Evaluation of Energy Homeostasis

Carmen A. Argmann1, Marie‐France Champy2, Johan Auwerx3

1 Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, 2 Institut Clinique de la Souris, Illkirch, 3 Institut de Génétique et de Biologie Moléculaire et Cellulaire and Institut Clinique de la Souris, Illkirch
Publication Name:  Current Protocols in Molecular Biology
Unit Number:  Unit 29B.1
DOI:  10.1002/0471142727.mb29b01s73
Online Posting Date:  February, 2006
GO TO THE FULL TEXT: PDF or HTML at Wiley Online Library

Abstract

Body mass and composition reflect the combined effects of three processes: energy intake, energy partitioning (storage), and energy expenditure. Energy is released from food as it is combusted to carbon dioxide and water, and is expended as heat and work within a cell. The energy stores, mainly in adipose tissue, represent the net balance between intake and expenditure. The methods outlined in this unit evaluate these three processes by measuring food intake and lipid absorption, body fat composition, and energy expenditure. Evaluation of food intake and fat mass is a useful firstā€line phenotyping test indicating altered energy homeostasis. Evaluation of energy expenditure in this unit addresses obligatory basal energy expenditure (for performance of cellular and organ functions), as measured by indirect calorimetry. The combined results of these tests provide indications of the metabolic defects in a mouse model and help to identify molecular targets that cause these abnormalities.

Keywords: energy homeostasis; glucose homeostasis; insulin sensitivity; pathophysiology

     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Table of Contents

  • Strategic Planning
  • Basic Protocol 1: Measurement of Body Weight, Food Intake, Body Mass Index, and Fecal Lipid Content
  • Basic Protocol 2: Assessment of Thermoregulation by the Cold Test
  • Basic Protocol 3: Determination of Body Composition by Dual‐Energy X‐ray Absorptiometry
  • Alternate Protocol 1: Determination of Body Composition by Quantitative Nuclear Magnetic Resonance
  • Basic Protocol 4: Indirect Calorimetry by Oxymax
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Materials

Basic Protocol 1: Measurement of Body Weight, Food Intake, Body Mass Index, and Fecal Lipid Content

  Materials
  • Mice
  • Mouse food
  • 0.1 mCi/ml [carboxyl‐14C]triolein (112 mCi/mmol)
  • 2:1 (v/v) chloroform/methanol
  • Container to accommodate mouse during weighing
  • Scale accurate to 0.01 g
  • Mouse cages (e.g., 24 × 38 × 20–cm) with metal floor grids (grids or screens can be custom cut to fit bottom of cage; optimal grid size is one that allows feces to pass through but is still comfortable for mouse to rest on)
  • Inhalation anesthesia unit (composed of a mixing and flow‐controlled system for O 2/isoflurane anesthesia; e.g., TEM, http://www.TEM.fr)
  • Digital caliper (range, 0 to 20 cm; accuracy, <0.02 mm; resolution, <0.01 mm; Ted Pella, Inc.)
  • Analytical balance accurate to 0.0001 g
  • 70°C vacuum oven
  • 10‐ml Erlenmeyer flasks
  • 60°C shaking water bath
  • Whatman no. 1 filter paper
  • 10‐ml glass test tubes with ground glass stoppers
  • Clinical tabletop centrifuge
  • Long‐stem glass Pasteur pipets
  • Nitrogen evaporator (e.g., N‐EVAP; Organomation Associates) with dry bath at 50°C
  • Preweighed glass scintillation vials
  • β‐counter and scintillation cocktail

Basic Protocol 2: Assessment of Thermoregulation by the Cold Test

  Materials
  • Mice
  • 70% (v/v) isopropanol
  • Mouse food
  • Container to accommodate mouse during weighing
  • Scale accurate to 0.1 g
  • Standard mouse cages (metal grid not required)
  • Thermometer with thermoprobe (accuracy <0.1°C) and rectal adapter (e.g., Bioseb; http://www.bioseb.com)

Basic Protocol 3: Determination of Body Composition by Dual‐Energy X‐ray Absorptiometry

  Materials
  • Anesthetic solution (see recipe)
  • Mice
  • PIXImus densitometer (0.18 × 0.18 mm; GE Medical Systems)
  • Computer with Windows PIXImus software (e.g., version 1.4x; GE Medical Systems)
  • Container to accommodate mouse during weighing
  • Scale accurate to 0.1 g
  • 1‐ml syringe with 25‐G, 0.5‐mm needle
  • Specimen tray for PIXImus (GE Medical Systems)

Alternate Protocol 1: Determination of Body Composition by Quantitative Nuclear Magnetic Resonance

  Materials
  • Mice
  • Control/calibration sample (Bruker)
  • Bruker Minispec NMR analyzer (http://www.minispec.com/mq/mice.html), interfaced with personal computer running MS Windows 2000 or higher with Minispec software package installed

Basic Protocol 4: Indirect Calorimetry by Oxymax

  Materials
  • Mice
  • Container to accommodate mouse during weighing
  • Scale (capable of weighing 1 to 50 g to an accuracy of 0.1 g)
  • Respirometer: Oxymax system (Columbus Instruments; http://www.respirometer.com/microoxy.html)
  • Computer with the Windows Oxymax software
  • Calibration gases: one tank with 20.5% O 2/0.8% CO 2 and a second tank with 100% N 2
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Figures

Videos

Literature Cited

Literature Cited
   Champy, M., Selloum, M., Piard, L., Zietler, V., Caradec, C., Chambon, P., and Auwerx, J. 2004. Mouse functional genomics requires standardization of mouse handling and housing conditions. Mamm. Genome 15:768‐783.
   Chen, A. and Innis, S. 2004. Assessment of phospholipid malabsorption by quantification of fecal phospholipid. J. Pediatr. Gastroenterol. Nutr. 39:85‐91.
   Cock, T.A., Back, J., Elefteriou, F., Karsenty, G., Kastner, P., Chan, S., and Auwerx, J. 2004. Enhanced bone formation in lipodystrophic PPARgamma(hyp/hyp) mice relocates haematopoiesis to the spleen. EMBO Rep. 5:1007‐1012.
   Coppari, R., Ichinose, M., Lee, C.E., Pullen, A.E., Kenny, C.D., McGovern, R.A., Tang, V., Liu, S.M., Ludwig, T., Chua, S.C. Jr, Lowell, B.B., and Elmquist, J.K. 2005. The hypothalamic arcuate nucleus: A key site for mediating leptin's effects on glucose homeostasis and locomotor activity. Cell. Metab. 1:63‐72.
   Flecknell, P.A. 1993. Anesthesia and perioperative care. Methods Enzymol. 225:16‐33.
   Folch, J., Ascoli, I., Lees, M., Meath, J.A., and LeBaron, N. 1951. Preparation of lipid extracts from brain tissue. J. Biol. Chem. 191:833‐841.
   Gordon, C.J. 1993. Temperature Regulation in Laboratory Rodents. Cambridge University Press, Cambridge, U.K.
   Grier, S., Turner, A., and Alvis, M. 1996. The use of dual‐energy X‐ray adsorptiometry in animals. Invest. Radiol. 31:50‐62.
   Havel, P.J. 2002. Control of energy homeostasis and insulin action by adipocyte hormones: Leptin, acylation stimulating protein, and adiponectin. Curr. Opin. Lipidol. 13:51‐59.
   Hrabe de Angeles, M., Flaswinkel, H., Fuchs, H., Rathkolb, B., Soewarto, D., Marschall, S., Heffner, S., Pargent, W., Wuensch, K., Jung, M., Reis, A., Richter, T., Alessandrini, F., Jakob, T., Fuchs, E., Kolb, H., Kremmer, E., Schaeble, K., Rollinski, B., Roscher, A., Peters, C., Meitinger, T., Strom, T., Steckler, T., Holsboer, F., Klopstock, T., Gekeler, F., Schindewolf, C., Jung, T., Avraham, K., Behrendt, H., Ring, J., Zimmer, A., Schughart, K., Pfeffer, K., Wolf, E., and Balling, R. 2000. Genome‐wide, large‐scale production of mutant mice by ENU mutagenesis. Nat. Genet. 25:444‐447.
   Ishibashi, S., Schwarz, M., Frykman, P.K., Herz, J., and Russell, D.W. 1996. Disruption of cholesterol 7α‐hydroxylase gene in mice. I. Postnatal lethality reversed by bile acid and vitamin supplementation. J. Biol. Chem. 271:18017‐18023.
   Koutnikova, H., Cock, T.A., Watanabe, M., Houten, S.M., Champy, M.F., Dierich, A., and Auwerx, J. 2003. Compensation by the muscle limits the metabolic consequences of lipodystrophy in PPAR gamma hypomorphic mice. Proc. Natl. Acad. Sci. U.S.A. 100:14457‐14462.
   Lowell, B.B. and Spiegelman, B.M. 2000. Towards a molecular understanding of adaptive thermogenesis. Nature 404:652‐660.
   Nagy, T. and Clair, A.L. 2000. Precision and accuracy of dual‐energy X‐ray absorptiometry for determining in vivo body composition of mice. Obes. Res. 8:392‐398.
   Nolan, P.M., Peters, J., Strivens, M., Rogers, D., Hagan, J., Spurr, N., Gray, I.C., Vizor, L., Brooker, D., Whitehill, E., Washbourne, R., Hough, T., Greenaway, S., Hewitt, M., Liu, X., McCormack, S., Pickford, K., Selley, R., Wells, C., Tymowska‐Lalanne, Z., Roby, P., Glenister, P., Thornton, C., Thaung, C., Stevenson, J.A., Arkell, R., Mburu, P., Hardisty, R., Kiernan, A., Erven, A., Steel, K.P., Voegeling, S., Guenet, J.L., Nickols, C., Sadri, R., Nasse, M., Isaacs, A., Davies, K., Browne, M., Fisher, E.M., Martin, J., Rastan, S., Brown, S.D., and Hunter, J. 2000. A systematic, genome‐wide, phenotype‐driven mutagenesis programme for gene function studies in the mouse. Nat. Genet. 25:440‐443.
   Picard, F., Annicotte, J., Rocchi, S., Champy, M.F., O'Malley, B.W., Chambon, P., and Auwerx, J. 2002. SRC1 and TIF2 control energy balance between white and brown adipose tissues. Cell 111:931‐941.
   Porter, R.K. 2001. Allometry of mammalian cellular oxygen consumption. Cell. Mol. Life Sci. 58:815‐822.
   Schwarz, M., Russell, D.W., Dietschy, J.M., and Turley, S.D. 2001. Alternate pathways of bile acid synthesis in the cholesterol 7α‐hydroxylase knockout mouse are not upregulated by either cholesterol or cholestyramine feeding. J. Lipid Res. 42:1594‐1603.
   Spiegelman, B.M. and Flier, J.S. 2001. Obesity and the regulation of energy balance. Cell 104:531‐543.
   Taicher, G.Z., Tinsley, F.C., Reiderman, A., and Heiman, M.L. 2003. Quantitative magnetic resonance (QMR) method for bone and whole‐body‐composition analysis. Anal. Bioanal. Chem. 377:990‐1002.
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library