Neurocircuitry Mapping and Genotyping Core

描述(由申请人提供)：这是一份名为“神经电路图谱和基因分型核心”的U01基金的竞争性续期申请；该申请是作为NIAAA赞助的“酒精中毒综合神经科学倡议(INIA)-West(G.Koob，Pi)”的成员提交的。这项申请延续了当前供资期间对研究和核心活动的重点。本供资期间的主要核心活动是：a)掌握对中、大样本使用加权基因协方差网络分析(WGCNA)(Lancu等人)。(B)制定和实施定量RNAseq战略(Bottomly等人，2011年；附录A)。有了这些工具，我们建议1)直接测序黑暗高饮酒(HDID)小鼠复制品系和HS/NPT对照动物的转录组(~25,000,000 75核苷酸读数/样本)，以及2)与适当的对照组一起对完成慢性间歇酒精(CIE)程序的HDID动物的转录组进行测序。这项分析所需的组织将由Crabbe U01提供。因为HDID和对照是从8向近交品系杂交中获得的(Hitzemann等人。1994)，考虑到掩蔽寡核苷酸阵列数据从来都不是最优的(见Walter等人)，RNAseq具有特殊性。2007、2009)。N=32个/组；以前的工作(Lancu等人2010年)表明，这种规模的样本足以进行拟议的分析。样本通过激光捕获显微解剖(LCM)收集；区域优先分析将是杏仁核(CEA)和杏仁核(GT)的中央核；边缘下皮质(IL)和边缘皮质(GT)；前皮质(PL)。枕叶皮质(OC)将被用作控制区。目标1侧重于酗酒，而目标2侧重于在有限获取的两瓶选择范例中，长期酒精暴露如何影响乙醇消费。我们的工作假设是，共表达网络之间的差异而不是单个基因的差异表达具有最大的翻译价值(参见Oti等人)。2008年；赵等人。2010)。在目标3中，来自酒精暴露的猕猴的样本(Grant U01-INLA-Stress)将被测序。来自CEA和大脑皮层第25和32区的数据将与具体目标1和2中的结果进行比较。

DESCRIPTION (provided by applicant): This is a competing renewal application for a U01 grant entitled "Neurocircuitry Mapping and Genotyping Core"; the application is submitted as a member of the NIAAA sponsored "Integrative Neuroscience Initiative on Alcoholism (INIA)-West (G. Koob, PI). The application continues the focus of the current funding period on both research and core activities. Key core activities of the current funding period were a) the mastery of the use of the Weighted Gene Co-variance Network Analysis (WGCNA) for moderate to large sample sizes (lancu et al. 2010) and b) the development of a strategy for and implementation of quantitative RNAseq (Bottomly et al, 2011; Appendix A). With these tools in hand, we propose 1) to directly sequence the transcriptome ( ~ 25,000,000 75 bp reads/sample) in both replicate High Drinking in the Dark (HDID) mouse lines and in the HS/NPT control animals and 2) to sequence the transcriptome HDID animals that have completed the chronic intermittent ethanol (CIE) procedure with the appropriate control groups. The tissues needed for this analysis will be provided by the Crabbe U01. As the HDID and controls are derived from a 8- way inbred strain cross (Hitzemann et al. 1994), RNAseq is particularity useful, given that masking oligonucleotide array data is never optimal (see Walter et al. 2007,2009). N = 32/group; previous work (lancu et al. 2010) has illustrated that samples of this size are adequate for the proposed analyses. Samples are collected by laser capture micro-dissection (LCM); the regional priority for analysis will be the central nucleus of the amygdala (CeA) > the infralimbic cortex (IL) > the prelimbic cortex (PL). The occipital cortex (OC) will be used as a control region. Aim 1 focuses on binge drinking whereas aim 2 focuses on how chronic ethanol exposure affects ethanol consumption in limited access 2-bottle choice paradigm. Our working hypothesis is that differences between co-expression networks and not the differential expression of individual genes have the greatest translational value (see e.g. Oti et al. 2008; Zhao et al. 2010). In Aim 3, samples from ethanol exposed macaques (Grant U01- INlA-Stress) will be sequenced. Data from the CeA and cortical areas 25 and 32 will be compared to the results obtained in specific aims 1 and 2.