Pharmacology of structural changes at the GABA(A) receptor transmitter binding site

Br J Pharmacol. 2011 Feb;162(4):840-50. doi: 10.1111/j.1476-5381.2010.01083.x.


Background and purpose: The binding of transmitter to specialized binding pockets leads to rearrangements in the structure of the receptor eventually resulting in channel opening. We used voltage-clamp fluorometry to investigate the pharmacological basis and biophysical processes that underlie structural changes at the transmitter binding site of the rat α1β2γ2L GABA(A) receptor.

Experimental approach: Simultaneous electrophysiological and site-specific fluorescence measurements were conducted on receptors expressed in Xenopus oocytes and labelled with an environmentally-sensitive fluorophore, Alexa 546 maleimide, at the α1L127C site.

Key results: Receptors activated by GABA demonstrate a concentration-dependent increase in fluorescence intensity, indicating that the environment surrounding the fluorophore becomes less polar upon activation. Qualitatively similar responses were observed with other GABA site ligands such as piperidine-4-sulphonic acid, muscimol, β-alanine and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol. Fluorescence changes were not affected by the direction of current flow. During long applications of GABA significant desensitization developed, which was not accompanied by additional changes in fluorescence. Pentobarbital was an efficacious agonist of the labelled mutant receptor but did not cause changes in fluorescence. Direct activation by etomidate or the steroid allopregnanolone also did not result in fluorescence changes. Functional potentiation of GABA-activated receptors by allopregnanolone or etomidate enhanced both the GABA-elicited functional response and the fluorescence change. In contrast, potentiation by pentobarbital was not accompanied by an enhanced fluorescence response.

Conclusions and implications: The data indicate that there is no direct correlation between current flow or position of the activation gate and the structural changes as detected by Alexa 546-labelled α1L127Cβ2γ2L GABA(A) receptors. Channel potentiation by pentobarbital qualitatively differs from potentiation by etomidate or allopregnanolone.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Binding Sites
  • Fluorescent Dyes / chemistry
  • GABA-A Receptor Agonists / pharmacology
  • GABA-A Receptor Antagonists / pharmacology
  • HEK293 Cells
  • Humans
  • Hydrophobic and Hydrophilic Interactions
  • Ligands
  • Mutagenesis, Site-Directed
  • Mutant Proteins / chemistry
  • Mutant Proteins / metabolism
  • Neurotransmitter Agents / metabolism*
  • Patch-Clamp Techniques
  • Protein Structure, Tertiary
  • Protein Subunits / chemistry*
  • Protein Subunits / genetics
  • Protein Subunits / metabolism*
  • Quinolinium Compounds / chemistry
  • Rats
  • Receptors, GABA-A / chemistry*
  • Receptors, GABA-A / genetics
  • Receptors, GABA-A / metabolism*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • gamma-Aminobutyric Acid / metabolism*


  • Alexa fluor 546
  • Fluorescent Dyes
  • GABA-A Receptor Agonists
  • GABA-A Receptor Antagonists
  • Ligands
  • Mutant Proteins
  • Neurotransmitter Agents
  • Protein Subunits
  • Quinolinium Compounds
  • Receptors, GABA-A
  • Recombinant Proteins
  • gamma-Aminobutyric Acid