Role of an arousal circuit in respiration and opioid-induced respiratory depression

项目摘要 这个K99/R 00独立之路奖旨在让候选人实现她的长期 我的目标是建立一个独立的研究生涯，专注于大脑中的唤醒回路如何控制 呼吸，以及阿片类药物如何影响这些神经机制。这项建议是针对 补充候选人在神经科学和呼吸神经生理学的背景， 知识和技术技能，研究呼吸控制电路在体内和体外，并将使她理想的 适合在她的职业道路上取得成功。阿片类药物过量死亡的主要原因是呼吸道疾病。 萧条阿片类药物的镇静作用进一步损害呼吸驱动， 机制等虽然过量通常用阿片类拮抗剂纳洛酮治疗， 逆转所有阿片类药物的作用，引起疼痛和戒断。我们目前对阿片类药物影响的理解 呼吸回路没有揭示出理想的治疗目标，以最小化呼吸抑制， 严重的副作用因此，必须确定新的策略来缓解呼吸抑制， 刺激呼吸。这项提案旨在揭示重要的新见解，即蓝斑（LC）， 一种因其对唤醒的贡献而广为人知的大脑结构，影响着K-100（KF），一个关键 呼吸区，这是高度敏感的阿片类药物和阿片类药物诱导的呼吸抑制的关键。 初步数据涵盖细胞类型特异性神经示踪、离体脑切片记录和体内 光遗传学和纤维光度学，支持LC输入KF在呼吸中的作用，这还没有被证实。 以前赞赏。根据初步数据，总体假设是LC/KF电路是关键的 呼吸功能调节剂，特别是在阿片类药物诱导的呼吸抑制期间。候选人的 在大脑切片记录、清醒动物体内电生理学和交叉病毒- 遗传学方法为最先进的离体和体内光遗传学和光纤技术提供了坚实的基础。 建议中的测光技术。这种多层次的方法将允许候选人测试假设 KF神经元从LC神经元接收阿片样物质敏感的单突触兴奋性输入（目的1），KF神经元 投射LC神经元活动与体内呼吸相耦合（目的2），LC-KF回路起着关键作用 在呼吸和阿片类药物诱导的呼吸抑制中的作用（目的3）。总之，这项建议的目的将 产生关于大脑控制呼吸的新信息，也将提供强有力的概念和 方法培训，使发展的影响力和成功的独立研究计划。 埃里卡莱维特博士和大卫富勒博士的强有力的指导，以及指导委员会，包括 资深教授，他们是所提出技术的专家，并有广泛的指导 经验，将有助于候选人实现本提案的研究和职业发展目标。

PROJECT SUMMARY This K99/R00 Pathway to Independence Award is designed to allow the candidate to achieve her long-term goal to establish an independent research career focused on how arousal circuits in the brain control breathing, and how these neural mechanisms are influenced by opioids. This proposal has been tailored to supplement the candidate’s background in neuroscience and respiratory neurophysiology with additional knowledge and technical skills to study respiratory control circuits in vivo and ex vivo, and will make her ideally suited to succeed on her career path. The primary cause of death from an opioid overdose is respiratory depression. The sedative effect of opioids further impairs respiratory drive through largely unexplored mechanisms. Although overdoses are routinely treated with the opioid antagonist, naloxone, naloxone reverses all opioid effects, causing pain and withdrawal. Our current understanding of opioid effects on the respiratory circuitry has not revealed an ideal therapeutic target to minimize respiratory depression without serious side effects. Therefore, it is essential to identify new strategies to alleviate respiratory depression and stimulate breathing. This proposal seeks to uncover important new insights whereby the locus coeruleus (LC), a brain structure widely known for its contributions to arousal, influences the Kӧlliker-Fuse (KF), a key respiratory region, which is highly sensitive to opioids and critical for opioid-induced respiratory depression. Preliminary data spanning cell-type specific neural tracing, ex vivo brain slice recordings, and in vivo optogenetics and fiber photometry, support a role for LC input to the KF in respiration, which has not been previously appreciated. Based on preliminary data, the overall hypothesis is that the LCKF circuit is a critical modulator of respiratory function, particularly during opioid-induced respiratory depression. The candidate’s prior training in brain slice recordings, in vivo electrophysiology in awake animals, and intersectional viral- genetic methods provides a solid foundation for the state-of-the-art ex vivo and in vivo optogenetics and fiber photometry techniques in the proposal. This multi-level approach will allow the candidate to test the hypothesis that KF neurons receive opioid-sensitive, monosynaptic excitatory input from LC neurons (Aim 1), that KF projecting LC neuron activity is coupled with respiration in vivo (Aim 2), and the LCKF circuit plays a key role in respiration and opioid-induced respiratory depression (Aim 3). Together, the aims of this proposal will yield novel information regarding the brain’s control of respiration and will also provide strong conceptual and methodological training, enabling development of an impactful and successful independent research program. Strong mentorship by Drs. Erica Levitt and David Fuller, as well as a Mentoring Committee comprised of established professors, who are experts in the proposed techniques and have extensive mentoring experience, will help the candidate achieve the research and career development goals of this proposal.