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，在遗传、神经元和运动水平上研究了线虫的温度反应。为了支持新的数量遗传学工作，我们发表了一份关于布里奇萨锥虫空间趋热行为的基础性报告，表明野生分离物在其趋性反应和热记忆整合方面存在巨大差异(Stegeman等人，2012年)。使用一种新的基于激光的加热仪器，我们展示了多参数表型在分析候选突变菌株文库的热“痛”行为方面的威力(Ghosh等人，2012年)。我们改进了这台仪器，用紧聚焦的红外激光光束程序化地刺激蠕虫身体的部分，以确定蠕虫热痛反应的空间感受野。使用多参数量化的“痛”行为，我们表明：1)秀丽线虫可以分辨热刺激的精度至少80微米时，集中在中体，2)神经元PVD是中体反应所需的感觉神经元，3)一些基因编码谷氨酸受体和TRPV样通道可能参与这种伤害性的热传导(Mohammadi等人，2013)。这项初步工作建立了线虫作为热疼痛转导的定量模型，其中线虫的疼痛水平是通过阅读其伤害性反应的“身体语言”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导致的疼痛转导的变化。**我们未来的项目通过应用系统级、多学科的方法和生物物理学、遗传学和神经科学领域的实施技术来继续这项先前的工作。目前，我们有三个主要项目属于NSERC的应用，旨在将线虫发展为伤害性/疼痛转导的模式生物。首先是使用我们的激光脉冲止痛法对含有止痛剂的线虫进行筛选，以量化疼痛，以了解这些止痛剂如何在分子水平上改变行为。第二个是研究一种未知功能的受体，该受体参与蠕虫中体的疼痛反应。第三个项目涉及开发PVD，一种具有高度分支突起的神经元，作为研究单个神经元如何处理空间感觉信息的生物物理学的模型神经元。

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.