Neuroactive steroids are potent modulators of ␥-aminobutyric acid type A receptors (GABA ARs), and their behavioral effects are generally viewed in terms of altered inhibitory synaptic transmission. Here we report that, at concentrations known to occur in vivo, neuroactive steroids specifically enhance a tonic inhibitory conductance in central neurons that is mediated by extrasynaptic ␦ subunit-containing GABAARs. The neurosteroid-induced augmentation of this tonic conductance decreases neuronal excitability. Fluctuations in the circulating concentrations of endogenous neuroactive steroids have been implicated in the genesis of premenstrual syndrome, postpartum depression, and other anxiety disorders. Recognition that ␦ subunit-containing GABAARs responsible for a tonic conductance are a preferential target for neuroactive steroids may lead to novel pharmacological approaches for the treatment of these common conditions. hippocampus ͉ cerebellum ͉ neurosteroids ͉ inhibitory postsynaptic currents ͉ ␦ knockout mice G ABA A Rs (␥-aminobutyric acid type A receptors) are pentameric proteins that form Cl Ϫ -permeable ion channels activated by the neurotransmitter GABA. To date, 19 mammalian GABA A subunit isoforms have been identified, and these assemble to produce the dozen or so different receptor subtypes most frequently found in the brain (1). The most potent positive endogenous modulators of GABA A R function are the 3␣-hydroxy ring A-reduced pregnane steroids, that have sedativehypnotic, anticonvulsant, and anxiolytic effects (2-4). Severe mood disorders that can occur during the menstrual cycle and after pregnancy are suggested to involve alterations in the function of synaptic GABA A Rs (2, 3, 5) triggered by rapid decreases in the concentrations of these progesterone-derived neuroactive steroids (6).Recently, it has become apparent that distinct GABA A Rs participate in two types of inhibitory control. Transient activation of synaptic GABA A Rs is responsible for conventional phasic inhibition, whereas the continuous activation of extrasynaptic GABA A Rs can generate a form of tonic inhibition (7-14). GABA A Rs containing the ␦ subunit are restricted to extrasynaptic locations (15) and have an unusually high affinity for GABA (16,17), making them likely mediators of the tonic GABA A conductance recorded in both cerebellar (7,8) and dentate gyrus granule cells (DGGC) (10, 11). In mice lacking the ␦ subunit of the GABA A R, the effects of neuroactive steroids are greatly reduced (18). Moreover, recent reports (17,19,20) have raised the possibility that the steroid sensitivity of ␦ subunitcontaining GABA A Rs may be much higher than previously thought (21). In light of these findings, and the possible involvement of ␦ subunit-containing receptors in generating tonic conductances (8-11), we recorded from wild-type and ␦Ϫ͞Ϫ mice, and examined the effects of the naturally occurring neuroactive steroid 3␣,21-dihydroxy-5␣-pregnan-20-one (allotetrahydrodeoxycorticosterone, THDOC) on the tonic GABA A R-mediated conduct...

Disturbances of neuronal excitability changes during the ovarian cycle may elevate seizure frequency in women with catamenial epilepsy and enhance anxiety in premenstrual dysphoric disorder (PMDD). The mechanisms underlying these changes are unknown, but they could result from the effects of fluctuations in progesterone-derived neurosteroids on the brain. Neurosteroids and some anxiolytics share an important site of action: tonic inhibition mediated by delta subunit-containing GABA(A) receptors (deltaGABA(A)Rs). Here we demonstrate periodic alterations in specific GABA(A)R subunits during the estrous cycle in mice, causing cyclic changes of tonic inhibition in hippocampal neurons. In late diestrus (high-progesterone phase), enhanced expression of deltaGABA(A)Rs increases tonic inhibition, and a reduced neuronal excitability is reflected by diminished seizure susceptibility and anxiety. Eliminating cycling of deltaGABA(A)Rs by antisense RNA treatment or gene knockout prevents the lowering of excitability during diestrus. Our findings are consistent with possible deficiencies in regulatory mechanisms controlling normal cycling of deltaGABA(A)Rs in individuals with catamenial epilepsy or PMDD.

A tonic inhibitory conductance mediated by GABA(A) receptors that supplements the phasic inhibition produced by IPSCs has been identified in the hippocampus and cerebellum. It is presently unknown whether tonic and phasic inhibitions are mediated by GABA(A) receptors with different subunit assemblies. Here we demonstrate that a low concentration (200 nm) of the highly specific competitive GABA(A) antagonist SR95531 (gabazine) reduces phasic inhibition in hippocampal granule cells by 71% but has no effect on tonic inhibition, whereas a high concentration (10 microm) of the antagonist blocked both conductances. These findings are consistent with tonic and phasic conductances being mediated by different GABA(A) receptor subtypes with different affinities for GABA.

␦ Subunit-containing GABA A receptors are located predominantly at nonsynaptic sites in the dentate gyrus where they may play important roles in controlling neuronal excitability through tonic inhibition and responses to GABA spillover. Immunohistochemical methods were used to determine whether ␦ subunit expression was altered after pilocarpine-induced status epilepticus in C57BL/6 mice in ways that could increase excitability of the dentate gyrus. In pilocarpine-treated animals, the normal diffuse labeling of the ␦ subunit in the dentate molecular layer was decreased by 4 d after status epilepticus (latent period) and remained low throughout the period of chronic seizures. In contrast, diffuse labeling of ␣4 and ␥2 subunits, potentially interrelated GABA A receptor subunits, was increased during the chronic period. Interestingly, ␦ subunit labeling of many interneurons progressively increased after pilocarpine treatment. Consistent with the observed changes in ␦ subunit labeling, physiological studies revealed increased excitability in the dentate gyrus of slices obtained from the pilocarpine-treated mice and demonstrated that physiological concentrations of the neurosteroid tetrahydrodeoxycorticosterone were less effective in reducing excitability in the pilocarpine-treated animals than in controls. The findings support the idea that alterations in nonsynaptic ␦ subunit-containing GABA A receptors in both principal cells and interneurons could contribute to increased seizure susceptibility in the hippocampal formation in a temporal lobe epilepsy model.