秀丽线虫伤害性感受调控的神经环路与分子机制研究

发布时间：2018-08-22 16:36

【摘要】：生物个体处于复杂多变的环境中,对环境做出正确的应答才能更好的生存。动物感觉调控对其感受、行为以及生存非常重要。目前对伤害性刺激感受与回避行为的调控知之甚少。研究应对外界有害刺激的反应机制对理解生物体如何应对复杂的环境具有重要意义。由于秀丽线虫(Caenorhabditis elegans)生长周期短、通体透明、神经系统简单、易于遗传操作等优势。在本文中,我们选取秀丽线虫为研究对象,研究其伤害性感受的神经回路以及分子机制。线虫有12对头感器感觉神经元,它们分工明确。其中多模感觉神经元ASH是感受伤害性和厌恶性刺激的主要神经元,它能感受水溶性的刺激物如重金属离子和奎宁、厌恶性气体、温度、机械刺激以及渗透压刺激等。关于ASH神经元介导的伤害性感受和回避行为的分子机制和神经回路已有大量研究。感觉神经元ASI是调节线虫是否进入休眠期的关键神经元,它能够整合环境中线虫生长密度、食物、C02的水平以及信息素等,有报道称它还参与温度以及厌恶性刺激反应的调控。嗅觉神经元AWC介导挥发性吸引气味的感受和温度感受,在AWCON神经元中gcy-28作为开关基因可以切换对气味的吸引性行为和排斥性行为。最近有文献报道显示AWCON作为中间神经元介导高盐感受。本文中我们对硫酸铜引起的线虫回避行为做了详细的运动学特性分析,结合在体钙成像记录、传统的基因操作、神经元特异性阻断和失活、神经元化学和遗传学操作以及神经元特异性敲低基因表达等方法,研究了线虫感觉神经元ASI和ASH介导伤害性感受和回避行为的分子机制和环路机制,以及嗅觉神经元AWC接受ASI的信号参与调节伤害性感受。研究中我们发现了ASH和ASI感觉神经元之间存在一个相互抑制的回路,即在硫酸铜刺激的初期,ASH起主导作用,它强烈兴奋抑制了ASI的活性,与此同时ASI也抑制ASH的活性,从而防止线虫的神经元过度兴奋以及产生过激的躲避行为。这种相互抑制的分子机制及神经回路机制为：感觉神经元ASI感受硫酸铜刺激后释放神经肽促进五羟色胺能感觉神经元ADF释放5-HT作用于感觉神经元ASH上的SER-5受体,进而抑制ASH的活性；同时,ASH通过电突触激活章鱼胺能中间神经元RIC释放章鱼胺作用于ASI上的SER-3受体,进而抑制ASI神经元的活动。这是一种外周水平上的感受调节,相比中枢神经系统水平的感觉调控,外周神经系统水平上的感受调节更加直接和快速,保证了线虫对伤害性刺激做出迅速而不过于激烈的行为反应,使线虫能够更好的适应环境,有利于它的生存。除此之外,我们还发现ASI通过AWC抑制对硫酸铜产生过度敏感的行为。这种调控的具体分子机制为：ASI感受硫酸铜刺激释放神经肽作用于AWCON上的DAF-2受体作用(其中AWCON起中间神经元的功能),激活AWCON释放谷氨酸和神经肽,通过中间神经元AIB或者其他神经元整合信号,最终抑制线虫对伤害性刺激的过度敏感,属于中枢神经系统水平的感觉调控。感觉神经元ASI在伤害性感受和躲避行为中起了非常重要的作用,它通过两条不同的信号通路抑制对伤害性刺激的过度敏感：一条是ASI通过ADF抑制ASH,降低ASH至AIB的信号,进而抑制对伤害刺激的过度敏感；另一条是ASI促进AWCON释放谷氨酸和神经肽抑制线虫的超敏感行为。
[Abstract]:Animal sensory regulation is very important to their feelings, behavior and survival. At present, little is known about the regulation of noxious stimulus perception and avoidance behavior. Complex environments are important. Because of the short growth cycle, transparent whole body, simple nervous system and easy genetic manipulation, Caenorhabditis elegans has 12 pairs of sensory nerves. Among them, multimodal sensory neurons (ASH) are the main neurons that sense nociceptive and aversive stimuli. It can sense water-soluble stimuli such as heavy metal ions and quinine, aversive gases, temperature, mechanical stimuli and osmotic pressure stimuli. Molecular mechanisms of nociceptive and avoidance behaviors mediated by ASH neurons Sensory neuron ASI is a key neuron that regulates whether a nematode enters dormancy. It integrates nematode growth density, food, C02 levels and pheromones in the environment. It has also been reported to be involved in the regulation of temperature and aversive stimulus responses. AWC mediates volatile attraction. In AWCON neurons, gcy-28 as a switch gene can switch the attractant and repulsive behavior to odor. Recently, it has been reported that AWCON acts as an intermediate neuron mediating hypersaline sensation. In vivo calcium imaging, traditional gene manipulation, neuron specific blockade and inactivation, neurochemical and genetic manipulation, and neuronal specific knockdown of gene expression were used to study the molecular mechanism and loop mechanism of ASI and ASH-mediated nociceptive and avoidance behavior in nematode sensory neurons, and the AWC junction of olfactory neurons. In this study, we found that there is a reciprocal inhibitory pathway between ASH and ASI sensory neurons, that is, in the early stage of copper sulfate stimulation, ASH plays a dominant role, which strongly excites and inhibits the activity of ASI. At the same time, ASI also inhibits the activity of ASH, thus preventing the over-excitation of nematode neurons. The molecular mechanism and neural circuit mechanism of this mutual inhibition are: sensory neuron ASI senses copper sulfate stimulation and releases neuropeptides to stimulate serotonin-responsive neuron ADF to release 5-HT on the SER-5 receptor of sensory neuron ASH, and then inhibits the activity of ASH; at the same time, ASH through the electrical synapse; Activating the RIC of the octophinergic intermediate neurons releases octophine on the SER-3 receptor on ASI and inhibits the activity of ASI neurons. This is a peripheral level sensory regulation, which is more direct and faster than the central nervous system level sensory regulation, and guarantees that the nematode infects the noxious spine. In addition, we also found that ASI inhibited excessive sensitivity to copper sulfate through AWC. The specific molecular mechanism of this regulation is that ASI sensed copper sulfate-stimulated release of neuropeptide DAF-2 on AWCON. AWCON acts as an intermediate neuron, activates AWCON to release glutamate and neuropeptides, integrates signals with AIB or other neurons, and ultimately inhibits the hypersensitivity of nematodes to noxious stimuli. It belongs to sensory regulation at the level of the central nervous system. Sensory neuron ASI is involved in noxious sensation and avoidance behavior. It plays an important role in inhibiting hypersensitivity to nociceptive stimuli through two different signaling pathways: one is that ASI inhibits ASH through ADF, lowers ASH to AIB signals, and then inhibits hypersensitivity to nociceptive stimuli; the other is that ASI promotes the release of glutamate and neuropeptides from AWCON to inhibit the hypersensitivity of nematodes.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：Q42

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