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.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Jones M.V., Westbrook G.L. 1995. Desensitized states prolong GABAA channel responses to brief agonist pulses. Neuron. 15(1), 181–191.
Jones M.V., Westbrook G.L. 1996. The impact of receptor desensitization on fast synaptic transmission. Trends Neurosci. 192(3), 96–101.
Bianchi M.T., Macdonald R.L. 2001. Agonist trapping by GABAA receptor channels. J. Neurosci. 21(23), 9083–9091.
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.
Vorobjev, V.S. 1991. Vibrodissociation of sliced mammalian nervous tissue. J. Neurosci. Methods. 38, 145–150.
Vorobjev V.S., Sharonova I.N., Haas H.L. 1996. A simple perfusion system for patch-clamp studies. J. Neurosci. Methods. 68, 303–307.
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.
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.
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.
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.
Adams P.R. 1976. Drug blockade of open end-plate channels. J. Physiol. (London). 260, 531–551.
Neher E., Steinbach J.H. 1978. Local anaesthetics transiently block currents through single acetylcholinereceptor channels. J. Physiol. (London). 277, 153–176.
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.
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.
Sobolevsky A.I., Koshelev S.G., Khodorov B.I., 1999. Probing of NMDA channels with fast blockers. J. Neurosci. 19, 10611–10626.
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.
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.
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.
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.
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.
Grosman C., Zhou M., Auerbach A. 2000. Mapping the conformational wave of acetylcholine receptor channel gating. Nature. 403(6771), 773–776.
Hilf R.J., Dutzler R. 2009. A prokaryotic perspective on pentameric ligand-gated ion channel structure. Curr. Opin. Struct. Biol. 19(4), 418–424.
Unwin N. 2005. Refined structure of the nicotinic acetylcholine receptor at 4 — resolution. J. Mol. Biol. 346(4), 967–989.
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.
Lee W.Y., Sine S.M. 2005. Principal pathway coupling agonist binding to channel gating in nicotinic receptors. Nature. 438(7065), 243–247.
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.
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.
Author information
Authors and Affiliations
Corresponding author
Additional information
Original Russian Text © I.N. Sharonova, A.Yu. Dvorzhak, 2012, published in Biologicheskie Membrany, 2012, Vol. 29, No. 6, pp. 414–421.
The article was translated by the authors.
Rights and permissions
About this article
Cite this article
Sharonova, I.N., Dvorzhak, A.Y. Blockade of GABAA receptor channels by niflumic acid prevents agonist dissociation. Biochem. Moscow Suppl. Ser. A 7, 37–44 (2013). https://doi.org/10.1134/S1990747812050169
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1134/S1990747812050169