Single‐cell RNA sequencing (scRNAseq) was performed on groups of iPSC‐derived neurons that were pooled and cultured in the same dish to reduce culture variability. 60 Comparing induced neurons from individuals with each haplotype, scRNAseq identified 1393 genes that were differentially expressed between affected and unaffected individuals, 797 up‐regulated, and 596 down‐regulated. 39 Gene ontology (GO) analysis of the up‐regulated genes revealed changes in pathways associated with protein targeting within the cells, catabolic metabolism, and nonsense‐mediated decay. Analysis of the downregulated genes predicted differences in pathways associated with nervous system development, axonal transport, and trans‐synaptic signaling. Neurons from individuals with the alternative haplotype and AUD had significantly lower levels of KCNJ6 expression, and ethanol exposure reversed these effects. Since most sequence variants in LD with the significant KCNJ6 SNPs were in the 3′ UTR of the mRNA (21 out of 22 SNPs), it was hypothesized that differences in KCNJ6 expression were due to differential mRNA stability, translation, or other post transcriptional processes.
