Assessment of ethanol-induced toxicity on iPSC-derived human dopaminergic neurons using a novel high-throughput mitochondrial neuronal health (MNH) assay

作者：Annika Zink, J. Vanessa Conrad, Narasimha Swami Telugu, Sebastian Diecke, Andreas Heinz, Erich E. Wanker, Josef Priller, Alessandro Prigione · 发表于：bioRxiv (Cold Spring Harbor Laboratory) · 年份：2020 · DOI：10.1101/2020.08.12.237461 · 研究领域：Neuroscience and Neuropharmacology Research、Neurogenesis and neuroplasticity mechanisms、Neuroscience and Neural Engineering

Abstract Excessive ethanol exposure can cause mitochondrial and cellular toxicity. In order to discover potential counteracting interventions, it is essential to develop assays capable of capturing the consequences of ethanol exposure in human dopaminergic (DA) neurons, which are crucial for the development and maintenance of alcohol use disorders (AUD). Here, we developed a novel high-throughput (HT) assay to quantify mitochondrial and neuronal toxicity in human DA neurons from induced pluripotent stem cells (iPSCs). The assay, dubbed m itochondrial n euronal h ealth (MNH) assay, combines live-cell measurement of mitochondrial membrane potential (MMP) with quantification of neuronal branching complexity post-fixation. Using the MNH assay, we demonstrated that chronic ethanol exposure in human iPSC-derived DA neurons decreases MMP and branching complexity in a dose-dependent manner. The toxic effect of ethanol on DA neurons was already detectable after 1 hour of exposure, and occurred similarly in DA neurons derived from healthy individuals and from patients with AUD. We next used the MNH assay to carry out a proof-of-concept compound screening using FDA-approved drugs. We identified potential candidate drugs modulating acute ethanol toxicity in human DA neurons. Among these drugs, flavoxate and disulfiram influenced mitochondrial neuronal health independently from ethanol, leading to amelioration and worsening, respectively. Altogether, we developed an HT assay to probe huma...