RESUMEN
We report the generation and analysis of single-cell RNA-Seq data (> 38,000 cells) from mouse native retinae and induced pluripotent stem cell (iPSC)-derived retinal organoids at four matched stages of development spanning the emergence of the major retinal cell types. We combine information from temporal sampling, visualization of 3D UMAP manifolds, pseudo-time and RNA velocity analyses, to show that iPSC-derived 3D retinal organoids broadly recapitulate the native developmental trajectories. However, we observe relaxation of spatial and temporal transcriptome control, premature emergence and dominance of photoreceptor precursor cells, and susceptibility of dynamically regulated pathways and transcription factors to culture conditions in retinal organoids. We demonstrate that genes causing human retinopathies are enriched in cell-type specifying genes and identify a subset of disease-causing genes with expression profiles that are highly conserved between human retinae and murine retinal organoids. This study provides a resource to the community that will be useful to assess and further improve protocols for ex vivo recapitulation and study of retinal development.
Asunto(s)
Células Madre Pluripotentes Inducidas , Ratones , Humanos , Animales , Transcriptoma , Retina/metabolismo , Células Fotorreceptoras , Organoides/metabolismo , Análisis de Secuencia de ARN , Diferenciación Celular/genéticaRESUMEN
In the forebrain, ventrally derived oligodendrocyte precursor cells (vOPCs) travel tangentially toward the cortex together with cortical interneurons. Here, we tested in the mouse whether these populations interact during embryogenesis while migrating. By coupling histological analysis of genetic models with live imaging, we show that although they are both attracted by the chemokine Cxcl12, vOPCs and cortical interneurons occupy mutually exclusive forebrain territories enriched in this chemokine. Moreover, first-wave vOPC depletion selectively disrupts the migration and distribution of cortical interneurons. At the cellular level, we found that by promoting unidirectional contact repulsion, first-wave vOPCs steered the migration of cortical interneurons away from the blood vessels to which they were both attracted, thereby allowing interneurons to reach their proper cortical territories.
Asunto(s)
Movimiento Celular , Corteza Cerebral , Interneuronas , Neurogénesis , Células Precursoras de Oligodendrocitos , Animales , Movimiento Celular/genética , Corteza Cerebral/citología , Corteza Cerebral/embriología , Quimiocina CXCL12/metabolismo , Interneuronas/fisiología , Ratones , Modelos Genéticos , Neurogénesis/genética , Células Precursoras de Oligodendrocitos/citología , Células Precursoras de Oligodendrocitos/fisiologíaRESUMEN
Interneuron migration involves repetitive cycles of pausing and motion that include nucleokinesis and dynamic branching of the leading process. Here, we provide a step-by-step description of how to culture and record the migration of cortical interneurons. We provide two culture models: the first includes organotypic brain slices and the second medial ganglionic eminence (MGE) explants. While organotypic brain slices provide a close-to-physiological context to analyze interneuron migration into cortical streams, MGE explants are appropriate to investigate the fine details of interneuron morphology remodeling during movement. For complete details on the use and execution of this protocol, please refer to Silva et al. (2018).
