title: Decreased agonist sensitivity of human GABA(A) receptors by an amino acid variant, isoleucine to valine, in the alpha1 subunit.
abstract: Recombinant human GABA(A) receptors were investigated in vitro by coexpression of cDNAs coding for alpha1, beta2, and gamma2 subunits in the baculovirus/Sf-9 insect cell system. We report that a single amino acid exchange (isoleucine 121 to valine 121) in the N-terminal, extracellular part of the alpha1 subunit induces a marked decrease in agonist GABA(A) receptor ligand sensitivity. The potency of muscimol and GABA to inhibit the binding of the GABA(A) receptor antagonist [3H]SR 95531 (2-(3-carboxypropyl)-3-amino-6-(4-methoxyphenyl)pyridazinium bromide) was higher in receptor complexes of alpha1(ile 121) beta2gamma2 than in those of alpha1(val 121) beta2gamma2 (IC50 values were 32-fold and 26-fold lower for muscimol and GABA, respectively). The apparent affinity of the GABA(A) receptor antagonist bicuculline methiodide to inhibit the binding of [3H]SR 95531 did not differ between the two receptor complex variants. Electrophysiological measurements of GABA induced whole-cell Cl- currents showed a ten-fold decrease in the GABA(A) receptor sensitivity of alpha1 (val 121) beta2gamma2 as compared to alpha1(ile 121) beta2gamma2 receptor complexes. Thus, a relatively small change in the primary structure of the alpha1 subunit leads to a decrease selective for GABA(A) receptor sensitivity to agonist ligands, since no changes were observed in a GABA(A) receptor antagonist affinity and benzodiazepine receptor binding.
authors.affiliation: The Research Institute of Biological Psychiatry, St. Hans Hospital, Roskilde, Denmark.
authors.forename: S E
authors.initials: SE
authors.lastname: Westh-Hansen
authors.forename: P B
authors.initials: PB
authors.lastname: Rasmussen
authors.forename: S
authors.initials: S
authors.lastname: Hastrup
authors.forename: J
authors.initials: J
authors.lastname: Nabekura
authors.forename: K
authors.initials: K
authors.lastname: Noguchi
authors.forename: N
authors.initials: N
authors.lastname: Akaike
authors.forename: M R
authors.initials: MR
authors.lastname: Witt
authors.forename: M
authors.initials: M
authors.lastname: Nielsen
mesh_terms: D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D017931:DNA Primers; D018755:GABA Agonists / Q000494:pharmacology; D018756:GABA Antagonists / Q000378:metabolism / Q000494:pharmacology; D058785:GABA-A Receptor Agonists; D006801:Humans; D007532:Isoleucine / Q000235:genetics*; D018408:Patch-Clamp Techniques; D017354:Point Mutation; D011724:Pyridazines / Q000378:metabolism; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000819:agonists / Q000235:genetics / Q000378:metabolism; D014633:Valine / Q000235:genetics*
chemical_list: D017931:DNA Primers; D018755:GABA Agonists; D018756:GABA Antagonists; D058785:GABA-A Receptor Agonists; D011724:Pyridazines; D011963:Receptors, GABA-A; D011994:Recombinant Proteins; D007532:Isoleucine; C049853:gabazine; D014633:Valine