Do somatosensory endings use axonal or dendritic regeneration pathways?

描述（由申请人提供）：支配皮肤的感觉神经元容易受损。当损伤超过这些细胞的修复能力，周围神经病变（PN）表现。化疗后幸存下来的癌症患者，或者努力控制血糖的糖尿病患者，可能会留下几个月甚至一辈子的麻木或剧痛。皮肤中的游离神经末梢介导疼痛和温度感觉，在大多数形式的PN中，这两者在早期都受到损害。这些感觉末梢的修复机制尚未阐明，尽管它是了解PN发病的关键，也可能是制定延迟或治疗PN策略的关键。轴突再生在所有被研究过的生物体和神经元中使用一套共同的机制。该核心机制包括由DLK启动的MAP激酶级联反应和AP-1转录因子的激活。当果蝇感觉神经元轴突受损时，这一核心通路介导再生，但同一细胞中外周感觉末梢的再生完全独立于这一机制。在果蝇中，细胞生物学研究将感觉末梢定义为树突，这表明DLK通路可能是轴突再生所特有的，而不是用于树突再生。确定脊椎动物的感觉末梢是通过保守的轴突再生途径还是完全未被探索的树突再生途径修复的，将是理解PN的关键基础。在这项探索性工作中，将采用两条主要的实验线来确定用于再生脊椎动物皮肤感觉末梢的修复途径。支配斑马鱼皮肤的感觉神经元将被用作所有这些实验的模型。罗洪-比尔德（RB）神经元是斑马鱼幼体的感觉神经元，可以用现有的方法进行成像和操作，因此这些细胞将被用作研究感觉末段细胞生物学和损伤反应的第一个模型。类似的方法随后将用于背根神经节神经元（DRGs），因为它们与理解PN最相关。为了确定脊椎动物的自由感觉末梢是像果蝇一样的树突，还是像脊椎动物历史上分类的轴突，我们将确定它们的微管的极性。所有已知的树突含有至少50%的负端外微管，而轴突含有接近100%的正端外微管。微管极性从未在脊椎动物感觉末梢中测定过，因此将在RB和DRG细胞中进行。为了确定感觉末梢损伤是否触发了已确定的轴突再生途径或新的树突再生途径，我们将开发DLK信号标志物，并比较斑马鱼感觉神经元对中央轴突损伤和外周感觉末梢损伤的反应。这两条实验路线将为理解感觉末梢如何修复以及该过程如何在PN患者中失败奠定基础。

DESCRIPTION (provided by applicant): Sensory neurons that innervate the skin are easily damaged. When damage exceeds the capacity of these cells for repair, peripheral neuropathy (PN) manifests. Cancer patients who survive chemotherapy, or diabetics who struggle to control blood sugars, may be left with months to a lifetime of numbness or shooting pain. Free nerve endings in the skin mediate pain and temperature sensation, both of which are compromised early in most forms of PN. The repair mechanism of these sensory endings has not been elucidated, although it is the key to understanding the onset of PN and may also be critical for developing strategies to delay or cure PN. Axon regeneration uses a common set of machinery in all organisms and neurons in which it has been studied. This core machinery includes a MAP kinase cascade initiated by DLK and the activation of AP-1 transcription factors. This core pathway mediates regeneration when axons of sensory neurons in Drosophila are injured, but regeneration of peripheral sensory endings in the same cells is completely independent of this machinery. In Drosophila, sensory endings have been defined as dendrites by cell biology studies, suggesting that the DLK pathway may be specific to axon regeneration and not used for dendrite regeneration. Determining whether sensory endings in vertebrates are repaired using the conserved axon regeneration pathway or a completely unexplored dendrite regeneration pathway will be a critical foundation for understanding PN. In this exploratory work, two major lines of experimentation will be used to determine the repair pathway used to regenerate vertebrate sensory endings in the skin. The sensory neurons that innervate the skin in zebrafish will be used as a model for all of these experiments. Zebrafish larval sensory neurons known as Rohon-Beard (RB) neurons are accessible for imaging and manipulation with current methodology, so these cells will be used as a first model in which to study sensory ending cell biology and injury responses. Analogous methods will then be developed for dorsal root ganglion neurons (DRGs), as these are the most relevant for understanding PN. To determine whether vertebrate free sensory endings are dendrites as in Drosophila, or axons as they are historically classified in vertebrates, we will determine the polarity of their microtubules. All known dendrites contain at least 50% minus-end-out microtubules while axons are close to 100% plus-end-out. Microtubule polarity has never been assayed in vertebrate sensory endings, so this will be done in both RB and DRG cells. To determine whether injury to sensory endings triggers the identified axon regeneration pathway or a novel dendrite regeneration pathway, we will develop markers for DLK signaling and compare responses to central axon injury and peripheral sensory ending injury in zebrafish sensory neurons. These two lines of experimentation will form a foundation for understanding how sensory endings are repaired, and how this process fails in patients with PN.