Cellular and Molecular Neurobiology

, Volume 8, Issue 1, pp 129–138 | Cite as

Dual regulation of ACTH secretion by guanine nucleotides in permeabilized AtT-20 cells

  • Alberto Luini
  • Maria Antonietta De Matteis


  1. 1.

    We have examined the effects of guanine nucleotides on ACTH secretion from digitonin-permeabilized AtT-20 cells, with the aim of analyzing the involvement of GTP-binding proteins (G proteins) in the secretory process.

  2. 2.

    AtT-20 cells permeabilized with 20µM digitonin displayed calciumdependent secretion. The EC50 of calcium was ~2µM and the maximal stimulation was 350% of basal release.

  3. 3.

    Nonhydrolyzable guanine nucleotides also stimulated ACTH release, in a virtually Ca2+-free medium. The EC50 of guanosine 5′-(3-O-thio)triphosphate (GTP γ S) was ~15µM and the maximal stimulation was ~230% of basal release. The effects of calcium and guanine nucleotides were not additive.

  4. 4.

    In the presence of the inhibitory hormone, somatostatin guanine nucleotides inhibited the calcium-stimulated secretion.

  5. 5.

    Both the stimulatory and the inhibitory effects on secretion of guanine nucleotides were independent of changes in cyclic AMP (cAMP) and calcium. It is suggested that G proteins influence an unknown step in the secretion process, which would be near or at the exocytotic site.

  6. 6.

    The results can be explained by assuming the existence of two types of G proteins, one with stimulatory effects on exocytotic release (GeS) and another with inhibitory effects (GeI).


Key words

GTP-binding proteins calcium exocytosis guanine nucleotides adrenocorticotropic hormone (ACTH) AtT-20 cells 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Axelrod, J., and Reisine T. D. (1984). Stress hormones: Their interaction and regulation.Science 224452–459.PubMedCrossRefGoogle Scholar
  2. Barrowman, M. M., Cockcroft, S., and Gomperts, B. D. (1986). Two roles for guanine nucleotides in the stimulus-secretion sequence of neutrophils.Nature 319504–507.PubMedCrossRefGoogle Scholar
  3. Bittner, M. A., Holz, R. W., and Neubig, R. R. (1986). Guanine nucleotide effects on catecholamine secretion from digitonin-permeabilized adrenal chromaffin cells.J. Biol. Chem. 26110182–10188.PubMedGoogle Scholar
  4. Bourne, H. R. (1986). One molecular machine can transduce diverse signals.Nature 321814–816.PubMedCrossRefGoogle Scholar
  5. Burch, R. M., Luini, A., and Axelrod, J. (1986). Phospholipase A2 and phospholipase C are activated by distinct GTP-binding proteins in response toα 1-adrenergic stimulation in FRTL5 thyroid cells.Proc. Natl. Acad. Sci USA 837201–7205.PubMedCrossRefGoogle Scholar
  6. Dunn, L. A., and Holz, R. W. (1983). Catecholamine secretion from digitonin-treated adrenal medullary chromaffin cells.J. Biol. Chem. 2584989–4993.PubMedGoogle Scholar
  7. Gierschik, P., Falloon, J., Milligan, G., Pines, M., Gallin, J. I., and Spiegel, A. (1986). Immunochemical evidence for a novel pertussis toxin substrate in human neutrophils.J. Biol. Chem. 2618058–8062.PubMedGoogle Scholar
  8. Gilman, A. G. (1984). G proteins and dual control of adenylate cyclase.Cell 36577–579.PubMedCrossRefGoogle Scholar
  9. Gomperts, B. D. (1986). Calcium shares the limelight in stimulus-secretion coupling.Trends Biochem. Sci. 11290.CrossRefGoogle Scholar
  10. Heisler, S. (1984). 12-O-Tetradecanoyl-phorbol-13-acetate-induced ACTH secretion in pituitary tumor cells.Eur. J. Pharmacol. 98177–183.PubMedCrossRefGoogle Scholar
  11. Heisler, S., Reisine, T. D., Hook, V. Y. H., and Axelrod, J. (1982). Somatostatin inhibits multireceptor stimulation of cyclic AMP formation and corticotropin secretion in mouse pituitary tumor cells.Proc. Natl. Acad. Sci. USA 796502–6506.PubMedCrossRefGoogle Scholar
  12. Hildebrandt, J. D., Hanoune, J., and Birnbaumer, L. (1982). Guanine nucleotide inhibition of cyc-S49 mouse lymphoma cell membrane adenylyl cyclase.J. Biol. Chem. 25714723–14725.PubMedGoogle Scholar
  13. Hook, V. Y. H., Heisler, S., Sabol, S. L., and Axelrod, J. (1982). Corticotropin releasing factor stimulates adrenocorticotropin andβ-endorphin release from AtT-20 mouse pituitary tumor cells.Biochem. Biophys. Res. Commun. 1061364–1371.PubMedCrossRefGoogle Scholar
  14. Illes, P. (1986). Mechanisms of receptor-mediated modulation of transmitter release in noradrenergic cholinergic and sensory neurones.Neuroscience 17909–928.PubMedCrossRefGoogle Scholar
  15. Irving, H. R., and Exton, J. H. (1987). Phosphatidylcholine breakdown in rat liver plasma membranes. Roles of guanine nucleotides and P2-purinergic agonists.J. Biol. Chem. 2623440–3443.PubMedGoogle Scholar
  16. Katada, T., Oinuma, M., and Ui, M. (1986a). Two guanine nucleotide-binding proteins in rat brain serving as the specific substrate of islet-activating protein, pertussis toxin. Interaction of theα-subunits withβ γ-subunits in development of their biological activities.J. Biol. Chem. 2618182–8191.PubMedGoogle Scholar
  17. Katada, T., Oinuma, M., and Ui, M. (1986b). Mechanisms for inhibition of the catalytic activity of adenylate cyclase by the guanine nucleotide-binding proteins serving as the substrate of islet-activating protein, pertussis toxin.J. Biol. Chem. 2615215–5221.PubMedGoogle Scholar
  18. Knight, D. E., and Baker, P. F. (1982). Calcium-dependence of catecholamine release from bovine adrenal medullary cells after exposure to intense electric fields.Membrane Biol. 68107–140.CrossRefGoogle Scholar
  19. Knight, D. E., and Baker, P. F. (1985). Guanine nucleotides and Ca-dependent exocytosis. Studies on two adrenal cell preparations.FEBS Lett. 189345.PubMedCrossRefGoogle Scholar
  20. Lewis, D. L., Weight, F. F., and Luini, A. (1986). A guanine nucleotide-binding protein mediates the inhibition of voltage-dependent calcium current by somatostatin in a pituitary cell line.Proc. Natl. Acad. Sci. USA 839035–9039.PubMedCrossRefGoogle Scholar
  21. Luini, A., and Axelrod, J. (1985). Inhibitors of the cytochrome P-450 enzymes block the secretagogue-induced release of corticotropin in mouse pituitary tumor cells.Proc. Natl. Acad. Sci. USA 821012–1014.PubMedCrossRefGoogle Scholar
  22. Luini, A., Lewis, D., Guild, S., Corda, D., and Axelrod, J. (1985). Hormone secretagogues increase cytosolic calcium by increasing cAMP in corticotropin-secreting cells.Proc. Natl. Acad. Sci. USA 828034–8038.PubMedCrossRefGoogle Scholar
  23. Metz, S., VanRollins, M., Strife, R., Fujimoto, W., and Robertson, R. P. (1983). Lipoxygenase pathway in islet endocrine cells. Oxidative metabolism of arachidonic acid promotes insulin release.J. Clin. Invest. 711191–1205.PubMedCrossRefGoogle Scholar
  24. Miyazaki, K., Reisine, T., and Kebabian, J. W. (1984). Adenosine 3′,5′-monophosphate (cAMP)-dependent protein kinase activity in rodent pituitary tissue: Possible role in cAMP-dependent hormone secretion.Endocrinology 1151933–1945.PubMedCrossRefGoogle Scholar
  25. Nishizuka, Y. (1986). Studies and perspectives of protein kinase C.Science 233305–312.PubMedCrossRefGoogle Scholar
  26. Oetting, M., LeBoff, M., Swiston, L., Preston, J., and Brown, E. (1986). Guanine nucleotides are potent secretagogues in permeabilized parathyroid cells.FEBS Lett. 20899–104.PubMedCrossRefGoogle Scholar
  27. Okano, Y., Yamada, K., Yano, K., and Nozawa, Y. (1987). Guanosine 5′-(γ-thio) triphosphate stimulates arachidonic acid liberation in permeabilized rat peritoneal mast cells.Biochem. Biophys. Res. Commun. 1451267–1275.PubMedCrossRefGoogle Scholar
  28. Portzehl, H., Caldwell, P. C., and Ruegg, J. C. (1964). The dependence of contraction and relaxation of muscle fibers from the crab Maia squinado on the internal concentration of free calcium ions.Biochem. Biophys. Acta 79581–591.PubMedGoogle Scholar
  29. Reisine, T. (1985). Multiple mechanisms of somatostatin inhibition of adrenocorticotropin release from mouse anterior pituitary tumor cells.Endocrinology 1162259–2266.PubMedCrossRefGoogle Scholar
  30. Terman, B. I., Slivka, S. R., Hughes, R. J., and Insel, P. A. (1987).α 1-Adrenergic receptor-linked guanine nucleotide-binding protein in muscle and kidney epithelial cells.Mol. Pharmacol. 3112–20.PubMedGoogle Scholar
  31. Vallar, L., Biden, T. J., and Wollheim, C. B. (1987). Guanine nucleotides induce Ca2+-independent insulin secretion from permeabilized RINm5F cells.J. Biol. Chem. 2625049–5056.PubMedGoogle Scholar
  32. Yamamoto, T., Furuki, Y., Guild, S., and Kebabian, J. W. (1987). Adenosine 3′,5′-cyclic monophosphate stimulates secretion ofα-melanocyte-stimulating hormone from permeabilized cells of the intermediate lobe of the rat pituitary gland.Biochem. Biophys. Res. Commun. 1431076–1084.PubMedCrossRefGoogle Scholar
  33. Zatz, M., and Reisine, T. D. (1985). Lithium induces corticotropin secretion and desensitization in cultured anterior pituitary cells.Proc. Natl. Acad. Sci. USA 821286–1290.PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Publishing Corporation 1988

Authors and Affiliations

  • Alberto Luini
    • 1
  • Maria Antonietta De Matteis
    • 1
  1. 1.Neuroendocrinology UnitIstituto di Ricerche Farmacologiche Mario NegriS. Maria ImbaroItaly

Personalised recommendations