Extended Data Figure 5: Representative OCR experiments of brown adipocytes and other cell types ± acute addition succinate and related substrates.

a, Representative OCR trace monitoring effect of acute addition of succinate or noradrenaline in the De2.3 immortalized brown adipocyte cell line³¹ (NE; vehicle n = 7; succinate n = 6; NE n = 7). b, Representative OCR trace monitoring dose-dependent effect of acute addition of succinate in the De2.3 immortalized brown adipocyte cell line (vehicle n = 7; 1 mM succinate n = 6; 5 mM succinate n = 7). c, Representative OCR trace monitoring dose-dependent effect of acute addition of succinate in various cell types (subcutaneous white adipocytes n = 6; 1 mM n = 6; 5 mM n = 5; myoblasts n = 7; 1 mM n = 6; 5 mM n = 7; HEK n = 7; 1 mM n = 6; 5 mM n = 7; hepatocytes n = 7; 1 mM n = 6; 5 mM n = 7; osteocytes n = 7; 1 mM n = 6; 5 mM n = 7; A549 lung n = 7; 1 mM n = 6; 5 mM n = 7; brown pre-adipocytes n = 7; 1 mM n = 6; 5 mM n = 7). d, Effect of acute addition of succinate on cellular OCR in human brown adipocytes (n = 6; 5 mM n = 7). e, Representative OCR trace monitoring dose-dependent effect of acute addition of succinate in human immortalized brown adipocytes (vehicle n = 7; 1 mM succinate n = 6). f, Inhibition of succinate-stimulated OCR in brown adipocytes by suppression of plasma membrane transport with DTNB. g, Representative oxygen consumption rate (OCR) experiment (n = 7; 0.1 mM DTNB n = 6). h,i, Inhibition of succinate-stimulated OCR in brown adipocytes by treatment with the plasma membrane transport inhibitor DIDS. (n = 12; 1 mM DIDS n = 8). j,k, Inhibition of succinate-stimulated OCR in brown adipocytes by treatment with the Na⁺/K⁺ ATPase inhibitor ouabain. (n = 5). Effects on respiration were determined by acute addition, ± oligomycin (oli) to determine leak respiration, ± 2,4-dinitrophenol (DNP) to determine chemically uncoupled maximal respiration, and rotenone + antimycin (r/a) to determine non-mitochondrial respiration. In all cases basal respiration in these cells is in the presence of 1 mM pyruvate. d, two-sided t-test; f,h,j two-way ANOVA; data are mean of biologically independent samples ± s.e.m.