Evaluation of Dbx1-derived neurons as the core rhythm generators in mammalian res

描述（申请人提供）：哺乳动物呼吸是一种重要的行为，其潜在的神经机制起源于脑干。该项目旨在确定产生和控制哺乳动物（包括人类）呼吸的脑干神经回路的组成部分，从而对人类健康产生重大影响。腹侧延髓中有一个叫做波青格前复合体（preBotzinger Complex，简称preBotC）的部位对呼吸至关重要。然而，preBotC的细胞组成，就其包含的不同细胞群的遗传和发育特性以及它们的生理特性而言，在很大程度上仍然未知。据推测，preBotC中一个遗传上不同的神经元亚群形成了产生呼吸节律的核心。为了评估这些关键神经元的作用，采用了分子遗传学和电生理学相结合的多学科研究方法。最近开发的技术通过遗传方法传递细胞谱系标记，如位点特异性重组和荧光标记，极大地影响了神经发育研究。用融合重组酶的细胞谱系示踪剂和报告基因改造的转基因敲入小鼠模型是该研究项目的重要组成部分。同样，这些遗传上不同的神经元群的特征是通过电生理记录进行的，使用独特的体外脑干切片制备，其中包含必要的呼吸神经回路，并允许细胞水平和系统水平的呼吸运动输出记录。因此，该研究项目可以用多层次的方法评估关键神经元群体在呼吸中的重要性：分子，细胞和系统水平的特性将被分析。特异性目的1将通过可逆的基因沉默和不可逆的激光损伤来评估这些神经元的节律性作用。Specific Aim 2将通过全细胞记录来评估关键神经元的膜特性及其节律性作用。这个项目将阐明哺乳动物呼吸的神经起源。本项目获得的新知识将促进我们对中枢神经系统功能障碍导致的呼吸系统疾病的诊断和治疗的认识，并提供关于节律产生的关键新知识，这些新知识普遍适用于理解脑功能。

DESCRIPTION (provided by applicant): Mammalian breathing is a vital behavior whose underlying neural mechanism originates in the brainstem. This project aims to determine the components of the brainstem neural circuits that generate and control respiration in mammals, including humans, and thus has significant implications for human health. A site in the ventral medulla called preBotzinger Complex (preBotC) is essential for breathing. However, the cellular composition of the preBotC, in terms of the genetic and developmental identity of the different cell populations it contains, and their physiological properties, remain largely unknown. A genetically distinct subpopulation of neurons in the preBotC is hypothesized to form the kernel that generates respiratory rhythm. To evaluate the role of these key neurons, a multidisciplinary research approach is employed that combines molecular genetics and electrophysiology. Recently developed technologies that deliver cell lineage markers via genetic methods, such as site-specific recombination and fluorescent tagging, have greatly impacted neural development studies. Transgenic knock-in mouse models engineered with recombinase-fused cell lineage tracers and reporter transgenes are essential components of the research project. Likewise, characterization of these genetically distinct neuronal populations is carried out through electrophysiological recordings using a unique in vitro brainstem slice preparation that contains essential respiratory neural circuits and allows both cellular-level and systems-level recordings of respiratory motor output. Thus, this research project can evaluate the importance of the key population of neurons in breathing with a multilevel approach: molecular, cellular and system-level properties will be analyzed. Specific Aim 1 will assess the rhythmogenic role of these neurons through reversible genetic silencing and irreversible laser lesioning. Specific Aim 2 will evaluate the membrane properties of the key neurons consistent with their rhythmogenic role through whole-cell recordings. This project will elucidate the neural origins of mammalian respiration. The new knowledge obtained in this project will advance our understanding in the diagnosis and treatment of respiratory disorders that result from dysfunctions in the central nervous system, and provide key new knowledge regarding rhythm generation, which is generally applicable to understanding brain function. PUBLIC HEALTH RELEVANCE: Breathing is a human behavior that is essential in maintaining life. This project aims to reveal the cellular composition of brainstem neural circuits that generate and control breathing rhythms, and to characterize the properties of these cells consistent with their role as rhythm generators. The new knowledge acquired will facilitate the diagnosis and treatment of respiratory disorders with a central neural etiology, and elucidate the neural mechanisms that underlie rhythmic motor behaviors in general.