Neurotherapeutics

, Volume 9, Issue 3, pp 658–672

Inactivation of the Constitutively Active Ghrelin Receptor Attenuates Limbic Seizure Activity in Rodents

  • Jeanelle Portelli
  • Leen Thielemans
  • Luc Ver Donck
  • Ellen Loyens
  • Jessica Coppens
  • Najat Aourz
  • Jeroen Aerssens
  • Katia Vermoesen
  • Ralph Clinckers
  • Anneleen Schallier
  • Yvette Michotte
  • Dieder Moechars
  • Graham L. Collingridge
  • Zuner A. Bortolotto
  • Ilse Smolders
Original Article

Abstract

Ghrelin is a pleiotropic neuropeptide that has been recently implicated in epilepsy. Animal studies performed to date indicate that ghrelin has anticonvulsant properties; however, its mechanism of anticonvulsant action is unknown. Here we show that the anticonvulsant effects of ghrelin are mediated via the growth hormone secretagogue receptor (GHSR). To our surprise, however, we found that the GHSR knockout mice had a higher seizure threshold than their wild-type littermates when treated with pilocarpine. Using both in vivo and in vitro models, we further discovered that inverse agonism and desensitization/internalization of the GHSR attenuate limbic seizures in rats and epileptiform activity in hippocampal slices. This constitutes a novel mechanism of anticonvulsant action, whereby an endogenous agonist reduces the activity of a constitutively active receptor.

Keywords

Epilepsy Seizures Ghrelin GHSR Hippocampus Pilocarpine 

Supplementary material

13311_2012_125_MOESM1_ESM.pdf (511 kb)
ESM 1(PDF 510 kb)

References

  1. 1.
    Kojima M, Hosoda H, Date Y, et al. Ghrelin is a growth-hormone-releasing acylated peptide from stomach. Nature 1999;402:656-660.PubMedCrossRefGoogle Scholar
  2. 2.
    Diano S, Farr SA, Benoit SC, et al. Ghrelin controls hippocampal spine synapse density and memory performance. Nat Neurosci 2006;9:381-388.PubMedCrossRefGoogle Scholar
  3. 3.
    Andrews ZB, Liu ZW, Wallingford N, et al. UCP2 mediates ghrelin's action on NPY/AgRP neurons by lowering free radicals. Nature 2008;454:846-851.PubMedCrossRefGoogle Scholar
  4. 4.
    Angelidis G, Valotassiou V, Georgoulias P. Current and potential roles of ghrelin in clinical practice. J Endocrinol Invest 2010;33:823-838.PubMedGoogle Scholar
  5. 5.
    Camina JP. Cell biology of the ghrelin receptor. J Neuroendocrinol 2006;18:65-76.PubMedCrossRefGoogle Scholar
  6. 6.
    Thielemans L, Peeters PJ, Jonckheere H, et al. The hepatocarcinoma cell line HepG2 does not express a GHS-R1a-type ghrelin receptor. J Recept Signal Transduct Res 2007;27:309-322.PubMedCrossRefGoogle Scholar
  7. 7.
    Chen L, Xing T, Wang M, et al. Local infusion of ghrelin enhanced hippocampal synaptic plasticity and spatial memory through activation of phosphoinositide 3-kinase in the dentate gyrus of adult rats. Eur J Neurosci 2011;33:266-275.PubMedCrossRefGoogle Scholar
  8. 8.
    Davis JF, Choi DL, Clegg DJ, Benoit SC. Signaling through the ghrelin receptor modulates hippocampal function and meal anticipation in mice. Physiol Behav 2011;103:39-43.PubMedCrossRefGoogle Scholar
  9. 9.
    Andrews ZB. The extra-hypothalamic actions of ghrelin on neuronal function. Trends Neurosci 2011;34:31-40.PubMedCrossRefGoogle Scholar
  10. 10.
    Lee J, Lim E, Kim Y, Li E, Park S. Ghrelin attenuates kainic acid-induced neuronal cell death in the mouse hippocampus. J Endocrinol 2010;205:263-270.PubMedCrossRefGoogle Scholar
  11. 11.
    Xu J, Wang S, Lin Y et al. Ghrelin protects against cell death of hippocampal neurons in pilocarpine-induced seizures in rats. Neurosci Lett 2009;453:58-61.PubMedCrossRefGoogle Scholar
  12. 12.
    Pitkanen A, Lukasiuk K. Mechanisms of epileptogenesis and potential treatment targets. Lancet Neurol 2011;10:173-186.PubMedCrossRefGoogle Scholar
  13. 13.
    Rogawski MA. Molecular targets versus models for new antiepileptic drug discovery. Epilepsy Res 2006; 68:22-28.PubMedCrossRefGoogle Scholar
  14. 14.
    Robertson CR, Flynn SP, White HS, Bulaj G. Anticonvulsant neuropeptides as drug leads for neurological diseases. Nat Prod Rep 2011;28:741-762.PubMedCrossRefGoogle Scholar
  15. 15.
    Berilgen MS, Mungen B, Ustundag B, Demir C. Serum ghrelin levels are enhanced in patients with epilepsy. Seizure 2006;15:106-111.PubMedCrossRefGoogle Scholar
  16. 16.
    Gungor S, Yucel G, Akinci A, et al. The role of ghrelin in weight gain and growth in epileptic children using valproate. J Child Neurol 2007;22:1384-1388.PubMedCrossRefGoogle Scholar
  17. 17.
    Aydin S, Dag E, Ozkan Y, et al. Time-dependent changes in the serum levels of prolactin, nesfatin-1 and ghrelin as a marker of epileptic attacks young male patients. Peptides 2011;32:1276-1280.PubMedCrossRefGoogle Scholar
  18. 18.
    Greco R, Latini G, Chiarelli F, Iannetti P, Verrotti A. Leptin, ghrelin, and adiponectin in epileptic patients treated with valproic acid. Neurology 2005;65:1808-1809.PubMedCrossRefGoogle Scholar
  19. 19.
    Aydin S, Dag E, Ozkan Y, et al. Nesfatin-1 and ghrelin levels in serum and saliva of epileptic patients: hormonal changes can have a major effect on seizure disorders. Mol Cell Biochem 2009;328:49-56.PubMedCrossRefGoogle Scholar
  20. 20.
    Dag E, Aydin S, Ozkan Y, et al. Alteration in chromogranin A, obestatin and total ghrelin levels of saliva and serum in epilepsy cases. Peptides 2010;31:932-937.PubMedCrossRefGoogle Scholar
  21. 21.
    Prodam F, Bellone S, Casara G, et al. Ghrelin levels are reduced in prepubertal epileptic children under treatment with carbamazepine or valproic acid. Epilepsia 2010;51:312-315.PubMedCrossRefGoogle Scholar
  22. 22.
    Cansu A, Serdaroglu A, Camurdan O, Hirfanoglu T, Cinaz P. Serum insulin, cortisol, leptin, neuropeptide y, galanin and ghrelin levels in epileptic children receiving valproate. Horm Res Paediatr 2011;76:65-71.PubMedCrossRefGoogle Scholar
  23. 23.
    Aslan A, Yildirim M, Ayyildiz M, Guven A, Agar E. The role of nitric oxide in the inhibitory effect of ghrelin against penicillin-induced epileptiform activity in rat. Neuropeptides 2009;43:295-302.PubMedCrossRefGoogle Scholar
  24. 24.
    Biagini G, Torsello A, Marinelli C, et al. Beneficial effects of desacyl-ghrelin, hexarelin and EP-80317 in models of status epilepticus. Eur J Pharmacol 2011;670:130-136.PubMedCrossRefGoogle Scholar
  25. 25.
    Obay BD, Tasdemir E, Tumer C, Bilgin HM, Sermet A. Antiepileptic effects of ghrelin on pentylenetetrazole-induced seizures in rats. Peptides 2007;28:1214-1219.PubMedCrossRefGoogle Scholar
  26. 26.
    Verhulst PJ, De Smet B, Saels I, et al. Role of ghrelin in the relationship between hyperphagia and accelerated gastric emptying in diabetic mice. Gastroenterology 2008;135:1267-1276.PubMedCrossRefGoogle Scholar
  27. 27.
    Portelli J, Aourz N, De Bundel D, et al. Intrastrain differences in seizure susceptibility, pharmacological response and basal neurochemistry of Wistar rats. Epilepsy Res 2009;87:234-246.PubMedCrossRefGoogle Scholar
  28. 28.
    Racine RJ. Modification of seizure activity by electrical stimulation. II. Motor seizure. Electroencephalogr Clin Neurophysiol 1972;32:281-294.PubMedCrossRefGoogle Scholar
  29. 29.
    Meurs A, Clinckers R, Ebinger G, Michotte Y, Smolders I. Seizure activity and changes in hippocampal extracellular glutamate, GABA, dopamine and serotonin. Epilepsy Res 2008;78:50-59.PubMedCrossRefGoogle Scholar
  30. 30.
    De Bundel D, Schallier A, Loyens E, et al. Loss of system x(c)- formula does not induce oxidative stress but decreases extracellular glutamate in hippocampus and influences spatial working memory and limbic seizure susceptibility. J Neurosci 2011;31:5792-5803.PubMedCrossRefGoogle Scholar
  31. 31.
    Bortolotto ZA, Amici M, Anderson WW, Isaac JT, Collingridge GL. Synaptic plasticity in the hippocampal slice preparation. Curr Protoc Neurosci 2011;6:Unit 6.13.Google Scholar
  32. 32.
    Anderson WW, Collingridge GL. Capabilities of the WinLTP data acquisition program extending beyond basic LTP experimental functions. J Neurosci Methods 2007;162:346-356.PubMedCrossRefGoogle Scholar
  33. 33.
    Carpino PA, Lefker BA, Toler SM, et al. Pyrazolinone-piperidine dipeptide growth hormone secretagogues (GHSs). Discovery of capromorelin. Bioorg Med Chem 2003;11:581-590.PubMedCrossRefGoogle Scholar
  34. 34.
    Holst B, Cygankiewicz A, Jensen TH, Ankersen M, Schwartz TW. High constitutive signaling of the ghrelin receptor--identification of a potent inverse agonist. Mol Endocrinol 2003;17:2201-2210.PubMedCrossRefGoogle Scholar
  35. 35.
    Serby MD, Zhao H, Szczepankiewicz BG, et al. 2,4-diaminopyrimidine derivatives as potent growth hormone secretagogue receptor antagonists. J Med Chem 2006;49:2568-2578.PubMedCrossRefGoogle Scholar
  36. 36.
    Bednarek MA, Feighner SD, Pong SS, et al. Structure-function studies on the new growth hormone-releasing peptide, ghrelin: minimal sequence of ghrelin necessary for activation of growth hormone secretagogue receptor 1a. J Med Chem 2000;43:4370-4376.PubMedCrossRefGoogle Scholar
  37. 37.
    Treiman DM. GABAergic mechanisms in epilepsy. Epilepsia 2001;42(suppl 3):8-12.PubMedCrossRefGoogle Scholar
  38. 38.
    Galanopoulou AS. Mutations affecting GABAergic signaling in seizures and epilepsy. Pflugers Arch 2010;460:505-523.PubMedCrossRefGoogle Scholar
  39. 39.
    Cowley MA, Smith RG, Diano S, et al. The distribution and mechanism of action of ghrelin in the CNS demonstrates a novel hypothalamic circuit regulating energy homeostasis. Neuron 2003;37:649-661.PubMedCrossRefGoogle Scholar
  40. 40.
    van Luijtelaar G, Sitnikova E. Global and focal aspects of absence epilepsy: the contribution of genetic models. Neurosci Biobehav Rev 2006;30:983-1003.PubMedCrossRefGoogle Scholar
  41. 41.
    Avoli M. The epileptic hippocampus revisited: back to the future. Epilepsy Curr 2007;7:116-118.PubMedCrossRefGoogle Scholar
  42. 42.
    Damian M, Marie J, Leyris JP, et al. High constitutive activity is an intrinsic feature of the ghrelin receptor protein: a study with a functional monomeric GHS-R1a receptor reconstituted in lipid discs. J Biol Chem 2012;287:3630-3641.PubMedCrossRefGoogle Scholar
  43. 43.
    Pantel J, Legendre M, Cabrol S, et al. Loss of constitutive activity of the growth hormone secretagogue receptor in familial short stature. J Clin Invest 2006;116:760-768.PubMedCrossRefGoogle Scholar
  44. 44.
    Camina JP, Carreira MC, El Messari S, et al. Desensitization and endocytosis mechanisms of ghrelin-activated growth hormone secretagogue receptor 1a. Endocrinology 2004;145:930-940.PubMedCrossRefGoogle Scholar
  45. 45.
    Delhanty PJ, van Kerkwijk A, Huisman M, et al. Unsaturated fatty acids prevent desensitization of the human growth hormone secretagogue receptor by blocking its internalization. Am J Physiol Endocrinol Metab 2010;299:E497-E505.PubMedCrossRefGoogle Scholar
  46. 46.
    Holliday ND, Holst B, Rodionova EA, Schwartz TW, Cox HM. Importance of constitutive activity and arrestin-independent mechanisms for intracellular trafficking of the ghrelin receptor. Mol Endocrinol 2007;21:3100-3112.PubMedCrossRefGoogle Scholar
  47. 47.
    Zhu X, Han X, Blendy JA, Porter BE. Decreased CREB levels suppress epilepsy. Neurobiol Dis 2012;45:253-263.PubMedCrossRefGoogle Scholar
  48. 48.
    Jin W, Sugaya A, Tsuda T, Ohguchi H, Sugaya E. Relationship between large conductance calcium-activated potassium channel and bursting activity. Brain Res 2000;860:21-28.PubMedCrossRefGoogle Scholar
  49. 49.
    Pal S, Sun D, Limbrick D, Rafiq A, DeLorenzo RJ. Epileptogenesis induces long-term alterations in intracellular calcium release and sequestration mechanisms in the hippocampal neuronal culture model of epilepsy. Cell Calcium 2001;30:285-296.PubMedCrossRefGoogle Scholar
  50. 50.
    Cataldi M, Lariccia V, Secondo A, di Renzo G, Annunziato L. The antiepileptic drug levetiracetam decreases the inositol 1,4,5-trisphosphate-dependent [Ca2+]I increase induced by ATP and bradykinin in PC12 cells. J Pharmacol Exp Ther 2005;313:720-730.PubMedCrossRefGoogle Scholar
  51. 51.
    Imazawa M, Kabuto Y, Miyamoto K, Nishimura S, Yagi K. Inositol trisphosphate (IP3) receptors and epileptic seizure. Jpn J Psychiatry Neurol 1989;43:465-468.PubMedGoogle Scholar
  52. 52.
    Zhang B, Wong M. Pentylenetetrazole-induced seizures cause acute, but not chronic, mTOR pathway activation in rat. Epilepsia 2012;53:506-511.PubMedCrossRefGoogle Scholar
  53. 53.
    Yokoyama N, Mori N, Kumashiro H. Chemical kindling induced by cAMP and transfer to electrical kindling. Brain Res 1989;492:158-162.PubMedCrossRefGoogle Scholar
  54. 54.
    Williams SF, Colling SB, Whittington MA, Jefferys JG. Epileptic focus induced by intrahippocampal cholera toxin in rat: time course and properties in vivo and in vitro. Epilepsy Res 1993;16:137-146.PubMedCrossRefGoogle Scholar
  55. 55.
    Chen G, Pan B, Hawver DB, Wright CB, Potter WZ, Manji HK. Attenuation of cyclic AMP production by carbamazepine. J Neurochem 1996;67:2079-2086.PubMedCrossRefGoogle Scholar
  56. 56.
    Higashima M, Ohno K, Koshino Y. Cyclic AMP-mediated modulation of epileptiform afterdischarge generation in rat hippocampal slices. Brain Res 2002;949:157-161.PubMedCrossRefGoogle Scholar
  57. 57.
    Rocha L, Orozco-Suarez S, Alonso-Vanegas M, et al. Temporal lobe epilepsy causes selective changes in mu opioid and nociceptin receptor binding and functional coupling to G-proteins in human temporal neocortex. Neurobiol Dis 2009;35:466-473.PubMedCrossRefGoogle Scholar
  58. 58.
    Caterina MJ, Leffler A, Malmberg AB, et al. Impaired nociception and pain sensation in mice lacking the capsaicin receptor. Science 2000;288:306-313.PubMedCrossRefGoogle Scholar
  59. 59.
    Szallasi A. Vanilloid (capsaicin) receptors in health and disease. Am J Clin Pathol 2002;118:110-121.PubMedCrossRefGoogle Scholar
  60. 60.
    Moran MM, McAlexander MA, Biro T, Szallasi A. Transient receptor potential channels as therapeutic targets. Nat Rev Drug Discov 2011;10:601-620.PubMedCrossRefGoogle Scholar
  61. 61.
    Fehrentz JA, Moulin A, Blayo L, et al. Ghrelin receptor ligands: from peptide to peptidomimetic, design and synthesis to clinical studies. Presented at the Belgian Peptide Group Meeting; February 10, 2012; Brussels.Google Scholar
  62. 62.
    Michel MC, Alewijnse AE. Ligand-directed signaling: 50 ways to find a lover. Mol Pharmacol 2007;72:1097-1099.PubMedCrossRefGoogle Scholar
  63. 63.
    Portelli J, Michotte Y, Smolders I. Ghrelin – an emerging new anticonvulsant neuropeptide. Epilepsia 2012;53:585-595.PubMedCrossRefGoogle Scholar
  64. 64.
    Atcha Z, Chen WS, Ong AB, et al. Cognitive enhancing effects of ghrelin receptor agonists. Psychopharmacology (Berl) 2009;206:415-427.CrossRefGoogle Scholar
  65. 65.
    Carlini VP, Monzon ME, Varas MM, et al. Ghrelin increases anxiety-like behavior and memory retention in rats. Biochem Biophys Res Commun 2002;299:739-743.PubMedCrossRefGoogle Scholar
  66. 66.
    Beck H, Goussakov IV, Lie A, Helmstaedter C, Elger CE. Synaptic plasticity in the human dentate gyrus. J Neurosci 2000;20:7080-7086.PubMedGoogle Scholar

Copyright information

© The American Society for Experimental NeuroTherapeutics, Inc. 2012

Authors and Affiliations

  • Jeanelle Portelli
    • 1
  • Leen Thielemans
    • 2
  • Luc Ver Donck
    • 2
  • Ellen Loyens
    • 1
  • Jessica Coppens
    • 1
  • Najat Aourz
    • 1
  • Jeroen Aerssens
    • 2
  • Katia Vermoesen
    • 1
  • Ralph Clinckers
    • 1
  • Anneleen Schallier
    • 1
  • Yvette Michotte
    • 1
  • Dieder Moechars
    • 2
  • Graham L. Collingridge
    • 3
    • 4
  • Zuner A. Bortolotto
    • 3
  • Ilse Smolders
    • 1
  1. 1.Center for Neurosciences, Department of Pharmaceutical Chemistry, Drug Analysis and Drug InformationVrije Universiteit BrusselBrusselsBelgium
  2. 2.Janssen Research and Development, a Division of Janssen Pharmaceutica NVBeerseBelgium
  3. 3.MRC Centre for Synaptic Plasticity, School of Physiology and PharmacologyUniversity of BristolBristolUnited Kingdom
  4. 4.Department of Brain and Cognitive Sciences, College of Natural SciencesSeoul National UniversitySeoulKorea

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