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Journal: bioRxiv
Article Title: Human neuromodulatory assembloids to study serotonin signaling and disease
doi: 10.64898/2026.03.08.710407
Figure Lengend Snippet: (A) Canonical markers of 5-HT neurons in single-cell RNA-seq data of hMHO. (B) Proportions of brain regions mapping to 5-HT neurons based on label transfer of PCW 6.9 primary developing human brain. (C) Tph2 expression in mouse E13 spatial atlas and projection of 5-HT neurons onto mouse reference using VoxHunt . (D) Immunohistochemical staining of TPH2 in hMHO (day 84, scale bar = 250 µ m (top) and 40 µ m (bottom)). (E) Immunohistochemical staining of 5-HT in hMHO (day 83). (F) HPLC measurements of 5-HT in hMHO and hCO (hMHO: 20 samples, 13 differentiation experiments, hCO: 9 samples, 4 differentiation experiments; 3 hiPS cell lines, day 91-183. Mann-Whitney test, ****p<0.0001). (G) Biocytin labeling of AAV-TPH2::EGFP labeled 5-HT neurons (day 157, one iPS cell line, one differentiation experiment). (H) Schematic of patch clamp recording of 5-HT neurons (TPH2::EGFP + cells) in hMHO at baseline and during addition of 5-HT and NAN-190. (I) Example trace of action potentials during addition of 5-HT (100 µ m) and NAN-190 (50 µ m, 5-HT1A receptor blocker). (J) Action potential frequencies of TPH2::EGFP + neurons during baseline and addition of 5-HT and NAN-190 (14 cells, 1 hiPS cell line, 3 differentiation experiments. Friedman test *p = 0.0124, with Dunn’s multiple comparison testing: baseline vs. 5-HT **p = 0.0092, baseline vs. NAN-190 p = 0.7902, ns: non-significant). (K) Structure of serotonin sensor PsychLight2 (adapted from Dong et al with permission from Elsevier publishing, License #: 6152850897994) and schematic of serotonin sensor imaging. (L) Example images of PsychLight2 imaging in hCO and hMHO during KCl stimulation at baseline (10s) and during KCl stimulation (60s) (day 172, scale bar = 200 µ m). (M) Quantification of temporal dynamics of serotonin sensor fluorescence changes in hMHO and hCO (hCO: 9 organoids, hMHO: 8 organoids, 2 hiPS cell lines, 3 differentiation experiments). (N) Change in fluorescence intensity for PsychLight2 in hCO and hMHO after 30 s of KCl stimulation across different iPS cell lines (hCO: n = 135 ROIs (left), 9 organoids (right), hMHO: 120 ROIs (left), 8 organoids (right), 2 hiPS cell lines, 3 differentiation experiments for hCO and hMHO. Mann-Whitney test, ****p<0.0001).
Article Snippet: To study the effect of 5-HT on evoked APs in hMHO, 100 μ m 5-HT (Sigma Aldrich, H9523, freshly prepared on day of recording) or 50 μ m
Techniques: Single Cell, RNA Sequencing, Expressing, Immunohistochemical staining, Staining, MANN-WHITNEY, Labeling, Patch Clamp, Comparison, Imaging, Fluorescence