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Confocal Microscopy of Adipocytes

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Adipose Tissue Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 155))

Abstract

The isolated rat adipose cell experimental system is the principal model for studies of the mechanism of insulin’s stimulatory action on glucose transport (reviewed in refs. 1–4). Despite the successful use of this preparation in biochemical studies, the unique structure of the adipose cell, with its large central triglyceride storage droplet (80 μm) and thin (1–2 μm) rim of cytoplasm, has caused special problems for morphological approaches. Several methods have been used for studies of protein localization and trafficking by immunocytochemistry (ICC) at both light microscopy (LM) and electron microscopy (EM) levels.

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References

  1. Simpson, I. A. and Cushman, S. W. (1986) Hormonal regulation of mammalian glucose transport. Annu. Rev. Biochem. 55, 1059–1089.

    Article  PubMed  CAS  Google Scholar 

  2. Birnbaum, M. J. (1992) The insulin-sensitive glucose transporter. Int. Rev. Cyto. 137A, 239–297.

    Article  CAS  Google Scholar 

  3. Holman, G. D. and Cushman, S. W. (1994) Subcellular localization and trafficking of the GLUT4 glucose transporter isoform in insulin-responsive cells. Bioessays 10, 753–759.

    Article  Google Scholar 

  4. Stephens, J. M. and Pilch, P. F. (1995) The metabolic regulation and vesicular transport of GLUT4, the major insulin-responsive glucose transporter. Endocr. Rev. 16, 529–546.

    PubMed  CAS  Google Scholar 

  5. Slot, J. W., Geuze, H. J., Gigengack, S., Lienhard, G. E., and James, D. E. (1991) Immuno-localization of the insulin regulatable glucose transporter in brown adipose tissue of the rat. J. Cell Biol. 113, 123–135.

    Article  PubMed  CAS  Google Scholar 

  6. Smith, R. M., Charron, M. J., Shah, N., Lodish, H. F., and Jarett, L. (1991) Immunoelectron microscopic demonstration of insulin-stimulated translocation of glucose transporters to the plasma membrane of isolated rat adipocytes and masking of the carboxyl-terminal epitope of intracellular GLUT4. Proc. Natl. Acad. Sci. USA 88, 6893–6897.

    Article  PubMed  CAS  Google Scholar 

  7. Blok, J., Gibbs, E. M., Lienhard, G. E., Slot, J. W., and Geuze, H. J. (1988) Insulin-induced translocation of glucose transporters from post-Golgi compartments to the plasma membrane of 3T3-L1 adipocytes. J. Cell Biol. 106, 69–76.

    Article  PubMed  CAS  Google Scholar 

  8. Martin, S., Slot, J. W., and James, D. E. (1999) GLUT4 trafficking in insulin-sensitive cells. A morphological review. Cell Biochem. Biophys. 30, 89–113.

    Article  PubMed  CAS  Google Scholar 

  9. Robinson, L. J., Pang, S., Harris, D. S., Heuser, J., and James, D. E. (1992) Translocation of the glucose transporter (GLUT4) to the cell surface in permeabilized 3T3-L1 adipocytes: effects of ATP, insulin, and GTPγS and localization of GLUT4 to clathrin lattices. J. Cell Biol. 117, 1181–1196.

    Article  PubMed  CAS  Google Scholar 

  10. Voldstedlund, M., Tranum-Jensen, J., and Vinten, J. (1993) Quantitation of Na+/ K+-ATPase and glucose transporter isoform in rat adipocyte plasma membrane by immunogold labeling. J. Membrane Biol. 136, 63–73.

    Article  CAS  Google Scholar 

  11. Weber, T. M., Joost, H. G., Simpson, I. A., and Cushman, S. W. (1988) Methods for assessment of glucose transport activity and the number of glucose transporters in isolated rat adipose cells and membrane fractions, in The Insulin Receptor Part B, (Kahn, C. R. and Harrison, L. C., eds.), Alan R Liss, New York, NY, pp. 171–187.

    Google Scholar 

  12. Omatsu-Kanbe, M., Zarnowski, M. J., and Cushman, S. W. (1996) Hormonal regulation of glucose transport in a brown adipose cell preparation isolated from rats that show a large response to insulin. Biochem. J. 315, 25–31.

    PubMed  CAS  Google Scholar 

  13. Malide, D., Dwyer, N. K., Blanchette-Mackie, E. J., and Cushman, S. W. (1997) Immunocytochemical evidence that GLUT4 resides in a specialized translocation post-endosomal, VAMP-positive compartment in rat adipose cells in the absence of insulin. J. Histochem. Cytochem. 8, 1083–1096.

    Article  Google Scholar 

  14. Malide, D. and Cushman, S. W. (1997) Morphological effects of wortmannin on the endosomal system and GLUT4-containing compartments in rat adipose cells. J. Cell Sci. 110, 2795–2806.

    PubMed  CAS  Google Scholar 

  15. Malide, D., St-Denis, J-F., Keller, S. R., and Cushman S. W. (1997) Vp165 and GLUT4 share similar vesicle pools along their trafficking pathways in rat adipose cells. FEBS Lett. 409, 461–468.

    Article  PubMed  CAS  Google Scholar 

  16. Barr, V. A., Malide, D., Zarnowski, M. J., Taylor, S. I., and Cushman, S. W. (1997) Insulin stimulates both leptin secretion and production by rat white adipose tissue. Endocrinology 138, 4463–4472.

    Article  PubMed  CAS  Google Scholar 

  17. Malide, D., Yewdell J. W., Bennink, J. R., and Cushman, S. W. MHC class I and GLUT4 define two distinct insulin-responsive compartments and exocytic pathways in rat adipose cells. In review.

    Google Scholar 

  18. Matsumoto, B., ed. (1993) Cell Biological Applications of Confocal Microscopy: Methods in Cell Biology, vol. 38, Academic Press, San Diego, CA.

    Google Scholar 

  19. Pawley, J., ed. (1995) The Handbook of Biological Confocal Microscopy. IMR Press, Madison, WI.

    Google Scholar 

  20. Oatey, P. B., Van Weering, D. H., Dobson, S. P., Gould, G. W., and Tavare, J. M. (1997) GLUT4 vesicle dynamics in living 3T3-L1 adipocytes visualized with green-fluorescent protein. Biochem. J. 327, 637–642.

    PubMed  CAS  Google Scholar 

  21. Venkateswarlu, K., Oatey, P. B., Tavare, J. M., and Cullen, P. J. (1998) Insulin-dependent translocation of ARNO to the plasma membrane of adipocytes requires phosphatidylinositol 3-kinase. Curr. Biol. 8, 463–466.

    Article  PubMed  CAS  Google Scholar 

  22. Blanchette-Mackie, E. J., Dwyer, N. K., Barber, T., Coxey, R. A., Takeda, T., Rondione, C. M., Theodorakis, J. L., Greenberg, A. S., and Londos, C. (1995) Perilipin is located on the surface layer of intracellular lipid droplets in adipocytes. J. Lipid Res. 36, 1211–1226.

    PubMed  CAS  Google Scholar 

  23. Martin, S., Reaves, B., Banting, G., and Gould, G. W. (1994) Analysis of the colocalization of the insulin-responsive glucose transporter (GLUT4) and the trans-Golgi network marker TGN38 within 3T3-L1 adipocytes. Biochem. J. 300, 743–749.

    PubMed  CAS  Google Scholar 

  24. Chakrabarti, R., Buxton, J., Joly, M., and Corvera, S. (1994) Insulin-sensitive association of GLUT-4 with endocytic clathrin-coated vesicles revealed with the use of Brefeldin-A. J. Biol. Chem. 269, 7926–7933.

    PubMed  CAS  Google Scholar 

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Author information

Authors and Affiliations

  1. EDMNS/DB/NIDDK/NIH, Bethesda, MD, USA

    Daniela Malide

Authors
  1. Daniela Malide
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Editor information

Editors and Affiliations

  1. Université de Nice-Sophia Antipolis, Nice, France

    Gérard Ailhaud

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© 2001 Humana Press Inc.

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Malide, D. (2001). Confocal Microscopy of Adipocytes. In: Ailhaud, G. (eds) Adipose Tissue Protocols. Methods in Molecular Biology™, vol 155. Springer, Totowa, NJ. https://doi.org/10.1385/1-59259-231-7:053

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  • DOI: https://doi.org/10.1385/1-59259-231-7:053

  • Publisher Name: Springer, Totowa, NJ

  • Print ISBN: 978-0-89603-747-2

  • Online ISBN: 978-1-59259-231-9

  • eBook Packages: Springer Protocols

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