Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1988 Jun;7(6):1627–1633. doi: 10.1002/j.1460-2075.1988.tb02989.x

Cyclic AMP-independent, dual regulation of voltage-dependent Ca2+ currents by LHRH and somatostatin in a pituitary cell line.

W Rosenthal 1, J Hescheler 1, K D Hinsch 1, K Spicher 1, W Trautwein 1, G Schultz 1
PMCID: PMC457146  PMID: 2458919

Abstract

Voltage-dependent Ca2+ currents appear to be involved in the actions of hormones that regulate pituitary secretion. In order to investigate modulation of Ca2+ currents by release-inducing and release-inhibiting hormones, we performed whole-cell clamp experiments in the pituitary cell line GH3. The resting potential was approximately -40 mV; spontaneous action potentials were observed in the majority of cells. Superfusion of cells with the stimulatory hormone, LHRH, depolarized the plasma membrane to approximately -10 mV, whereas the inhibitory hormone, somatostatin, caused hyperpolarization to approximately -60 mV; both hormones suppressed spontaneous action potentials. Under voltage clamp conditions, GH3 cells exhibited slowly and fast inactivating Ca2+ currents. LHRH increased whereas somatostatin decreased the slowly inactivating currents; fast inactivating currents were not affected by these hormones. The stimulatory effect of LHRH was not mimicked by intracellularly applied cAMP. In contrast to vasoactive intestinal peptide and forskolin, LHRH did not activate adenylate cyclase in membranes of GH3 cells, but rather appeared to cause inhibition of the enzyme. Hormonal stimulation and inhibition of inward currents were abolished by pretreatment of the cells with pertussis toxin. In membranes of GH3 cells, we identified a pertussis toxin-sensitive G-protein of the Gi-type and Go. We conclude that LHRH and somatostatin modulate voltage-dependent Ca2+ currents via cAMP-independent mechanisms involving pertussis toxin-sensitive G-proteins. The occurrence of both pertussis toxin-sensitive hormonal stimulation and inhibition of voltage-dependent Ca2+ currents in one cell type suggest that these opposite regulations are mediated by distinct G-proteins.

Full text

PDF
1627

Images in this article

1629Image
on p.1629
1630Image
on p.1630

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aktories K., Schultz G., Jakobs K. H. Adenylate cyclase inhibition and GTPase stimulation by somatostatin in S49 lymphoma cyc- variants are prevented by islet-activating protein. FEBS Lett. 1983 Jul 11;158(1):169–173. doi: 10.1016/0014-5793(83)80701-7. [DOI] [PubMed] [Google Scholar]
  2. Armstrong C. M., Matteson D. R. Two distinct populations of calcium channels in a clonal line of pituitary cells. Science. 1985 Jan 4;227(4682):65–67. doi: 10.1126/science.2578071. [DOI] [PubMed] [Google Scholar]
  3. Armstrong D., Eckert R. Voltage-activated calcium channels that must be phosphorylated to respond to membrane depolarization. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2518–2522. doi: 10.1073/pnas.84.8.2518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cassel D., Pfeuffer T. Mechanism of cholera toxin action: covalent modification of the guanyl nucleotide-binding protein of the adenylate cyclase system. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2669–2673. doi: 10.1073/pnas.75.6.2669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chang J. P., McCoy E. E., Graeter J., Tasaka K., Catt K. J. Participation of voltage-dependent calcium channels in the action of gonadotropin-releasing hormone. J Biol Chem. 1986 Jul 15;261(20):9105–9108. [PubMed] [Google Scholar]
  6. Codina J., Grenet D., Yatani A., Birnbaumer L., Brown A. M. Hormonal regulation of pituitary GH3 cell K+ channels by Gk is mediated by its alpha-subunit. FEBS Lett. 1987 May 25;216(1):104–106. doi: 10.1016/0014-5793(87)80765-2. [DOI] [PubMed] [Google Scholar]
  7. Conn P. M., Rogers D. C., Seay S. G. Structure-function relationship of calcium ion channel antagonists at the pituitary gonadotrope. Endocrinology. 1983 Nov;113(5):1592–1595. doi: 10.1210/endo-113-5-1592. [DOI] [PubMed] [Google Scholar]
  8. Draznin B., Steiner C., Sherman N., Sussman K. Somatostatin inhibits fusion of pituitary secretion vesicles with the plasma membranes. Biochem Biophys Res Commun. 1986 Sep 14;139(2):673–678. doi: 10.1016/s0006-291x(86)80043-2. [DOI] [PubMed] [Google Scholar]
  9. Gross R. A., Macdonald R. L. Dynorphin A selectively reduces a large transient (N-type) calcium current of mouse dorsal root ganglion neurons in cell culture. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5469–5473. doi: 10.1073/pnas.84.15.5469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hagiwara S., Ohmori H. Studies of single calcium channel currents in rat clonal pituitary cells. J Physiol. 1983 Mar;336:649–661. doi: 10.1113/jphysiol.1983.sp014603. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F. J. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 1981 Aug;391(2):85–100. doi: 10.1007/BF00656997. [DOI] [PubMed] [Google Scholar]
  12. Hamprecht B., Glaser T., Reiser G., Bayer E., Propst F. Culture and characteristics of hormone-responsive neuroblastoma X glioma hybrid cells. Methods Enzymol. 1985;109:316–341. doi: 10.1016/0076-6879(85)09096-6. [DOI] [PubMed] [Google Scholar]
  13. Hescheler J., Kameyama M., Trautwein W. On the mechanism of muscarinic inhibition of the cardiac Ca current. Pflugers Arch. 1986 Aug;407(2):182–189. doi: 10.1007/BF00580674. [DOI] [PubMed] [Google Scholar]
  14. Hescheler J., Rosenthal W., Hinsch K. D., Wulfern M., Trautwein W., Schultz G. Angiotensin II-induced stimulation of voltage-dependent Ca2+ currents in an adrenal cortical cell line. EMBO J. 1988 Mar;7(3):619–624. doi: 10.1002/j.1460-2075.1988.tb02855.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hescheler J., Rosenthal W., Trautwein W., Schultz G. The GTP-binding protein, Go, regulates neuronal calcium channels. 1987 Jan 29-Feb 4Nature. 325(6103):445–447. doi: 10.1038/325445a0. [DOI] [PubMed] [Google Scholar]
  16. Hess P., Lansman J. B., Tsien R. W. Different modes of Ca channel gating behaviour favoured by dihydropyridine Ca agonists and antagonists. Nature. 1984 Oct 11;311(5986):538–544. doi: 10.1038/311538a0. [DOI] [PubMed] [Google Scholar]
  17. Holz G. G., 4th, Rane S. G., Dunlap K. GTP-binding proteins mediate transmitter inhibition of voltage-dependent calcium channels. Nature. 1986 Feb 20;319(6055):670–672. doi: 10.1038/319670a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Huff R. M., Axton J. M., Neer E. J. Physical and immunological characterization of a guanine nucleotide-binding protein purified from bovine cerebral cortex. J Biol Chem. 1985 Sep 5;260(19):10864–10871. [PubMed] [Google Scholar]
  19. Johnson R. A., Walseth T. F. The enzymatic preparation of [alpha-32P]ATP, [alpha-32P]GTP, [32P]cAMP, and [32P]cGMP, and their use in the assay of adenylate and guanylate cyclases and cyclic nucleotide phosphodiesterases. Adv Cyclic Nucleotide Res. 1979;10:135–167. [PubMed] [Google Scholar]
  20. Kameyama M., Hescheler J., Hofmann F., Trautwein W. Modulation of Ca current during the phosphorylation cycle in the guinea pig heart. Pflugers Arch. 1986 Aug;407(2):123–128. doi: 10.1007/BF00580662. [DOI] [PubMed] [Google Scholar]
  21. Kameyama M., Hofmann F., Trautwein W. On the mechanism of beta-adrenergic regulation of the Ca channel in the guinea-pig heart. Pflugers Arch. 1985 Oct;405(3):285–293. doi: 10.1007/BF00582573. [DOI] [PubMed] [Google Scholar]
  22. Koch B. D., Blalock J. B., Schonbrunn A. Characterization of the cyclic AMP-independent actions of somatostatin in GH cells. I. An increase in potassium conductance is responsible for both the hyperpolarization and the decrease in intracellular free calcium produced by somatostatin. J Biol Chem. 1988 Jan 5;263(1):216–225. [PubMed] [Google Scholar]
  23. Kojima I., Shibata H., Ogata E. Pertussis toxin blocks angiotensin II-induced calcium influx but not inositol trisphosphate production in adrenal glomerulosa cell. FEBS Lett. 1986 Aug 18;204(2):347–351. doi: 10.1016/0014-5793(86)80841-9. [DOI] [PubMed] [Google Scholar]
  24. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  25. Lewis D. L., Weight F. F., Luini A. A guanine nucleotide-binding protein mediates the inhibition of voltage-dependent calcium current by somatostatin in a pituitary cell line. Proc Natl Acad Sci U S A. 1986 Dec;83(23):9035–9039. doi: 10.1073/pnas.83.23.9035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Luini A., Lewis D., Guild S., Schofield G., Weight F. Somatostatin, an inhibitor of ACTH secretion, decreases cytosolic free calcium and voltage-dependent calcium current in a pituitary cell line. J Neurosci. 1986 Nov;6(11):3128–3132. doi: 10.1523/JNEUROSCI.06-11-03128.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Luqman W. A., Matej L. A., Smith M. L. Comparison of prolactin levels in human semen and seminal plasma. J Endocrinol. 1979 Apr;81(1):131–133. doi: 10.1677/joe.0.0810131. [DOI] [PubMed] [Google Scholar]
  28. Marian J., Conn P. M. Gonadotropin releasing hormone stimulation of cultured pituitary cells requires calcium. Mol Pharmacol. 1979 Jul;16(1):196–201. [PubMed] [Google Scholar]
  29. McArdle C. A., Huckle W. R., Conn P. M. Phorbol esters reduce gonadotrope responsiveness to protein kinase C activators but not to Ca2+-mobilizing secretagogues. Does protein kinase C mediate gonadotropin-releasing hormone action? J Biol Chem. 1987 Apr 15;262(11):5028–5035. [PubMed] [Google Scholar]
  30. McCleskey E. W., Fox A. P., Feldman D. H., Cruz L. J., Olivera B. M., Tsien R. W., Yoshikami D. Omega-conotoxin: direct and persistent blockade of specific types of calcium channels in neurons but not muscle. Proc Natl Acad Sci U S A. 1987 Jun;84(12):4327–4331. doi: 10.1073/pnas.84.12.4327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Milligan G., Gierschik P., Spiegel A. M., Klee W. A. The GTP-binding regulatory proteins of neuroblastoma x glioma, NG108-15, and glioma, C6, cells. Immunochemical evidence of a pertussis toxin substrate that is neither Ni nor No. FEBS Lett. 1986 Jan 20;195(1-2):225–230. doi: 10.1016/0014-5793(86)80165-x. [DOI] [PubMed] [Google Scholar]
  32. Mumby S. M., Kahn R. A., Manning D. R., Gilman A. G. Antisera of designed specificity for subunits of guanine nucleotide-binding regulatory proteins. Proc Natl Acad Sci U S A. 1986 Jan;83(2):265–269. doi: 10.1073/pnas.83.2.265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Naor Z., Azrad A., Limor R., Zakut H., Lotan M. Gonadotropin-releasing hormone activates a rapid Ca2+-independent phosphodiester hydrolysis of polyphosphoinositides in pituitary gonadotrophs. J Biol Chem. 1986 Sep 25;261(27):12506–12512. [PubMed] [Google Scholar]
  34. Nowycky M. C., Fox A. P., Tsien R. W. Three types of neuronal calcium channel with different calcium agonist sensitivity. Nature. 1985 Aug 1;316(6027):440–443. doi: 10.1038/316440a0. [DOI] [PubMed] [Google Scholar]
  35. Reuter H. Calcium channel modulation by neurotransmitters, enzymes and drugs. Nature. 1983 Feb 17;301(5901):569–574. doi: 10.1038/301569a0. [DOI] [PubMed] [Google Scholar]
  36. Rosenthal W., Binder T., Schultz G. NADP efficiently inhibits endogenous but not pertussis toxin-catalyzed covalent modification of membrane proteins incubated with NAD. FEBS Lett. 1987 Jan 26;211(2):137–143. doi: 10.1016/0014-5793(87)81424-2. [DOI] [PubMed] [Google Scholar]
  37. Rosenthal W., Koesling D., Rudolph U., Kleuss C., Pallast M., Yajima M., Schultz G. Identification and characterization of the 35-kDa beta subunit of guanine-nucleotide-binding proteins by an antiserum raised against transducin. Eur J Biochem. 1986 Jul 15;158(2):255–263. doi: 10.1111/j.1432-1033.1986.tb09745.x. [DOI] [PubMed] [Google Scholar]
  38. Schimmer B. P. Isolation of ACTH-resistant Y1 adrenal tumor cells. Methods Enzymol. 1985;109:350–356. doi: 10.1016/0076-6879(85)09099-1. [DOI] [PubMed] [Google Scholar]
  39. Schlegel W., Winiger B. P., Mollard P., Vacher P., Wuarin F., Zahnd G. R., Wollheim C. B., Dufy B. Oscillations of cytosolic Ca2+ in pituitary cells due to action potentials. Nature. 1987 Oct 22;329(6141):719–721. doi: 10.1038/329719a0. [DOI] [PubMed] [Google Scholar]
  40. Scott R. H., Dolphin A. C. Regulation of calcium currents by a GTP analogue: potentiation of (-)-baclofen-mediated inhibition. Neurosci Lett. 1986 Aug 15;69(1):59–64. doi: 10.1016/0304-3940(86)90414-3. [DOI] [PubMed] [Google Scholar]
  41. Sternweis P. C., Robishaw J. D. Isolation of two proteins with high affinity for guanine nucleotides from membranes of bovine brain. J Biol Chem. 1984 Nov 25;259(22):13806–13813. [PubMed] [Google Scholar]
  42. Suki W. N., Abramowitz J., Mattera R., Codina J., Birnbaumer L. The human genome encodes at least three non-allellic G proteins with alpha i-type subunits. FEBS Lett. 1987 Aug 10;220(1):187–192. doi: 10.1016/0014-5793(87)80900-6. [DOI] [PubMed] [Google Scholar]
  43. Suzuki N., Yoshioka T. Differential blocking action of synthetic omega-conotoxin on components of Ca2+ channel current in clonal GH3 cells. Neurosci Lett. 1987 Mar 31;75(2):235–239. doi: 10.1016/0304-3940(87)90303-x. [DOI] [PubMed] [Google Scholar]
  44. Ui M., Katada T., Murayama T., Kurose H., Yajima M., Tamura M., Nakamura T., Nogimori K. Islet-activating protein, pertussis toxin: a specific uncoupler of receptor-mediated inhibition of adenylate cyclase. Adv Cyclic Nucleotide Protein Phosphorylation Res. 1984;17:145–151. [PubMed] [Google Scholar]
  45. Yajima Y., Akita Y., Saito T. Pertussis toxin blocks the inhibitory effects of somatostatin on cAMP-dependent vasoactive intestinal peptide and cAMP-independent thyrotropin releasing hormone-stimulated prolactin secretion of GH3 cells. J Biol Chem. 1986 Feb 25;261(6):2684–2689. [PubMed] [Google Scholar]
  46. Yamashita N., Shibuya N., Ogata E. Hyperpolarization of the membrane potential caused by somatostatin in dissociated human pituitary adenoma cells that secrete growth hormone. Proc Natl Acad Sci U S A. 1986 Aug;83(16):6198–6202. doi: 10.1073/pnas.83.16.6198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Yatani A., Codina J., Sekura R. D., Birnbaumer L., Brown A. M. Reconstitution of somatostatin and muscarinic receptor mediated stimulation of K+ channels by isolated GK protein in clonal rat anterior pituitary cell membranes. Mol Endocrinol. 1987 Apr;1(4):283–289. doi: 10.1210/mend-1-4-283. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES