Genetic Basis of Opioid Reward and Aversion in Mice

描述(申请人提供)：处方类阿片类药物的非医疗滥用在美国是一个主要的社会经济问题。众所周知，药物滥用有遗传因素，流行病学研究表明，报告愉快经历的人最有可能过渡到药物滥用。因此，这项应用试图确定阿片类药物奖赏和厌恶的遗传基础。我们将使用一个提供遗传信息的小鼠品系小组和一个行为模型-位置条件作用试验-来筛选对常见滥用的处方阿片激动剂羟考酮(Oxy)的奖赏反应和对阿片拮抗剂纳洛酮(NAL)的厌恶反应的差异。在目标1中，我们将确定导致Oxy奖赏和NAL厌恶差异的基因组区域。在目标2中，我将接受信使核糖核酸测序(RNA-seq)方面的培训，以确定这些基因组区域中差异表达的基因和剪接变体可能导致氧奖赏和NAL厌恶的变异。在目标3中，我们将产生携带这些基因组区域的一小部分的同源小鼠，以缩小潜在相关基因的清单。在这些区域中，我们将选择差异表达的候选基因，通过分子遗传学技术进行进一步分析。这一应用的结果将有助于识别导致阿片类药物奖赏和厌恶的基因，这将对理解和治疗阿片类药物滥用产生影响。我的导师亚伯拉罕·帕尔默博士在这个项目的各个方面都有多年的经验，包括QTL定位、转录组分析和大规模的小鼠表型研究。Palmer博士在转录组分析方面的专业经验涉及微阵列；因此，我聘请了Jonathan Pritchard博士(共同导师)和Yoav Gilad博士(咨询委员会成员)来提供RNA-seq分析和技术方面的指导和培训。此外，我还成立了一个顾问委员会，由帕尔默博士、普里查德博士、吉拉德博士、哈里特·德维特博士和詹姆斯·扎克尼博士组成。De Wit博士在药物滥用的啮齿动物和人类研究方面拥有广泛的背景和专业知识，并对拟议研究的翻译方面特别感兴趣。德维特博士进行的基因关联研究涉及对滥用药物的敏感性，特别是与药物奖励等主观表型的关系。因此，De Wit博士很有能力就这一应用的行为、遗传和药理学方面提供专家建议，并可以提供应用结果的翻译上下文。扎克尼博士是阿片类药物药理学和人类阿片类药物特征表型方面的专家，并发表了多篇关于阿片类药物--特别是Oxy--的生理和主观反应的变异性的论文。扎克尼博士有兴趣将这一应用的结果应用于关于人类阿片类药物敏感性遗传基础的翻译研究。支持和补充这一应用的制度环境非常好。在该项目的基因和基因组方面，人类遗传学系有几位优秀的教员与我互动，包括我的委员会成员。我们有几个附属于该部门的研讨会系列，我将参加，包括正在进行的工作，每月研讨会系列，杂志俱乐部，和模式生物的遗传学。至少有六台Illumina GA2机器可以运行我的RNA-Seq样本，其中三台位于加州大学校园。在神经科学方面，Paul Vezina博士领导一个由NIDA资助的T32培训计划，“药物滥用研究的神经精神药理学培训”，以及每月两次的研讨会系列。德维特博士和帕尔默博士都参加了这个项目，私家侦探S和其他几名对成瘾生物学都感兴趣的神经科学教职员工也参加了这个项目。我的长期目标是在一所主要的研究型大学建立我自己的学术研究项目，旨在识别与药物滥用相关的表型相关基因，从本申请中概述的位置条件反射试验开始。在当前的QTL定位时代，转录组分析现在是寻找导致复杂表型(如药物敏感性)的基因的主要补充工具。因此，我在RNA-seq分析方面所接受的培训将帮助我实现这个职业目标。在成瘾生物学领域，关于地点偏好的遗传决定因素，存在着巨大的差距。由于这一分析方法在成瘾生物学中得到了广泛的应用，这一领域的研究无疑将有助于我们理解现有的众多研究，并为药物奖励和滥用的机制提供新的见解。我的实验室未来的应用将致力于测试这项研究中确定的基因对与药物滥用相关的其他行为的多效性影响，包括戒断、自我管理、灭绝、重新巩固和恢复。我还对奖励/厌恶和痛苦路径之间的交集感兴趣，并希望进一步探索这一领域的研究，作为未来的应用。

DESCRIPTION (provided by applicant): The nonmedical abuse of prescription opioids is a major socioeconomic problem in the United States. Drug abuse is known to have a genetic component and epidemiological studies indicate that individuals reporting a pleasurable experience are most likely to transition to drug abuse. Accordingly, this application seeks to identify the genetic basis of opioid reward and aversion. We will use a genetically informative panel of mouse strains and a behavioral model- the place conditioning assay- to screen for differences in the rewarding response to the commonly abused prescription opioid agonist oxycodone (OXY) and the aversive response to the opioid antagonist naloxone (NAL). In Aim 1, we will identify genomic regions that are responsible for differences in OXY reward and NAL aversion. In Aim 2, I will receive training in mRNA sequencing (RNA-seq) to identify differentially expressed genes and splice variants in these genomic regions as potentially causal for variation in OXY reward and NAL aversion. In Aim 3, we will generate congenic mice carrying small portions of these genomic regions to narrow the list of potential genes responsible. Within these regions, we will choose candidate genes that are differentially expressed for further analysis via molecular genetics techniques. The outcome of this application will aid in identifying genes contributing to opioid reward and aversion which will have implications for understanding and treating opioid abuse. My mentor, Dr. Abraham Palmer, has several years of experience with regard to all aspects of this project, including QTL mapping, transcriptome analysis, and large-scale mouse phenotyping studies. Dr. Palmer's history of expertise for transcriptome analysis has involved microarrays; thus, I have recruited Dr. Jonathan Pritchard (Co-Mentor) and Dr. Yoav Gilad (advisory committee member) to provide mentorship and training in the analytical and technical aspects of RNA-seq. Additionally, I have formed an advisory committee consisting of Dr. Palmer, Dr. Pritchard, Dr. Gilad, Dr. Harriet de Wit, and Dr. James Zacny. Dr. de Wit has extensive background and expertise in both rodent and human studies of drug abuse and is particularly interested in the translational aspect of the proposed research. Dr. de Wit conducts genetic association studies involving sensitivity to drugs of abuse, notably with subjective phenotypes such as drug reward. Thus, Dr. de Wit is well-positioned to provide expert advice on the behavioral, genetic, and pharmacological aspects of this application and can offer a translational context from which to apply the results. Dr. Zacny is an expert in opioid pharmacology and human phenotyping of opioid traits and has published multiple papers regarding variability in the physiological and subjective response to opioids- in particular OXY. Dr. Zacny is interested in applying the results of this application to translational studies regarding the genetic basis of opioid sensitivity in humans. The institutional environment for supporting and complementing this application is excellent. On the genetic and genomic end of the project, there are several excellent faculty in the Department of Human Genetics with whom I interact, including members of my committee. We have several seminar series affiliated with the department that I will participate in, including Work in Progress, Monthly Seminar Series, Journal Club, and Genetics of Model Organisms. There will be at least six Illumina GA2 machines in place for my RNA-seq samples to be run, three of which are located on the U of C campus. On the neuroscience end, Dr. Paul Vezina heads a NIDA-funded T32 training program, "Neuropsychopharmacology Training for Drug abuse Research" and seminar series that meets twice per month. Dr. de Wit and Dr. Palmer are both participating P.I.'s in this program, along with several other neuroscience faculty members who are all interested in addiction biology. My long-term objective is to establish my own academic research program at a major research university aimed at identifying genes involved with phenotypes associated with drug abuse, starting with the place conditioning assay as outlined in this application. In the current era of QTL mapping, transcriptome analysis is now a mainstay complementary tool for finding genes contributing to complex phenotypes such as drug sensitivity. Thus, the training I receive in RNA-seq analysis will help me to realize this career goal. There is a huge gap in the field of addiction biology regarding the genetic determinants of place preference. Because this assay is so widely used in addiction biology, this area of research will undoubtedly contribute to our understanding of the multitude of existing studies and provide novel insight into the mechanisms of drug reward and abuse. Future applications from my laboratory will be aimed at testing genes identified in this study for pleiotropic effects on other behaviors associated with drug abuse, including withdrawal, self-administration, extinction, reconsolidation, and reinstatement. I am also interested in the intersection between reward/aversion and pain pathways and would like to further explore this area of research as a future application.