Fragile X syndrome (FXS) is the most common form of inherited intellectual disability and a major cause for a diagnosis of autism spectrum disorders. The symptoms of FXS include hypersensitivity to sensory stimuli, and seizures. The cause of FXS is a loss of the fragile X mental retardation protein (FMRP) yet the underlying pathways and mechanisms of these symptoms are poorly described. GABAA receptors (GABAARs) are major inhibitory ion channels in the brain. Studies from both FXS patients and animal models have revealed altered expression levels of GABAAR α4 subunits with a concomitant reduced efficacy of tonic inhibition, a non-synaptic type of inhibition important for determining the gain of the neuronal output, thus regulating the excitability and activity of neuronal circuits. Our preliminary data suggest that in Fmr1 KO mice there is a decrease in the phosphorylation of α4 subunits leading to decreased tonic current, and an increased excitation. The deficits of tonic inhibition are due to a disruption in trafficking of α4-containing GABAARs from extrasynaptic to synaptic sites. Both FXS patients and Fmr1 KO mice show auditory processing deficits making them hypersensitive to sound. Acoustic information is processed by thalamic auditory neurons in the medial geniculate body (MGB) as it ascends to the auditory cortex. Excitation of auditory thalamic neurons is controlled by tonic inhibition dependent upon GABAARs containing α4 subunits. Reduced tonic inhibition in MGB of FXS patients could explain the increase in auditory sensitivity and be a target for therapy to reduce sensory hypersensitivity. Studies from our laboratory have revealed that neuro-active steroids (NASs) act via a metabotropic, kinase dependent mechanism to increase the phosphorylation of serine 443 in the α4 subunit to promote GABAAR insertion into the plasma membrane leading to sustained elevations in their accumulation on the cell surface and increases in the efficacy of tonic inhibition. These preliminary studies have allowed us to formulate a central hypothesis that will be tested here; Hypersensitivity to sensory stimuli in FXS is caused by reduced tonic inhibition in the central auditory system due to mis-trafficking of α4 subunits to GABAergic synapses. Exposure to neuroactive steroids will increase the trafficking of α4 subunit containing GABAARs to the membrane to boost tonic inhibition and diminish the severity of the disorder. Aim 1. Characterize the phosphorylation and sub-cellular localization of α4 subunit containing GABAARs in Fmr1 KO mice. Aim 2. Examine the electroencephalographic (EEG) activity of Fmr1 KO mice. Aim 3. Determine the efficacy of NAS treatment to reverse deficits in GABAergic inhibition and neuronal excitability. The result of this study will provide new insights into the mechanisms that regulate the efficacy of tonic inhibition and if that alterations in these contribute to the pathophysiology of FXS. Such insights may promote the development...