Fig. 5: Neuronal metabolic response to GSK3β inhibition with LiCl.

A Rank order plot of enriched pathways detected by GSEA of RNA-sequencing of neurons treated for 24 h with 15 mM LiCl (n = 4 replicates). Pathways are adjusted p-value determined by FDR. B, C Heatmap of the significantly changing genes in the oxidative phosphorylation (B) and ribosome (C) pathways enriched in LiCl-treated neurons. D Heatmap of PGC-1α responsive genes detected in the LiCl-treated neuron RNA-seq data. E Oxygen consumption of primary neurons treated with LiCl or a control media change measured by RESIPHER monitor for 24 h (n = 6 replicates). F Mitochondrial membrane potential assessed by JC-1 assay in primary neurons treated with LiCl or GSK3β inhibitor (Inhibitor VIII) for 24 h. G Representative images and quantification of mitochondria detected with TOMM20 antibody after 24 h treatment with LiCl or Inhibitor VIII. Mitochondrial size was determined through particle size detection in ImageJ (n = 25 control, n = 26 LiCl, n = 22 inhibitor VIII). H Representative images (left) and distributions (right) of NAD(P)H mean fluorescence lifetime of control or LiCl-treated primary neurons. I Representative images (left) and quantitation (right) of NAD(P)H fluorescence intensity of control or LiCl-treated primary neurons (n = 32 control neurons, n = 28 LiCl neurons). J NAD/NADH and NADP/NADPH ratios determined by biochemical assay (NAD/NADH: n = 18 control, n = 11 LiCl; NADP/NADPH: n = 16 control, n = 24 LiCl). Data shown with error bars noting mean ± SEM (F, G, I, J). Asterisks indicate p-value < 0.05 by multiple two-tailed t-tests (E), two-tailed unpaired student’s t-test (F, I, J), or Brown-Forsythe and Welch ANOVA with Dunnett’s test (G). See Source data file for exact p-values. See methods for details on biological replicates for western blots.