Identification of AgRP cells in the murine hindbrain that drive feeding [scRNA-seq]

Objective: The central melanocortin system is essential for the regulation of food intake and body weight. Agouti-related protein (AgRP) is the sole orexigenic component of the central melanocortin system and is conserved across mammalian species. AgRP is currently known to be expressed exclusively in the mediobasal hypothalamus, and hypothalamic AgRP-expressing neurons are essential for feeding. Here we characterized a previously unknown population of AgRP cells in the mouse hindbrain. Methods: Expression of AgRP in the hindbrain was investigated using gene expression analysis, single-cell RNA sequencing, immunofluorescent analysis and multiple transgenic mice with reporter expressions. Activation of AgRP neurons was achieved by Designer Receptors Exclusively Activated by Designer Drugs (DREADD) and by transcranial focal photo-stimulation using a step-function opsin with ultra-high light sensitivity (SOUL). Results: AgRP expressing cells were present in the area postrema (AP) and the adjacent subpostrema area (SubP) and commissural nucleus of the solitary tract (cNTS) of the mouse hindbrain (termed AgRPHind herein). AgRPHind cells consisted of locally projecting neurons as well as tanycyte-like cells. Food deprivation stimulated hindbrain Agrp expression as well as neuronal activity of subsets of AgRPHind cells. In adult mice that lacked hypothalamic AgRP neurons, chemogenetic activation of AgRP neurons resulted in hyperphagia and weight gain. In addition, transcranial focal photo-stimulation of hindbrain AgRP cells increased food intake in adult mice with or without hypothalamic AgRP neurons. Conclusions: Our study indicates that the central melanocortin system in the hindbrain possesses an orexigenic component, and that AgRPHind neurons stimulate feeding independently of hypothalamic AgRP neurons. Single cell RNA Sequencing of murine Area Postrema/NTS region研究目的：中枢黑皮质素系统（central melanocortin system）对食物摄入与体重稳态调控至关重要。刺豚鼠相关蛋白（Agouti-related protein, AgRP）是中枢黑皮质素系统中唯一的促食性效应成分，且在所有哺乳动物物种中均高度保守。目前已知AgRP仅表达于正中基底部下丘脑，而下丘脑表达AgRP的神经元对摄食行为不可或缺。本研究对小鼠后脑内此前未被发现的AgRP细胞群进行了系统表征。 研究方法：本研究通过基因表达分析、单细胞RNA测序（single-cell RNA sequencing）、免疫荧光分析，以及多株携带报告基因表达的转基因小鼠模型，对AgRP在小鼠后脑内的表达分布进行了探究。AgRP神经元的激活可通过设计受体仅由设计药物（Designer Receptors Exclusively Activated by Designer Drugs, DREADD）技术，以及搭载超高光敏感度阶梯功能视蛋白（step-function opsin with ultra-high light sensitivity, SOUL）的经颅局部光刺激手段实现。 研究结果：表达AgRP的细胞存在于小鼠后脑的最后区（area postrema, AP）、相邻的亚最后区（subpostrema area, SubP）以及孤束连合核（commissural nucleus of the solitary tract, cNTS）中，本文将此类细胞命名为AgRPHind。AgRPHind细胞包含局部投射神经元与室管膜细胞样细胞两类。食物剥夺可上调后脑Agrp的基因表达，同时激活部分AgRPHind细胞的神经元活性。在缺失下丘脑AgRP神经元的成年小鼠中，化学遗传学激活AgRP神经元可引发过度摄食与体重增加。此外，无论成年小鼠是否缺失下丘脑AgRP神经元，对其后脑AgRP细胞进行经颅局部光刺激均可显著提升其食物摄入量。 研究结论：本研究证实，后脑内的中枢黑皮质素系统同样具备促食性效应成分，且AgRPHind神经元可独立于下丘脑AgRP神经元发挥促摄食作用。 小鼠最后区/孤束核区域单细胞RNA测序数据集