Chronic cold exposure enhances glucose oxidation in brown adipose tissue

Abstract The cultured brown adipocytes can oxidize glucose in vitro, but it is still not fully clear whether brown adipose tissue (BAT) could completely oxidize glucose in vivo. Although positron emission tomography (PET) with 18F‐fluorodeoxyglucose (18F‐FDG) showed a high level of glucose uptake in the activated BAT, the non‐metabolizable 18F‐FDG cannot fully demonstrate intracellular glucose metabolism. Through in vivo [U‐13C]glucose tracing, here we show that chronic cold exposure dramatically activates glucose oxidation in BAT and the browning/beiging subcutaneous white adipose tissue (sWAT). Specifically, chronic cold exposure enhances glucose flux into the mitochondrial TCA cycle. Metabolic flux analysis models that β3‐adrenergic receptor (β3‐AR) agonist significantly enhances the flux of mitochondrial pyruvate uptake through mitochondrial pyruvate carrier (MPC) in the differentiated primary brown adipocytes. Furthermore, in vivo MPC inhibition blocks cold‐induced glucose oxidation and impairs body temperature maintenance in mice. Together, mitochondrial pyruvate uptake and oxidation serve an important energy source in the chronic cold exposure activated BAT and beige adipose tissue, which supports a role for glucose oxidation in brown fat thermogenesis.

Mice, housed at 30°C or 6°C for 10 days, were administered with [U-13 C]glucose (2 g/kg, IP). 15 minutes after injection, liver and muscle were harvested for metabolic enrichment assay.
A m+6 glucose enrichment in liver. B Metabolic 13 C enrichments in liver are shown as m+3 glycolysis intermediates, m+2 TCA cycle intermediates. GAP, glyceraldehyde 3-phosphate; DHAP, dihydroxyacetone phosphate. C After normalizing to the glucose enrichment in the liver of each mouse, the relative metabolic 13 C enrichments were shown as m+3 glycolysis intermediates and m+2 TCA cycle intermediates. D m+6 glucose enrichment in muscle. E Metabolic 13 C enrichments in muscle are shown as m+3 glycolysis intermediates, m+2 TCA cycle intermediates.
Data information: n = 10, data are represented as the mean AE SD. Statistical analysis was performed using two-tailed Student's t-test, *P < 0.05. Mice, housed at 30 or 6°C for 10 days, were administered with [U-13 C]glucose (2 g/kg, IP). 15 minutes after injection, BAT, sWAT, and gWAT were harvested for metabolic enrichment assay.
A-C Metabolic 13 C enrichments in BAT of female mice are shown as m+6 glucose and m+3 glycolytic intermediates (A), m+2 TCA cycle intermediates (B), and the enrichment of G3P (C). D, E Metabolic 13 C enrichments in sWAT of female mice are shown as m+6 glucose and m+3 glycolysis intermediates (D), m+2 TCA cycle intermediates (E). F The m+3 enrichment of G3P in sWAT of both female and male mice. G, H Metabolic 13 C enrichments in gWAT of female mice are shown as m+6 glucose and m+3 glycolysis intermediates (G), m+2 TCA cycle intermediates (H). I The m+3 enrichment of G3P in gWAT of both female and male mice.
Data information: n = 6-8 female mice, and n = 10 male mice, data are represented as the mean AE SD. Statistical analysis was performed using two-tailed Student's ttest, *P < 0.05.
Source data are available online for this figure. A B C Figure EV4. b3-AR agonist activates glucose oxidation in differentiated primary brown adipocytes.
A-C In the sample [U-13 C]glucose experiment as shown in Fig 5, the enrichments of other metabolites were used for MFA modeling. n = 3 biological repeats, data are represented as the mean AE SD. Statistical analysis was performed using two-way ANOVA followed by Tukey's multiple comparisons test, *P < 0.05.
Source data are available online for this figure.

EV4
A D E B C Figure EV5. CHC represses glucose metabolism in multiple adipose tissues.
A Relative mRNA levels of Mpc1/2 expression were measured by qPCR in BAT of the mice, housed at 30 or 6°C for 10 days. n = 5-6 biological replicates. B Relative mRNA levels of Mpc1/2 expression were measured by qPCR in the pre-differentiated day 0 and fully differentiated brown adipocytes day 6. n = 4 biological replicates. C Oxygen consumption rate (OCR) of mouse brown adipocytes treated with MPC inhibitor CHC (2 mM) or UK5099 (2 lM), n = 6-7 biological repeats. CL, CL316,243. D, E Mice were housed at 6°C for 10 days, and mice were IP injected with PBS or CHC (500 mg/kg). 30 minutes after CHC treatment, mice were administered with [U-13 C]glucose (2 g/kg, IP). Metabolic 13 C enrichments in sWAT (D) and gWAT (E) of male mice are shown as m+2 and m+3 TCA cycle intermediates. n = 7 biological replicates.
Data information:, data are represented as the mean AE SD, except that (C) is represented as the mean AE SEM. Statistical analysis was performed using two-tailed Student's t-test, *P < 0.05.
Source data are available online for this figure.