Molecular and spatial mapping of hypothalamic cell-types using the Xenium analyzer

项目摘要 人类和其他动物的类固醇激素协调生理和行为过程， 对社会环境的最佳反应。大脑是类固醇激素作用的主要场所;然而，我们的 类固醇激素在调节基因表达和大脑神经可塑性中的作用的知识， 它的婴儿期。我的研究项目旨在揭示类固醇激素基因表达 在大脑中，和特定的细胞功能使用Astatotilapia burtoni，一种慈鲷鱼， 社会等级制度在野外和实验室中，雄性A. burtoni沿着一个社会等级划分， 占主导地位的男性拥有明亮的色彩，积极捍卫领土，并与女性交配，而非 占统治地位的雄性则不然女性A. burtoni没有形成一个社会等级，但行为积极， 另一个是交配机会。社会等级是不断变化的，因为占主导地位的和非占主导地位的男性可以改变他们的地位。 社会地位取决于社会环境。这些复杂的社会互动与一个阶层的水平紧密相连， 叫做雄激素的类固醇激素我的研究项目利用了A.布尔托尼在 该实验室旨在发现雄激素在控制大脑基因和神经可塑性中的作用。在我们正在进行的 工作，我们已经产生了一个细胞类型特异性的分子图谱的A。Burtoni下丘脑单细胞 RNA测序（scRNAseq）。下丘脑是一个保守的大脑区域， 脊椎动物的行为。虽然scRNAseq提供了特定基因表达的细胞类型分辨率， 模式，它不会告诉你特定的细胞在解剖学上在感兴趣的区域内的位置。当前 一项提案旨在使用10 X Genomics Xenium原位分析仪解决这一问题，这是一种端到端的基因分析仪。 表达分析平台，可以同时标记新鲜冷冻或福尔马林固定的数百个基因， 切片组织。使用Xenium将允许不同细胞类型的空间分辨率 在我们目前的scRNAseq数据集中发现。我们将联合收割机结合所有细胞类型的遗传标记， 基因，如我们的Xenium分析中存在的不同雄激素受体（AR）基因，精确揭示了 哪些细胞和哪里是AR+，为与分子相关的可验证假设提供了精确的路线图。 以及细胞对社会行为的控制。我们的下丘脑空间转录组将与目前的 只有可用的小鼠下丘脑空间转录组，揭示了新的见解的遗传特征， AR+细胞类型和其他细胞群体在两个物种中大量用于社会行为研究。与 产生A. burtoni下丘脑，我们将能够解决 关于大脑和社会行为的分子和神经控制的基本问题。的确， 这些问题可能自然地与其他物种的社会行为控制有关， 人类以及社会系统如何在进化过程中出现。

PROJECT SUMMARY Steroid hormones in humans and other animals coordinate physiological and behavioral processes underlying optimal responses to the social environment. The brain is a major site of steroid hormone action; however, our knowledge of the role of steroid hormones in regulating gene expression and neuroplasticity in the brain is in its infancy. My research program aims to uncover the connections between steroid hormones, gene expression in the brain, and specific cellular functions using Astatotilapia burtoni, a cichlid fish that exhibits sophisticated social hierarchies. In the wild as in the laboratory, male A. burtoni stratify along a social hierarchy where dominant males possess bright coloration, aggressively defend a territory, and mate with females, while non- dominant males do not. Female A. burtoni do not form a social hierarchy but behave aggressively towards one another for mating opportunities. Social rank is in flux, as dominant and non-dominant males can change their status depending on the social milieu. These complex social interactions are tightly linked to levels of a class of steroid hormones called androgens. My research program leverages the social dynamics of A. burtoni in the laboratory to discover the role of androgens in controlling genes in the brain and neuroplasticity. In our ongoing work, we have generated a cell-type specific molecular atlas of the A. burtoni hypothalamus using single-cell RNA-sequencing (scRNAseq). The hypothalamus is a conserved brain region that coordinates adaptive social behaviors across vertebrates. While scRNAseq provides cell-type resolution of specific gene expression patterns, it does not tell you where the specific cells are anatomically within the region of interest. The current proposal aims to address this issue using the 10X Genomics Xenium in situ analyzer, an end-to-end gene expression analysis platform that can label hundreds of genes simultaneously in fresh-frozen or formalin-fixed, sectioned tissue. Using the Xenium will allow for the spatial resolution of the distinct cell-types we have identified in our current scRNAseq dataset. We will combine genetic markers of all cell-types with candidate genes such as distinct androgen receptor (AR) genes present in our Xenium analysis, revealing precisely which cells and where are AR+, providing a precise road-map for testable hypotheses related to the molecular and cellular control of social behavior. Our hypothalamic spatial transcriptome will be compared to the currently only available hypothalamic spatial transcriptome in mice to reveal novel insights into the genetic signatures of AR+ cell types and other cell populations in two species used heavily in social behavior studies. With the generation of a molecular and spatial map of the A. burtoni hypothalamus, we will be able to address fundamental questions regarding the molecular and neural control of the brain and social behavior. Indeed, these questions may connect naturally to those on the control of social behavior in other species such as humans and how social systems emerge throughout evolution.