A zebrafish model to study functional regeneration of motor circuits

项目摘要 损伤后功能的恢复是运动控制中一个重要但难以实现的目标。脊髓运动神经元 由于长轴突支配周围肌肉，特别容易受到损伤。运动神经元 也有影响运动输出的中央轴突侧支，但我们对它们对损伤的反应知之甚少。这 是重要的，因为运动神经元兴奋性的变化伴随着脊髓损伤，有助于短期 痉挛和长期恢复。这个探索性的R21提案的目标是开发一个新的模型系统 为了了解中央脊髓回路在受伤后如何重组，从一个被忽视的运动神经元开始， 对伤害的反应。像所有的脊椎动物一样，斑马鱼也有慢、中、快三种类型的脊髓运动单位 由它们的目标肌肉组织定义，并且每种类型都表现出中央轴突侧支。然而，与所有其他 脊椎动物可以追踪相同的脊髓运动神经元及其中枢和外周连接 在受伤之前和之后，活的，生长中的斑马鱼。在使用双光子轴切术的试点实验中，我们发现 一些运动神经元再生并成功地重新支配目标肌肉组织，而另一些则未能退出 而不是长出精致的中央轴突并形成突触。这也反映了一个现象， 在成年猫的轴突切断的脊髓运动神经元中观察到。这些观察结果表明，一些轴突切断， 运动神经元正在产生可能影响运动控制的异位回路。但是什么样的条件能预测 外周与中枢再生以及异位中枢运动轴突整合到功能回路中？在 目的1，我们将评估发育时间和运动单位特性对轴突切断的相对贡献 应答在目标2中，我们将确定轴突切断的运动神经元是否被募集并连接到其他神经元。 脊髓神经元这些目标将描述运动神经元形成异位循环回路的能力 并将为探索损伤后脊髓回路结构的适应不良和适应性变化的研究提供信息。

Project Summary The recovery of function after damage is an important, yet elusive goal in motor control. Spinal motor neurons are particularly vulnerable to damage thanks to long axons that innervate peripheral muscles. Motor neurons also have central axon collaterals that impact motor output, but we know little of their response to injury. This is important because changes in motor neuron excitability accompany spinal injury, contributing to short-term spasticity and long-term recovery. The goal of this exploratory R21 proposal is to develop a new model system to understand how central spinal circuits reorganize after injury, beginning with an overlooked motor neuron response to injury. Like all vertebrates, zebrafish have slow, intermediate and fast types of spinal motor units defined by their target musculature and each type exhibits central axon collaterals. However, unlike all other vertebrates it is possible to track the same spinal motor neurons and their central and peripheral connections before and after injury in living, growing zebrafish. In pilot experiments using two-photon axotomies, we found that some motor neurons regrow and successfully re-innervate target musculature, while others fail to exit spinal cord and instead grow elaborate central axons and form synapses. This mirrors a phenomenon also observed in axotomized spinal motor neurons of adult cats. These observations suggest that some axotomized motor neurons are creating ectopic circuits that could impact motor control. But what conditions predict peripheral versus central regeneration and do ectopic central motor axons integrate into functional circuits? In Aim 1, we will assess the relative contribution of time of development and motor unit identity to axotomy responses. In Aim 2, we will determine if axotomized motor neurons are recruited and connected to other spinal neurons. These aims will characterize the capacity for motor neurons to form ectopic recurrent circuits and will inform studies exploring maladaptive and adaptive changes in spinal circuit structure following injury.