(a) Decreased nuclear TBR1 (red) expression in the upper cortical zone of 5-week schizophrenia organoidsNuclei were stained with DAPI. Images show representative sections of control (iPSC line BJ1) and schizophrenia (iPSC line 2038) organoids. Total number of DAPI-stained nuclei and the number of nuclei expressing TBR1 were counted in multiple randomly selected ROI (5 × 103 μm2, ∼50 cells/ ROI) within the upper cortical layers (*6 cells deep) of three control individuals and three patients. Percent of (TBR1 + DAPI)/DAPI-stained nuclei was determined for each ROI. Graph shows distribution of the % of TBR1 expressing cells in the individual ROIs (26 control and 33 schizophrenia ROIs). The difference between control and schizophrenia mean values was significant (t-test). Individual value plots are shown in Supplementary Fig. 5b. b Decreased reelin expression in schizophrenia organoid cortex. Images show control (BJ1) and schizophrenia (1835) organoids. Note the lack of reelin staining in 2-week organoids. In 5-week organoids, reelin immunofluorescence intensity was determined in randomly selected ROIs (3 × 103 μm2) in the upper CZ (*) and in the IZ (**) regions of three control individuals and three patients using Zen 2.0 Blue Imaging software (22 control and 17 schizophrenia upper CZ ROIs and the same number of IZ ROIs). ANOVA of four groups followed by Tukey posthoc test showed a significant decrease in the reelin expression in the schizophrenia upper CZ and a lack of significant differences between control and schizophrenia in the IZ. Individual value plots are shown in Supplementary Fig. 5. c Morphology and orientation of cortical calretinin interneurons. c1—images of control and schizophrenia organoids. A total of 770 control and 547 schizophrenia calretinin interneurons were measured in 20 and 16 ROIs, respectively, in the organoids from three control and three schizophrenia patients. The average cell density (d = number of cells/ROI) was not significantly different between control and schizophrenia (Supplementary Fig. 3a, b). c2—graph shows cell distribution (cumulative frequency) relative to their total length, including the cell body and neurites. An average cell body had a length of ∼50 pixels, 18 μm. A two-sample Kolmogorov–Smirnov test of cumulative density function (CDF shown in the inset) of control and schizophrenia groups found no significant difference between the length of control and schizophrenia interneurons. c3—angles between the long axis of each cell and the cortical surface organoids were computed as described in the Supplementary Methods. Graph shows distribution of cells (cumulative frequency) in bins corresponding to the deviation angles from the cortical surface. A two-sample Kolmogorov–Smirnov CDF test (CDFs shown in the inset) of control and schizophrenia groups yielded a highly significant difference (p-value of <13.9 × 10−7) between the orientation of control and schizophrenia interneurons, relative to the cortical surface
