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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1991 Aug 1;88(15):6790–6794. doi: 10.1073/pnas.88.15.6790

Refractoriness to growth hormone is associated with increased intracellular calcium in rat adipocytes.

Y Schwartz 1, H M Goodman 1, H Yamaguchi 1
PMCID: PMC52174  PMID: 1862102

Abstract

In adipocytes that have been deprived of growth hormone (GH) for at least 3 hr, GH elicits a transient insulin-like response that is followed by a period of refractoriness to further insulin-like stimulation. Exposure of adipocytes to GH in the first hour of a 3-hr incubation prevents the appearance of insulin-like sensitivity. Intracellular Ca2+ concentration [( Ca2+]i) was measured in individual adipocytes that were loaded with fura-2 hexakis(acetoxymethyl) ester after preincubation in the presence (refractory) or absence (sensitive) of recombinant human GH at 100 ng/ml. Using a dual nitrogen laser imaging microscope with computer-assisted image processing to measure fluorescence changes, we observed that resting [Ca2+]i was 220 +/- 10 nM in refractory adipocytes and 110 +/- 6 nM in sensitive adipocytes (P less than 0.001). GH had no acute effect on [Ca2+]i in sensitive adipocytes but caused a sustained 3-fold increase in [Ca2+]i in refractory cells within 3 min (P less than 0.001). Insulin did not change [Ca2+]i in either sensitive or refractory adipocytes. In refractory cells treated with insulin and GH simultaneously, insulin completely blocked the rise in [Ca2+]i due to GH. Oxytocin elicited a prompt increase in [Ca2+]i followed by a quick return to resting levels in both sensitive and refractory cells. These findings indicate that basal [Ca2+]i is increased in refractory cells and that GH produces a sustained rise in [Ca2+]i only in refractory adipocytes. We suggest that the sustained increase in [Ca2+]i produced by GH in refractory cells prevents the expression of the insulin-like response.

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Selected References

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  1. Augert G., Exton J. H. Insulin and oxytocin effects on phosphoinositide metabolism in adipocytes. J Biol Chem. 1988 Mar 15;263(8):3600–3609. [PubMed] [Google Scholar]
  2. Birnbaum R. S., Goodman H. M. Studies on the mechanism of the antilipolytic effects of growth hormone. Endocrinology. 1976 Nov;99(5):1336–1345. doi: 10.1210/endo-99-5-1336. [DOI] [PubMed] [Google Scholar]
  3. Black B. L., Jarett L., McDonald J. M. The regulation of endoplasmic reticulum calcium uptake of adipocytes by cytoplasmic calcium. J Biol Chem. 1981 Jan 10;256(1):322–329. [PubMed] [Google Scholar]
  4. Blinks J. R., Wier W. G., Hess P., Prendergast F. G. Measurement of Ca2+ concentrations in living cells. Prog Biophys Mol Biol. 1982;40(1-2):1–114. doi: 10.1016/0079-6107(82)90011-6. [DOI] [PubMed] [Google Scholar]
  5. Cherqui G., Caron M., Wicek D., Lascols O., Capeau J., Picard J. Insulin stimulation of glucose metabolism in rat adipocytes: possible implication of protein kinase C. Endocrinology. 1986 May;118(5):1759–1769. doi: 10.1210/endo-118-5-1759. [DOI] [PubMed] [Google Scholar]
  6. Davidson M. B. Effect of growth hormone on carbohydrate and lipid metabolism. Endocr Rev. 1987 May;8(2):115–131. doi: 10.1210/edrv-8-2-115. [DOI] [PubMed] [Google Scholar]
  7. Draznin B., Lewis D., Houlder N., Sherman N., Adamo M., Garvey W. T., LeRoith D., Sussman K. Mechanism of insulin resistance induced by sustained levels of cytosolic free calcium in rat adipocytes. Endocrinology. 1989 Nov;125(5):2341–2349. doi: 10.1210/endo-125-5-2341. [DOI] [PubMed] [Google Scholar]
  8. Draznin B., Sussman K., Kao M., Lewis D., Sherman N. The existence of an optimal range of cytosolic free calcium for insulin-stimulated glucose transport in rat adipocytes. J Biol Chem. 1987 Oct 25;262(30):14385–14388. [PubMed] [Google Scholar]
  9. Edén S., Schwartz J., Kostyo J. L. Effects of preincubation on the ability of rat adipocytes to bind and respond to growth hormone. Endocrinology. 1982 Nov;111(5):1505–1512. doi: 10.1210/endo-111-5-1505. [DOI] [PubMed] [Google Scholar]
  10. Glynn B. P., Colliton J. W., McDermott J. M., Witters L. A. Phorbol esters, but not insulin, promote depletion of cytosolic protein kinase C in rat adipocytes. Biochem Biophys Res Commun. 1986 Mar 28;135(3):1119–1125. doi: 10.1016/0006-291x(86)91044-2. [DOI] [PubMed] [Google Scholar]
  11. Goodman H. M., Coiro V. Induction of sensitivity to the insulin-like action of growth hormone in normal rat adipose tissue. Endocrinology. 1981 Jan;108(1):113–119. doi: 10.1210/endo-108-1-113. [DOI] [PubMed] [Google Scholar]
  12. Goodman H. M. Effects of growth hormone on the lipolytic response of adipose tissue to theophylline. Endocrinology. 1968 May;82(5):1027–1034. doi: 10.1210/endo-82-5-1027. [DOI] [PubMed] [Google Scholar]
  13. Goodman H. M. Growth hormone and the metabolism of carbohydrate and lipid in adipose tissue. Ann N Y Acad Sci. 1968 Feb 5;148(2):419–440. doi: 10.1111/j.1749-6632.1968.tb20367.x. [DOI] [PubMed] [Google Scholar]
  14. Gorin E., Goodman H. M. Turnover of growth hormone receptors in rat adipocytes. Endocrinology. 1985 May;116(5):1796–1805. doi: 10.1210/endo-116-5-1796. [DOI] [PubMed] [Google Scholar]
  15. Graves C. B., Gale R. D., Laurino J. P., McDonald J. M. The insulin receptor and calmodulin. Calmodulin enhances insulin-mediated receptor kinase activity and insulin stimulates phosphorylation of calmodulin. J Biol Chem. 1986 Aug 5;261(22):10429–10438. [PubMed] [Google Scholar]
  16. Grichting G., Levy L. K., Goodman H. M. Relationship between binding and biological effects of human growth hormone in rat adipocytes. Endocrinology. 1983 Sep;113(3):1111–1120. doi: 10.1210/endo-113-3-1111. [DOI] [PubMed] [Google Scholar]
  17. Grynkiewicz G., Poenie M., Tsien R. Y. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem. 1985 Mar 25;260(6):3440–3450. [PubMed] [Google Scholar]
  18. KNOBIL E., BEST G. R., GOODMAN H. M. The hypoglycemic action of simian and bovine growth hormone in hypophysectomized rhesus monkeys. Endocrinology. 1961 Apr;68:723–725. doi: 10.1210/endo-68-4-723. [DOI] [PubMed] [Google Scholar]
  19. Kelly K. L., Deeney J. T., Corkey B. E. Cytosolic free calcium in adipocytes. Distinct mechanisms of regulation and effects on insulin action. J Biol Chem. 1989 Aug 5;264(22):12754–12757. [PubMed] [Google Scholar]
  20. Konishi M., Olson A., Hollingworth S., Baylor S. M. Myoplasmic binding of fura-2 investigated by steady-state fluorescence and absorbance measurements. Biophys J. 1988 Dec;54(6):1089–1104. doi: 10.1016/S0006-3495(88)83045-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lee H. M., Fain J. N. Regulation of oxytocin-induced phosphoinositide breakdown in adipocytes by adenosine, isoproterenol and insulin. Biochim Biophys Acta. 1989 Sep 4;1013(1):73–79. doi: 10.1016/0167-4889(89)90130-4. [DOI] [PubMed] [Google Scholar]
  22. McDonald J. R., Lawrence J. C., Jr Identification of an adipocyte protein that binds to calmodulin in the absence of Ca2+ and is phosphorylated in response to insulin and tumor-promoting phorbol esters. J Biol Chem. 1989 Jun 5;264(16):9611–9618. [PubMed] [Google Scholar]
  23. Moody A. J., Stan M. A., Stan M., Gliemann J. A simple free fat cell bioassay for insulin. Horm Metab Res. 1974 Jan;6(1):12–16. doi: 10.1055/s-0028-1093895. [DOI] [PubMed] [Google Scholar]
  24. Moore E. D., Becker P. L., Fogarty K. E., Williams D. A., Fay F. S. Ca2+ imaging in single living cells: theoretical and practical issues. Cell Calcium. 1990 Feb-Mar;11(2-3):157–179. doi: 10.1016/0143-4160(90)90068-6. [DOI] [PubMed] [Google Scholar]
  25. Ozaki H., Satoh T., Karaki H., Ishida Y. Regulation of metabolism and contraction by cytoplasmic calcium in the intestinal smooth muscle. J Biol Chem. 1988 Oct 5;263(28):14074–14079. [PubMed] [Google Scholar]
  26. Pershadsingh H. A., Landt M., McDonald J. M. Calmodulin-sensitive ATP-dependent Ca2+ transport across adipocyte plasma membranes. J Biol Chem. 1980 Oct 10;255(19):8983–8986. [PubMed] [Google Scholar]
  27. Pershadsingh H. A., Lee L. Y., Snowdowne K. W. Evidence for a sodium/calcium exchanger and voltage-dependent calcium channels in adipocytes. FEBS Lett. 1989 Feb 13;244(1):89–92. doi: 10.1016/0014-5793(89)81169-x. [DOI] [PubMed] [Google Scholar]
  28. RABEN M. S. Growth hormone. 1. Physiologic aspects. N Engl J Med. 1962 Jan 4;266:31–35. doi: 10.1056/NEJM196201042660109. [DOI] [PubMed] [Google Scholar]
  29. RODBELL M. METABOLISM OF ISOLATED FAT CELLS. I. EFFECTS OF HORMONES ON GLUCOSE METABOLISM AND LIPOLYSIS. J Biol Chem. 1964 Feb;239:375–380. [PubMed] [Google Scholar]
  30. Scanlon M., Williams D. A., Fay F. S. A Ca2+-insensitive form of fura-2 associated with polymorphonuclear leukocytes. Assessment and accurate Ca2+ measurement. J Biol Chem. 1987 May 5;262(13):6308–6312. [PubMed] [Google Scholar]
  31. Schwartz Y., Goodman H. M. Refractoriness to the insulin-like effects of growth hormone depends upon calcium. Endocrinology. 1990 Jul;127(1):170–176. doi: 10.1210/endo-127-1-170. [DOI] [PubMed] [Google Scholar]
  32. Smal J., Closset J., Hennen G., De Meyts P. Receptor binding properties and insulin-like effects of human growth hormone and its 20 kDa-variant in rat adipocytes. J Biol Chem. 1987 Aug 15;262(23):11071–11079. [PubMed] [Google Scholar]
  33. Smal J., De Meyts P. Role of kinase C in the insulin-like effects of human growth hormone in rat adipocytes. Biochem Biophys Res Commun. 1987 Sep 30;147(3):1232–1240. doi: 10.1016/s0006-291x(87)80202-4. [DOI] [PubMed] [Google Scholar]
  34. Smal J., De Meyts P. Sphingosine, an inhibitor of protein kinase C, suppresses the insulin-like effects of growth hormone in rat adipocytes. Proc Natl Acad Sci U S A. 1989 Jun;86(12):4705–4709. doi: 10.1073/pnas.86.12.4705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Thomas A. P., Martin-Requero A., Williamson J. R. Interactions between insulin and alpha 1-adrenergic agents in the regulation of glycogen metabolism in isolated hepatocytes. J Biol Chem. 1985 May 25;260(10):5963–5973. [PubMed] [Google Scholar]
  36. Uto A., Arai H., Ogawa Y. Reassessment of Fura-2 and the ratio method for determination of intracellular Ca2+ concentrations. Cell Calcium. 1991 Jan;12(1):29–37. doi: 10.1016/0143-4160(91)90082-p. [DOI] [PubMed] [Google Scholar]
  37. Williams D. A., Fogarty K. E., Tsien R. Y., Fay F. S. Calcium gradients in single smooth muscle cells revealed by the digital imaging microscope using Fura-2. Nature. 1985 Dec 12;318(6046):558–561. doi: 10.1038/318558a0. [DOI] [PubMed] [Google Scholar]

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