Prenatal Alcohol and Anxiety: An Ontogenetic Role for CRF

摘要 怀孕期间饮酒是一种常见的现象，可以产生无数的损害，称为胎儿 酒精谱系障碍（FASD）。焦虑和酒精滥用的发展通常见于 产前酒精暴露（PAE）的个体，这两种适应不良的行为被认为是驱动 通过重叠的神经回路，即促肾上腺皮质激素释放因子受体（CRFR 1）系统， 内侧中央杏仁核（CeM）。在父母补助（R 01 AA 028566）中，我们假设中度PAE （mPAE）在妊娠第12天（杏仁核出现的胚胎期）降低CRF 1 R 通过个体发育发挥作用，导致急性酒精和CRF 1 R相互作用减少。为支持这一 中心假设，在父母补助金资助的第一年，我们表明G12 mPAE产生性- 在青少年野生型后代的CeM中CRF 1 R表达和功能的特异性改变， mPAE雄性后代也表现出焦虑样行为增加的发育期。然而，在这方面， 考虑到CeM复杂的微电路和最近的报告表明细胞特异性反应急性 乙醇在这一地区，这一多样性补充的研究目标是建立和利用 转基因大鼠系（CRF 1 R-Cre-tdTomato）中直接测试G12 mPAE的作用，并且所得的结果显示， 急性乙醇作用的改变，特别是对CRFR 1+和CRFR 1-神经元在CeM内， 发展具体来说，目标1将研究mPAE如何改变焦虑样行为和随后的 乙醇的摄入量。目的2将研究mPAE如何影响急性乙醇对CeM的作用 GABA在CRFR 1+和CRFR 1-神经元中的跨发育传递。目标3将决定 mPAE对急性乙醇和CRFR 1激活对CRFR 1+中CeM GABA传递的相互作用的影响， CRFR 1-个体发育中的神经元。对这些新假设的测试将促进我们对 神经生物学机制潜在的焦虑样行为和乙醇摄入量在整个发展和作为 这是PAE的后果。重要的是，在本申请中使用转基因模型的建议将补充 来自父母资助的拟议实验是基于使用野生型受试者，这限制了我们的研究。 解剖脑细胞膜微电路的能力该多元化补充计划的指导部分包括 直接培训1）繁殖和建立转基因大鼠群体，2）行为评估， 焦虑样行为和乙醇摄入，和3）全细胞膜片钳电生理技术， 转基因老鼠这项培训将使温彻斯特女士掌握开发新假设所需的工具， 联合收割机与其他方法在她的研究生培训和以后。此外，一个强大的， 制定了严格的个人发展计划，以确保专业和职业培训，包括 1)强调写作（包括手稿和赠款），2）统计分析，3）接触一般 酒精，胎儿酒精，和更大的神经科学领域，4）科学文献的解释，5）口头 演讲，6）网络，7）通过指导和管理本科生和大三学生的领导力 研究生。总体而言，拟议的研究和指导计划将为温彻斯特女士成为一名 成功的酒精研究者和未来的独立科学家。

Abstract Alcohol use during pregnancy is a common occurrence that can produce a myriad of impairments termed Fetal Alcohol Spectrum Disorder (FASD). The development of anxiety and alcohol misuse are commonly seen in individuals with prenatal alcohol exposure (PAE), two maladaptive behaviors that are considered to be driven by overlapping neural circuits, namely the corticotrophin releasing factor receptor (CRFR1) system within the medial central amygdala (CeM). In the parent grant (R01AA028566), we hypothesized that moderate PAE (mPAE) during gestational day (G) 12, an embryological period when the amygdala emerges, reduces CRF1R function through ontogeny, resulting in reduced acute alcohol and CRF1R interactions. In support of this central hypothesis, in the first year of funding of the parent grant, we showed that G12 mPAE produced sex- specific alterations in CRF1R expression and function in the CeM of adolescent wildtype offspring, a developmental period when mPAE male offspring also show heightened anxiety-like behaviors. However, given the intricate microcircuitry of the CeM and recent reports demonstrating cell-specific responses to acute ethanol within this region, the research goals of this Diversity Supplement are to establish and utilize a transgenic rat line (CRF1R-Cre-tdTomato) to directly test the effects of G12 mPAE and the resulting modifications of acute ethanol action specifically on CRFR1+ and CRFR1- neurons within the CeM across development. Specifically, Aim 1 will investigate how mPAE may alter anxiety-like behaviors and subsequent ethanol intake across development. Aim 2 will examine how mPAE affects the actions of acute ethanol on CeM GABA transmission in CRFR1+ and CRFR1- neurons across development. Aim 3 will determine the impact of mPAE on the interaction of acute ethanol and CRFR1 activation on CeM GABA transmission in CRFR1+ and CRFR1- neurons across ontogeny. Testing of these novel hypotheses will advance our understanding of neurobiological mechanisms underlying anxiety-like behaviors and ethanol intake across development and as a consequence of PAE. Importantly, the proposed use of a transgenic model in this application will supplement the proposed experiments from the parent grant that were based on using wildtype subjects which restrict our ability to dissect the microcircuitry of the CeM. The mentoring components of this Diversity Supplement include direct training on 1) breeding and establishment of a transgenic rat colony, 2) behavioral assessment of anxiety-like behaviors and ethanol intake, and 3) whole-cell patch-clamp electrophysiological techniques in transgenic rats. This training will equip Ms. Winchester with tools necessary to develop novel hypotheses and combine with additional approaches later in her graduate training and beyond. Additionally, a strong and rigorous individual development plan has been designed to assure professional and career training, including 1) emphasis on writing (both manuscripts and grants), 2) statistical analyses, 3) exposure to the general alcohol, fetal alcohol, and larger neuroscience field, 4) interpretation of the scientific literature, 5) oral presentations, 6) networking, and 7) leadership through mentoring and managing undergraduate and junior graduate students. Overall, the proposed research and mentoring plan will prepare Ms. Winchester to be a successful alcohol researcher and future independent scientist.