Unraveling respiratory rhythm generation in the medullary network

描述(申请人提供)：呼吸是由沿延髓腹外侧吻-尾轴分布的神经元网络产生的(“腹侧呼吸柱”，VRC)。这个网络产生了三个不同的阶段：灵感(I)、后灵感(Post-I)和主动终止(AE)。许多疾病与不同形式的呼吸障碍和对低氧的反应有关。因此，了解在缺氧等不同条件下这些呼吸时相是如何产生和动态调节的，具有重要的基础科学和临床意义。在这个项目中，我们介绍了两种新的有节奏的活性脑干切片制备方法，使我们能够研究更广泛的延髓网络中突触、内在和调制属性的整合，这是以前不可能达到的程度。基于我们的初步数据，我们提出了这样的假设，即后I和灵感是由一个从前Bötzinger复合体向Bötzinger复合体延伸的兴奋柱产生的。这种分布的兴奋网络与GABA能和甘氨酸能机制以及固有的膜特性相互作用，这些将在水平切片制备中使用各种电生理、药理学和光遗传学方法来探索(目标1)。将从水平切片制备中获得的见解与从分离该兴奋柱的尾部和吻部的两个横向切片制备中获得的数据进行比较(目标2)。这种方法将使我们能够区分发生在这一柱的尾端和吻端的节律机制。这些体外发现中揭示的概念随后将在体内自发呼吸制剂中进行测试(目标3)。我们期望，这些新的体外制剂的引入，结合现代光遗传学技术，以及与体内方法的严格整合，将使我们能够重新审视现有的呼吸节律产生模型。这可能会导致更好地理解在呼吸神经控制领域目前的知识状态下仍未解决和解释的各种问题。

DESCRIPTION (provided by applicant): Breathing is generated by a neuronal network that is distributed along the rostro-caudal axis of the ventrolateral medulla ("ventral respiratory column", VRC). This network gives rise to three distinct phases: inspiration (I), post-inspiration (Post-I), and active expiration (AE). Many disorders are associated with disturbances in different forms of breathing and the response to hypoxia. Thus understanding how these phases of breathing are generated and dynamically regulated under various conditions such as hypoxia is of great basic scientific and clinical interest. In this project we introduce two novel rhythmicall active brainstem slice preparations that allow us to study the integration of synaptic, intrinsic and modulatory properties in the wider medullary network to an extent that was not possible before. Based on our preliminary data we propose the hypothesis that post-I and inspiration are generated by an excitatory column that extends rostrally from the pre-Bötzinger complex into the Bötzinger complex. This distributed excitatory network interacts with GABAergic and glycinergic mechanisms as well as intrinsic membrane properties that will be explored in the horizontal slice preparation using a variety of electrophysiological, pharmacological, and optogenetic approaches (Aim 1). Insights gained in this horizontal slice preparation will be compared with data obtained from two transverse slice preparations that isolate the caudal and rostral portion of this excitatory column (Aim 2). This approach will allow us to differentiate rhythmogenic mechanisms occurring at the caudal and rostral end of this column. Concepts revealed in these in vitro findings will then be tested in a spontaneously breathing in vivo preparation (Aim 3). We expect that the introduction of these novel in vitro preparations, combined with modern optogenetic techniques and a rigorous integration with in vivo approaches will allow us to revisit existing models of respiratory rhythm generation. This may lead to a better understanding of various issues that remain unresolved and unexplained at the current state of knowledge in the field of neural control of breathing.