Fig. 7: Inhibition of mitochondrial permeability transition in NCLX-null mice restores thermogenic function.

a, b OCR of NCLX-null and WT primary BA that were pretreated either with the mPTP inhibitor, NIM811, or with vehicle control, DMSO. a Representative OCR traces in BA from NCLX KO and WT pretreated with NIM811 (500 nM) or vehicle control. b Quantification of NE response in NCLX KO and WT BA pretreated with NIM811 or vehicle control (n = 28–36 per condition from N = 8–9 independent experiments; See Supplementary Fig. 10 for WT treatment traces). c Representative mitochondrial Ca²⁺ transients of NCLX KO and WT BA pretreated with NIM811 or vehicle control and stimulated by NE. d Quantification of Ca²⁺ amplitude, influx, and efflux rates (n = 4–11 independent experiments per condition). e Representative images of cells stained for TOM20 to label mitochondrial localization and assess mitochondrial swelling. Scale bar, 10 and 1 μm for the zoom images. f Quantifications of mitochondrial swelling in NE activated WT and NCLX KO BA pretreated with NIM811 or vehicle control (n = 68–118 cells). g Cell death in WT and NCLX KO BA pretreated with NIM811 or vehicle control after 72 h of NE stimulation. Cell death was measured in vitro by TUNEL assay (n = 77–83 per condition from N = 5–6 independent experiments). h Representative images of WT and NCLX KO BA after 72 h of NE stimulation. Cells were pretreated with NIM811 or vehicle control. TOM20 was used to label mitochondrial localization while DAPI was used to label the nuclei. Scale bar, 10 μm. i Schematic graph for in vivo NIM811 experimental procedure. Mice were pretreated for 5 days with a 50 mg/kg dose NIM811 or a vehicle introduced subcutaneously. On the day of the experiment, mice were acutely cold stressed (6–8 h) and then immediately PET-imaged with the glucose analog ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) to measure BAT activity under 4 °C. j Survival curves of 8–10-week-old male NCLX KO and WT mice pretreated with or without NIM811 and cold stressed at 4 °C. Mice reaching 28 °C or lower were returned to room temperature for recovery (n = 5–10 mice per group). k Representative PET-CT images of ¹⁸F-FDG uptake in cold-stressed mice. Each image is a composition of a CT image superimposed on a PET image. In the CT image, radio-opacity is reflected in grayscale. In the PET image, ¹⁸F-FDG intensity is reflected in a hot-metal scale. BAT is marked with blue arrows. ¹⁸F-FDG uptake is presented as percent injected dose per gram (%ID/g). l Quantification of ¹⁸F-FDG uptake in BAT and normalized to liver uptake as a reference tissue (n = 3–11 mice per group). One-way ANOVA with Tukey’s post hoc test (b, d, f, g, l); log-rank (j). Data are expressed as means ± SEM. ^nsp > 0.05, *p < 0.05, **p < 0.01, ***p < 0.0001. Replicates are indicated by individual dots shown for each group in all graphs. Source data are available as a Source Data file.