Thermal pain transduction in C. elegans

在过去的5年里，我们利用新的行为表型、仪器和分析方法，在遗传、神经元和运动尺度上研究了秀丽隐杆线虫的热反应。为了支持C. briggsae新的定量遗传学研究，我们发表了一份关于C. briggsae空间趋热行为的基础报告，表明野生菌株在趋热反应和热记忆整合方面差异很大（Stegeman et al., 2012）。使用一种新型的基于激光的加热仪器，我们证明了多参数表型在候选突变菌株库的热“疼痛”行为分析中的作用（Ghosh等，2012）。我们改进了这个仪器，用紧密聚焦的红外激光光束程序化地刺激蠕虫身体的某些部位，以确定蠕虫热痛反应的空间接受场。通过对“疼痛”行为的多参数量化，我们发现：1)秀丽隐杆线虫在聚焦于其中体时，可以识别至少80微米精度的热刺激；2)神经元PVD是中体反应所需的感觉神经元；3)编码谷氨酸受体和trpv样通道的许多基因可能参与这种热的伤害性传导（Mohammadi et al., 2013）。这项初步的工作建立了秀丽隐杆线虫作为热痛转导的定量模型，其中蠕虫的疼痛水平是通过阅读其伤害性反应的“肢体语言”来统计推断的，这使我们能够客观地确定由于基因、神经元或经验的扰动而引起的疼痛转导的变化。**我们未来的项目通过应用系统级、多学科的方法和实施生物物理学、遗传学和神经科学领域的技术来跟进之前的工作。目前，我们有三个主要项目属于NSERC申请，旨在开发秀丽隐杆线虫作为伤害/疼痛转导的模式生物。首先是用激光脉冲法对秀丽隐杆线虫进行疼痛量化，以了解这些镇痛药如何在分子水平上改变行为。第二个是研究一种功能未知的受体，它参与了蠕虫中体的疼痛反应。第三个项目涉及PVD的发展，PVD是一种具有高度分支过程的神经元，作为模型神经元来研究单个神经元如何处理空间感觉信息的生物物理学。

Over the last 5 years, we have studied the thermal response of C. elegans at the genetic, neuronal, and locomotory scales utilizing novel behavioral phenotyping, instrumentation, and analysis. In support of new quantitative genetics work with C. briggsae we have published a foundational report on C. briggsae spatial thermotaxis behavior showing that wild isolates wildly vary in their taxis response and their integration of thermal memory (Stegeman et al., 2012). Using a novel laser-based heating instrument, we demonstrated the power of multi-parameter phenotyping on analysis of thermal "pain" behavior of a candidate mutant strain library (Ghosh et al., 2012). We improved this instrument to programmatically stimulate parts of the worm's body with tightly focused beams of infrared laser light, to determine the spatial receptive field of the worm's thermal pain response. Using multi-parameter quantification of the "pain" behavior we showed that: 1) C. elegans can discriminate thermal stimuli to a precision of at least 80 microns when focused on its mid-body, 2) the neuron PVD is the sensory neuron that is required for the mid-body response, and 3) a number of genes encoding glutamate receptors and TRPV-like channels are likely involved in this nociceptive transduction of heat (Mohammadi et al., 2013). This preliminary work establishes C. elegans as a quantitative model for thermal pain transduction, where the worm's level of pain is statistically inferred by reading the "body language" of its nociceptive response, which allows us to objectively determine changes in pain transduction due to perturbations of genes, neurons, or experience.**Our future projects follows up on this previous work by applying a systems-level, multi-disciplinary approach and implementing techniques from the areas of biophysics, genetics, and neuroscience. Currently we have three major projects that fall under this NSERC application toward the development of C. elegans as a model organism for nociceptive/pain transduction. The first is to screen C. elegans with analgesics using our laser pulse assay for pain quantification in order to understand how these analgesics modify behavior at the molecular level. The second is to investigate a receptor with unknown function that is involved in the pain response at the midbody of the worm. The third project involves the development of PVD, a neuron with highly branched processes, as a model neuron to study the biophysics of how a single neuron processes spatial sensory information.