We thus examined the impact of microglial cells with high and low AUD-PRS on human iPSC-derived neurons. We individually cocultured each line of microglial cells (six lines of high AUD PRS and seven lines of low AUD PRS; see table S1 for detail) with induced human iNs. The iNs were induced from an iPSC line with no diagnosed AUD, as used in a previous study (58). Thereby, we were able to evaluate the differential effects between high-PRS and low-PRS microglia on a common set of “neutral” neurons. To facilitate synapse formation, these iNs were cultured on monolayer mouse glial cells (59, 60). High-PRS or low-PRS microglial cells were added to iN cultures on day 14 after induction. The final microglial density in culture (days 35 to 40 after induction) is around 7 to 9% of total cell numbers, including mouse glial cells (Fig. 8, A and B), which is consistent with the microglia proportion in the human brain (61). We first analyzed synapse formation after a 7-day intermittent ethanol exposure with 75 mM ethanol. After IHC staining with the presynaptic
