Fig. 4: Cellular landscape of fluoxetine action in dorDG and venDG.

a Rank-rank hypergeometric overlap (RRHO) maps show the threshold-free differential expression comparison between dorsal and ventral DG in this study. Pixels represent the overlap between the transcriptome of each comparison, with the significance of overlap (–log₁₀ (P-val) of a hypergeometric test) colour coded. Genes along each axis are sorted from most significantly up-regulated (lower left) to most down-regulated (top right). Venn diagram shows overlap of significant DEGs in dorDG and venDG. b Heatmap showing 12 cell types obtained by clustering 3,517 single cells from 5 replicates per treatment group. Two-step clustering using supervised RCA2 clustering, followed by Seurat unsupervised clustering was performed. Top 10 markers specific for each cluster are plotted. c Cell types with significant changes in proportion between Sham and FT groups in dorDG or venDG (both regions shown to highlight region-specific differences). *: Wilcoxon P-val ≤ 0.05. d Treatment-specific DEGs (FT vs Sham) for dorDG and venDG cell types. Single cells from each cell type were aggregated by replicate into an averaged pseudo-bulk expression profile. Pseudo-bulk expression profiles were then used to calculate DEGs (absolute log₂FC ≥ log₂(1.25), FDR Q-val < 0.2). e Top GO terms following gene set enrichment analysis of cell-type-specific pseudo-bulk DEGs (FT vs Sham). f Module-score analysis for the oxidative phosphorylation gene set in dorDG and venDG cell types. Cell types with significant FT vs Sham module score are plotted. **** indicates: FDR Q-val < 3e-05, ***: FDR Q-val < 5e-03, **: FDR Q-val < 0.05. g Left, representative images of CRL-2199 glial line treated with vehicle control, corticosterone (CORT), fluoxetine (FT) and CORT + FT, stained with DAPI (blue) and MitoTracker Red (red) and immunostained against cytochrome c (green). Scale bars, 20 μm. Right, quantification of average MitoTracker Red (top) and cytochrome c (bottom) fluorescence signals normalised to vehicle control (n = ~200 cells per group using three independent experiments; values for individual cells are shown as open circles; error bars denote SEM). SEM, standard error of the mean. h Oxygen consumption rate (OCR) measurements normalised to cell number measured by Hoechst 33342 fluorescence signal (mean of 3 biological replicates ±SD, technical replicates =34). SD, standard deviation.