Hypocretin and Locus Ceruleus Interactions in Zebrafish Development and Behavior

下丘脑分泌素和蓝斑在斑马鱼发育和行为中的相互作用

• 批准号: 8566608
• 负责人: Grigorios Oikonomou
• 金额: $ 5.33万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-02-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8566608
• 关键词: Ablation; Address; Affect; Alzheimer' s Disease; Anatomy; Animal Model; Animals; Anxiety Disorders; Arousal; Attention deficit hyperactivity disorder; Bathing; Behavior; Behavioral; Brain; Brain Stem; Bypass; Cells; Confocal Microscopy; Cues; Development; Down Syndrome; Drug Addiction; Electrophysiology (science); Exhibits; Exposure to; Health; Human; Hypothalamic structure; Image; In Vitro; Individual; Injection of therapeutic agent; Larva; Life; Location; Mammals; Mediating; Microscopy; Monitor; Morphogenesis; Motor Activity; Mus; Mutation; Narcolepsy; Neurons; Parkinson Disease; Patients; Pattern; Peptides; Pharmaceutical Preparations; Phenotype; Probability; Process; Rattus; Resolution; Rett Syndrome; Signal Transduction; Sleep; Sleep Disorders; System; Systems Development; Techniques; Testing; Time; Wakefulness; Zebrafish; hypocretin; in vivo; insight; nerve supply; nervous system disorder; neural circuit; neuron development; noradrenergic; optogenetics; overexpression; receptor; relating to nervous system; research study; sleep regulation; teleost; time use; zebrafish development

DESCRIPTION (provided by applicant): The hypocretin (Hcrt)-expressing neurons of the hypothalamus and the noradrenergic (NA) neurons of the locus ceruleus (LC) in the brainstem form two major centers of sleep regulation. Activity in these centers is correlated with wakefulness and stimulation of either cluster increases the probability of waking in sleeping mice. Importantly, disruption of either cluster can give rise to debilitating neurological disorder. Mutation of the hcrt locus, or loss of hcrt expressing neurons, results in the sleep disorder narcolepsy. Disruption of the LC has been implicated in Parkinson's and Alzheimer's diseases, Down and Rett syndromes, drug addiction, attention- deficit hyperactivity disorder and anxiety disorders. In vitro studies have provided strong evidence for interactions between Hcrt neurons and the LC. Hcrt neurons densely innervate the LC, LC neurons express the Hcrt receptor (HcrtR) and Hcrt can activate the LC in vitro. However, our understanding of the developmental, neuronal and behavioral interactions between these two systems in vivo is limited due to the large number of these neurons in mammals, their location deep within the mammalian brain, the complexity of their projections, the difficulty of imaging neural circuits in live mammals and the difficulty of analyzing mammalian behavior in a high-throughput fashion. Danio rerio (zebrafish) larvae are optically transparent and have only ~10 Hcrt and ~6 LC neurons per hemisphere. The mammalian pattern of hcrt expression and Hcrt neuron anatomy, activity and function are conserved in zebrafish. Furthermore, drugs that impinge upon the NA system similarly affect sleep/wake behaviors in zebrafish and mammals, suggesting that the function of the LC is conserved. This proposal exploits the advantageous features of zebrafish larvae to investigate Hcrt-LC interactions in development, neuronal activity and behavior. The development of Hcrt neuron projections to the LC and the arborization of these projections will be characterized at single neuron resolution in live animals using time-lapse microscopy. To test the hypothesis that LC neurons provide cues necessary for the normal extension and arborization of Hcrt projections, the LC will be ablated and silenced. To test the hypothesis that the LC provides cues that are sufficient to instruct Hcrt neuron morphogenesis, ectopic LC-like neurons will be generated. Hcrt overexpression, as well as optogenetic stimulation of Hcrt neurons, will be used to assess whether Hcrt neurons can drive LC activation in vivo. Hcrt overexpression in zebrafish larvae has been shown to increase locomotor activity, decrease the amount of time spent in a sleep-like state and induce hyperarousal. Cell ablation and neural silencing techniques will be used to assess the extent to which the LC mediates these phenotypes. These in vivo studies should yield insights into the developmental and functional interactions between two neuronal circuits of fundamental importance in sleep regulation and human health.

描述（由申请人提供）：在脑干中，下丘脑的表达神经元和下甲肾上腺素能（Na）神经元的神经元形成了睡眠调节的两个主要中心。这些中心的活性与清醒和刺激两组的刺激增加了沉睡小鼠醒来的可能性。重要的是，这两个集群的破坏都会导致神经系统疾病令人衰弱。 HCRT基因座的突变或表达神经元的HCRT的丧失导致睡眠障碍性睡病。 LC的破坏与帕金森氏症和阿尔茨海默氏症，疾病和RETT综合征，药物成瘾，注意力缺陷多动障碍和焦虑症有关。体外研究为HCRT神经元与LC之间的相互作用提供了有力的证据。 HCRT神经元密集地支配LC，LC神经元表达HCRT受体（HCRTR），HCRT可以在体外激活LC。但是，由于这些两个系统在体内之间的发育，神经元和行为相互作用的理解由于哺乳动物中的这些神经元数量，其位置在哺乳动物大脑内的深处，其预测的复杂性，对神经的难度，因此，这两个系统之间的发育，神经元和行为相互作用受到限制。活哺乳动物的电路和以高通量方式分析哺乳动物行为的困难。 Danio Rerio（斑马鱼）幼虫在光学上是透明的，每个半球只有〜10 hcrt和〜6 LC神经元。 HCRT表达和HCRT神经元解剖学，活性和功能的哺乳动物模式在斑马鱼中保守。此外，对NA系统造成的药物类似地影响斑马鱼和哺乳动物的睡眠/唤醒行为，这表明LC的功能是保守的。该建议利用了斑马鱼幼虫的有利特征，以研究发育，神经元活性和行为中的HCRT-LC相互作用。 HCRT神经元对LC的开发和这些投影的树博化的发展将在活动物中使用延时显微镜在单个神经元分辨率上进行表征。为了测试LC神经元提供的假设，为HCRT投影的正常延伸和树皮提供了必要的线索，LC将被消融和沉默。为了检验LC提供足以指导HCRT神经元形态发生的线索的假设，将产生异位LC样神经元。 HCRT过表达以及HCRT神经元的光遗传学刺激将用于评估HCRT神经元是否可以在体内驱动LC激活。斑马鱼幼虫中的HCRT过表达已被证明会增加运动活性，减少在睡眠状态下所花费的时间并诱导高伴侣。细胞消融和神经沉默技术将用于评估LC介导这些表型的程度。这些体内研究应洞悉对睡眠调节和人类健康中基本重要性的两个神经元回路之间的发展和功能相互作用。