Chronic opioid treatment arrests neurodevelopment and alters synaptic activity in human midbrain organoids
Understanding the impact of long-term opioid exposure on the embryonic brain is critical due to the surging number of pregnant mothers with opioid dependency, but has been limited by human brain inaccessibility and cross-species differences in animal models. Here we establish a human midbrain model that uses hiPSC-derived midbrain organoids and apply it to assess cell-type-specific responses to acute and chronic fentanyl treatment and fentanyl withdrawal. Single-cell mRNA sequencing of 25,510 cells from organoids in different treatment groups indicated that chronic fentanyl treatment arrests neuronal subtype specification during early midbrain development and alters synaptic activity and neuron projection. In contrast, acute fentanyl treatment increases dopamine release but does not significantly alter gene expression related to cell lineage development. These results provide the first examination of the effects of opioid exposure on human midbrain development at the single cell level. Overall design: Midbrain organoids derived from human FA11 iPSC cells were cultured and later treated with or without fentanyl. The acute treatment with Fentanyl was administered at a concentration of 74nM for a duration of 4 hours , on day 53 (KH002_D53_AC_FTY). The chronic treatment with Fentanyl was administered at a concentration of 74nM for a duration of 77 days, from Day 1 to Day 77 (KH004_D77_FTY). Withdrawal sample was treated with Fentanyl (74nM for 77 days, Day 1 to Day 77) and then replaced with fresh media for 2 days (KH005_D79_WD).