Early postnatal interactions between beige adipocytes and sympathetic neurites regulate innervation of subcutaneous fat
- Laboratory of Molecular Metabolism, The Rockefeller University, United States
- Laboratory of Metabolic Regulation and Genetics, The Rockefeller University, United States
Figures
Figure 1 with 5 supplements
Sympathetic innervation of iWAT is established during early postnatal development.
(A) Schematic view of iWAT. Red lines represent main blood vessels. Using the lymph node and blood vessels as landmarks, iWAT depot is divided into the inguinal and dorsolumbar regions. Dorsolumbar …
Figure 1—figure supplement 1
Organization of sympathetic nervous system in iWAT at P2.
(A–B) Representative whole-tissue maximum intensity projections (MIPs) of 50 μm z-stacks of iWAT from a P2 mouse. (A) TH (green) and CD31 (magenta). (B) TH only. (C–F) High-magnification images …
Figure 1—figure supplement 2
Organization of sympathetic nervous system in iWAT at P6.
Figure 1—figure supplement 3
Development of sympathetic parenchymal innervation in iWAT.
High-magnification images corresponding to Figure 1G,H,I & J. MIPs of 50 μm z-stacks from iWAT of (A–B) P6, (C–D) P10, (E–F) P14, (G–H) P21, and (I–J) P28 immunolabeled with TH. (A, C, E, G, and I) …
Figure 1—video 1
Corresponding to Figure 1B–C.
Organization of sympathetic nervous system in iWAT at P6. The video first shows a rotating 3D projection of a whole iWAT from a P6 C57BL/6J mouse immunolabeled with TH, followed by a fly-through of …
Figure 1—video 2
Corresponding to Figure 1—figure supplement 3.
Development of sympathetic parenchymal innervation in iWAT. The video shows fly-throughs of optical sections from iWAT samples of P6, P10, and P14 C57BL/6J mice immunolabeled with TH, followed by …
Figure 2 with 3 supplements
UCP1+ beige adipocytes emerge during early postnatal development.
(A–J) Representative optical sections of iWAT from P6, P10, P14, and P21 C57BL6/J mice immunolabeled with UCP1. (A) Whole-tissue optical section of a P6 iWAT. (B) High-magnification view of the …
Figure 2—figure supplement 1
UCP1+ beige adipocytes emerge during early postnatal development.
(A–C) Representative optical sections of iWAT from a P28 C57BL6/J mouse immunolabeled with UCP1. (A) Whole-tissue optical section of a P28 iWAT. (B) High-magnification view of the boxed dorsolumbar …
Figure 2—figure supplement 2
UCP1+ beige adipocytes emerge during early postnatal development.
(A–C) Normalized gene expression of dorsolumbar vs. inguinal regions in iWAT from P6, P8, P10, P12, P14, P16, P21, and P28 C57BL6/J mice, n = 4. Representative genes involved in (A) the thermogenic …
Figure 2—figure supplement 3
UCP1+ beige adipocytes and dense sympathetic parenchymal innervation emerge together during early postnatal development.
(A–J) High-magnification MIPs of 50 μm z-stacks from iWAT of (A–B) P6, (C–D) P10, (E–F) P14, (G–H) P21, and (I–J) P28 immunolabeled with TH (green) and UCP1 (magenta), corresponding to Figure …
Figure 3 with 2 supplements
Prdm16 regulates the emergence of early postnatal beige adipocytes and dense sympathetic parenchymal innervation.
(A–D) Representative images of iWAT from (A–B) control and (C–D) cPrdm16KO mice at P14, immunolabeled with TH. MIPs from 50 μm z-stacks are shown. (E–H) Representative images of iWAT from (E–F) …
Figure 3—figure supplement 1
Prdm16 regulates the emergence of early postnatal beige adipocytes and dense sympathetic parenchymal innervation.
(A–D) Representative images of iWAT from (A–B) control and (C–D) cPrdm16KO mice at P6, immunolabeled with TH. MIPs from 50 μm z-stacks are shown. (E–H) Overlay of TH (green) and UCP1 (magenta), …
Figure 3—figure supplement 2
Prdm16 regulates the emergence of early postnatal beige adipocytes and dense sympathetic parenchymal innervation.
(A–D) Overlay of TH (green) and UCP1 (magenta), corresponding to Figure 3A–D. (E–H) Overlay of TH (green) and UCP1 (magenta), corresponding to Figure 3E–H. (I) qPCR analysis of Prdm16 mRNA levels in …
Figure 4 with 5 supplements
Prdm16 deletion during early development causes decreased sympathetic parenchymal innervation.
(A) Schematic representation of the genetic components of the control and iPrdm16KO mice. iPrdm16KO mice carry floxed Prdm16 alleles (Prdm16lox/lox), two copies of AdipoqrtTA transgene, and one copy …
Figure 4—figure supplement 1
Prdm16 deletion during early development blocks cold-induced thermogenic gene expression and causes decreased sympathetic parenchymal innervation in iWAT.
(A) Normalized Prdm16 mRNA expression of the indicated tissue types from control and iPrdm16KO (perinatal, 5 weeks) mice maintained at RT, n = 4–5. Data are presented as mean + SEM and analyzed by …
Figure 4—figure supplement 2
Neurite density quantification in 3D images.
(A–F) Illustration of sympathetic parenchymal neurite density quantification with random sampling and filament tracing. (A) In each inguinal region as shown in Figure 4H–I and Figure 5G–H, small …
Figure 4—figure supplement 3
Prdm16 deletion during early development does not affect thermogenic gene program or sympathetic innervation in iBAT.
(A–B) Normalized gene expression of iBAT from control and iPrdm16KO (perinatal, 5 weeks) mice exposed to RT or cold, n = 3–5. (A) Representative genes involved in the thermogenic program and (B) the …
Figure 4—figure supplement 4
Prdm16 deletion during early development causes lasting defects in sympathetic parenchymal innervation.
(A) Control (Cre-) and iPrdm16KO (Cre+) mice housed at RT (23°C) were kept on a doxycycline-containing chow diet from E14 until P21 before being switched to a regular chow diet and maintained until …
Figure 4—video 1
Corresponding to Figure 4.
Sympathetic neurite density following perinatal Prdm16 deletion. The video shows fly-throughs of optical sections from in the inguinal regions of representative control and iPrdm16KO (perinatal, 5 we…
Figure 5 with 3 supplements
Prdm16 is not required for maintaining sympathetic parenchymal innervation in mature iWAT.
(A) Control (Cre-) and iPrdm16KO (Cre+) mice housed at RT (23°C) were kept on a regular chow diet until 8 weeks of age before being switched to a doxycycline-containing chow diet for 4 weeks. …
Figure 5—figure supplement 1
Prdm16 deletion in adulthood attenuates cold-induced thermogenic gene expression but does not affect sympathetic parenchymal innervation in iWAT.
(A) Western blot of PRDM16 and lamin A/C (loading control) from nuclear extracts of iWAT from control (Cre-) and iPrdm16KO (adult) (Cre+) samples. (B) Body weights of control and iPrdm16KO (adult) …
Figure 5—figure supplement 2
Additional tissue structures in the core of the inguinal region.
(A–E) Representative images of iWAT from a P8 C57BL6/J mouse immunolabeled with LYVE1 (magenta) and TH (green). MIPs of 50 μm z-stacks are shown. (A) LYVE1. (B) TH. (C) Autofluorescence. (D) Overlay …
Figure 5—video 1
Corresponding to Figure 5.
Sympathetic neurite density following adult Prdm16 deletion. The video shows fly-throughs of optical sections from in the inguinal regions of representative control and iPrdm16KO (adult deletion) …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain, strain background (Mus musculus) | Adipoq-Cre | Jackson Laboratory | RRID:IMSR_JAX:028020 | |
| Strain, strain background (Mus musculus) | Adipoq-rtTA | PMID:22451920 | RRID:IMSR_JAX:033448 | |
| Strain, strain background (Mus musculus) | TRE-Cre | Jackson Laboratory | RRID:IMSR_JAX:006234 | |
| Strain, strain background (Mus musculus) | Prdm16lox/lox | PMID:24439384 | RRID:IMSR_JAX:024992 | |
| Antibody | Anti-tyrosine hydroxylase (Rabbit polyclonal) | Millipore | Cat# AB152, RRID:AB_390204 | IF(1:200) |
| Antibody | Anti-tyrosine hydroxylase (Sheep polyclonal) | Millipore | Cat# AB1542, RRID:AB_90755 | IF(1:200) |
| Antibody | Anti-CD31/PECAM-1 (Goat polyclonal) | R and D Systems | Cat# AF3628, RRID:AB_2161028 | IF(1:200) |
| Antibody | Anti-UCP1 (Rabbit polyclonal) | Abcam | Cat# ab10983, RRID:AB_2241462 | IF(1:200) |
| Antibody | Anti-LYVE1 (Rabbit polyclonal) | Abcam | Cat# ab14917, RRID:AB_301509 | IF(1:200) |
| Antibody | Anti-PRDM16 (Sheep polyclonal) | R and D Systems | Cat# AF6295, RRID:AB_10717965 | WB(1:500) |
| Antibody | Anti-Lamin A/C (Mouse monoclonal) | Santa Cruz Biotechnology | Cat# sc-376248, RRID:AB_10991536 | WB(1:2000) |
| Antibody | Anti-Rabbit IgG (H+L), Alexa Fluor 568 (Donkey polyclonal) | Thermo Fisher Scientific | Cat# A10042, RRID:AB_2534017 | IF(1:200) |
| Antibody | Anti-Rabbit IgG (H+L), Alexa Fluor 647 (Donkey polyclonal) | Thermo Fisher Scientific | Cat# A32795, RRID:AB_2762835 | IF(1:200) |
| Antibody | Anti-Sheep IgG (H+L), Alexa Fluor 647 (Donkey polyclonal) | Thermo Fisher Scientific | Cat# A-21448, RRID:AB_2535865 | IF(1:200) |
| Antibody | Anti-Goat IgG (H+L), Alexa Fluor 568 (Donkey polyclonal) | Thermo Fisher Scientific | Cat# A-11057, RRID:AB_2534104 | IF(1:200) |
| Software, algorithm | Imaris | Bitplane | http://www.bitplane.com/imaris/imaris; RRID:SCR_007370 | |
| Software, algorithm | FilamentTracer | Bitplane | http://www.bitplane.com/imaris/filamenttracer; RRID:SCR_007366 |
Additional files
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Supplementary file 1
qPCR primer sequences used in this study.
- https://cdn.elifesciences.org/articles/64693/elife-64693-supp1-v2.docx
-
Transparent reporting form
- https://cdn.elifesciences.org/articles/64693/elife-64693-transrepform-v2.docx
