Generation of mice to selectively mark a subset of spinal interneurons

项目摘要 现代神经科学的一个主要目标是了解行为的神经基础。运动是一种 由复杂的神经回路在脊髓中产生的行为。它被定义为精确、协调和 在相对肢体之间以及在同一肢体的拮抗肌之间交替活动。 此外，运动行为对于实验研究是有吸引力的，因为它可以很容易地获得， 定义和量化。被称为中央模式发生器（CPG）的中间神经元网络被认为是 负责节奏活动的起源。CPG神经元激活运动神经元， 激活周围肌肉导致运动。运动神经元还具有轴突侧支， 这是一种专门的抑制性中间神经元，称为Renshaw细胞。我们最近的研究挑战了 传统观念认为运动神经元只是脊髓的运动输出。运动刺激 新生小鼠神经元轴突在胆碱能受体存在下可触发自发活动 对手。这表明，运动神经元轴突侧支释放的乙酰胆碱不是运动神经元的必需物质。 节律发生此外，我们发现新生的运动神经元释放第二个快速兴奋性神经元。 除了乙酰胆碱之外，神经递质还来自其轴突侧支（谷氨酸或天冬氨酸）。的 刺激运动轴突如何触发新生儿脊髓中的运动样活动的机制是 不理解。参与运动CPG的神经元及其连接的识别是 对于阐明参与运动活动的机制至关重要，因此将代表 这是该领域的一个根本性进步。在这个提议中，我们提供了强有力的初步证据，表明一个新的类 兴奋性中间神经元是腹侧脊髓回路中的关键组成部分，它可能参与了 运动中枢模式发生器我们的目标是产生新的小鼠品系，表达选择基因， 类脊髓中间神经元来测试我们的主要假设，这些脊髓中间神经元在神经系统中起着积极的作用。 自发活动的产生。 在目标1中，我们将生成两个敲入鼠标线，用作交叉方法，以便标记 选择性地一类脊髓中间神经元。在目标2中，我们将验证两种小鼠的选择性和效率， 当它们彼此交叉时，也是如此。总之，这个为期两年的研究项目 描述了产生敲入小鼠品系的综合设计，所述小鼠品系具有鉴定 一类新的兴奋性中间神经元作为一个关键的神经元的球员在运动活动的节奏。

Project Summary A major goal of modern Neuroscience is to understand the neuronal basis of behavior. Locomotion is a behavior generated in the spinal cord by complex neuronal circuits. It is defined as precise, coordinated and alternating activity between opposing limbs as well as between antagonistic muscles of the same limb. Furthermore, locomotor behavior is attractive for experimental study because it can be easily accessed, defined, and quantified. A network of interneurons, known as the central pattern generator (CPG), is thought to be responsible for the genesis of rhythmic activity. CPG neurons activate motor neurons which in turn activate peripheral muscles resulting in movement. Motor neurons also possess axon collaterals which target exclusively a class of inhibitory interneurons, known as Renshaw cells. Our recent studies have challenged the traditional idea that motor neurons are simply the motor output from the spinal cord. Stimulation of motor neuron axons in neonatal mice can trigger locomotor activity in the presence of cholinergic receptor antagonists. This suggests that acetylcholine released by motor neuron axon collaterals is not required for rhythmogenesis. Furthermore, we discovered that neonatal motor neurons release a second fast excitatory neurotransmitter from their axon collaterals (glutamate or aspartate) in addition to acetylcholine. The mechanisms of how stimulation of motor axons can trigger locomotor-like activity in neonatal spinal cords are not understood. The identification of neurons and their connectivity that participate in the locomotor CPG is critically needed for the elucidation of mechanisms involved in locomotor activity and would therefore represent a fundamental advance in the field. In this proposal, we provide strong preliminary evidence that a novel class of excitatory interneurons is a critical component in a ventral spinal cord circuit and it may be involved in the locomotor central pattern generator. We aim to generate novel mouse lines that express select genes in this class of spinal interneurons to test our main hypothesis, that these spinal interneurons play an active role in the generation of locomotor activity. In Aim 1, we will generate two knock-in mouse lines to be used as an intersection approach in order to label selectively a class of spinal interneurons. In Aim 2, we will validate the selectivity and efficiency of both mouse lines individually as well as when they are crossed amongst them. In summary, this two-year research project describes a comprehensive design of generation of knock-in mouse lines that have the potential to identify a novel class of excitatory interneurons as a key neuronal player in the rhythmogenesis of locomotor activity.