Single-cell transcriptomics reveals multiple neuronal cell types in human midbrain-specific organoids

Abstract

Human stem cell-derived organoids have great potential for modelling physiological and pathological processes. They recapitulate in vitro the organization and function of a respective organ or part of an organ. Human midbrain organoids (hMOs) have been described to contain midbrain-specific dopaminergic neurons that release the neurotransmitter dopamine. However, the human midbrain contains also additional neuronal cell types, which are functionally interacting with each other. Here, we analysed hMOs at high-resolution by means of single-cell RNA sequencing (scRNA-seq), imaging and electrophysiology to unravel cell heterogeneity. Our findings demonstrate that hMOs show essential neuronal functional properties as spontaneous electrophysiological activity of different neuronal subtypes, including dopaminergic, GABAergic, glutamatergic and serotonergic neurons. Recapitulating these in vivo features makes hMOs an excellent tool for in vitro disease phenotyping and drug discovery.

Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids

“…In this study, we patterned hPSC-derived brain organoids into a ventral midbrain (VM) identity using a protocol that results in the formation of midbrain dopamine (DA) progenitors and of functionally mature DA neurons after transplantation in xenograft models of Parkinson's disease (PD) 14 . DA neurons in these organoids exhibited mature electrophysiological properties, neuromelanin production, and the ability to release DA, confirming the long-term maintenance of functionally mature human DA neurons in 3D culture as previously reported in both hPSCderived [15][16][17][18] and neural progenitor-derived [19][20][21] midbrain organoids. A time-course transcriptional analysis of human VM development and DA neuron differentiation at single-cell level revealed four populations of cells with high transcriptional similarity to VM floor-plate cells, followed by the stepwise emergence of neurons, vascular leptomeningeal cells (VLMCs), astrocytes, and oligodendrocytes.…”

Midbrain organoids with anSNCAgene triplication model key features of synucleinopathy

“…As hMOs are composed of multiple cell types, 23 , 26 , 27 , 58 they offer a unique model to explore the contribution of different cell populations on the ability of α-syn to aggregate. Our earlier findings with PLA and imaging of hMO cryosections showed aggregates were detected in both neurons and glia, albeit to varying degrees.…”

“… 22 Recent protocols to generate human midbrain organoids (hMOs) from iPSCs have made it possible to generate 3D human tissue in vitro that more closely resembles the native environment and cellular diversity, including DNs, found in vivo in the SN. 23 Neurons within hMOs form interconnected networks, are organized in multiple layers, and exhibit a number of functional properties normally displayed by neurons, that include synapse formation and spontaneous electrophysiological activity. 24 Thus, hMOs provides a human model in a dish with the potential to capture the 3D architecture, cellular diversity and connectivity found in the midbrain in vivo .…”

Modeling Rett Syndrome with Human Pluripotent Stem Cells: Mechanistic Outcomes and Future Clinical Perspectives

“…Benefiting from the knowledge of neurodevelopmental biology principles, directed differentiation processes using the appropriate growth factors and small molecules have been able to induce the generation of region-specific organoids, decreasing inter-organoid variability [ 125 ]. Several methodologies have been able to generate organoids resembling, for example, the cerebral cortex [ 127 , 128 ], the ventral forebrain [ 91 , 129 ], the cerebellum [ 130 , 131 , 132 ], the midbrain with functional dopaminergic neurons [ 133 , 134 ] and the spinal cord [ 135 ]. Moreover, the fusion of different regional organoids [ 91 , 136 ] can be used to model the kinetic processes of cell migration, cell–cell interactions and neural circuit formation.…”