Coupling neuroimaging with CLARITY and single cell genomics to dissect sex differences in the developing brain

摘要 许多神经行为疾病对女性和男性的影响不同，出现的时间也不同。 生命的各个阶段，导致得出这样的结论，即一个性别受到先天性别因素的保护，不受疾病的影响， 比如性腺激素和性染色体。这些对大脑的影响的部位和时间 发展是未知的。高通量高精度全脑MRI成像方法 无论是在动物模型还是在人类身上，都能非常灵敏地检测到这种变化。这 项目将利用这些强大的方法来分离性腺激素和性染色体的影响。 在小鼠模型“性染色体三体”(SCT)中，在9个不同发育年龄，发现 这些因素在何时何地导致大脑发育的性别差异(目标1)。SCT模型 比较具有不同数量和类型的性染色体(XX，XY，XXY，XYY)的小鼠，每种基因 存在于雄性或雌性性腺中。然后，一条新的分析管道将比较老鼠和人类 大脑发育的许多阶段和信息丰富的群体(不同的年龄，性别，荷尔蒙状况，和 性染色体互补)，以确定老鼠大脑中的哪些变化也在人类中发现，在 与不同疾病相关的特定脑区(目标2)。分析将指向鼠标的变化 模拟人脑发育的大脑。这些研究还将确定新的性别偏见效应 荷尔蒙和性染色体在小鼠特定发育阶段的局部大脑区域， 导致进一步研究在这些地点发生性行为引起的细胞和分子变化(目标3)。 性别偏见因素的细胞效应(细胞大小、数量、确定的细胞群体密度)将是 使用Clarity(透明脂交换丙烯酰胺杂交刚性成像相容组织- 水凝胶)。个体细胞类型中对激素和性染色体效应作出反应的基因途径 将使用单细胞RNA-seq来测量特定脑区的。这些研究，结合了高- 分辨率神经成像、清晰度和单细胞测序将提供概念的基础 在发育过程中，特定的细胞群在大脑的什么地方以及何时对性作出反应 作为假设的前奏，性别差异是性别保护作用的基础-- 细胞中的偏向因素。

Abstract Many neurobehavioral diseases affect females and males differently, and emerge at different stages of life, leading to the conclusion that one sex is protected from diseases by inherent sex factors, such as gonadal hormones and sex chromosomes. The sites and timing of these effects on brain development are unknown. The method of high-throughput high-precision whole-brain MRI imaging has the power to detect such changes with great sensitivity, in both animal models and humans. This project will exploit these powerful methods to separate gonadal hormonal and sex chromosome effects in the mouse model “Sex Chromosome Trisomy” (SCT), at 9 different ages of development, to discover where and when these factors cause sex differences in brain development (Aim 1). The SCT model compares mice with different numbers and types of sex chromosomes (XX, XY, XXY, XYY), each genotype present in gonadal males or females. Then, a novel pipeline of analysis will compare mouse and human brain development at many stages and in informative groups (differing by age, sex, hormonal status, and sex chromosome complement) to determine which changes in mouse brain are also found in humans, in specific brain regions related to different diseases (Aim 2). The analysis will point to changes in mouse brain that model human brain development. These studies will also pinpoint new sex-biasing effects of hormones and sex chromosomes at localized brain regions at specific developmental stages in mice, leading to further investigation of cellular and molecular changes caused by sex at those sites (Aim 3). Cellular effects (cell size, number, density of defined cell populations) of sex-biasing factors will be measured using CLARITY (Clear Lipid-exchanged Acrylamide-hybridized Rigid Imaging-compatible Tissue- hYdrogel). Gene pathways responding to hormonal and sex chromosome effects in individual cell types in specific brain regions will be measured using single cell RNA-seq. These studies, combining high- resolution neuroimaging, CLARITY, and single cell sequencing, will provide a foundation of concepts about where in the brain, and when during development, specific cell populations respond to sex factors, as a prelude for hypothesizing which sex differences underlie the protective effects of sex- biased factors in cells.