Three-dimensional (3D) in vitro models have become increasingly popular as systems to study cell-cell and cell-ECM interactions dependent on the spatial, mechanical, and chemical cues within the environment of the tissue, which is limited in traditional two-dimensional (2D) models. Although electrophysiological recordings of neuronal action potentials through 2D microelectrode arrays (MEAs) are a common and trusted method of evaluating neuronal function, network communication, and response to chemicals and biologicals, there are currently limited options for measuring electrophysiological activity from many locations simultaneously throughout a 3D network of neurons in vitro. Here, we have developed a thin-film, 3D flexible microelectrode array (3DMEA) that non-invasively interrogates a 3D culture of neurons and can accommodate 256 channels of recording or stimulation. Importantly, the 3DMEA is straightforward to fabricate and integrates with standard commercially available electrophysiology hardware. Polyimide probe arrays were microfabricated on glass substrates and mechanically actuated to collectively lift the arrays into a vertical position, relying solely on plastic deformation of their base hinge regions to maintain vertical alignment. Human induced pluripotent stem cell (hiPSC)-derived neurons and astrocytes were entrapped in a collagen-based hydrogel and seeded onto the 3DMEA, enabling growth of suspended cells in the matrix and the formation and maturation of a neural network around the 3DMEA probes. The 3DMEA supported the growth of functional neurons in 3D with action potential spike and burst activity recorded over 45 days in vitro. This platform is an important step in facilitating noninvasive electrophysiological characterization of 3D networks of electroactive cells in vitro.

中文翻译：

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用于体外脑模型的灵活的3维微电极阵列。

三维（3D）体外模型已成为越来越受欢迎的系统，用于研究取决于组织环境中空间，机械和化学线索的细胞-细胞和细胞-ECM相互作用，而这在传统二维环境中是有限的（2D）模型。尽管通过2D微电极阵列（MEA）记录神经元动作电位的电生理学记录是评估神经元功能，网络通讯以及对化学物质和生物学反应的普遍且值得信赖的方法，但目前在整个过程中同时从多个位置测量电生理学活动的选择有限。体外神经元3D网络。在这里，我们开发了一个薄膜，3D柔性微电极阵列（3DMEA），可以无创地询问神经元的3D文化，并可以容纳256个记录或刺激通道。重要的是，3DMEA易于制造并与标准的市售电生理硬件集成。聚酰亚胺探针阵列在玻璃基板上进行微加工，然后机械驱动以将阵列共同提升到垂直位置，这完全依赖于其基础铰链区的塑性变形来维持垂直对齐。将人类诱导的多能干细胞（hiPSC）衍生的神经元和星形胶质细胞包埋在基于胶原的水凝胶中，并接种到3DMEA上，从而使悬浮细胞能够在基质中生长，并使3DMEA探针周围的神经网络形成和成熟。3DMEA支持3D中功能神经元的生长，并在体外45天记录了动作电位峰值和爆发活性。该平台是促进体外电活性细胞的3D网络无创电生理表征的重要步骤。