Figure 1.

Automated midbrain organoids enable toxicity screening in human tissue-like aggregates. (A) Tyrosine hydroxylase (TH) and MAP2 expression declined with exposure to the toxic compound rotenone (bottom) in comparison to control AMOs of the same age (top, both at day 56 of differentiation). The AMO on the bottom was treated with 100 μM rotenone for 48 h starting on day 48 of differentiation and afterward kept under standard culture conditions for six additional days before fixation and whole-mount staining (see “Materials and Methods” section for more details). Representative scanning confocal images of a single optical slice of whole-mount stained and cleared AMOs labeled for the general neuron marker Map2 (green) and dopaminergic neuron marker TH (red). Nuclei were counterstained with DAPI (blue). The inlays in the bottom right corner of each image display enlargements of the sections marked by the dotted squares in the respective composite images. Scale bar = 100 μm (entire AMO), 20 μm (inlays). (B) Schematic overview of the high throughput screening workflow for the evaluation of overall and cell type-specific toxicity.