conducted. However, published data indicate that alterations in 5-HT signaling may play a role in humans. In addition, visceral pain i more common in women than in men, suggesting that there are interactions between 5-HT and gender in the genesis of visceral pain. The overall hypothesis will be tested in 3 specific aims. Specific aim 1 will test the hypothesis that there is increased extracellular availability of 5-HT n vitro in the colon of wild type (WT) and SERT KO rats and that 5-HT release from enterochromaffin (EC) cells is unaffected by the SERT KO. Amperometry will measure 5-HT near the mucosa in response to mechanical and chemical mucosal stimulation. Immunohistochemical (IHC) and Western blot techniques will be used to verify SERT deletion and to assess 5-HT-containing EC cells in the gut of WT and SERT KO rats. IHC localization of the dopamine and norepinephrine transporters and organic cation transporters will be assessed to determine if their expression increases in SERT KO rats. Specific aim 2 will test the hypothesis that estrogen interacts with 5-HT to cause visceral hypersensitivity in female SERT KO rats. The visceromotor response to colorectal balloon distention will be used to measure visceral sensitivity in intact and ovariectomized female WT and SERT KO rats with and without estrogen replacement. In Specific Aim 3, the functional properties of colon projecting sensory neurons maintained in short term primary culture will be studied. These studies will test the hypothesis that colon projecting sensory nerves from female but not male SERT KO rats exhibit increased excitability when studied using whole cell patch clamp methods in vitro. The increased excitability is proposed to be due to interactions between 5-HT and estrogen on sensory neurons that lead to upregulation of tetrodotoxin-resistant sodium channels. These studies will show that increased 5-HT availability in female SERT KO rats alters visceral sensitivity as occurs in female human irritable bowel syndrome patients. The data would indicate that the SERT KO rat is a model for studying changes in the sensory nerve supply of the gut that leads to visceral hypersensitivity. PUBLIC HEALTH RELEVANCE: Gender-related visceral pain associated with gut motility disturbances affects up to 20% of the U.S. population. The proposed studies will attempt to establish an animal model of gender-related visceral pain that can be used to identify the pathophysiological mechanisms responsible for this common disorder. The animal model could also be used to develop new drug treatments for visceral pain.

描述（由申请人提供）：该项目将检验以下假设：5-羟色胺（5-HT，5-羟色胺）和雌激素信号传导相互作用以增加提供结肠的原发性传入神经元的兴奋性。兴奋性提高是由于离子通道表达的变化引起的，导致雌性大鼠内脏超敏反应。这个假设将是 使用雄性和雌性5-羟色胺转运蛋白（SERT）敲除（KO）大鼠进行测试，我们建议这是一种独特的动物模型，是性别特定的内脏超敏反应。内脏疼痛的潜在病理生理尚不清楚，这部分是由于缺乏可以进行机械和介入研究的动物模型。但是，已发布的数据表明，5-HT信号的变化可能在人类中起作用。此外，在女性中，内脏疼痛I在女性中比男性更普遍，这表明5-HT和性别之间存在相互作用。总体假设将以3个特定目的进行检验。具体目标1将检验以下假设：在野生型（WT）和SERT KO大鼠的结肠中，细胞外可用性增加，并且从Enterochromaffin（EC）细胞中释放了5-HT释放，不受Sert KO的影响。响应机械和化学粘膜刺激的粘膜附近的安培法将测量5-HT。免疫组织化学（IHC）和蛋白质印迹技术将用于验证SERT缺失并评估WT和Sert KO大鼠肠道中含5-HT的EC细胞。将评估多巴胺和去甲肾上腺素转运蛋白和有机阳离子转运蛋白的IHC定位，以确定其表达在SERT KO大鼠中是否增加。特定的目标2将检验雌激素与5-HT相互作用的假设，导致雌性Sert KO大鼠内脏性超敏反应。内脏运动对结直肠球囊延伸的反应将用于测量完整和卵巢切除的雌性WT和SERT KO大鼠的内脏敏感性，并没有替代雌激素。在特定的目标3中，将研究在短期原发性培养中维持的结肠投射感觉神经元的功能特性。这些研究将检验以下假设：在体外使用全细胞斑块方法研究时，使用全细胞贴剂方法研究了女性而不是雄性SERT KO大鼠的结肠突出感官神经表现出更大的兴奋性。提高的兴奋性被认为是由于5-HT与雌激素之间的相互作用在感觉神经元上的相互作用，从而导致抗四毒素耐钠通道的上调。这些研究将表明，雌性Sert KO大鼠的5-HT可用性增加了肠易激综合症患者的内脏敏感性。数据将表明SERT KO大鼠是研究肠道的感觉神经供应变化的模型，从而导致内脏超敏反应。 公共卫生相关性：与肠道运动障碍相关的与性别相关的内脏疼痛影​​响多达20％的美国人群。拟议的研究将试图建立与性别相关的内脏疼痛的动物模型，该模型可用于识别导致这种常见疾病的病理生理机制。该动物模型也可用于开发新药物治疗内脏疼痛。