Fig. 8

Glial distribution and morphology over time. GFAP + astrocytes (green), Iba1 + microglia (pink), and MAP2 + neurons (cyan) are present in culture at 3 DIV, 5 DIV, 10 DIV, and 12 DIV. At 3 DIV, large clusters of individual astrocytes are seen. Astrocytes are relatively simplistic in shape mostly with a branch extending on each end on the nucleus. Microglia at 3 DIV have different degrees of branching. At 5 DIV, cells begin to have > 2 branches while large clusters of individual astrocytes remain. Simplistic astrocytes are seen along with more mature astrocytes with thicker, more fibrous cell bodies with multiple branches. Microglia at both 3 DIV and 5 DIV present with various degrees of branching. At 10 DIV, classical astrocyte morphology is seen, and cells are evenly distributed over the growth area instead of in clusters. Astrocytes appear thick with projections of various sizes while microglia are less numerous and rounder. At 12 DIV, cells begin to grow to confluency, yet the number of cells is lower as each cell is larger and takes up more space than the cells in 3 DIV and 5 DIV. Astrocytes have a mix of thick and fibrous and thin stars-like shapes but none of the earlier more simplistic shapes. Microglia only appear in round shapes. All visible nuclei are DAPI+ (blue). Scale bars in all images in left column: 1000 μm. Scale bars in all images in right column: 50 μm