Mechanisms underlying diverse effects of low-dose embryonic ethanol on development and function of hypocretin/orexin neurons

低剂量胚胎乙醇对下丘脑分泌素/食欲素神经元发育和功能的多种影响的机制

• 批准号: 9886626
• 负责人: SARAH F LEIBOWITZ
• 金额: $ 38.14万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9886626
• 关键词: Ablation; Adult; Affect; Age; Agonist; Alcohol abuse; Alcohol consumption; Anatomy; Anxiety; Appetite Stimulants; Area; Behavior; Behavioral; Biological Models; Brain; CCL2 gene; Calcium; Cells; Chronic; Clinical; Communication; Consumption; Date of birth; Development; Dose; Embryo; Embryonic Development; Ethanol; Exhibits; Exposure to; Fetal Alcohol Exposure; Fetal Alcohol Spectrum Disorder; Foundations; Genetic; Human; Hypothalamic structure; Image; Individual; Inflammatory; Injections; Knock-out; Laboratories; Lasers; Lead; Left; Life; Lifestyle-related condition; Location; Long-Term Effects; Measures; Mediating; Mental disorders; Methodology; Modeling; Molecular; Neuroimmune; Neuroimmune system; Neurons; Neuropeptides; Optics; Pattern; Peptides; Physiological; Play; Pregnancy; Prevention strategy; Property; Publications; Rattus; Research; Resolution; Risk; Rodent; Rodent Model; Role; Side; Signal Transduction; Site; System; Techniques; Testing; Time; Transgenic Organisms; Zebrafish; alcohol effect; alcohol exposure; alcohol risk; alcohol use disorder; base; behavior test; behavioral pharmacology; chemokine; cost; cytotoxic; density; embryonic alcohol exposure; experimental study; fetal; hypocretin; improved; in utero; in vivo; migration; monocyte chemoattractant protein 1 receptor; mutant; neurodevelopment; neurogenesis; neuron development; novel; offspring; optogenetics; pre-clinical; prenatal; prenatal exposure; receptor; relating to nervous system

PROJECT SUMMARY Alcohol use disorder (AUD) is a chronic psychiatric disorder with severe societal consequences that impacts over 15 million adults in the USA and 76 million worldwide. Ethanol exposure early in life, particularly in utero, strikingly alters fetal brain development and increases the risk for AUD. Even exposure to low concentrations of ethanol can produce the most prevalent, milder forms of fetal alcohol spectrum disorder. Our recent studies in the rat demonstrate that prenatal exposure to low levels of ethanol, in addition to increasing ethanol consumption and anxiety in the offspring, stimulates neurogenesis with negligible cytotoxic effects. Interestingly, prenatal ethanol stimulates the density of neurons expressing hypocretin/orexin (Hcrt), an orexigenic neuropeptide located almost exclusively in the hypothalamus that has a major role in promoting AUD-related behaviors. These compelling effects observed in our rodent model led us to seek a simpler vertebrate model to investigate in real- time, at cellular and anatomical levels, the mechanisms involved in ethanol’s effects on embryonic development of Hcrt neurons. The zebrafish (ZF) is a perfect vertebrate model system for our studies, due to its optical accessibility and external development, its small size and low cost, and its comparable CNS that develops early and rapidly alongside a relatively sophisticated behavioral repertoire. With our recent publications showing a conservation of ethanol’s effects across species, we established in our laboratory a variety of techniques particularly suited for comprehensive studies of the ZF brain, including time-lapse live imaging, optogenetics, calcium imaging, and in vivo targeted laser ablation, and obtained preliminary results revealing robust and unexpected changes in Hcrt neurons and neuroimmune systems that may lead to the formation of specific asymmetric Hcrt subpopulations. Based on these new findings, we propose to test the following hypothesis: Embryonic exposure to ethanol at low doses has diverse effects on the development of Hcrt neurons, which are mediated by dynamic changes in local inflammatory chemokine systems and contribute to the formation of asymmetric, abnormally-located Hcrt subpopulations that exhibit altered neural activity and are causally related to disturbances in ethanol consumption and associated behaviors. In 3 specific aims, we plan in ZF embryonically exposed to low-dose ethanol: 1) to thoroughly characterize Hcrt neuronal development along with behavior, under normal conditions and after ethanol exposure, and directly test the behavioral functions of these neurons; 2) to precisely determine, at a single-cell level, if Hcrt neurons in specific subpopulations are unique in their birth date, site of origin, migratory path, and signaling activity, and if they are causally related to behavioral disturbances; and 3) to test the possibility that effects of embryonic ethanol on Hcrt neuronal development and behavior are mediated by local neuroimmune systems, specifically CXCL12a/CXCR4b/CXCR7b and CCL2/CCR2. This research aims to elucidate in depth the dynamic and diverse neuropeptide, neuroimmune and behavioral changes caused by low levels of in utero ethanol exposure that increase the risk for AUD.

项目摘要 酒精使用障碍（AUD）是一种慢性精神疾病，具有严重的社会后果，影响 美国有超过1500万成年人，全球有7600万人。在生命早期，尤其是在子宫内， 显著改变胎儿大脑发育并增加AUD风险。即使暴露在低浓度的 乙醇可产生最普遍的、较温和形式的胎儿酒精谱系障碍。我们最近的研究 大鼠证明，除了增加乙醇消耗量之外，产前暴露于低水平乙醇 和焦虑，刺激神经发生，细胞毒性作用可以忽略不计。有趣的是， 乙醇刺激表达下丘脑泌素/食欲素（Hcrt）的神经元密度， 几乎完全位于下丘脑，在促进AUD相关行为中起主要作用。这些 在我们的啮齿动物模型中观察到的令人信服的效果使我们寻求一种更简单的脊椎动物模型来研究真实的- 在细胞和解剖学水平上，乙醇对胚胎发育影响的机制 Hcrt神经元。斑马鱼（zebrafish，ZF）是一种理想的脊椎动物模型系统，由于其光学特性， 可访问性和外部开发，其体积小，成本低，其可比的中枢神经系统，开发早， 并迅速地与相对复杂的行为库一起。我们最近的出版物显示， 为了保护乙醇对物种的影响，我们在实验室建立了各种技术， 特别适用于ZF大脑的全面研究，包括延时实时成像，光遗传学， 钙成像和体内靶向激光消融，并获得了初步结果，揭示了鲁棒性和 Hcrt神经元和神经免疫系统的意外变化，可能导致特异性 不对称Hcrt亚群。基于这些新的发现，我们建议检验以下假设： 胚胎暴露于低剂量乙醇对Hcrt神经元的发育有不同的影响， 通过局部炎性趋化因子系统的动态变化介导，并有助于 不对称的、位置异常的Hcrt亚群，表现出改变的神经活动，并且与 乙醇消耗和相关行为的干扰。在3个具体目标，我们计划在ZF胚胎 暴露于低剂量乙醇：1）彻底表征Hcrt神经元发育沿着行为， 在正常条件下和乙醇暴露后，直接测试这些神经元的行为功能; 2)为了在单细胞水平上精确地确定特定亚群中的Hcrt神经元是否在其出生时是独特的， 日期，起源地，迁移路径和信号活动，以及它们是否与行为相关 3）测试胚胎乙醇对Hcrt神经元发育和 行为由局部神经免疫系统介导，特别是CXCL 12 a/CXCR 4 b/CXCR 7 b， CCL2/CCR2。本研究旨在深入阐明动态和多样的神经肽，神经免疫和 子宫内低水平乙醇暴露引起的行为变化，增加了AUD的风险。