Linking the molecular logic of sensory neuron diversity and somatosensory circuitry in mouse spinal cord: development of novel tools for viral tracing and 3D analysis

项目摘要/摘要 脊髓背侧感觉神经元是中枢神经系统中第一个接受躯体感觉的部位。 来自外设的信息，是控制姿势和运动的感应器电路中的关键部件 行为。确定躯体感觉回路形成的发育原理和分子同一性 服务于特定感觉功能的神经元是我们理解感觉运动整合逻辑的关键 以及受伤或疾病后的战略干预。所有DIN都来自分子定义的小阵列 胚胎细胞群体，但标记只是瞬时表达，以及这些早期神经元是如何多样化的 并整合到为成熟动物遗骸中的不同感觉模式提供服务的感觉回路中 未知。迫切需要基因途径来表达示踪剂，并操纵个别类别的 DIN在胚胎期出现时就会发育，但在出生后就会形成回路。在这里，我们将 第一次创造出小鼠，通过提供 在整个发育过程中，有选择地获得一组DIN，称为DI1。我们将创建鼠标线 引入顺行和逆行病毒示踪剂以揭示和定位脑和脑内DI1s的突触靶点。 并识别DRG、脑和脊髓中DI1s的输入神经元。这些线将延伸到时间 通过第一种依赖于Cre的Cre方法，选择性地获得发育到出生后阶段的DI1的遗传途径， 允许依赖Cre的轴突和突触示踪剂进入，第二，表达这些蛋白的小鼠 使用狂犬病病毒进行跨突触追踪所需的。我们还将开发平台来有效地分析 电路的解剖和连通性。此外，我们将创建一个广泛需要的3D脊髓地图集 神经元特性和标测电路的评估。要了解调节dI1的程序 在开发过程中，我们还将识别和跟踪dI1中转录签名的动态变化 单细胞RNA测序的胚胎和出生后个体发育。建议中建立的工具 实验和分析将适用于所有DIN子集，并将提供资源和信息 并为健康体感通路的操纵和功能分析提供了模板 在疾病中。

Project Summary/Abstract Sensory neurons in the dorsal spinal cord (dINs) represent the first site in the CNS to receive somatosensory information from the periphery and are a crucial component in sensorimotor circuits that control posture and behavior. Defining the developmental principles of somatosensory circuit formation and the molecular identity of neurons that serve specific sensory functions is key to our understanding of the logic of sensorimotor integration and for strategic interventions following injury or disease. All dINs arise from a small array of molecularly defined embryonic cell populations, but markers are expressed only transiently and how these early neurons diversify and integrate into sensory circuits serving distinct sensory modalities in the mature animal remains largely unknown. There is an urgent need for genetic access to express tracers and to manipulate individual classes of developing dINs as they emerge during the embryonic period but establish circuits postnatally. Here, we will generate for the first time, mice that bridge the gap between early identity and mature function by providing access selectively to one population of dINs, called dI1s, throughout development. We will create mouse lines to introduce anterograde and retrograde viral tracers to reveal and map the synaptic targets of dI1s in brain and spinal cord, and to identify input neurons to dI1s, in DRG, brain and spinal cord. These lines will extend temporal genetic access selectively to developing dI1s into postnatal stages, by, first, a Cre-dependent Cre approach, permitting access by Cre-dependent axonal and synaptic tracers, and second, mice that express the proteins required for transsynaptic tracing using rabies virus. We will also develop platforms to analyze efficiently the anatomy and connectivity of the circuitry. Furthermore, we will create a widely needed 3D spinal cord atlas for assessment of neuronal identity and mapping circuitry. To understand the programs that regulate dI1 development, we will also identify and follow dynamic changes of transcriptional signatures in dI1s throughout embryonic and postnatal ontogeny by single cell RNA sequencing. The tools established in the proposed experiments and analyses will be applicable to all dIN subsets and will contribute both resources and information to the field, and provide a template for manipulation and functional analysis of somatosensory pathways in health and in disease.