ABSTRACT
Human organoids recapitulating the cell-type diversity and function of their target organ are valuable for basic and translational research. We developed light-sensitive human retinal organoids with multiple nuclear and synaptic layers and functional synapses. We sequenced the RNA of 285,441 single cells from these organoids at seven developmental time points and from the periphery, fovea, pigment epithelium and choroid of light-responsive adult human retinas, and performed histochemistry. Cell types in organoids matured in vitro to a stable "developed" state at a rate similar to human retina development in vivo. Transcriptomes of organoid cell types converged toward the transcriptomes of adult peripheral retinal cell types. Expression of disease-associated genes was cell-type-specific in adult retina, and cell-type specificity was retained in organoids. We implicate unexpected cell types in diseases such as macular degeneration. This resource identifies cellular targets for studying disease mechanisms in organoids and for targeted repair in human retinas.
Subject(s)
Cell Differentiation/genetics , Organoids/cytology , Organoids/metabolism , Retina/cytology , Retina/metabolism , Single-Cell Analysis/methods , Synapses/physiology , Transcriptome/genetics , Cell Culture Techniques/methods , Cell Line , Electrophysiology , Female , Gene Expression Regulation, Developmental/genetics , Genetic Predisposition to Disease/genetics , Humans , In Situ Hybridization , Induced Pluripotent Stem Cells/cytology , Induced Pluripotent Stem Cells/metabolism , Microscopy, Electron , Multigene Family , Naphthoquinones , Organoids/radiation effects , Organoids/ultrastructure , Retina/pathology , Retina/radiation effectsABSTRACT
Targeting genes to specific neuronal or glial cell types is valuable for both understanding and repairing brain circuits. Adeno-associated viruses (AAVs) are frequently used for gene delivery, but targeting expression to specific cell types is an unsolved problem. We created a library of 230 AAVs, each with a different synthetic promoter designed using four independent strategies. We show that a number of these AAVs specifically target expression to neuronal and glial cell types in the mouse and non-human primate retina in vivo and in the human retina in vitro. We demonstrate applications for recording and stimulation, as well as the intersectional and combinatorial labeling of cell types. These resources and approaches allow economic, fast and efficient cell-type targeting in a variety of species, both for fundamental science and for gene therapy.
