Perifusion model system to culture bovine hypothalamic slices in series with dispersed anterior pituitary cells

  • H. A. Hassan
  • R. A. Merkel
Cellular Models

Summary

Dispersed bovine anterior pituitary cells were incubated either in static or perifusion cultures to assess basal growth hormone release as well as stimulatory and inhibitory effects of growth hormone-releasing hormone and somatostatin, respectively, on growth hormone release. Total concentrations of growth hormones over a 12-hour incubation period were fivefold greater in perifused than in static cultures (2034 ± 160 vs. 387 ± 33 ng/12 h). A dose-dependent increase in growth hormone secretion in response to challenge with growth hormone-releasing hormone (10−12 to 10−8M) for 1 h was observed in both static and perifusion cultures; however, perifused cells were more responsive to the same concentration of neuropeptide than those in static culture. Concentrations of somatostatin (10−12 to 10−8M) for 1 h did not inhibit basal growth hormone secretion in either static or perifusion cultures. To establish model, slices of the hypothalamus, immediately adjacent to the sagittal midline, were perifused in series with anterior pituitary cells, and media effluent was assayed for growth hormone concentrations. Release of growth hormone was pulsatile and seemed to mimic the episodic pattern of bovine secretion. Hypothalamic slices were placed in one chamber of the perifusion system, and basal secretion of growth hormone-releasing hormone and somatostatin was pulsatile in media effluent. Tissue viability of hypothalamic slices and anterior pituitary cells was evaluated by KCl depolarization. Tissues were viable for at least 120 h. Thus, this hypothalamo-pituitary dual chamber perifusion system is a valid in vitro model to study regulation of growth hormone secretion.

Key words

perifusion system hypothalamic slices anterior pituitary cells bovine 

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References

  1. 1.
    Baes, M.; Denef, C. Evidence that stimulation of growth hormone release by epinephrine and vasoactive intestinal peptide is based on cell-to-cell communication in the pituitary. Endocrinology 120:280–290; 1987.PubMedGoogle Scholar
  2. 2.
    Becker, K.; Conway, S. A novel hypothalamic-dispersed pituitary coperifusion model for the study of growth hormone secretion. Brain Res. 578:107–114; 1992.PubMedCrossRefGoogle Scholar
  3. 3.
    Brazeau, P.; Ling, N.; Bohlen, P., et al. Growth hormone releasing factor, somatocrinin, release pituitary growth hormone in vitro. Proc. Natl. Acad. Sci. USA 79:7909–7913; 1982.PubMedCrossRefGoogle Scholar
  4. 4.
    Brazeau, P.; Vale, W.; Burgus, B., et al. Hypothalamic peptide that inhibits the secretion of immunoreactive pituitary growth hormone. Science 179:77–79; 1973.PubMedCrossRefGoogle Scholar
  5. 5.
    Brownstein, M.; Arimura, A.; Sato, H., et al. The regional distribution of somatostatin in the rat brain. Endocrinology 96:1456–1461; 1975.PubMedGoogle Scholar
  6. 6.
    Cuttler, L.; Glaum, S. R.; Collins, B. A., et al. Calcium signalling in single growth hormone-releasing factor-responsive pituitary cells. Endocrinology 130:945–953; 1992.PubMedCrossRefGoogle Scholar
  7. 7.
    Fagin, J. A.; Brown, A.; Melmed, S. Regulation of pituitary insulin-like growth factor-I messenger ribonucleic acid levels in rats harboring somatomammotropic tumors: implications for growth hormone autoregulation. Endocrinology 122:2204–2210; 1988.PubMedGoogle Scholar
  8. 8.
    Folkman, J.; Moscona, A. Role of cell shape in growth control. Nature 273:347–349; 1978.CrossRefGoogle Scholar
  9. 9.
    Glenn, K. C. Regulation of release of somatotropin from in vitro cultures of bovine and porcine pituitary cells. Endocrinology 118:2450–2457; 1986.PubMedGoogle Scholar
  10. 10.
    Goodyer, C. G.; DeStephano, L.; Guyda, H. J., et al. Effects of insulinlike growth factors on adult male rat pituitary function in tissue culture. Endocrinology 115:1568–1576; 1984.PubMedGoogle Scholar
  11. 11.
    Guillemin, R. Peptides in the brain: the new endocrinology of neurons. Science 202:390–402; 1978.PubMedCrossRefGoogle Scholar
  12. 12.
    Guillemin, R.; Brazeau, P.; Bohlen, P., et al. Somatocrinin, the growth hormone releasing factor. Rec. Prog. Horm. Res. 40:233–299; 1984.PubMedGoogle Scholar
  13. 13.
    Ham, R. G. Nutritional requirements of primary cultures. In Vitro 10:119–125; 1974.PubMedCrossRefGoogle Scholar
  14. 14.
    Hassan, H. A. Effects of testosterone and estradiol-17β on growth hormone release from dispersed bovine anterior pituitary cells in vitro. East Lansing: Michigan State Univ.; 1990. Thesis.Google Scholar
  15. 15.
    Hassan, H. A.; Merkel, R. A.; Enright, W. J., et al. Androgens modulate growth hormone-releasing factor-induced GH release from bovine anterior pituitary cells in static culture. Domest. Anim. Endocrinol. 9:37–45; 1992.CrossRefGoogle Scholar
  16. 16.
    Heiman, M. L.; Nekola, M. V.; Murphy, W. A., et al. An extremely sensitive in vitro model for elucidating structure-activity relationships of growth hormone-releasing factor analogs. Endocrinology 116:410–415; 1985.PubMedGoogle Scholar
  17. 17.
    Henquin, J. C. Relative importance of extracellular and intracellular calcium for the two phases of glucose-stimulated insulin release: studies of the theophylline. Endocrinology 102:723–730; 1978.PubMedGoogle Scholar
  18. 18.
    Horng, C. B.; McLimans, W. Primary suspension culture of calf anterior pituitary cells on a microcarrier surface. Biotechnol. Bioeng. 17:713–732; 1975.CrossRefGoogle Scholar
  19. 19.
    Lapp, C. A.; Stachura, M. E.; Tyler, J. M., et al. GH3 cell secretion of growth hormone and prolactin increases spontaneously during perifusion. In Vitro Cell. Dev. Biol. 23:686–690; 1987.PubMedCrossRefGoogle Scholar
  20. 20.
    Leidy, J. W.; Robbins, J. Regional distribution of human growth hormone-releasing hormone in the human hypothalamus by radioimmunoassay. J. Clin. Endocrinol. & Metab. 62:372–378; 1986.CrossRefGoogle Scholar
  21. 21.
    McDonald, J. A. Matrix regulation of cell shape and gene expression. Curr. Opin. Cell. Biol. 1:995–999; 1989.PubMedCrossRefGoogle Scholar
  22. 22.
    Merriam, G. R.; Wachter, K. W. Algorithms for the study of episodic hormone secretion. Am. J. Physiol. 243:E310-E318; 1982.PubMedGoogle Scholar
  23. 23.
    Moss, G. E.; Adams, T. E.; Niswender, G. D., et al. Effects of parturition and suckling on concentrations of pituitary gonadotropins, hypothalamic GnRH and pituitary responsiveness to GnRH in ewes. J. Anim. Sci. 50:496–502; 1980.PubMedGoogle Scholar
  24. 24.
    Ohlsson, L.; Lindstrom, P. The correlation between calcium outflow and growth hormone release in perifused rat somatotrophs. Endocrinology 126:488–497; 1990.PubMedGoogle Scholar
  25. 25.
    Oosterom, R.; Verlenn, T.; Zuierwijz, J., et al. GH secretion by cultured rat anterior pituitary cells: effects of culture conditions and dexamethasone. Endocrinology 113:735–741; 1983.PubMedGoogle Scholar
  26. 26.
    Padmanabhan, V.; Enright, W. J.; Zinn, S. A., et al. Modulation of growth hormone-releasing factor-induced release of growth hormone from bovine pituitary cells. Domest. Anim. Endocrinol. 4:243–252; 1987.PubMedCrossRefGoogle Scholar
  27. 27.
    Plotsky, P. M.; Vale, W. Patterns of growth hormone-releasing factor and somatostatin secretion into hypophyseal-portal circulation of the rat. Science 230:461–463; 1985.PubMedCrossRefGoogle Scholar
  28. 28.
    Purchas, R. W.; Macmillan, K. L.; Hafs, H. D. Pituitary and plasma growth hormone levels in bulls from birth to one year of age. J. Anim. Sci. 31:358–363; 1970.PubMedGoogle Scholar
  29. 29.
    Richardson, S. B.; Twente, S. Pre-exposure of rat anterior pituitary cells to somatostatin enhances subsequent growth hormone secretion. J. Endocrinol. 128:91–95; 1991.PubMedGoogle Scholar
  30. 30.
    Rosenthal, S. M.; Silverman, B. L.; Wehrenberg, W. B. Exogenous growth hormone inhibits bovine but not murine pituitary growth hormone secretion in vitro: evidence against negative feedback in rat pituitary cells. Endocrinology 122:1511–1514; 1991.Google Scholar
  31. 31.
    Schoenemann, H. M.; Humphrey, W. D.; Crowder, M. E., et al. Pituitary luteinizing hormone-releasing hormone receptors in ovariectomized cows after challenge with ovarian steroids. Biol. Reprod. 32:574–583; 1985.PubMedCrossRefGoogle Scholar
  32. 32.
    Stachura, M. E.; Tyler, J. M.; Farmer, P. K. Combined effects of human growth hormone (GH)-releasing factor-44 (GRF) and somatostatin (SRIF) on post-SRIF rebound release of GH and prolactin: a model for GRF-SRIF modulation of secretion. Endocrinology 123:1476–1482; 1988.PubMedGoogle Scholar
  33. 33.
    Stachura, M. E.; Tyler, J. M.; Kent, P. G. Effect of growth hormone-releasing factor-44 upon release of concurrently synthesized hormone by perifused rat pituitary tissue. Endocrinology 119:1245–1253; 1986.PubMedCrossRefGoogle Scholar
  34. 34.
    Stern, J. E.; Mitchell, T.; Herzberg, V. L., et al. Secretion of vasopressin, oxytocin and two neurophysins from rat hypothalamo-neurohypophyseal explants in organ culture. Neuroendocrinology 43:252–258; 1986.PubMedGoogle Scholar
  35. 35.
    Tsagarakis, F. G. S.; Rees, L. H.; Besser, G. M., et al. Relationship between growth hormone-releasing hormone and somatostatin in the rat: effects of age and sex on content and in vitro release from the hypothalamic explants. J. Endocrinol. 123:53–58; 1989.PubMedCrossRefGoogle Scholar
  36. 36.
    Vale, W.; Vaughan, J.; Jolley, D., et al. Assay of growth hormone-releasing factor. Methods Enzymol. 124:389–401; 1986.PubMedCrossRefGoogle Scholar

Copyright information

© Tissue Culture Association 1994

Authors and Affiliations

  • H. A. Hassan
    • 1
  • R. A. Merkel
    • 1
  1. 1.Growth Biology Program, Department of Animal Science, and Food Science and Human NutritionMichigan State UniversityEast Lansing

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