Advertisement

Purinergic signalling in rat GFSHR-17 granulosa cells: an in vitro model of granulosa cells in maturing follicles

  • Willem Bintig
  • Judith Baumgart
  • Wilhelm J. Walter
  • Alexander Heisterkamp
  • Holger Lubatschowski
  • Anaclet Ngezahayo
Article

Abstract

Purinergic signalling in rat GFSHR-17 granulosa cells was characterised by Ca2+-imaging and perforated patch-clamp. We observed a resting intracellular Ca2+-concentration ([Ca2+]i) of 100 nM and a membrane potential of −40 mV. This was consistent with high K+− and Cl permeability and a high intracellular Cl concentration of 40 mM. Application of ATP for 5–15 s every 3 min induced repeated [Ca2+]i increases and a 30 mV hyperpolarization. The phospholipase C inhibitor U73122 or the IP3-receptor antagonist 2-aminoethoethyl diphenyl borate suppressed ATP responses. Further biochemical and pharmacological experiments revealed that ATP responses were related to stimulation of P2Y2 and P2Y4 receptors and that the [Ca2+]i increase was a prerequisite for hyperpolarization. Inhibitors of Ca2+-activated channels or K+ channels did not affect the ATP-evoked responses. Conversely, inhibitors of Cl channels hyperpolarized cells to −70 mV and suppressed further ATP-evoked hyperpolarization. We propose that P2Y2 and P2Y4 receptors in granulosa cells modulate Cl permeability by regulating Ca2+-release.

Keywords

Granulosa cells Purinergic receptors Cl channels Follicle maturation Perforated patch-clamp Ca2+-imaging 

References

  1. Abbracchio MP, Burnstock G, Boeynaems JM, Barnard EA, Boyer JL, Kennedy C, Knight GE, Fumagalli M, Gachet C, Jacobson KA, Weisman GA (2006) Pharmacol Rev 58:281–341CrossRefGoogle Scholar
  2. Aguado LI (2002) Microsc Res Tech 59:462–473CrossRefGoogle Scholar
  3. Arellano RO, Martinez-Torres A, Garay E (2002) Biol Reprod 67:837–846CrossRefGoogle Scholar
  4. Bintig W, Buchholz V, Schlie S, Baumgart J, Heisterkamp A, Ngezahayo A (2007) Acta Physiologica 188(Suppl. 653):P10Google Scholar
  5. Burnstock G (2007a) Physiol Rev 87:659–797CrossRefGoogle Scholar
  6. Burnstock G (2007b) Cell Mol Life Sci 64:1471–1483CrossRefGoogle Scholar
  7. Burnstock G, Williams MJ (2002) Pharmacol Exp Ther 295:862–869Google Scholar
  8. Gomes P, Srinivas SP, Van Driessche W, Vereecke J, Himpens B (2005) Invest Ophthalmol Vis Sci 46:1208–1216CrossRefGoogle Scholar
  9. Grynkiewicz G, Poenie M, Tsien RY (1985) J Biol Chem 260:3440–3450Google Scholar
  10. Keren TI, Dantes Sprengel AR, Amsterdam A (1993) Mol Cell Endocrinol 95:R1–R10CrossRefGoogle Scholar
  11. Kim SJ, Shin SY, Lee JE, Kim JH, Uhm DY (2003) Prostate 55:118–127CrossRefGoogle Scholar
  12. King BF, Townsend-Nicholson A (2003) Tocris Reviews 23:1–12Google Scholar
  13. King BF, Townsend-Nicholson A (2008) J Pharmacol Exp Ther 324:1055–1063CrossRefGoogle Scholar
  14. Ngezahayo A, Altmann B, Kolb HA (2003) J Membr Biol 194:165–176CrossRefGoogle Scholar
  15. Romanello M, Pani B, Bicego M, D’Andrea P (2001) Biochem Biophys Res Commun 289:1275–1281CrossRefGoogle Scholar
  16. Sage CL, Marcus DC (2002) J Membr Biol 185:103–115CrossRefGoogle Scholar
  17. Tai CJ, Kang SK, Cheng KW, Choi KC, Nathwani PS, Leung PCK (2000) J Clin Endocrinol Metab 85:591–597CrossRefGoogle Scholar
  18. Tong D, Li TY, Naus KE, Bai D, Kidder GM (2007) J Cell Sci 120:4016–4024CrossRefGoogle Scholar
  19. von Kügelgen I (2000) Naunyn Schmiedebergs Arch Pharmacol 362:310–323CrossRefGoogle Scholar
  20. von Kügelgen I (2006) Pharmacol Ther 110:415–432CrossRefGoogle Scholar
  21. White PJ, Webb TE, Boarder MR (2003) Mol Pharmacol 63:1356–1363CrossRefGoogle Scholar
  22. Wildman SS, Unwin RJ, King BF (2003) Br J Pharmacol 140:1177–1186CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Willem Bintig
    • 1
  • Judith Baumgart
    • 2
  • Wilhelm J. Walter
    • 3
  • Alexander Heisterkamp
    • 2
  • Holger Lubatschowski
    • 2
  • Anaclet Ngezahayo
    • 1
  1. 1.Institute of BiophysicsLeibniz University HannoverHannoverGermany
  2. 2.Laser Zentrum Hannover e.V.HannoverGermany
  3. 3.Molecular and Cell PhysiologyHannover Medical SchoolHannoverGermany

Personalised recommendations