Competing effects of AgRP and POMC neurons on cAMP signaling in downstream neurons in vivo

AgRP 和 POMC 神经元对体内下游神经元 cAMP 信号传导的竞争作用

• 批准号: 10572109
• 负责人: Xingjie Zhang
• 金额: $ 9.15万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10572109
• 关键词: Adenylate Cyclase; Adult; Affect; Animals; Axon; Behavior; Biochemical; Biochemical Process; Body Weight; Brain; Cell Compartmentation; Cells; Cilia; Concentration Camps; Cyclic AMP; Data; Dendrites; Drug Delivery Systems; Eating; Electrophysiology (science); Energy Metabolism; Engineering; Fasting; Feeding behaviors; Felis catus; Genetic study; Goals; Grant; Histology; Human; Hunger; Hypothalamic structure; Image; Individual; Influentials; Intraventricular; Knowledge; Learning; Light; Measurement; Measures; Mentors; Modeling; Molecular; Monitor; Morbid Obesity; Mus; Mutation; Nature; Neurons; Neuropeptides; Obesity; Obesity Epidemic; Optics; PDE4D3; Partner in relationship; Pathway interactions; Peptide Receptor; Peptide Signal Sequences; Peptides; Pharmacologic Substance; Physiology; Production; Protein Engineering; Research; Research Personnel; Satiation; Second Messenger Systems; Signal Transduction; Slice; Synapses; Synaptic plasticity; Techniques; Technology; Testing; Time; Titrations; Training; Viral; career development; design; energy balance; experimental study; feeding; fluorescence lifetime imaging; in vivo; in vivo imaging; insight; meetings; neuronal cell body; novel; obesity treatment; optical sensor; optogenetics; paraventricular nucleus; patch clamp; phosphoric diester hydrolase; prevent; programs; real time monitoring; receptor; receptor function; sensor; skills; subcellular targeting; tool; two-photon

PROJECT SUMMARY Hypothalamic AgRP hunger neurons and POMC satiety neurons use neuropeptides AgRP, NPY, and αMSH to regulate food intake, energy expenditure, and body weight. One important downstream target is the satiety- promoting MC4R receptor-expressing neurons in the paraventricular nucleus of the hypothalamus (PVHMC4R), which receive converging peptidergic signals from AgRP and POMC neurons. However, exactly how PVHMC4R neurons interpret and integrate these peptide signals remains unclear, especially regarding the involvement of the downstream second messenger cAMP. The central unknowns can be summarized into three questions: (i) How do hunger and satiety peptides alter cAMP in PVHMC4R neurons? (ii) How does cAMP regulate the spiking of PVHMC4R neurons? (iii) Where in a PVHMC4R neuron is cAMP signaling most influential in regulating PVHMC4R activity? The main barrier to answering these questions is the lack of tools to measure and manipulate cAMP in PVHMC4R neurons. I have recently overcome this barrier by helping develop and apply a molecular and optical toolset to measure, make, and degrade cAMP during behaviors. The studies and career development activity in this K99/R00 proposal are designed to provide me with the necessary training to initiate an independent research program that applies the new cAMP tools to understand peptide signaling in PVHMC4R neurons. I hypothesize that coordinated, opposing changes in AgRP and POMC neuron activity regulate spiking in PVHMC4R neurons via competing actions on cAMP signaling. To test this hypothesis, I propose the following aims: Aim 1: To image cAMP dynamics in PVHMC4R neurons with the optical sensor cADDis during experimentally induced and feeding-induced peptide release, and to compare cAMP signaling between fasted and fed states. Aim 2: To measure how directly increasing cAMP (using a new optogenetic tool, biPAC) and degrading cAMP (using an engineered phosphodiesterase, PDE4D3-Cat) changes plasticity and spiking of PVHMC4R neurons. Aim 3: To engineer and use subcellularly targeted cADDis, biPAC, PDE4D3-Cat in order to understand local cAMP signaling in PVHMC4R neurons. The proposed research will be mentored by Drs. Mark Andermann and Bradford Lowell, who will also provide guidance on becoming an independent researcher. I will learn new techniques in intraventricular drug delivery, whole-cell patch-clamp, and measurements of energy expenditure from my mentors as well as Drs. Alex Banks, Maria Lehtinen, and Joseph Majzoub. I will receive training in protein engineering from Drs. Shiqiang Gao, Georg Nagel, Bernardo Sabatini, and Gary Yellen. I will acquire new skills from the above experts and from courses and scientific meetings, combined with my existing expertise in cAMP research. This will enable me to understand when, where, and how cAMP is used to titrate PVHMC4R neurons activity. These results will fill an important knowledge gap in developing and understanding treatments for obesity.

项目摘要 下丘脑AgRP饥饿神经元和POMC饱腹神经元使用神经肽AgRP、NPY和αMSH来调节 调节食物摄入、能量消耗和体重。一个重要的下游目标是饱腹感- 促进下丘脑室旁核中表达MC 4 R受体的神经元（PVHMC 4 R）， 其接收来自AgRP和POMC神经元的会聚肽能信号。然而，PVHMC 4 R 神经元解释和整合这些肽信号仍然不清楚，特别是关于参与 下游第二信使cAMP。核心的未知数可以归纳为三个问题： (i)饥饿和饱足肽如何改变PVHMC 4 R神经元中的cAMP？ (ii)cAMP如何调节PVHMC 4 R神经元的发放？ (iii)在PVHMC 4 R神经元中，cAMP信号在调节PVHMC 4 R活性方面最有影响力？ 回答这些问题的主要障碍是缺乏测量和操纵PVHMC 4 R中cAMP的工具。 神经元我最近通过帮助开发和应用分子和光学工具集来克服这一障碍， 在行为过程中测量、制造和降解cAMP。在这方面的研究和职业发展活动 K99/R 00提案旨在为我提供必要的培训，以启动独立研究 该计划应用新的cAMP工具来理解PVHMC 4 R神经元中的肽信号传导。我假设 AgRP和POMC神经元活动的协调、对立变化调节PVHMC 4 R神经元的尖峰发放， 通过对cAMP信号的竞争作用。为了验证这个假设，我提出了以下目标： 目的1：利用光学传感器cADDis在实验过程中成像PVHMC 4 R神经元中的cAMP动态 诱导的和进食诱导的肽释放，并比较禁食和进食状态之间的cAMP信号传导。 目的2：测量如何直接增加cAMP（使用新的光遗传学工具biPAC）和降解cAMP （使用工程化的磷酸二酯酶，PDE 4D 3-Cat）改变PVHMC 4 R神经元的可塑性和尖峰。 目的3：设计和使用亚细胞靶向cADDis、biPAC、PDE 4D 3-Cat，以了解局部 PVHMC 4 R神经元中的cAMP信号传导。 拟议中的研究将由Mark Andermann博士和Bradford Lowell博士指导，他们还将提供 成为独立研究员的指导。我将学习脑室内药物输送的新技术， 全细胞膜片钳，以及我的导师和亚历克斯·班克斯博士对能量消耗的测量， Maria Lehtinen和Joseph Majzoub。我将接受高世强博士的蛋白质工程培训， 内格尔、贝尔纳多·萨巴蒂尼和加里·耶伦。我将从上述专家和课程中获得新的技能 和科学会议，结合我在cAMP研究方面的现有专业知识。这将使我能够 了解cAMP何时、何地以及如何用于滴定PVHMC 4 R神经元活性。这些结果将填补一个 在开发和理解肥胖治疗方面存在重要的知识差距。