RESUMEN
GABAergic interneurons are essential for neural circuit function, and their loss or dysfunction is implicated in human neuropsychiatric disease. In vitro methods for interneuron generation hold promise for studying human cellular and functional properties and, ultimately, for therapeutic cell replacement. Here we describe a protocol for generating cortical interneurons from hESCs and analyze the properties and maturation time course of cell types using single-cell RNA-seq. We find that the cell types produced mimic in vivo temporal patterns of neuron and glial production, with immature progenitors and neurons observed early and mature cortical neurons and glial cell types produced late. By comparing the transcriptomes of immature interneurons to those of more mature neurons, we identified genes important for human interneuron differentiation. Many of these genes were previously implicated in neurodevelopmental and neuropsychiatric disorders.
Asunto(s)
Diferenciación Celular/fisiología , Movimiento Celular/fisiología , Neuronas GABAérgicas/citología , Interneuronas/citología , Proteínas del Tejido Nervioso/metabolismo , Neuroglía/citología , Células Cultivadas , Humanos , Neurogénesis/fisiología , Análisis de la Célula Individual , Factores de Transcripción/metabolismoRESUMEN
During human brain development, multiple signaling pathways generate diverse cell types with varied regional identities. Here, we integrate single-cell RNA sequencing and clonal analyses to reveal lineage trees and molecular signals underlying early forebrain and mid/hindbrain cell differentiation from human embryonic stem cells (hESCs). Clustering single-cell transcriptomic data identified 41 distinct populations of progenitor, neuronal, and non-neural cells across our differentiation time course. Comparisons with primary mouse and human gene expression data demonstrated rostral and caudal progenitor and neuronal identities from early brain development. Bayesian analyses inferred a unified cell-type lineage tree that bifurcates between cortical and mid/hindbrain cell types. Two methods of clonal analyses confirmed these findings and further revealed the importance of Wnt/ß-catenin signaling in controlling this lineage decision. Together, these findings provide a rich transcriptome-based lineage map for studying human brain development and modeling developmental disorders.
