Division of Pediatric Neurology, Department of Pediatrics, Michigan Medicine, Ann Arbor; Michigan Neuroscience Institute, University of Michigan, Ann Arbor.
Division of Pediatric Neurology, Department of Pediatrics, Michigan Medicine, Ann Arbor; Michigan Neuroscience Institute, University of Michigan, Ann Arbor;
J Vis Exp. 2024 Sep 27(211). doi: 10.3791/67396.
Human brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that recapitulate aspects of fetal brain development. The fusion of dorsal with ventral regionally specified brain organoids in vitro generates assembloids, which have functionally integrated microcircuits with excitatory and inhibitory neurons. Due to their structural complexity and diverse population of neurons, assembloids have become a useful in vitro tool for studying aberrant network activity. Multi-electrode array (MEA) recordings serve as a method for capturing electrical field potentials, spikes, and longitudinal network dynamics from a population of neurons without compromising cell membrane integrity. However, adhering assembloids onto the electrodes for long-term recordings can be challenging due to their large size and limited contact surface area with the electrodes. Here, we demonstrate a method to plate assembloids onto MEA plates for recording electrophysiological activity over a 2-month span. Although the current protocol utilizes human cortical organoids, it can be broadly adapted to organoids differentiated to model other brain regions. This protocol establishes a robust, longitudinal, electrophysiological assay for studying the development of a neuronal network, and this platform has the potential to be used in drug screening for therapeutic development in epilepsy.
人类脑类器官是源自人类多能干细胞（hPSC）的三维（3D）结构，可再现胎儿大脑发育的某些方面。在体外将背侧与腹侧区域特化的脑类器官融合会产生 assembloids，其具有功能上整合的兴奋性和抑制性神经元的微电路。由于其结构复杂性和神经元的多样性，assembloids 已成为研究异常网络活动的有用体外工具。多电极阵列（MEA）记录可作为一种从神经元群体中捕获电场电位、尖峰和纵向网络动力学的方法，而不会损害细胞膜完整性。然而，由于其体积大且与电极的接触表面积有限，将 assembloids 粘附到电极上进行长期记录可能具有挑战性。在这里，我们展示了一种将 assembloids 接种到 MEA 板上以记录电生理活性的方法，时间跨度为 2 个月。尽管当前方案利用了人类皮质类器官，但它可以广泛适用于分化为模拟其他脑区的类器官。该方案建立了一种稳健的、纵向的电生理测定方法，用于研究神经网络的发育，并且该平台有可能用于癫痫症治疗开发的药物筛选。
