Blockade of GABAA receptor channels by niflumic acid prevents agonist dissociation

Articles

Abstract

The modulation by the nonsteroidal anti-inflammatory drug niflumic acid (NFA) of the GABAA receptor-mediated currents was studied in acutely isolated cerebellar Purkinje cells using the whole-cell recording and fast drug application system. At concentrations of 3–300 μM NFA potentiated GABA (2 μM)-activated currents, and at concentrations of 1–3 mM NFA blocked these responses. The NFA-induced block was strongly voltage-dependent. Analysis of the voltage dependence of the block suggests that the blocking action of NFA is a result of NFA binding at the site located within GABAA channel pore. The termination of GABA and NFA application was followed by a transient increase of the inward current — “tail” current. These data suggest that NFA acts as a sequential open channel blocker, which prevents dissociation of agonist while the channel is blocked. The tail current develops because, prior to dissociation of agonist, the channels that are in the blocked state must return to the close state via the open state. The tail currents were compared in the presence and absence of gabazine, a competitive antagonist that also allosterically inhibits GABAA receptors. Application of gabazine only during development of tail current did not change neither amplitude nor time course of this current, while noncompetitive antagonists picrotoxin and penicillin blocked it. Protection of tail current from gabazine block indicates that GABA cannot dissociate from the open-blocked state and the agonist was trapped on the receptor while the channel was open. Trapping was specific for the agonist, because the positive allosteric modulator zolpidem (benzodiazepine site agonist) was able to potentiate the tail current in the absence of GABA in the external solution. Our observations provide a model-independent functional support of the hypothesis that open channel block of GABAA channels by NFA prevents an escape of the agonist from its binding sites.

Keywords

GABAA receptors niflumic acid patch-clamp method rat brain neurons 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Jones M.V., Westbrook G.L. 1995. Desensitized states prolong GABAA channel responses to brief agonist pulses. Neuron. 15(1), 181–191.PubMedCrossRefGoogle Scholar
  2. 2.
    Jones M.V., Westbrook G.L. 1996. The impact of receptor desensitization on fast synaptic transmission. Trends Neurosci. 192(3), 96–101.CrossRefGoogle Scholar
  3. 3.
    Bianchi M.T., Macdonald R.L. 2001. Agonist trapping by GABAA receptor channels. J. Neurosci. 21(23), 9083–9091.PubMedGoogle Scholar
  4. 4.
    Kolbaev S.N., Sharonova I.N., Vorobjev V.S., Skrebitsky V.G. 2002. Mechanisms of GABA(A) receptor blockade by millimolar concentrations of furosemide in isolated rat Purkinje cells. Neuropharmacol. 42(7), 913–921.CrossRefGoogle Scholar
  5. 5.
    Vorobjev, V.S. 1991. Vibrodissociation of sliced mammalian nervous tissue. J. Neurosci. Methods. 38, 145–150.PubMedCrossRefGoogle Scholar
  6. 6.
    Vorobjev V.S., Sharonova I.N., Haas H.L. 1996. A simple perfusion system for patch-clamp studies. J. Neurosci. Methods. 68, 303–307.PubMedCrossRefGoogle Scholar
  7. 7.
    MacDonald J.F. Bartlett M.C., Mody I., Pahapill P., Reynolds J.N., Salter M.W., Schneiderman J.H., Pennefather, P.S. 1991. Actions of ketamine, phencyclidine and MK-801 on NMDA receptor currents in cultured mouse hippocampal neurones. J. Physiol. 432, 483–508.PubMedGoogle Scholar
  8. 8.
    MacDonald J.F., Miljkovic Z., Pennefather P. 1987. Use-dependent block of excitatory amino acid currents in cultured neurons by ketamine. J. Neurophysiol. 58, 251–266.PubMedGoogle Scholar
  9. 9.
    Blanpied T.A., Boeckman F.A., Aizenman E., Johnson J.W., 1997. Trapping channel block of NMDA-activated responses by amantadine and memantine. J. Neurophysiol. 77, 309–323.PubMedGoogle Scholar
  10. 10.
    Samoilova M.V., Buldakova S.L., Vorobjev V.S., Sharonova I.N., Magazanik L.G. 1999. The open channel blocking drug, IEM-1460, reveals functionally distinct α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors in rat brain neurons. Neuroscience. 94, 261–268.PubMedCrossRefGoogle Scholar
  11. 11.
    Adams P.R. 1976. Drug blockade of open end-plate channels. J. Physiol. (London). 260, 531–551.Google Scholar
  12. 12.
    Neher E., Steinbach J.H. 1978. Local anaesthetics transiently block currents through single acetylcholinereceptor channels. J. Physiol. (London). 277, 153–176.Google Scholar
  13. 13.
    Neher E., 1983. The charge carried by single-channel currents of rat cultured muscle cells in the presence of local anaesthetics. J. Physiol. (London). 339, 663–678.Google Scholar
  14. 14.
    Antonov S.M., Johnson, J.W. 1996. Voltage-dependent interaction of open-channel blocking molecules with gating of NMDA receptors in rat cortical neurons. J. Physiol. (London). 493, 425–445.Google Scholar
  15. 15.
    Sobolevsky A.I., Koshelev S.G., Khodorov B.I., 1999. Probing of NMDA channels with fast blockers. J. Neurosci. 19, 10611–10626.PubMedGoogle Scholar
  16. 16.
    Vorobjev V.S., Sharonova I.N. 1994. Tetrahydroaminoacridine blocks and prolongs NMDA receptormediated responses in a voltage-dependent manner. Europ. J. Pharmacol. 253, 1–8.CrossRefGoogle Scholar
  17. 17.
    Costa A.C., Albuquerque E.X., 1994. Dynamics of the actions of tetrahydro-9-aminoacridine and 9-aminoacridine on glutamatergic currents: Concentrationjump studies in cultured rat hippocampal neurons. J. Pharmacol. Exp. Therapeutics. 268, 503–514.Google Scholar
  18. 18.
    Benveniste M., Mayer M.L. 1995. Trapping of glutamate and glycine during open channel block of rat hippocampal neuron NMDA receptors by 9-aminoacridine. J. Physiol. (London). 483, 367–384.Google Scholar
  19. 19.
    Koshelev S.G., Khodorov B.I., 1995. Blockade of open channels by tetrabutylammonium, 9-aminoacridine and tacrine prevents channels closing and desensitization. Membrane Cell Biol. 9, 93–109.Google Scholar
  20. 20.
    Ueno S., Bracamontes J., Zorumski C., Weiss D.S., Steinbach J.H. 1997. Bicuculline and gabazine are allosteric inhibitors of channel opening of the GABAA receptor. J.Neurosci. 17(2), 625–634.PubMedGoogle Scholar
  21. 21.
    Grosman C., Zhou M., Auerbach A. 2000. Mapping the conformational wave of acetylcholine receptor channel gating. Nature. 403(6771), 773–776.PubMedCrossRefGoogle Scholar
  22. 22.
    Hilf R.J., Dutzler R. 2009. A prokaryotic perspective on pentameric ligand-gated ion channel structure. Curr. Opin. Struct. Biol. 19(4), 418–424.PubMedCrossRefGoogle Scholar
  23. 23.
    Unwin N. 2005. Refined structure of the nicotinic acetylcholine receptor at 4 — resolution. J. Mol. Biol. 346(4), 967–989.PubMedCrossRefGoogle Scholar
  24. 24.
    Brejc K., van Dijk W.J., Klaassen R.V., Schuurmans M., van Der Oost J., Smit A.B., Sixma T.K. 2001. Crystal structure of an ACh-binding protein reveals the ligand-binding domain of nicotinic receptors. Nature. 411(6835), 269–276.PubMedCrossRefGoogle Scholar
  25. 25.
    Lee W.Y., Sine S.M. 2005. Principal pathway coupling agonist binding to channel gating in nicotinic receptors. Nature. 438(7065), 243–247.PubMedCrossRefGoogle Scholar
  26. 26.
    Lee W.Y., Free C.R., Sine S.M. 2009. Binding to gating transduction in nicotinic receptors: Cys-loop energetically couples to pre-M1 and M2-M3 regions. J. Neurosci. 29(10), 3189–3199.PubMedCrossRefGoogle Scholar
  27. 27.
    Hilf R.J., Bertozzi C., Zimmermann I., Reiter A., Trauner D., Dutzler R. 2010. Structural basis of open channel block in a prokaryotic pentameric ligand-gated ion channel. Nat. Struct. Mol. Biol. 17(11), 1330–1336.PubMedCrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2013

Authors and Affiliations

  1. 1.Research Center of NeurologyRussian Academy of Medical Sciences, Brain Research DepartmentMoscowRussia

Personalised recommendations