A survey of the mouse hindbrain in the fed and fasted state using single-nucleus RNA sequencing.

Objective: The area postrema (AP) and the nucleus tractus solitaris (NTS), located in the hindbrain, are key nuclei that sense and integrate peripheral nutritional signals and, consequently, regulate feeding behaviour. While single cell transcriptomics have been used in mice to reveal the gene expression profile and heterogeneity of key hypothalamic populations, similar in-depth studies have not yet been performed in the hindbrain. Methods: Using single-nucleus RNA sequencing, we provide a detailed survey of 16,034 cells within the AP and NTS of the mouse, in the fed and fasted state. Results: Of these, 8910 are neurons that group into 30 clusters, with 4289 coming from mice fed ad libitum and 4621 from overnight fasted mice. 7124 nuclei are from non-neuronal cells, including oligodendrocytes, astrocytes and microglia. Interestingly, we identified that the oligodendrocyte population was particularly transcriptionally sensitive to an overnight fast. The receptors GLP1R, GIPR, GFRAL and CALCR, which bind GLP1, GIP, GDF15 and amylin respectively, are all expressed in the hindbrain and are major targets for anti-obesity therapeutics. We characterise the transcriptomes of these four populations and show that their gene expression profiles are not dramatically altered by an overnight fast. Notably, we find that roughly half of cells that express GIPR are oligodendrocytes. Additionally, we profile POMC expressing neurons within the hindbrain and demonstrate that 84% of POMC neurons express either PCSK1, PSCK2 or both, implying that melanocortin peptides are likely produced by these neurons. Conclusion: We provide a detailed single-cell level characterisation of AP and NTS cells expressing receptors for key anti-obesity drugs that are either already approved for human use or are in clinical trials. This resource will help delineate the mechanisms underlying the effectiveness of these compounds, and also prove useful in the continued search for other novel therapeutic targets. Single-nucleus RNA sequencing data from 6 mice fed ad libitum and 6 mice fasted overnight, aged 6-8 weeks.

研究目的：最后区（area postrema, AP）与孤束核（nucleus tractus solitaris, NTS）位于后脑，是感知并整合外周营养信号、进而调控摄食行为的关键核团。目前已有研究通过单细胞转录组学（single-cell transcriptomics）在小鼠中揭示了关键下丘脑核团群体的基因表达谱与细胞异质性，但针对后脑的类似深入研究尚未开展。
研究方法：本研究采用单细胞核RNA测序技术，对摄食与禁食状态下小鼠后脑内AP与NTS的16034个细胞进行了全面解析。
研究结果：其中8910个为神经元，可分为30个细胞聚类，其中4289个来自自由进食小鼠，4621个来自整夜禁食小鼠；剩余7124个细胞核来自非神经元细胞，包括少突胶质细胞、星形胶质细胞与小胶质细胞。值得注意的是，本研究发现少突胶质细胞群体对整夜禁食的转录应答尤为显著。分别结合胰高血糖素样肽-1（GLP1）、葡萄糖依赖性促胰岛素多肽（GIP）、生长分化因子15（GDF15）与胰岛淀粉样多肽（amylin）的受体GLP1R、GIPR、GFRAL及CALCR均在后脑表达，且是抗肥胖治疗的主要靶点。本研究对这四类受体阳性细胞群体的转录组进行了表征，结果显示整夜禁食并未显著改变它们的基因表达谱；同时发现约有一半表达GIPR的细胞为少突胶质细胞。此外，本研究对后脑内表达阿黑皮素原（POMC）的神经元进行了转录组分析，结果显示84%的POMC神经元表达PCSK1、PCSK2或两者兼具，提示这些神经元很可能产生促黑素皮质素肽。
研究结论：本研究对表达关键抗肥胖药物靶点受体的AP与NTS细胞进行了单细胞水平的详细表征，这些药物要么已获批用于人类临床，要么正处于临床试验阶段。本数据集将有助于阐明这些化合物发挥疗效的潜在机制，同时也可为后续探索其他新型治疗靶点提供重要支撑。本研究的单细胞核RNA测序数据来源于6只自由进食小鼠与6只整夜禁食的6~8周龄小鼠。