Abstract

The compounds CGP7930 [2,6-Di-tert-butyl-4-(3-hydroxy-2,2-dimethyl-propyl)-phenol] and its close analog CGP13501 were identified as positive modulators of γ-aminobutyric acid_B (GABA_B) receptor function. They potentiate GABA-stimulated guanosine 5′-O-(3-[³⁵S]thiotriphosphate) (GTPγ[³⁵S]) binding to membranes from a GABA_B(1b/2) expressing Chinese hamster ovary (CHO) cell line at low micromolar concentrations and are ineffective in the absence of GABA. The structurally related compounds propofol and malonoben are inactive. Similar effects of CGP7930 are seen in a GTPγ[³⁵S] binding assay using a native GABA_B receptor preparation (rat brain membranes). Receptor selectivity is demonstrated because no modulation of glutamate-induced GTPγ[³⁵S] binding is seen in a CHO cell line expressing the metabotropic glutamate receptor subtype 2. Dose-response curves with GABA in the presence of different fixed concentrations of CGP7930 reveal an increase of both the potency and maximal efficacy of GABA at the GABA_B(1b/2) heteromer. Radioligand binding studies show that CGP7930 increases the affinity of agonists but acts at a site different from the agonist binding site. Agonist affinity is not modulated by CGP7930 at homomeric GABA_B(1b) receptors. In addition to GTPγ[³⁵S] binding, we show that CGP7930 also has modulatory effects in cellular assays such as GABA_B receptor-mediated activation of inwardly rectifying potassium channels in Xenopus laevis oocytes and Ca²⁺ signaling in human embryonic kidney 293 cells. Furthermore, we show that CGP7930 enhances the inhibitory effect ofl-baclofen on the oscillatory activity of cultured cortical neurons. This first demonstration of positive allosteric modulation at GABA_B receptors may represent a novel means of therapeutic interference with the GABA-ergic system.