Optogenetic Stimulation of Nociceptive Escape Behaviors in Drosophila Larvae.

Stephanie E Mauthner, W Daniel Tracey
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引用次数: 0

Abstract

In animals, noxious stimuli activate a neural process called nociception. Drosophila larvae perform a rolling escape locomotion behavior in response to nociceptive sensory stimuli. Noxious mechanical, thermal, and chemical stimuli each trigger this same escape response in larvae. The polymodal sensory neurons that initiate the rolling response have been identified based on the expression patterns of genes that are known to be required for nociception responses. The synaptic output of these neurons, known as class IV multidendritic sensory neurons, is required for behavioral responses to thermal, mechanical, and chemical triggers of the rolling escape locomotion. Importantly, optogenetic stimulation of the class IV multidendritic neurons has also shown that the activation of those cells is sufficient to trigger nociceptive rolling. Optogenetics uses light-activated ion channels expressed in neurons of interest to bypass the normal physiological transduction machinery so that the cell may be activated in response to light that is applied by the investigator. This protocol describes an optogenetic technique that uses channelrhodopsin-2 (ChR2) to activate larval nociceptors and trigger nociceptive rolling. First, we explain how to set up the necessary genetic crosses and culture the larval progeny. Next, we describe how to perform the optogenetic nociception assay on third-instar larvae.

光遗传刺激果蝇幼虫的痛觉逃逸行为
在动物体内,有害刺激会激活一种称为痛觉的神经过程。果蝇幼虫在痛觉刺激下会做出滚动逃逸的运动行为。有害的机械、热和化学刺激都会触发幼虫的这种逃逸反应。根据已知的痛觉反应所需基因的表达模式,已经确定了启动滚动反应的多模态感觉神经元。这些神经元被称为第四类多树突感觉神经元,其突触输出是对滚动逃逸运动的热、机械和化学触发行为反应所必需的。重要的是,对第四类多树突神经元的光遗传学刺激也表明,激活这些细胞足以触发痛觉滚动。光遗传学利用在相关神经元中表达的光激活离子通道,绕过正常的生理传导机制,使细胞对研究人员施加的光做出反应而被激活。本方案介绍了一种光遗传学技术,该技术利用通道色素-2(ChR2)激活幼虫痛觉感受器并触发痛觉滚动。首先,我们解释了如何建立必要的遗传杂交和培养幼虫后代。接下来,我们将介绍如何在三龄幼虫身上进行光遗传痛觉检测。
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来源期刊
Cold Spring Harbor protocols
Cold Spring Harbor protocols Biochemistry, Genetics and Molecular Biology-Biochemistry, Genetics and Molecular Biology (all)
CiteScore
3.00
自引率
0.00%
发文量
163
期刊介绍: Cold Spring Harbor Laboratory is renowned for its teaching of biomedical research techniques. For decades, participants in its celebrated, hands-on courses and users of its laboratory manuals have gained access to the most authoritative and reliable methods in molecular and cellular biology. Now that access has moved online. Cold Spring Harbor Protocols is an interdisciplinary journal providing a definitive source of research methods in cell, developmental and molecular biology, genetics, bioinformatics, protein science, computational biology, immunology, neuroscience and imaging. Each monthly issue details multiple essential methods—a mix of cutting-edge and well-established techniques.
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