Mature spinal networks in rodents: electrophysiological investigations combined with optogenetic methods to define neuron and circuit function

哺乳动物运动所需的有节奏的运动输出可以完全由称为中央模式发生器（CPG）的特殊脊髓（间）神经元回路产生。识别参与CPG活动的中间神经元是理解脊神经群的关键，这些脊神经群是启动持续模式肢体运动所必需的，那些通过感觉输入（即反射回路）启动运动输出的脊神经细胞与那些通过椎上下降输入（即自愿回路）启动运动的脊神经细胞，以及CPG神经元的相互连接如何导致有节奏的运动输出。***使用乙酰胆碱作为神经递质的中间神经元- CPG的胆碱能神经元成分-已被认为以状态依赖的方式控制运动神经元的兴奋性，并改变运动节律的频率。沿几乎整个脊髓的中央管附近的胆碱能中间神经元群已被描述为通过专门的突触支配脊髓运动神经元。这些胆碱能层X神经元（ChATX INs）是本研究的重点。***用于选择性激活不同神经元回路的光遗传学工具将在体内神经生理学实验室中用于成熟的小鼠脊髓网络，该实验室具有在集成立体定向脑和脊柱框架中研究功能成熟的小鼠脊髓的能力。在失智的小鼠中，运动输出可以通过几种方式产生，包括通过反射和自愿循环。通过对ChATX INs进行光学刺激，研究其产生运动输出或改变已经存在的运动活动和自主或反射回路诱导活动输出的能力。电生理技术的光刺激将与解剖学技术和活动依赖标记相结合，以确定ChATX INs的功能作用和连通性。在提出的目标中，概述了培养五名本科生和两名研究生水平的学员。由于我的项目专门研究成熟的哺乳动物网络，基本功能图将连接来自简化系统（例如新生动物）和来自人类的知识。通过定义ChATX IN神经元在运动中的作用，有可能在CPG网络中识别一类与运动神经元直接连接的新型兴奋性末阶中间神经元。如果ChATX IN神经元的选择性激活导致持续的CPG输出，那么它可能是第一批可以被视为CPG节律产生层神经元的细胞

The rhythmic patterned motor output required for locomotion in mammals can be produced entirely by a specialized spinal (inter)neuronal circuit called the central pattern generator (CPG). Identification of interneurons that contribute to CPG activity is pivotal understanding spinal neuronal populations that are necessary to initiate sustained patterned limb movements and those spinal neurons that initiate motor output via sensory-input (i.e., reflex loops) vs. those that initiate locomotion via supraspinal descending input (i.e., voluntary loops), and how the interconnections of CPG neurons lead to rhythmic motor output.***Interneurons using acetylcholine as a neurotransmitter - cholinergic neuronal components of the CPG - have been suggested to control motoneuronal excitability in a state-dependent manner and to modify the frequency of the locomotor rhythm. A cholinergic interneuron population near the central canal along nearly the entire spinal cord has been described to innervate spinal motoneurons by specialized synapses. These cholinergic lamina X neurons (ChATX INs) are the focus of this proposal. ***Optogenetic tools for selective activation of distinct neuronal circuits will be used in mature, murine spinal networks in an in vivo neurophysiology laboratory with capabilities for studying functionally mature mouse spinal cords in an integrated stereotaxic brain and spinal frame. In decerebrated mice, motor output can be generated in several ways, including via reflex and voluntary loops. By the use of optical stimulation applied to the ChATX INs, their ability to generate locomotor output or to modify already existing locomotor activity and the output of voluntary or reflex loop induced activity will be investigated. Optical stimulation with electrophysiological techniques will be used in combination with anatomical techniques and activity dependent labelling to determine the functional roles and connectivity of ChATX INs. During the proposed objectives, training of five undergraduate students and two graduate level trainees is outlined.***As my program specifically aims to study mature, mammalian networks, the basic functional maps will bridge knowledge derived from simplified systems (for example, neonatal animals) and those derived from humans. By defining the role of ChATX IN neurons in locomotion, it may be possible to identify a novel class of excitatory last order interneurons within the CPG network with direct connections to motoneurons The ChATX IN neurons may be the first population of cells which could be viewed as CPG rhythm generation layer neurons if their selective activation leads to sustained CPG output.**