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Site-specific fluorescence reveals distinct structural changes with GABA receptor activation and antagonism

Nature Neuroscience volume 5, pages 1163–1168 (2002)Cite this article

Abstract

Neurotransmitter-operated ion channels, such as the GABA (γ-aminobutyric acid) receptor, are important in fast synaptic transmission between neurons. Using site-specific fluorescent labeling and simultaneous electrophysiological analysis in Xenopus laevis oocytes expressing recombinant ρ1 GABA receptors, we identified agonist-mediated molecular rearrangements at three positions within and near the agonist-binding pocket that were highly correlated with receptor activation. We also show that competitive antagonists induced distinct rearrangements on their own that stabilized the receptor in a closed state. Finally, the allosteric antagonist picrotoxin induced a global conformational change that was sensed in the subunit–subunit interface of the amino (N)-terminal domain, distant from its presumed site of action within the transmembrane domains. This first detection in real time of molecular rearrangements of a ligand-activated receptor provides insights into the structural correlates of activation, antagonism and allosteric modulation.

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Figure 1: Location of cysteine mutants, magnitude of oocyte labeling by fluorophore, and the recording apparatus.
Figure 2: Simultaneously recorded GABA-induced ΔF and current were closely correlated.
Figure 3: Changes in fluorescence in response to the competitive antagonist 3-APMPA.
Figure 4: Actions of the agonist TACA and competitive antagonist 3-APA.
Figure 5: Actions of the antagonist picrotoxin on GABA-mediated ΔF and I.

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Acknowledgements

We acknowledge R. Llinas, who encouraged us to undertake these studies. We also thank P. Richardson, A. Klon and S. Harvey for assistance with Fig. 1a; C. Garner, R.A.J. Lester, M. Quick and S. Silberberg for comments on the manuscript; J. Claude for carrying out the measurements of the emission spectra and quantum yield of AF546 and E. Isacoff for providing advice on the fluorescence detection apparatus. The work was supported by NINDS 35291 and 36195 (D.W.) and NIDDK 07545 (Y.C.).

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  1. Departments of Neurobiology and Physiology & Biophysics, University of Alabama at Birmingham School of Medicine, 1719 Sixth Avenue South CIRC410, Birmingham, 35294-0021, Alabama, USA

    Yongchang Chang & David S. Weiss

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Correspondence to David S. Weiss.

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Chang, Y., Weiss, D. Site-specific fluorescence reveals distinct structural changes with GABA receptor activation and antagonism. Nat Neurosci 5, 1163–1168 (2002). https://doi.org/10.1038/nn926

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