Preventing Phosphorylation of the GABA_AR β3 Subunit Compromises the Behavioral Effects of Neuroactive Steroids

Neuroactive steroids (NASs) have potent anxiolytic, anticonvulsant, sedative, and hypnotic actions, that reflect in part their efficacy as GABA_AR positive allosteric modulators (PAM). In addition to this, NAS exert metabotropic effects on GABAergic inhibition via the activation of membrane progesterone receptors (mPRs), which are G-protein coupled receptors. mPR activation enhances the phosphorylation of residues serine 408 and 409 (S408/9) in the β3 subunit of GABA_ARs, increasing their accumulation in the plasma membrane leading to a sustained increase in tonic inhibition. To explore the significance of NAS-induced phosphorylation of GABA_ARs, we used mice in which S408/9 in the β3 subunit have been mutated to alanines, mutations that prevent the metabotropic actions of NASs on GABA_AR function while preserving NAS allosteric potentiation of GABAergic current. While the sedative actions of NAS were comparable to WT, their anxiolytic actions were reduced in S408/9A mice. Although the induction of hypnosis by NAS were maintained in the mutant mice the duration of the loss of righting reflex was significantly shortened. Finally, ability of NAS to terminate diazepam pharmacoresistant seizures was abolished in S408/9A mice. In conclusion, our results suggest that S408/9 in the GABA_AR β3 subunit contribute to the anxiolytic and anticonvulsant efficacy of NAS, in addition to their ability to regulate the loss of righting reflex.