Neurons of the medial habenula regulate behavioral responses to nicotine in mouse

内侧缰核神经元调节小鼠对尼古丁的行为反应

基本信息

批准号：
10017027
负责人：
Christian Peters
金额：
$ 24.9万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-05-01 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10017027
关键词：
Acute Affect Affinity Agonist Amino Acids Area Award Behavior Behavioral Binding Biological Assay Biological Models Biophysics Blood Brain Calcium Cannulas Cause of Death Cell Nucleus Chloride Channels Chloride Ion Chronic Collaborations Complement Complex Cystic Fibrosis Data Dependence Dyes Epithalamic structure Exhibits Exposure to Family Fluorescence Genes Genetic Goals Habenula Hour Human Image Imaging Techniques Implant Infusion procedures Injections Intake Integral Membrane Protein Intravenous Ion Channel Ion Channel Gating Ion Channel Protein Ions Journals Knock-out Knockout Mice Lead Link Mastication Medial Mediating Membrane Mentors Microinjections Modeling Molecular Molecular Probes Motivation Mucous body substance Mus Neuromuscular Junction Neurons Nicotine Nicotine Dependence Nicotine Withdrawal Nicotinic Receptors Output Pathway interactions Pharmaceutical Preparations Pharmacology Phase Phenotype Physiological Physiology Play Postdoctoral Fellow Prevalence Property Publications Publishing RNA Interference Regulation Research Rewards Role Self Administration Slice Smoker Structure Sucrose Symptoms System Techniques Technology Testing Tissues Tobacco smoking behavior Training Translating Viral Wild Type Mouse Withdrawal Withdrawal Symptom addiction avoidance behavior behavior influence behavioral phenotyping behavioral response brain circuitry brain tissue deprivation design drug seeking behavior electronic cigarette use experience experimental study fluorescence imaging genetic analysis imaging study in vivo in vivo imaging interest interpeduncular nucleus knock-down microendoscope mouse model mutant neuronal circuitry neurophysiology neurotransmission nicotine cessation nicotine craving nicotine exposure nicotine seeking behavior nicotine use novel post-doctoral training preventable death programs receptor response reward processing transcriptome sequencing voltage

Acute Affect Affinity Agonist Amino Acids Area Award Behavior Behavioral Binding Biological Assay Biological Models Biophysics Blood Brain Calcium Cannulas Cause of Death Cell Nucleus Chloride Channels Chloride Ion Chronic Collaborations Complement Complex Cystic Fibrosis Data Dependence Dyes Epithalamic structure Exhibits Exposure to Family Fluorescence Genes Genetic Goals Habenula Hour Human Image Imaging Techniques Implant Infusion procedures Injections Intake Integral Membrane Protein Intravenous Ion Channel Ion Channel Gating Ion Channel Protein Ions Journals Knock-out Knockout Mice Lead Link Mastication Medial Mediating Membrane Mentors Microinjections Modeling Molecular Molecular Probes Motivation Mucous body substance Mus Neuromuscular Junction Neurons Nicotine Nicotine Dependence Nicotine Withdrawal Nicotinic Receptors Output Pathway interactions Pharmaceutical Preparations Pharmacology Phase Phenotype Physiological Physiology Play Postdoctoral Fellow Prevalence Property Publications Publishing RNA Interference Regulation Research Rewards Role Self Administration Slice Smoker Structure Sucrose Symptoms System Techniques Technology Testing Tissues Tobacco smoking behavior Training Translating Viral Wild Type Mouse Withdrawal Withdrawal Symptom addiction avoidance behavior behavior influence behavioral phenotyping behavioral response brain circuitry brain tissue deprivation design drug seeking behavior electronic cigarette use experience experimental study fluorescence imaging genetic analysis imaging study in vivo in vivo imaging interest interpeduncular nucleus knock-down microendoscope mouse model mutant neuronal circuitry neurophysiology neurotransmission nicotine cessation nicotine craving nicotine exposure nicotine seeking behavior nicotine use novel post-doctoral training preventable death programs receptor response reward processing transcriptome sequencing voltage

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项目摘要

Project Summary/Abstract The long-term goal of my research is to determine the mechanism by which the brain responds to the presence of nicotine and leads to voluntary regulation of its intake. The ultimate purpose is to develop a better understanding of how positively and negatively rewarding effects of this drug can lead to nicotine craving or nicotine aversive behavior, and how these two opposing effects are both regulated by the habenulo- interpeduncular circuitry. The studies detailed in this proposal will focus specifically on the neurophysiology of the medial habenula and the behaviors associated with activity of neurons in that region. Nicotine addiction, fed by regular tobacco smoking or chewing, or more recently by e-cigarette use, is a leading cause of death in both the developed and developing world. Nicotine acts in the body as an extremely potent agonist of the eponymous nicotinic acetylcholine receptor family (nAChR), which are ion channel proteins with functions in neurotransmission in the brain and at neuromuscular junctions in the periphery. NAChr are ubiquitous throughout the brain, and the mechanisms by which nicotine influences behavior to produce physiological dependency are complex. A specific nucleus in the epithalamus called the medial habenula has been implicated as a locus where circulating nicotine binds directly to a specific subtype of nAChR, in which genetic mutants have been found to be upregulated in heavy smokers, to produce downstream behavioral responses regulating voluntary nicotine intake. Intriguingly, a recently identified calcium-activated chloride channel called TMEM16A (Transmembrane protein of unknown function 16A) is very highly expressed in the medial habenula but almost nowhere else in the brain, and it is likely to contribute strongly to the firing properties of mHb neurons, though the mechanism by which nicotine produces its effects on the habenula and its associated circuitry is not known. I propose to investigate the mechanism of nicotine aversion in medial habenula neurons, and to begin by studying the TMEM16A channel as a functional contributor to nicotine aversion. In the first aim pursued during the K99 phase, I will use self-administration assays with direct habenular nicotine microinjections, as well as withdrawal assays with long term exposure followed by deprivation, to examine how acute and chronic nicotine affects behavioral responses mediated in the medial habenula and how knockout of the Tmem16a gene in mice affects those effects. In the second aim, I will begin to more generally probe the medial habenula's function by implanting microendoscopes and using in vivo fluorescence imaging experiments to directly visualize neuronal activity in response to acute and chronic nicotine exposure. In the third aim, taking place following the transition to independence, I will combine the in vivo imaging and self-administration paradigms to rigorously investigate medial habenular neuron function in regulating nicotine self-administration in wild-type mice and models with established nicotine- seeking phenotypes traceable to the medial habenula. I will also probe the cellular basis of plasticity during onset of nicotine dependence using RNAseq analysis of medial habenula neurons in naïve and dependent mice. Finally, in aim 4, I will perform paired slice recordings of the medial habenula and the interpeduncular nucleus to investigate the input/output relationship of firing in the mHb, and how these are affected by focal applications of nicotine or other specific agonists, as well as by pre-exposure of the mice to chronic nicotine. Since joining the Jan lab in 2012, my research has focused specifically on the biophysics and pharmacology of the TMEM16A channel. Directly upon joining the lab, I undertook a short collaboration with Jan lab postdoc Fen Huang, Jason Rock, and others to explore the role of TMEM16A in airway mucus secretion and the ability of TMEM16A blockers to alleviate this in a cystic fibrosis model. Having a background in the structural biophysics of ion channel gating, I quickly became interested in how the TMEM16A channel is opened in response to elevation of intracellular calcium concentration (as would happen as a result of activation of a nicotine receptor, for instance), a study which resulted in a publication in the journal eLife, where I was co-first author. More recently, I performed a study to identify amino acids in TMEM16A important for chloride ion flux, and to characterize two novel inhibitory compounds with a high affinity for the TMEM16A ion pore. That study was published in PNAS in 2015. Over the remainder of my postdoctoral training, I would like to gain experience in experimental approaches to translate my expertise in the study of ion channel function into better understanding of how they contribute to the physiology of neuronal circuitry of the brain, with a specific focus on nicotine-sensitive circuitry involved in addiction and associated behavioral phenotypes. I believe the K99/R00 Pathway to Independence award is an ideal program for this goal, as it allows me to use both my specific areas of strength, and to acquire my desired training in the K99 phase to develop long term projects studying the neurophysiology of nicotine addiction and aversion in the R00 phase.

项目摘要/摘要我的研究的长期目标是确定大脑对存在的反应的机制尼古丁的摄入量并导致自愿调节。最终目的是发展更好了解该药物对这种药物的积极和负面奖励的影响如何导致尼古丁渴望或尼古丁厌恶行为，以及这两个相反的作用如何受到habenulo-的调节枝条回路。该提案中详细介绍的研究将专门关注内侧Habenula及其与该区域神经元活性相关的行为。尼古丁成瘾是由常规烟草吸烟或咀嚼喂养的，或者最近通过电子烟的使用是一种发达国家和发展中国家的主要死亡原因。尼古丁在体内起作用有效的烟碱乙酰胆碱受体家族（NACHR）的有效激动剂，它是离子通道蛋白质具有神经传递在大脑和周围神经肌肉连接处的蛋白质。 NACHR在整个大脑中无处不在，尼古丁影响行为的机制产生身体依赖是复杂的。上皮中的特定核称为培养基 Habenula被认为是一个基因座，循环尼古丁直接与特定亚型结合 NACHR，其中发现遗传突变体已在浓烟中进行更新，以生产下游行为反应调节自愿尼古丁的摄入。有趣的是，最近确定的称为TMEM16A的钙活化的氯化物通道（未知功能16a的跨膜蛋白）为在内侧的Habenula中非常高度表达，但在大脑中几乎没有其他地方，它很可能有助于强烈地具有MHB神经元的发射特性，尽管尼古丁产生其作用的机制在Habenula及其相关电路上尚不清楚。我建议研究尼古丁的机制内侧Habenula神经元的厌恶，首先研究TMEM16A通道作为功能否认尼古丁的贡献者。在K99阶段的第一个目标中，我将使用自我管理直接超级尼古丁微型注射和长期暴露的提款测定法其次是剥夺，以检查急性和慢性尼古丁如何影响介导的行为反应小鼠中的内侧Habenula以及TMEM16A基因的敲除如何影响这些作用。在第二个目标中我将通过植入微观镜检查并使用的体内荧光成像实验，以直接对急性和慢性尼古丁暴露。在第三个目标中，发生在过渡到独立之后，我将结合严格研究培养基神经元的体内成像和自我管理范例在调节野生型小鼠中调节尼古丁自我给药的功能以及具有既定尼古丁的模型寻求可追溯到内侧Habenula的表型。我还将探测可塑性的细胞基础使用RNASEQ内侧habenula神经元的RNASEQ分析在幼稚和依赖的内侧Habenula神经元的依赖性发作老鼠。最后，在AIM 4中，我将执行内侧Habenula的配对切片录音调查MHB发射的输入/输出关系的细胞核，以及它们如何受到焦点的影响尼古丁或其他特定激动剂的应用，以及小鼠前暴露于慢性尼古丁。自2012年加入Jan实验室以来，我的研究专门针对生物物理学和 TMEM16A通道的药理学。直接加入实验室，我与 Jan Lab Postdoc Fen Huang，Jason Rock等人探索TMEM16A在气道粘液中的作用分泌和TMEM16A阻滞剂在囊性纤维化模型中减轻这种情况的能力。有背景在离子通道门控的结构生物物理学中，我很快就对TMEM16A通道的感兴趣响应于细胞内钙浓度升高而开放的（由于例如，激活尼古丁受体的研究），这是一项研究，导致了《埃里夫》期刊上的出版物我是联合第一作者。最近，我进行了一项研究，以鉴定TMEM16A中的氨基酸对氯离子通量，并表征两种新型抑制性化合物，对TMEM16A离子具有高亲和力毛孔。该研究于2015年在PNAS上发表。在我的博士后培训的其余部分中，我想要获得实验方法的经验，以翻译我在离子渠道功能研究方面的专业知识更好地了解它们如何促进大脑神经元电路的生理学对成瘾和相关行为表型涉及的尼古丁敏感电路的具体关注。我相信K99/R00独立奖奖是该目标的理想计划，因为它允许我使用我的特定优势领域，并在K99阶段获得我所需的培训以发展长期研究R00阶段中尼古丁成瘾和厌恶的神经生理学的项目。