Prenatal Alcohol and Anxiety: An Ontogenetic Role for CRF

摘要 怀孕期间饮酒是一种常见的情况，会导致许多称为胎儿的损害 酒精谱系障碍(FASD)。焦虑和酗酒的发展通常见于 有产前酒精暴露(PAE)的个人，两种被认为是驱动因素的适应不良行为 通过重叠神经回路，即促肾上腺皮质激素释放因子受体(CRFR1)系统在 杏仁中央内侧核(CEM)。在父授权(R01AA028566)中，我们假设适度的PAE (MPAE)在妊娠12天，杏仁核出现的胚胎期，减少CRF1R 通过个体发育发挥作用，导致急性酒精和CRF1R相互作用减少。为了支持这一点 中心假设，在父母资助的第一年，我们发现G12 mPAE产生了性行为- 青少年野生型子代CEM中CRF1R表达和功能的特异性变化 当mPAE雄性后代也表现出高度焦虑样行为的发育期。然而， 鉴于CEM错综复杂的微电路和最近的报告显示细胞对急性白血病的特异性反应 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 转基因大鼠品系(CRF1R-Cre-tdTomato)直接检测G12 mPAE的作用及其结果 急性乙醇对跨中脑皮层内CRFR1和CRFR1阳性神经元的影响 发展。具体地说，目标1将调查mPAE如何改变焦虑样行为和随后的 不同发育阶段的乙醇摄入量。目标2将研究mPAE如何影响急性乙醇对CEM的作用 CRFR1和CRFR1神经元发育过程中的GABA传递。目标3将确定 急性乙醇和CRFR1激活对CRFR1和CRFR1细胞CEM-GABA传递的影响 CRFR1--个体发育中的神经元。检验这些新颖的假说将促进我们对 焦虑样行为和酒精摄入在发育和AS中的神经生物学机制 这是PAE的后果。重要的是，在这一应用中建议使用转基因模型将补充 来自父母拨款的拟议实验是基于使用野生型受试者的，这限制了我们的 能够剖析CEM的微电路。这份多样性补充资料的指导部分包括 直接培训1)培育和建立转基因大鼠群体，2)行为评估 焦虑样行为和酒精摄入，以及3)全细胞膜片钳电生理技术 转基因大鼠。这一培训将使温彻斯特女士掌握必要的工具，以开发新的假设和 在她后来的研究生培训和以后的培训中，结合其他方法。此外，一个强大的和 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(手稿和补助金)，2)统计分析，3)了解一般情况 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，以及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 成功的酒精研究人员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.