title: A Single phenylalanine residue in the main intracellular loop of α1 γ-aminobutyric acid type A and glycine receptors influences their sensitivity to propofol.
abstract: BACKGROUND
  The intravenous anesthetic propofol acts as a positive allosteric modulator of glycine (GlyRs) and γ-aminobutyric acid type A (GABAARs) receptors. Although the role of transmembrane residues is recognized, little is known about the involvement of other regions in the modulatory effects of propofol. Therefore, the influence of the large intracellular loop in propofol sensitivity of both receptors was explored.
  
  
  METHODS
  The large intracellular loop of α1 GlyRs and α1β2 GABAARs was screened using alanine replacement. Sensitivity to propofol was studied using patch-clamp recording in HEK293 cells transiently transfected with wild type or mutant receptors.
  
  
  RESULTS
  Alanine mutation of a conserved phenylalanine residue within the α1 large intracellular loop significantly reduced propofol enhancement in both GlyRs (360 ± 30 vs. 75 ± 10%, mean ± SEM) and GABAARs (361 ± 49% vs. 80 ± 23%). Remarkably, propofol-hyposensitive mutant receptors retained their sensitivity to other allosteric modulators such as alcohols, etomidate, trichloroethanol, and isoflurane. At the single-channel level, the ability of propofol to increase open probability was significantly reduced in both α1 GlyR (189 ± 36 vs. 22 ± 13%) and α1β2 GABAAR (279 ± 29 vs. 29 ± 11%) mutant receptors.
  
  
  CONCLUSIONS
  In this study, it is demonstrated that the large intracellular loop of both GlyR and GABAAR has a conserved single phenylalanine residue (F380 and F385, respectively) that influences its sensitivity to propofol. Results suggest a new role of the large intracellular loop in the allosteric modulation of two members of the Cys-loop superfamily. Thus, these data provide new insights into the molecular framework behind the modulation of inhibitory ion channels by propofol.
authors.affiliation: Department of Physiology, University of Concepción, Concepción, Chile.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Moraga-Cid
authors.forename: Gonzalo E
authors.initials: GE
authors.lastname: Yevenes
authors.forename: Günther
authors.initials: G
authors.lastname: Schmalzing
authors.forename: Robert W
authors.initials: RW
authors.lastname: Peoples
authors.forename: Luis G
authors.initials: LG
authors.lastname: Aguayo
mesh_terms: D000409:Alanine / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D018686:Anesthetics, Intravenous / Q000494:pharmacology*; D004247:DNA / Q000235:genetics; D055724:Electrophysiological Phenomena; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000502:physiology; D018408:Patch-Clamp Techniques; D010649:Phenylalanine / Q000737:chemistry* / Q000235:genetics*; D015742:Propofol / Q000494:pharmacology*; D011963:Receptors, GABA-A / Q000737:chemistry* / Q000187:drug effects* / Q000235:genetics; D018009:Receptors, Glycine / Q000737:chemistry* / Q000187:drug effects* / Q000235:genetics; D014162:Transfection
chemical_list: D018686:Anesthetics, Intravenous; D011963:Receptors, GABA-A; D018009:Receptors, Glycine; D010649:Phenylalanine; D004247:DNA; D000409:Alanine; D015742:Propofol