title: Contrasting Effects of the γ-Aminobutyric Acid Type A Receptor β3 Subunit N265M Mutation on Loss of Righting Reflexes Induced by Etomidate and the Novel Anesthetic Barbiturate R-mTFD-MPAB.
abstract: Previous studies have shown that etomidate modulates γ-aminobutyric acid type A receptors by binding at the β-α subunit interface within the transmembrane domain of receptors that incorporate β2 or β3 subunits. Introducing an asparagine-to-methionine (N265M) mutation at position 265 of the β3 subunit, which sits within the etomidate-binding site, attenuates the hypnotic effect of etomidate in vivo. It was reported recently that the photoactivatable barbiturate R-mTFD-MPAB also acts on γ-aminobutyric acid type A receptors primarily by binding to a homologous site at the γ-β interface. Given this difference in drug-binding sites established by the in vitro experiments, we hypothesized that the β3-N265M-mutant mice would not be resistant to the anesthetic effects of R-mTFD-MPAB in vivo, whereas the same mutant mice would be resistant to the anesthetic effects of R-etomidate.
  
  
  
  We measured the effects of IV injection of etomidate and R-mTFD-MPAB on loss and recovery of righting reflex in wild-type mice and in mice carrying the β3-N265M mutation.
  
  
  
  Etomidate-induced hypnosis, as measured by the duration of loss of righting reflex, was attenuated in the N265M knock-in mice, confirming prior results. By contrast, recovery of balance and coordinated movement, as measured by the ability to maintain all 4 paws on the ground, was unaffected by the mutation. Neither hypnosis nor impairment of coordinated movement produced by the barbiturate R-mTFD-MPAB was affected by the mutation.
  
  
  
  The findings confirmed our hypothesis that mutating the etomidate-binding site would not alter the response to the barbiturate R-mTFD-MPAB. Furthermore, we confirmed previous studies indicating that etomidate-induced hypnosis is mediated in part by β3-containing receptors. We also extended previous findings by showing that etomidate-impaired balance and coordinated movement are not mediated by β3-containing receptors, thus implicating β2-containing receptors in this end point.
authors.affiliation: From the *Department of Anesthesiology, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin; and †Department of Anesthesia, Critical Care and Pain Medicine, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Corey A
authors.initials: CA
authors.lastname: Amlong
authors.forename: Mark G
authors.initials: MG
authors.lastname: Perkins
authors.forename: Timothy T
authors.initials: TT
authors.lastname: Houle
authors.forename: Keith W
authors.initials: KW
authors.lastname: Miller
authors.forename: Robert A
authors.initials: RA
authors.lastname: Pearce
mesh_terms: D000818:Animals; D001463:Barbiturates / Q000378:metabolism / Q000494:pharmacology*; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005045:Etomidate / Q000378:metabolism / Q000494:pharmacology*; D005260:Female; D006993:Hypnotics and Sedatives / Q000378:metabolism / Q000494:pharmacology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation / Q000187:drug effects / Q000502:physiology*; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011963:Receptors, GABA-A / Q000235:genetics* / Q000378:metabolism; D057897:Reflex, Righting / Q000187:drug effects / Q000502:physiology*
chemical_list: D001463:Barbiturates; D006993:Hypnotics and Sedatives; D021122:Protein Subunits; D011963:Receptors, GABA-A; C032232:barbituric acid; D005045:Etomidate