Cold induces brain region-selective cell activity-dependent lipid metabolism
- Department of Medicine, Division of Endocrinology, Albert Einstein College of Medicine, United States
- Friends Seminary, United States
- Department of Neuroscience, Albert Einstein College of Medicine, Bronx, United States
- Einstein-Mount Sinai Diabetes Research Center, Albert Einstein College of Medicine, Bronx, United States
- The Fleischer Institute for Diabetes and Metabolism, Albert Einstein College of Medicine, Bronx, United States
Figures
Figure 1 with 1 supplement
Brain region-selective responses to cold.
Micro-punches of paraventricular nucleus of the hypothalamus (PVH), lateral hypothalamus (LH), dorsomedial hypothalamus (DMH), ventromedial hypothalamus (VMH), and arcuate nucleus (ARC) were made from male mice exposed to a cold chamber for 4–6 hr, which were directly used for real-time qPCR (RT-qPCR) of the gene markers of lipolysis and thermogenesis. Group data of the lipolytic marker (A1) Pnpla2 (Ctrl, n = 6; Cold, n = 11) and (A2) Lipe (Ctrl, n = 6; Cold, n = 12) as well as thermogenic marker (A3) Ucp2 (Ctrl, n = 7; Cold, n = 8), (A4) Cidea (Ctrl, n = 7; Cold, n = 8), and (A5) Prdm16 (Ctrl, n = 7; Cold, n = 8) in the PVH. Group data of the lipolytic marker (B1) Pnpla2 (Ctrl, n = 7; Cold, n = 8) and (B2) Lipe (Ctrl, n = 7; Cold, n = 8) as well as thermogenic marker (B3) Ucp2 (n = 7 each group), (B4) Cidea (n = 7 each group), and (B5) Prdm16 (n = 7 each group) in the LH. Group data of the lipolytic marker (C1) Pnpla2 (Ctrl, n = 7; Cold, n = 8) and (C2) Lipe (n = 7 each group), and thermogenic marker (C3) Ucp2 (Ctrl, n = 7; Cold, n = 8), (C4) Cidea (Ctrl, n = 7; Cold, n = 8), and (C5) Prdm16 (Ctrl, n = 7; Cold, n = 8) in the DMH. Group data of the lipolytic marker (D1) Pnpla2 (Ctrl, n = 7; Cold, n = 8) and (D2) Lipe (Ctrl, n = 7; Cold, n = 8), and thermogenic marker (D3) Ucp2 (Ctrl, n = 7; Cold, n = 8), (D4) Cidea (Ctrl, n = 7; Cold, n = 8), and (D5) Prdm16 (Ctrl, n = 7; Cold, n = 8) in the VMH. Group data of the lipolytic marker (E1) Pnpla2 (Ctrl, n = 7; Cold, n = 8) and (E2) Lipe (Ctrl, n = 7; Cold, n = 8), and thermogenic marker (E3) Ucp2 (Ctrl, n = 7; Cold, n = 8), (E4) Cidea (Ctrl, n = 7; Cold, n = 8), and (E5) Prdm16 (Ctrl, n = 7; Cold, n = 8) in the ARC. Data represent mean ± SEM. Student t tests were performed. *p < 0.05, **p < 0.01. Each dot represents one animal in each group of all the panels.
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Figure 1—source data 1
Lipid metabolic gene markes in different hypothalamic areas in male mice.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig1-data1-v1.xlsx
Figure 1—figure supplement 1
Cold did not affect gene markers in paraventricular nucleus of the hypothalamus (PVH) in female mice.
Micro-punches of PVH were made from female mice exposed to a cold chamber for 4–6 hr for real-time qPCR (RT-qPCR) of the gene markers of lipolysis and thermogenesis. Group data of the lipolytic marker (A) Pnpla2 (Ctrl, n = 6; Cold, n = 12) and (B) Lipe (Ctrl, n = 6; Cold, n = 12) as well as thermogenic marker (C) Ucp2 (Ctrl, n = 4; Cold, n = 8), (D) Cidea (Ctrl, n = 4; Cold, n = 8), and (E) Prdm16 (Ctrl, n = 4; Cold, n = 8) in the PVH. Data represent mean ± SEM. Student t tests were performed. Each dot represents one animal in each group of all the panels.
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Figure 1—figure supplement 1—source data 1
Lipid metabolic gene markes in the PVH in female mice.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig1-figsupp1-data1-v1.xlsx
Figure 2
Cold increases protein expressions of ATGL in neurons and astrocytes in paraventricular nucleus of the hypothalamus (PVH).
Control or cold (4–6 hr)-challenged mice were perfused and fixed. Mouse brains were sectioned. ATGL, NSE, and S100b were stained using the relevant antibodies, respectively. (A) Representative images of ATGL (green), NSE (red), and ATGL/NSE overlay (yellow) signals in PVH sections from cold-challenged mouse. (B) Group data of relative ATGL/NSE overlay signals to total NSE signals in control and cold-challenged mice (n = 5 each group). (C) Group data of relative ATGL/S100b overlay signals to total S100b signals in control and cold-challenged mice (n = 5 each group). Data represent mean ± SEM. Student t tests were performed. ***p < 0.001, *p < 0.05. Scale bar, 100 μm for (A). PVH, paraventricular of hypothalamus; 3rd V, third ventricle.
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Figure 2—source data 1
Group data of ATGL expressing neurons and astrocyets.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig2-data1-v1.xlsx
Figure 3
Cold increases HSL protein expressions in both neurons and astrocytes in paraventricular nucleus of the hypothalamus (PVH).
Control or cold (4–6 hr)-challenged mouse brains were sectioned. HSL, NSE, and S100b were, respectively, co-stained using relevant antibodies. (A) Representative images of HSL (green), NSE (red), and HSL/NSE overlay (yellow) signals in PVH sections from cold-challenged mouse. (B) Group data of relative HSL/NSE overlay signals to total NSE signals in control and cold-challenged mice (n = 5 each group). (C) Group data of relative HSL/S100b overlay signals to total S100b signals in control and cold-challenged mice (n = 5 each group). Data represent mean ± SEM. Student t tests were performed. ***p < 0.001, *p < 0.05. Scale bar, 100 μm for (A). PVH, paraventricular of hypothalamus; 3rd V, third ventricle.
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Figure 3—source data 1
Group data of HSL expressing neurons and astrocyets.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig3-data1-v1.xlsx
Figure 4
Cold increases the level of phosphorylated HSL in paraventricular nucleus of the hypothalamus (PVH).
Micro-punches of PVH were collected in control and cold (4–6 hr)-challenged mice, which were used for (A) western blots of p-HSL (Ser660), HSL, and β-actin. (B) Group data of p-HSL fold change to HSL (n = 6 each group). Data represent mean ± SEM. Student t tests were performed. **p < 0.01. Each dot represents one mouse.
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Figure 4—source data 1
Western blots of the HSL proteins in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig4-data1-v1.zip
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Figure 4—source data 2
Gels of western blots of HSL, p-HSL, and actin.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig4-data2-v1.zip
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Figure 4—source data 3
Group data of western blots of HSL proteins in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig4-data3-v1.xlsx
Figure 5 with 2 supplements
Cold-induced lipid droplet (LD) accumulation.
Control or cold (30 min to 1 hr)-challenged mice were perfused and fixed. Mouse brains were sectioned. LDs in the paraventricular nucleus of the hypothalamus (PVH) were labeled using BODIPY493 (BD493). Representative images of BD493 signals in PVH sections from one control (A) and one cold-challenged mouse (B). (C) Group data of relative BD493-labeled area in PVH in control and cold-challenged mice (n = 6 each group). Sample images of BD493 (D) and perilipin2 (PLIN2) (E) and overlay (F) signals in the PVH. Data represent mean ± SEM. Student t tests were performed. ****p < 0.0001. Scale bars, 50 μm for (A, B) and 1 μm for (D–F). PVH, paraventricular of hypothalamus; 3rd V, third ventricle.
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Figure 5—source data 1
Group data of BD493 area in cold-challenged mice and control mice.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig5-data1-v1.xlsx
Figure 5—figure supplement 1
Cold increases lipid droplet (LD) accumulation in astrocytes but not neurons.
Control or cold (4–6 hr)-challenged mouse brains were sectioned. BD493, S100b, and NSE were, respectively, co-stained using relevant antibodies. (A) Representative images of BD493 (green), S100b (red), and BD493/S100b overlay (yellow) signals in paraventricular nucleus of the hypothalamus (PVH) sections from cold-challenged mouse. (B) Group data of relative BD493/S100b overlay signals to total S100b signals in control and cold-challenged mice (n = 4 each group). (C) Group data of relative BD493/NSE overlay signals to total NSE signals in control and cold-challenged mice (n = 4 each group). Data represent mean ± SEM. Student t tests were performed. *p < 0.05. Scale bar, 100 μm for (A). 3rd V, third ventricle.
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Figure 5—figure supplement 1—source data 1
Group data of BD493 areas in neurons and astrocytes.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig5-figsupp1-data1-v1.xlsx
Figure 5—figure supplement 2
Group data of relative BD493-labeled area in paraventricular nucleus of the hypothalamus (PVH) in control or cold-challenged (4–6 hr) mice (n = 3 each group).
Data represent mean ± SEM. Student t tests were performed.
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Figure 5—figure supplement 2—source data 1
Relative LD area in the PVH in control mice and 4-6 hr cold challenged mice.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig5-figsupp2-data1-v1.xlsx
Figure 6
Cold increased Fos expressions in the paraventricular nucleus of the hypothalamus (PVH).
Control or cold-challenged mouse brains were perfused and fixed and sectioned. Fos in the PVH were labeled using anti-Fos antibodies. (A) Representative images of Fos signals in PVH from one control (top) and one cold-challenged mouse (bottom). (B) Group data of Fos-positive cells in PVH in control and cold-challenged mice (n = 5 each group). Data represent mean ± SEM. Student t tests were performed. **p < 0.01. Scale bars, 100 μm for (A). 3rd V, third ventricle.
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Figure 6—source data 1
Group data of Fos-positive cells in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig6-data1-v1.xlsx
Figure 7 with 1 supplement
Cold-induced cell activity-dependent lipid peroxidation in the paraventricular nucleus of the hypothalamus (PVH).
Mice were injected with the lipid peroxidation indicator BD-C11 in the PVH via the implanted optical fiber multiple fluid injection cannula (OmFC) 4 hr before placing them in a temperature-controlled chamber. Two-color fiber photometry was applied for time-lapse monitoring of BD-C11 conversion via the optic fiber of the OmFC. (A) Representative traces of real-time photometry monitoring of red and green signals simultaneously. Group data of relative BD-C11 ratio to baseline in mice treated with vehicle (B), n = 6, α-TP (C), n = 4, and KYN + MUS (D), n = 4. Data represent mean ± SEM.
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Figure 7—source data 1
Group data of photometry monitoring of lipid peroxidation in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig7-data1-v1.xlsx
Figure 7—figure supplement 1
Neuronal inhibition by KYN + MUS.
(A) Representative images of GCaMP6f expressions in paraventricular nucleus of the hypothalamus (PVH) neurons and cannula track. (B) Representative trace of neuronal GCaMP6f signals recorded in a time-lapse manner in a cold chamber. (C) Group data of the experiment performed in the B (n = 4). (D) Representative traces of neuronal GCaMP6f signals recorded from vehicle (top) and KYN + MUS (bottom)-injected mice, and stars indicate Ca2+ spikes. (E) Group data of relative GCaMP6f-labeled Ca2+ spikes in PVH neurons in vehicle or KYN + MUS-injected mice (n = 3 each group). Data represent mean ± SEM. Student t tests were performed. *p < 0.05 for (E).
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Figure 7—figure supplement 1—source data 1
Group data of photometry monitoring of PVH neuronal activity.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig7-figsupp1-data1-v1.xlsx
Figure 8
Cold-induced cell activity-dependent lipid lipolysis.
Mice were injected with the lipase substrate in the paraventricular nucleus of the hypothalamus (PVH) via the implanted optical fiber multiple fluid injection cannula (OmFC) 4 hr before placing them in a temperature-controlled chamber. Two-color fiber photometry was used for time-lapse monitoring of lipase substrate conversion via the optic fiber of the OmFC. (A) Representative traces of real-time photometry monitoring of green signals. Group data of lipase substrate conversion in mice treated with vehicle (B), n = 4, α-TP (C), n = 4, and KYN + MUS (D), n = 4. Data represent mean ± SEM.
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Figure 8—source data 1
Group data of photometry monitoring of lipid lipolysis in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig8-data1-v1.xlsx
Figure 9
Cold-induced cell activity-dependent lipid droplet (LD) accumulation.
Mice were injected with the LD marker BODIPY493 in the paraventricular nucleus of the hypothalamus (PVH) via the optical fiber multiple fluid injection cannula (OmFC) 4 hr before placing them in a temperature-controlled chamber. Fiber photometry was used for time-lapse monitoring of BODIPY493 signals via the optic fiber of the OmFC. (A) Representative traces of real-time photometry monitoring of BODIPY493 signals. Group data of BODIPY493 in mice treated with vehicle (B), n = 4, α-TP (C), n = 4, and KYN + MUS (D), n = 6. Data represent mean ± SEM.
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Figure 9—source data 1
Group data of photometry monitoring of LD dynamics in the PVH.
- https://cdn.elifesciences.org/articles/98353/elife-98353-fig9-data1-v1.xlsx
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Cold induces brain region-selective cell activity-dependent lipid metabolism
eLife 13:RP98353.
https://doi.org/10.7554/eLife.98353.3
