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Abstract Multiphoton microscopy has emerged as the primary imaging tool for studying the structural and functional dynamics of neural circuits in brain tissue, which is highly scattering to light. Recently, three-photon microscopy has enabled high-resolution fluorescence imaging of neurons in deeper brain areas that lie beyond the reach of conventional two-photon microscopy, which is typically limited to ~ 450 µm. Three-photon imaging of neuronal calcium signals, through the genetically-encoded calcium indicator GCaMP6, has been used to successfully record neuronal activity in deeper neocortical layers and parts of the hippocampus in rodents. Bulk-loading cells in deeper cortical layers with synthetic calcium indicators could provide an alternative strategy for labelling that obviates dependence on viral tropism and promoter penetration, particularly in non-rodent species. Here we report a strategy for visualized injection of a calcium dye, Oregon Green BAPTA-1 AM (OGB-1 AM), at 500–600 µm below the surface of the mouse visual cortex in vivo. We demonstrate successful OGB-1 AM loading of cells in cortical layers 5–6 and subsequent three-photon imaging of orientation- and direction- selective visual responses from these cells.
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Multimodal monitoring of human cortical organoids implanted in mice reveal functional connection with visual cortex
Abstract Human cortical organoids, three-dimensional neuronal cultures, are emerging as powerful tools to study brain development and dysfunction. However, whether organoids can functionally connect to a sensory network in vivo has yet to be demonstrated. Here, we combine transparent microelectrode arrays and two-photon imaging for longitudinal, multimodal monitoring of human cortical organoids transplanted into the retrosplenial cortex of adult mice. Two-photon imaging shows vascularization of the transplanted organoid. Visual stimuli evoke electrophysiological responses in the organoid, matching the responses from the surrounding cortex. Increases in multi-unit activity (MUA) and gamma power and phase locking of stimulus-evoked MUA with slow oscillations indicate functional integration between the organoid and the host brain. Immunostaining confirms the presence of human-mouse synapses. Implantation of transparent microelectrodes with organoids serves as a versatile in vivo platform for comprehensive evaluation of the development, maturation, and functional integration of human neuronal networks within the mouse brain.
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- Award ID(s):
- 1752241
- PAR ID:
- 10395620
- Date Published:
- Journal Name:
- Nature Communications
- Volume:
- 13
- Issue:
- 1
- ISSN:
- 2041-1723
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1038/s41467-022-35536-3
