Fig. 2

Morphologically complex astrocytes interact with neurons. HiPSC-derived neurons and human primary astrocytes were co-cultured in 3D for 4 weeks, and astrocyte morphology and interaction with neurons was assessed by confocal microscopy. A 160 µm-thick maximum intensity confocal image projection of neurons and astrocytes in 3D co-culture. Immunostaining was performed for astrocytes (GFAP, magenta; CD44, green). Single channels are shown in greyscale. A zoom of the boxed area indicated in the merged image is shown. B 112 µm-thick maximum intensity confocal image projection of neurons and astrocytes in 3D co-culture. Immunostaining was performed for neurons (NeuroChrom, green) and astrocytes (GFAP, magenta). Single channels are shown in greyscale. A zoom of the boxed area indicated in the merged image is shown. See Supplementary Video 1 for a confocal z-stack. C 105 µm-thick maximum intensity confocal image projection of neurons and astrocytes in 3D co-culture. Immunostaining was performed for neurons (NeuroChrom, green) and astrocytes (GFAP, magenta). A merged image is shown, in which three numbered boxed areas are shown as zoomed areas. In the zoomed images, the arrows indicate astrocytes extending upon neuronal processes. D 78 µm-thick maximum intensity confocal image projection of axons, dendrites and astrocytes in a 3D co-culture. Immunostaining was performed for axons (SMI312, green), dendrites (MAP2, blue) and astrocytes (GFAP, magenta). Single channels are maximum intensity projections and shown in greyscale. The boxed area indicated in the MAP2 channel is shown as a merged image and the arrows indicate an astrocyte interacting with axons and dendrites