Structural evidence that propofol stabilizes different GABAA receptor states at potentiating and activating concentrations

Daniel B. Williams, Myles Akabas

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

The GABAA receptor is a target of many general anesthetics, such as propofol. General anesthetic binding sites are distinct from the GABA binding sites. At low concentrations, the anesthetics potentiate the currents induced by submaximal GABA concentrations. At higher concentrations the anesthetics directly activate GABAA receptors. In contrast, benzodiazepines, such as diazepam, only potentiate currents induced by submaximal GABA concentrations. Channel kinetic studies suggest that these drugs stabilize different receptor states. We previously showed that the accessibility of the anionic sulfhydryl reagent p-chloromercuribenzenesulfonate (pCMBS-) applied extracellularly to cysteines substituted for residues in the GABAA α1 subunit M3 membrane-spanning segment was state-dependent. The subset of pCMBS--accessible, M3 segment cysteine mutants acts as a reporter for receptor conformation. Here we show that pCMBS-, applied in the presence of a potentiating concentration of propofol, reacts with a subset of α1 subunit, M3 segment, cysteine-substitution mutants (Y294C, V297C, I302C, F304C). In the presence of a directly activating concentration of propofol pCMBS- reacts with a different subset of the M3 cysteine-substitution mutants (Y294C, S299C, I302C, E303C, A305C). These subsets are distinct from the subsets of M3 cysteine-substitution mutants that are reactive with pCMBS- in the absence and presence of GABA and in the presence of diazepam. We hypothesize that distinct subsets of reactive residues represent distinct conformations or ensembles of conformations of the receptor. These results provide structural evidence for at least five distinct receptor states, three nonconducting states, resting, diazepam-bound and potentiating propofol-bound, and two conducting-desensitized states, the activating propofol-bound and GABA-bound states.

Original languageEnglish (US)
Pages (from-to)7417-7424
Number of pages8
JournalJournal of Neuroscience
Volume22
Issue number17
StatePublished - Sep 1 2002

Fingerprint

Propofol
GABA-A Receptors
gamma-Aminobutyric Acid
Cysteine
Diazepam
General Anesthetics
Anesthetics
4-Chloromercuribenzenesulfonate
Binding Sites
Sulfhydryl Reagents
Benzodiazepines
Membranes
Pharmaceutical Preparations

Keywords

  • Acetylcholine
  • Anesthesia
  • Benzodiazepine
  • Ethanol
  • Glycine
  • Isoflurane

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Structural evidence that propofol stabilizes different GABAA receptor states at potentiating and activating concentrations. / Williams, Daniel B.; Akabas, Myles.

In: Journal of Neuroscience, Vol. 22, No. 17, 01.09.2002, p. 7417-7424.

Research output: Contribution to journalArticle

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abstract = "The GABAA receptor is a target of many general anesthetics, such as propofol. General anesthetic binding sites are distinct from the GABA binding sites. At low concentrations, the anesthetics potentiate the currents induced by submaximal GABA concentrations. At higher concentrations the anesthetics directly activate GABAA receptors. In contrast, benzodiazepines, such as diazepam, only potentiate currents induced by submaximal GABA concentrations. Channel kinetic studies suggest that these drugs stabilize different receptor states. We previously showed that the accessibility of the anionic sulfhydryl reagent p-chloromercuribenzenesulfonate (pCMBS-) applied extracellularly to cysteines substituted for residues in the GABAA α1 subunit M3 membrane-spanning segment was state-dependent. The subset of pCMBS--accessible, M3 segment cysteine mutants acts as a reporter for receptor conformation. Here we show that pCMBS-, applied in the presence of a potentiating concentration of propofol, reacts with a subset of α1 subunit, M3 segment, cysteine-substitution mutants (Y294C, V297C, I302C, F304C). In the presence of a directly activating concentration of propofol pCMBS- reacts with a different subset of the M3 cysteine-substitution mutants (Y294C, S299C, I302C, E303C, A305C). These subsets are distinct from the subsets of M3 cysteine-substitution mutants that are reactive with pCMBS- in the absence and presence of GABA and in the presence of diazepam. We hypothesize that distinct subsets of reactive residues represent distinct conformations or ensembles of conformations of the receptor. These results provide structural evidence for at least five distinct receptor states, three nonconducting states, resting, diazepam-bound and potentiating propofol-bound, and two conducting-desensitized states, the activating propofol-bound and GABA-bound states.",
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