下丘脑是大脑的一个区域，对调节身体功能和行为起着重要作用。人类多能干细胞(hPSCs)在模拟影响下丘脑的疾病方面越来越受关注。本研究建立了一种hPSC派生的下丘脑器官样体分化方案，以模拟该脑区的细胞多样性。我们使用一种带有酪氨酸羟化酶(TH)-TdTomato报道基因以标记多巴胺能神经元(DNs)和其他表达TH的细胞的hPSC株系，对电生理活性的下丘脑器官样体中DN特定的途径和功能进行了研究。单细胞RNA测序(scRNA-seq)揭示了成熟下丘脑器官样体中多样的神经元和非神经元细胞类型。我们确定了几种分子特异性不同的下丘脑DN亚型，它们具有不同的发育成熟度。我们的体外三维下丘脑分化方案可用于研究这一关键脑结构的发育，并可应用于疾病模型，为围绕下丘脑的疾病提供新的治疗方法。© 2023 The Authors.

The hypothalamus is a region of the brain that plays an important role in regulating body functions and behaviors. There is a growing interest in human pluripotent stem cells (hPSCs) for modeling diseases that affect the hypothalamus. Here, we established an hPSC-derived hypothalamus organoid differentiation protocol to model the cellular diversity of this brain region. Using an hPSC line with a tyrosine hydroxylase (TH)-TdTomato reporter for dopaminergic neurons (DNs) and other TH-expressing cells, we interrogated DN-specific pathways and functions in electrophysiologically active hypothalamus organoids. Single-cell RNA sequencing (scRNA-seq) revealed diverse neuronal and non-neuronal cell types in mature hypothalamus organoids. We identified several molecularly distinct hypothalamic DN subtypes that demonstrated different developmental maturities. Our in vitro 3D hypothalamus differentiation protocol can be used to study the development of this critical brain structure and can be applied to disease modeling to generate novel therapeutic approaches for disorders centered around the hypothalamus.© 2023 The Authors.