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α4βδ GABA receptors are high-affinity targets for γ-hydroxybutyric acid (GHB)

Research output: Contribution to journalJournal articleResearchpeer-review

N. Absalom, N. Karim, L.F. Eghorn, I.S. Villumsen, T. Bay, H. Bräuner-Osborne, B. Frølund, R.P. Clausen, J.V. Olsen, G.M. Knudsen, M. Chebib, P. Wellendorph

γ-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 receptor subunits as possible candidates. A subsequent functional screening of various recombinant GABA receptors in Xenopus laevis oocytes using the two-electrode voltage clamp technique showed GHB to be a partial agonist at αβδ- but not αβγ-receptors, proving that the δ-subunit is essential for potency and efficacy. GHB showed preference for α4 over α(1,2,6)-subunits and preferably activated α4β1δ (EC = 140 nM) over α4β(2/3)δ (EC = 8.41/1.03 mM). Introduction of a mutation, α4F71L, in α4β 1(δ)-receptors completely abolished GHB but not GABA function, indicating nonidentical binding sites. Radioligand binding studies using the specific GHB radioligand [ 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 α4 KO brain tissue compared with WT controls, corroborating the direct involvement of the α4-subunit in high-affinity GHB binding. Our data link specific GHB forebrain binding sites with α4-containing GABA receptors and postulate a role for extrasynaptic α4β-containing GABA 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.
Original languageEnglish
JournalProceedings of the National Academy of Sciences USA (PNAS)
Issue number33
Pages (from-to)13404-13409
Number of pages6
Publication statusPublished - 14 Aug 2012

Bibliographical note

Keywords: gamma-hydroxybutyric acid receptor, gamma-hydroxybutyric acid high-affinity binding sites, alpha4-subunit knockout, photoaffinity ligand

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