Neuroactive steroids (NASs) such as allopregnanolone (ALLO) play a central role in regulating behavior via
their potent anxiolytic, anticonvulsant, sedative, and hypnotic actions. Accordingly, modifications in the levels of
NASs contribute to anxiety, autism spectrum disorders, depression, epilepsy, and premenstrual syndrome.
Classically, NASs are thought to act by rapidly boosting neuronal inhibition by positive allosteric modulation of
the activity of ¿-aminobutyric acid type A receptors (GABAARs). In addition to their allosteric actions, we have
recently shown that NASs act via a protein kinase C-dependent mechanism to enhance the phosphorylation of
residues including Serine’s 408 and 409 in the ß3 subunit (S408/9), a process that increases GABAAR number
on the plasma membrane leading to a sustained increase in the efficacy of GABAergic inhibition. Although we
have shown that NASs do not directly activate PKC, the mechanism by which NASs lead to changes in
phosphorylation of GABAAR subunits are unknown. It is emerging that in addition to their positive allosteric
modulation of GABAARs, NASs can directly activate membrane progesterone receptors (mPRs); G-protein
coupled receptors that regulate PKC signaling. However, no information is available on the role that mPRs play
in regulating GABAAR activity. Likewise, the behavioral significance of the sustained mPR-mediated
metabotropic actions of NASs remains unexplored. To address these issues we have created mice in which
S408/9 in the ß3 subunit have been mutated to alanines, mutations that are predicted to reduce the
metabotropic actions of NASs on GABAAR function. Preliminary studies using these tools have allowed us to
formulate a central hypothesis that will be tested here; NASs activate mPRs to enhance the
phosphorylation of GABAARs on residues including S408/9 in the ß3 subunit, a mechanism that
underlies their anticonvulsant efficacy. In contrast, their anxiolytic efficacy is mediated via allosteric
potentiation of GABAAR activity, a process dependent upon Q241 in the a2 subunit. Our experiments will
focus on the following aims.
Aim 1. To test the hypothesis that the ability of NAS to induce sustained effects on GABAergic
inhibition is dependent upon S408/9A in the ß3 subunit.
Aim 2. To test the hypothesis that the anxiolytic, and anticonvulsant efficacy of NASs is dependent
upon S408/9 in the ß3 subunit.
Aim 3. To test the hypothesis that NASs mediate their metabotropic effects on GABAARs via the
activation of mPRs.
Collectively, our proposal will identify the molecular mechanisms by which NAS exert their therapeutic actions.
This information may aid the development of new therapeutic strategies to alleviate the burdens of anxiety,
autism spectrum disorders, depression, epilepsy, and premenstrual syndrome.