Petersilie et al. 2023 - Figure 7

Brain organoids derived from human pluripotent stem cells (hPSCs) are a promising tool for studying human neurodevelopment and related disorders. Here, we generated long-term cultures of cortical brain organoid slices (cBOS) grown at the air-liquid interphase from regionalized cortical organoids. We demonstrate that cBOS host mature neurons and astrocytes organized in complex architecture. By leveraging the fact that cBOS allow direct access to the developing neural cells, we carried out an array of functional analyses. Whole-cell patch-clamp demonstrated subthreshold synaptic inputs and action potential firing of neurons. Spontaneous intracellular calcium signals turned into synchronous large scale calcium oscillations upon combined disinhibition of NMDA receptors and blocking of GABAA receptors. Lastly, FRET-based imaging with the genetically-encoded nanosensor ATeam1.03YEMK revealed the high sensitivity of neurons to acute metabolic inhibition. Altogether, cBOS represent a powerful platform for assessing the morphological and functional aspects of organized human neural cells in intact minimal networks.

由人类多能干细胞（human pluripotent stem cells, hPSCs）诱导获得的脑类器官，是研究人类神经发育及相关疾病的极具潜力的实验工具。本研究从区域化皮层类器官出发，制备了可在气液界面（air-liquid interphase）进行长期培养的皮层脑类器官切片（cortical brain organoid slices, cBOS）。研究证实，cBOS中含有结构复杂的成熟神经元与星形胶质细胞。依托cBOS可直接获取发育中神经细胞的技术优势，我们开展了一系列功能分析实验：全细胞膜片钳记录显示，神经元可产生阈下突触输入与动作电位发放；在联合NMDA受体（NMDA receptors）去抑制与GABAA受体（GABAA receptors）阻断的条件下，自发细胞内钙信号可转化为同步大规模钙振荡；最后，借助基因编码纳米传感器ATeam1.03YEMK开展的荧光共振能量转移（Fluorescence Resonance Energy Transfer, FRET）成像实验显示，神经元对急性代谢抑制具有高敏感性。综上，cBOS可作为强大的研究平台，用于评估完整小型神经网络中人类有序神经细胞的形态与功能特征。