?4?? GABA(A) receptors are high-affinity targets for ?-hydroxybutyric acid (GHB).

ABSTRACT: ?-Hydroxybutyric acid (GHB) binding to brain-specific high-affinity sites is well-established and proposed to explain both physiological and pharmacological actions. However, the mechanistic links between these lines of data are unknown. To identify molecular targets for specific GHB high-affinity binding, we undertook photolinking studies combined with proteomic analyses and identified several GABA(A) receptor subunits as possible candidates. A subsequent functional screening of various recombinant GABA(A) receptors in Xenopus laevis oocytes using the two-electrode voltage clamp technique showed GHB to be a partial agonist at ???- but not ???-receptors, proving that the ?-subunit is essential for potency and efficacy. GHB showed preference for ?4 over ?(1,2,6)-subunits and preferably activated ?4?1? (EC(50) = 140 nM) over ?4?(2/3)? (EC(50) = 8.41/1.03 mM). Introduction of a mutation, ?4F71L, in ?4?1(?)-receptors completely abolished GHB but not GABA function, indicating nonidentical binding sites. Radioligand binding studies using the specific GHB radioligand [(3)H](E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene)acetic acid showed a 39% reduction (P = 0.0056) in the number of binding sites in ?4 KO brain tissue compared with WT controls, corroborating the direct involvement of the ?4-subunit in high-affinity GHB binding. Our data link specific GHB forebrain binding sites with ?4-containing GABA(A) receptors and postulate a role for extrasynaptic ?4?-containing GABA(A) receptors in GHB pharmacology and physiology. This finding will aid in elucidating the molecular mechanisms behind the proposed function of GHB as a neurotransmitter and its unique therapeutic effects in narcolepsy and alcoholism.

SUBMITTER: Absalom N 

PROVIDER: S-EPMC3421209 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature