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. 2019 Nov 28;7:303. doi: 10.3389/fcell.2019.00303

FIGURE 5.

FIGURE 5

Calcium response to ATP and KCl stimulation of cerebral organoids. PSC-derived cerebral organoids grown for (A–C) 30 or (G–I) 45 days had their cells dissociated and plated for 10 days on 15 mm coverslips. Cultured cells shown in (A,G) bright field and (B,H) under fluorescence. (C,I) Selected cells are shown in the same microscope field under fura-2 fluorescence in SCCI experiments. Typical responses are shown for cells when stimulated with (D) 100 μM ATP or (E,J) 50 mM KCl. As shown in (C–E), only one cell (#74) responded exclusively to ATP (yellow arrow), whereas several cells (# 3, 31, 82, 83, 84) responded to KCl (pink arrows), in 30-day cerebral organoids. As shown in (I,J) selected cells (# 21, 36, 38, 40, and 41) were only responsive to 50 mM KCl (pink arrows) in 45-day cerebral organoids. Post-stimulation, coverslips were fixed and cells labeled for (F) nestin, (K) TUJ1/tubulin βIII (TUBB3), or MAP2 (L) scale bars 50 μm. Areas under the curve are representative of calcium responses to ATP or KCl obtained from 30-day (M) or 45-day organoids (N).