The supraoptic nucleus (SON) of the hypothalamus contains magnocellular neurosecretory cells that secrete the hormones vasopressin and oxytocin. Sex differences in SON gene expression have been relatively unexplored. Our study used spatially resolved transcriptomics to visualize gene expression profiles in the SON of adult male (n = 4) and female (n = 4) Sprague-Dawley rats using Visium Spatial Gene Expression (10x Genomics). Briefly, 10-μm coronal sections (~ 4 × 4 mm) containing the SON were collected from each rat and processed using Visium slides and recommended protocols. Data were analyzed using 10x Genomics’ Space Ranger and Loupe Browser applications and other bioinformatic tools. Two unique differential expression (DE) analysis methods, Loupe Browser and DESeq2, were used. Loupe Browser DE analysis of the SON identified 116 significant differentially expressed genes (DEGs) common to both sexes (e.g., Avp and Oxt), 31 significant DEGs unique to the males, and 73 significant DEGs unique to the females. DESeq2 analysis revealed 183 significant DEGs between the two groups. Gene Ontology (GO) enrichment and pathway analyses using significant genes identified via Loupe Browser revealed GO terms and pathways related to (1) neurohypophyseal hormone activity, regulation of peptide hormone secretion, and regulation of ion transport for the significant genes common to both males and females, (2) Gi signaling/G-protein mediated events for the significant genes unique to males, and (3) potassium ion transport/voltage-gated potassium channels for the significant genes unique to females, as some examples. GO/pathway analyses using significant genes identified via DESeq2 comparing female vs. male groups revealed GO terms/pathways related to ribosomal structure/function. Ingenuity Pathway Analysis (IPA) identified additional sex differences in canonical pathways (e.g., ‘Mitochondrial Dysfunction’, ‘Oxidative Phosphorylation’) and upstream regulators (e.g., CSF3, NFKB complex, TNF, GRIN3A). There was little overlap in the IPA results for the two different DE methods. These results suggest sex differences in SON gene expression that are associated with cell signaling and ribosomal pathways. The brain releases the hormones oxytocin and vasopressin from the supraoptic nucleus. Oxytocin is involved in maternal behaviors, lactation, and childbirth. Vasopressin is involved in sex-based differences in social behavior and body fluid regulation. However, how the brain contributes to sex-based differences in vasopressin and oxytocin release is poorly understood. This study aimed to address this knowledge gap using spatial transcriptomics to test for sex-based differences in gene expression in the supraoptic nucleus. Spatial transcriptomics combines anatomy with gene sequencing technology, allowing us to identify groups of genes that are expressed in specific locations in the brain. We applied this approach to brain sections containing the supraoptic nucleus from four adult male and four adult female rats. Using a data analysis workflow specifically for spatial transcriptomics, we identified genes that are significantly expressed in the supraoptic nuclei of both males and females (116 genes), primarily males (31 genes), and primarily females (73 genes). Genes enriched in the supraoptic nucleus of both males and females are related to the synthesis and release of peptides like vasopressin and oxytocin. Genes specific to the male supraoptic nucleus are broadly related to cell signaling, while the female-specific genes are related to ion transporters/channels. Results from a more traditional data analysis workflow identified sex-based differences in the expression of genes related to cell metabolism and protein synthesis. Together these results may provide a mechanistic foundation that can be used to better understand how differences in gene expression related to biological sex influence brain function.

中文翻译：

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空间转录组学揭示成年大鼠视上核基因表达的基础性别差异，与细胞信号传导和核糖体途径相关

下丘脑的视上核 (SON) 含有巨细胞神经分泌细胞，可分泌加压素和催产素。SON 基因表达的性别差异相对尚未被探索。我们的研究使用空间分辨转录组学，使用 Visium 空间基因表达 (10x Genomics) 来可视化成年雄性 (n = 4) 和雌性 (n = 4) Sprague-Dawley 大鼠 SON 中的基因表达谱。简而言之，从每只大鼠收集包含 SON 的 10 μm 冠状切片（~ 4 × 4 mm），并使用 Visium 载玻片和推荐方案进行处理。使用 10x Genomics 的 Space Ranger 和 Loupe Browser 应用程序以及其他生物信息学工具对数据进行分析。使用了两种独特的差异表达 (DE) 分析方法：Loupe Browser 和 DESeq2。SON 的 Loupe Browser DE 分析确定了两性共有的 116 个显着差异表达基因 (DEG)（例如 Avp 和 Oxt）、男性特有的 31 个显着差异表达基因和女性特有的 73 个显着差异表达基因。DESeq2 分析显示两组之间存在 183 个显着 DEG。使用通过放大浏览器识别的重要基因进行基因本体论 (GO) 富集和通路分析，揭示了与 (1) 神经垂体激素活性、肽激素分泌调节以及男性和女性共有的重要基因的离子转运调节相关的 GO 术语和通路。作为一些例子，(2)男性特有的重要基因的Gi信号传导/G蛋白介导的事件，以及(3)女性特有的重要基因的钾离子转运/电压门控钾通道。使用通过 DESeq2 识别的重要基因进行 GO/通路分析，比较女性与男性组，揭示与核糖体结构/功能相关的 GO 术语/通路。Ingenuity Pathway Analysis (IPA) 发现了经典途径（例如，“线粒体功能障碍”、“氧化磷酸化”）和上游调节因子（例如，CSF3、NFKB 复合体、TNF、GRIN3A）中的其他性别差异。两种不同 DE 方法的 IPA 结果几乎没有重叠。这些结果表明 SON 基因表达的性别差异与细胞信号传导和核糖体途径相关。大脑从视上核释放催产素和加压素激素。催产素参与母亲行为、哺乳和分娩。加压素参与社会行为和体液调节的性别差异。然而，人们对大脑如何影响加压素和催产素释放的性别差异知之甚少。本研究旨在利用空间转录组学来测试视上核基因表达的性别差异，以解决这一知识差距。空间转录组学将解剖学与基因测序技术相结合，使我们能够识别在大脑特定位置表达的基因组。我们将这种方法应用于包含四只成年雄性和四只成年雌性大鼠的视上核的脑切片。使用专门针对空间转录组学的数据分析工作流程，我们鉴定了在男性和女性视上核中显着表达的基因（116 个基因），主要是男性（31 个基因），主要是女性（73 个基因）。男性和女性视上核中富含的基因与加压素和催产素等肽的合成和释放有关。男性视上核特有的基因与细胞信号传导广泛相关，而女性特有的基因与离子转运蛋白/通道相关。更传统的数据分析工作流程的结果确定了与细胞代谢和蛋白质合成相关的基因表达的性别差异。这些结果共同提供了一个机制基础，可用于更好地理解与生物性别相关的基因表达差异如何影响大脑功能。