Thermoneutral housing does not influence fat mass or glucose homeostasis in C57BL/6 mice

in Journal of Endocrinology
Correspondence should be addressed to A E Brandon:

*(L Small and H Gong contributed equally to this work)

One major factor affecting physiology often overlooked when comparing data from animal models and humans is the effect of ambient temperature. The majority of rodent housing is maintained at ~22°C, the thermoneutral temperature for lightly clothed humans. However, mice have a much higher thermoneutral temperature of ~30°C, consequently data collected at 22°C in mice could be influenced by animals being exposed to a chronic cold stress. The aim of this study was to investigate the effect of housing temperature on glucose homeostasis and energy metabolism of mice fed normal chow or a high-fat, obesogenic diet (HFD). Male C57BL/6J(Arc) mice were housed at standard temperature (22°C) or at thermoneutrality (29°C) and fed either chow or a 60% HFD for 13 weeks. The HFD increased fat mass and produced glucose intolerance as expected but this was not exacerbated in mice housed at thermoneutrality. Changing the ambient temperature, however, did alter energy expenditure, food intake, lipid content and glucose metabolism in skeletal muscle, liver and brown adipose tissue. Collectively, these findings demonstrate that mice regulate energy balance at different housing temperatures to maintain whole-body glucose tolerance and adiposity irrespective of the diet. Despite this, metabolic differences in individual tissues were apparent. In conclusion, dietary intervention in mice has a greater impact on adiposity and glucose metabolism than housing temperature although temperature is still a significant factor in regulating metabolic parameters in individual tissues.


      Society for Endocrinology

TrendMD Widget

Index Card


All Time Past Year Past 30 Days
Abstract Views 436 436 60
Full Text Views 419 419 17
PDF Downloads 145 145 13

Altmetric Badge


  • View in gallery

    Thermoneutrality reduces energy expenditure and energy intake without changing body or fat mass. 22°C = black. 29°C = red. Solid line/circles = chow. Dashed line/crosses = HFD. Mean ± s.e.m. (A) Weekly body mass. (B) Fat and (C) lean mass determined by echo MRI. n = 12. Analysed by repeated measures 2-way ANOVA for an effect of temperature and an effect of diet. (D) Diurnal oxygen consumption and (E) total over 24 h. (F) Diurnal respiratory exchange ratio and (G) average over 24 h. (H) Total energy intake over 24 h. (I) Total activity (beam breaks) over 24 h. n = 4. Analysed by 2-way ANOVA for an effect of temperature and an effect of diet. ***P < 0.0005. A full colour version of this figure is available at

  • View in gallery

    Effect of diet but no effect of temperature on glucose tolerance. 22°C = black. 29°C = red. Solid line/ circles = chow. Dashed line/crosses = HFD. Mean ± s.e.m. Oral glucose tolerance curves at (A) 8 and (E) 13 weeks from the start of study. Area under the curve of the glucose tolerance at (B) 8 and (F) 13 weeks. Blood insulin during the oGTT at (C) 8 and (G) 13 weeks. Area under the curve of the blood insulin at (D) 8 and (H) 13 weeks. Analysed by 2-way ANOVA for an effect of temperature and an effect of diet. n = 10–12. A full colour version of this figure is available at

  • View in gallery

    HFD-feeding increases UCP1 abundance in BAT while thermoneutrality reduces it. 22°C = black. 29°C = red. Mean ± s.e.m. (A) Representative immunoblot and (B) quantitation of UCP1 in BAT normalised to 14-3-3. n = 12 (C) 2-DG uptake (phosphorylated [3H]2-deoxyglucose) in BAT. n = 7–12. (D) Representative immunoblot and quantitation of subunits of mitochondrial OXPHOS Complex I-V in BAT, (E) Complex I (NDUFB8), (F) Complex II (SDHB), (G) Complex III (UQCRC2), (H) Complex IV (MTCO1) and (I) Complex V (ATP5A) normalised to 14-3-3. (J) BAT protein content. n = 10–12. Analysed by 2-way ANOVA for an effect of temperature and an effect of diet. ***P < 0.0005. (K) Hematoxylin & Eosin staining of BAT sections, scale bar = 25 µm. A full colour version of this figure is available at

  • View in gallery

    Increased triglyceride content and expression of lipogenic proteins in liver from mice at 29°C. 22°C = black. 29°C = red. Mean ± s.e.m. (A) Liver triglyceride content. (B) Liver glycogen content. (C) Representative immunoblots and quantitation of (D) ACC, (E) FAS, (F) SCD1 and (G) PEPCK normalised to 14-3-3. Analysed by 2-way ANOVA for an effect of temperature and an effect of diet. n = 11–12. *P < 0.05, **P < 0.005, ***P < 0.0005. (H) Hematoxylin & Eosin staining of liver sections, scale bar = 50 µm. A full colour version of this figure is available at

  • View in gallery

    Thermoneutrality reduces triglyceride content and increases basal glucose uptake in the quadriceps muscle of HFD mice. 22°C = black. 29°C = red. Mean ± s.e.m. (A) Quadriceps triglyceride content (B) Quadriceps β-HAD activity. n = 12. (C) 2-DG uptake (phosphorylated [3H]2-deoxyglucose) in the quadriceps. n = 7–12. (D) Quadriceps glycogen content. n = 11–12. (E) Linear regression of quadriceps triglyceride content and 2-DG uptake, n = 39, circles = chow, crosses = HFD, dashed lines = 95% confidence intervals. Representative immunoblot (F) and quantitation (G) of UCP3 in the quadriceps muscle normalised to β-Actin. n = 12. (E) Analysed by 2-way ANOVA for an effect of temperature and an effect of diet. **P < 0.005, ***P < 0.0005, #P = 0.051. A full colour version of this figure is available at


Abreu-VieiraGXiaoCGavrilovaOReitmanML 2015 Integration of body temperature into the analysis of energy expenditure in the mouse. Molecular Metabolism 4 461470. (

AndrikopoulosSBlairARDelucaNFamBCProiettoJ 2008 Evaluating the glucose tolerance test in mice. American Journal of Physiology. Endocrinology and Metabolism 295 E1323E1332. (

BarneaMShamayAStarkAHMadarZ 2006 A high-fat diet has a tissue-specific effect on adiponectin and related enzyme expression. Obesity 14 21452153. (

BonnardCDurandAVidalHRieussetJ 2008 Changes in adiponectin, its receptors and AMPK activity in tissues of diet-induced diabetic mice. Diabetes and Metabolism 34 5261. (

BrandonAETid-AngJWrightLEStuartESuryanaEBentleyNTurnerNCooneyGJRudermanNBKraegenEW 2015 Overexpression of SIRT1 in rat skeletal muscle does not alter glucose induced insulin resistance. PLoS ONE 10 e0121959. (

CannonBNedergaardJ 2011 Nonshivering thermogenesis and its adequate measurement in metabolic studies. Journal of Experimental Biology 214 242253. (

CastilloMHallJACorrea-MedinaMUetaCKangHWCohenDEBiancoAC 2011 Disruption of thyroid hormone activation in type 2 deiodinase knockout mice causes obesity with glucose intolerance and liver steatosis only at thermoneutrality. Diabetes 60 10821089. (

CooneyGJCatersonIDNewsholmeEA 1985 The effect of insulin and noradrenaline on the uptake of 2-[1–14C]deoxyglucose in vivo by brown adipose tissue and other glucose-utilising tissues of the mouse. FEBS Letters 188 257261 (

CuiXNguyenNLTZarebidakiECaoQLiFZhaLBartnessTShiHXueB 2016 Thermoneutrality decreases thermogenic program and promotes adiposity in high-fat diet-fed mice. Physiological Reports 4. (

FeldmannHMGolozoubovaVCannonBNedergaardJ 2009 UCP1 ablation induces obesity and abolishes diet-induced thermogenesis in mice exempt from thermal stress by living at thermoneutrality. Cell Metabolism 9 203209. (

FreglyMJMarshallNBMayerJ 1957 Effect of changes in ambient temperature on spontaneous activity, food intake and body weight of goldthioglucose-obese and nonobese mice. American Journal of Physiology 188 435438. (

FrommeTKlingensporM 2011 Uncoupling protein 1 expression and high-fat diets. American Journal of Physiology. Regulatory Integrative and Comparative Physiology 300 R1R8. (

GilesDARamkhelawonBDonelanEMStankiewiczTEHutchisonSBMukherjeeRCappellettiMKarnsRKarpCLMooreKJ 2016 Modulation of ambient temperature promotes inflammation and initiates atherosclerosis in wild type C57BL/6 mice. Molecular Metabolism 5 11211130. (

GilesDAMoreno-FernandezMEStankiewiczTEGraspeuntnerSCappellettiMWuDMukherjeeRChanCCLawsonMJKlarquistJ 2017 Thermoneutral housing exacerbates nonalcoholic fatty liver disease in mice and allows for sex-independent disease modeling. Nature Medicine 23 829838. (

HoevenaarsFPMBekkenkamp-GrovensteinMJanssenRJHeilSGBunschotenAHoek-van den HilEFSnaas-AldersSTeerdsKvan SchothorstEMKeijerJ 2014 Thermoneutrality results in prominent diet-induced body weight differences in C57BL/6J mice, not paralleled by diet-induced metabolic differences. Molecular Nutrition and Food Research 58 799807. (

HoyAJBrandonAETurnerNWattMJBruceCRCooneyGJKraegenEW 2009 Lipid and insulin infusion-induced skeletal muscle insulin resistance is likely due to metabolic feedback and not changes in IRS-1, Akt, or AS160 phosphorylation. American Journal of Physiology. Endocrinology and Metabolism 297 E67E75. (

HylanderBLRepaskyEA 2016 Thermoneutrality, mice, and cancer: a heated opinion. Trends in Cancer 2 166175. (

KingmaBRFrijnsAJSchellenLvan Marken LichtenbeltWD 2014 Beyond the classic thermoneutral zone: including thermal comfort. Temperature 1 142149. (

KowalskiGMBruceCR 2014 The regulation of glucose metabolism: implications and considerations for the assessment of glucose homeostasis in rodents. American Journal of Physiology. Endocrinology and Metabolism 307 E859E871. (

Lara-CastroCLuoNWallacePKleinRLGarveyWT 2006 Adiponectin multimeric complexes and the metabolic syndrome trait cluster. Diabetes 55 249259 (

LeeYSLiPHuhJYHwangIJLuMKimJIHamMTalukdarSChenALuWJ 2011 Inflammation is necessary for long-term but not short-term high-fat diet-induced insulin resistance. Diabetes 60 24742483. (

LiuXRossmeislMMcClaineJRiachiMHarperM-EKozakLP 2003 Paradoxical resistance to diet-induced obesity in UCP1-deficient mice. Journal of Clinical Investigation 111 399407. (

LodhiIJSemenkovichCF 2009 Why we should put clothes on mice. Cell Metabolism 9 111112. (

MaloneySKFullerAMitchellDGordonCOvertonJM 2014 Translating animal model research: does it matter that our rodents are cold? Physiology 29 413420. (

MasSMartínez-PinnaRMartín-VenturaJLPérezRGomez-GarreDOrtizAFernandez-CruzAVivancoFEgidoJ 2010 Local non-esterified fatty acids correlate with inflammation in atheroma plaques of patients with type 2 diabetes. Diabetes 59 12921301. (

OanaFTakedaHHayakawaKMatsuzawaAAkahaneSIsajiMAkahaneM 2005 Physiological difference between obese (fa/fa) Zucker rats and lean Zucker rats concerning adiponectin. Metabolism: Clinical and Experimental 54 9951001. (

OvertonJM 2010 Phenotyping small animals as models for the human metabolic syndrome: thermoneutrality matters. International Journal of Obesity 34(Supplement 2) S53S58. (

RippeCBergerKBöiersCRicquierDErlanson-AlbertssonC 2000 Effect of high-fat diet, surrounding temperature, and enterostatin on uncoupling protein gene expression. American Journal of Physiology. Endocrinology and Metabolism 279 E293E300. (

SchrauwenPHoeksJHesselinkMKC 2006 Putative function and physiological relevance of the mitochondrial uncoupling protein-3: involvement in fatty acid metabolism? Progress in Lipid Research 45 1741. (

ShabalinaIGPetrovicNde JongJMAKalinovichAVCannonBNedergaardJ 2013 UCP1 in Brite/beige adipose tissue mitochondria is functionally thermogenic. Cell Reports 5 11961203. (

StemmerKKotzbeckPZaniFBauerMNeffCMüllerTDPflugerPTSeeleyRJDivanovicS 2015 Thermoneutral housing is a critical factor for immune function and diet-induced obesity in C57BL/6 nude mice. International Journal of Obesity 39 791797. (

StorlienLHBaurLAKriketosADPanDACooneyGJJenkinsABCalvertGDCampbellLV 1996 Dietary fats and insulin action. Diabetologia 39 621631 (

SwoapSJLiCWessJParsonsADWilliamsTDOvertonJM 2008 Vagal tone dominates autonomic control of mouse heart rate at thermoneutrality. American Journal of Physiology. Heart and Circulatory Physiology 294 H1581H1588. (

TianXYGaneshanKHongCNguyenKDQiuYKimJTangiralaRKTontonozPTonotonozPChawlaA 2016 Thermoneutral housing accelerates metabolic inflammation to potentiate atherosclerosis but not insulin resistance. Cell Metabolism 23 165178. (

TurnerNBruceCRBealeSMHoehnKLSoTRolphMSCooneyGJ 2007 Excess lipid availability increases mitochondrial fatty acid oxidative capacity in muscle: evidence against a role for reduced fatty acid oxidation in lipid-induced insulin resistance in rodents. Diabetes 56 20852092. (

TurnerNKowalskiGMLeslieSJRisisSYangCLee-YoungRSBabbJRMeiklePJLancasterGIHenstridgeDC 2013 Distinct patterns of tissue-specific lipid accumulation during the induction of insulin resistance in mice by high-fat feeding. Diabetologia 56 16381648. (

UchidaKShiuchiTInadaHMinokoshiYTominagaM 2010 Metabolic adaptation of mice in a cool environment. Pflugers Archiv 459 765774. (

Cited By


Google Scholar