Chunk #15 — Materials and methods — Differentiation of hESCs into cortical interneurons and analysis of ethanol-induced miRNA and mRNA transcriptomic changes by RNA-seq
hESC-derived cortical interneurons were used as cellular models for analyzing ethanol-induced miRNA and mRNA transcriptomic changes. H1 hESCs (WiCell Research Institute, Madison, USA) were differentiated into cortical interneurons as previously described [32]. Briefly, H1 hESCs were dissociated with accutase (STEMCELL Technologies, Vancouver, Canada) and cultured in mTeSR1 (STEMCELL Technologies, Vancouver, Canada) on Matrigel (Corning Life Science, Tewksbury, USA) coated plate until 95% confluence. For neural induction (from Day 1 to Day 10), hESCs were cultured in the neural induction medium containing three inhibitors including 100 nM of LDN-193189 (Stemgent, Cambridge, MA, USA), 10 μM of SB-431542 (Tocris Bioscience, Bristol, UK), and 2 μM of XAV-939 (Stemgent, Cambridge, MA, USA), and the neural induction medium was changed daily. For ventral patterning (Day 11–Day 18), the cells were cultured in neural induction media containing 100 ng/ml of SHH (R&D Systems, Minneapolis, MN, USA) and 1 μM of purmorphamine (Stemgent, Cambridge, MA, USA), and the medium was changed every other day. For final neuronal differentiation and maturation (Day 19 and after), the cells were cultured in the neuronal maturation medium supplemented with 20
