Neuropharmacology 39 (2000) 1523–1535 www.elsevier.com/locate/neuropharm Modulation of GABAergic synaptic transmission by the non- benzodiazepine anxiolytic etifoxine Re ´my Schlichter a,* , Volodymyr Rybalchenko a , Pierrick Poisbeau a , Marc Verleye b , Jean-Marie Gillardin b a Laboratoire de Neurophysiologie cellulaire et inte ´gre ´e, UMR 7519 CNRS, Universite ´ Louis Pasteur, 21 rue Rene ´ Descartes, 67084 Strasbourg Cedex, France b Laboratoires Biocodex, Centre de recherche, 60200 Compie `gne, France Accepted 10 November 1999 Abstract We have investigated the effects of 2-ethylamino-6-chloro-4-methyl-4-phenyl-4H-3,1-benzoxazine hydrochloride (etifoxine) on GABA A receptor function. Etifoxine displaced [ 35 S]TBPS (t-butylbicyclophosphorothionate) from GABA A receptors of rat cortical membranes with an IC 50 of 6.7±0.8 μM and [ 3 H]PK11195 from peripheral (mitochondrial)-type benzodiazepine receptors (PBRs) of rat heart homogenates with an IC 50 of 27.3±1.0 μM. Etifoxine displayed anxiolytic properties in an anticonflict test in rats, and potentiated GABA A receptor-mediated membrane currents elicited by submaximal (5–10 μM) but not saturating (0.5 mM) concen- trations of GABA in cultured rat hypothalamic and spinal cord dorsal horn neurones. In hypothalamic cultures, etifoxine induced a dose-dependent inward current for concentrations 1 μM which reflected the post-synaptic potentiation of a small (20 pA) tonic and bicuculline-sensitive GABA A receptor-gated Cl - current. Etifoxine also increased the frequency of spontaneous and miniature GABAergic inhibitory post-synaptic currents without changing their amplitude and kinetic characteristics. Both effects of etifoxine were insensitive to flumazenil (10 μM), an antagonist of central-type benzodiazepine sites present at GABA A receptors, but were partly inhibited by PK11195 (10 μM) an antagonist of PBRs which control the synthesis of neurosteroids. Our results indicate that etifoxine potentiates GABA A receptor-function by a direct allosteric effect and by an indirect mechanism involving the activation of PBRs.  2000 Elsevier Science Ltd. All rights reserved. Keywords: Peripheral benzodiazepine receptor (PBR); Inhibitory post-synaptic current (IPSC); Anxiolytic; Neurosteroid; PK11195 1. Introduction γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the central nervous system (CNS) of vertebrates and activates both ionotropic (GABA A ) and G-protein-coupled (GABA B ) receptors. GABA A recep- tors (GABA A Rs) are ligand-gated Cl - channels formed by the pentameric assembly of distinct subunits and mediate the fast component of GABAergic synaptic transmission (Macdonald and Olsen, 1994; Sieghart, 1995; Mehta and Ticku, 1999). To date, 15 different GABA A R subunits have been identified. Among these, α and β subunits are of fundamental importance for the * Corresponding author. Tel.: + 33-3-88-41-60-18; fax: + 33-3-88- 61-33-47. E-mail address: schlichter@neurochem.u-strasbg.fr (R. Schlichter). 0028-3908/00/$ - see front matter  2000 Elsevier Science Ltd. All rights reserved. PII:S0028-3908(99)00253-1 formation of the Cl - channel and the binding of the agonist, whereas other subunits confer on GABA A Rs a sensitivity/insensitivity to endogenous and/or exogenous allosteric modulators (Macdonald and Olsen, 1994; Sieghart, 1995; Mehta and Ticku, 1999). For example, the presence of γ subunits confers GABA A R sensitivity to benzodiazepines (Pritchett et al., 1989; Sigel and Buhr, 1997; Mehta and Ticku, 1999), whereas the pres- ence of δ (Zhu et al., 1996) or ε (Davies et al., 1997) subunits renders GABA A Rs insensitive to the modu- latory action of neuroactive steroids, i.e. potent allosteric modulators of GABA A Rs which are synthesised within the CNS by neurones and glial cells (Majewska, 1992; Robel and Beaulieu, 1994; Lambert et al., 1995; Men- sah-Nyagan et al., 1999). Both benzodiazepines (Defazio and Hablitz, 1998; Perrais and Ropert, 1999) and neurosteroids (Harrison et al., 1987; Poisbeau et al., 1997) were shown to potentiate