PR-independent neurosteroid regulation of α2-GABA-A receptors in the hippocampus subfields

Abstract

Progesterone (P) binding to the intracellular progesterone receptors (PRs) plays a key role in epilepsy via modulation of GABA-A receptor plasticity in the brain. This is thought to occur via conversion of P to neurosteroids such as allopregnanolone, an allosteric modulator of GABA-A receptors. In the female brain, the composition of GABA-A receptors is not static and undergoes dynamic spatial changes in response to fluctuations in P and neurosteroid levels. Synaptic α2-containing GABA-A receptors contribute to phasic neuronal excitability and seizure susceptibility. However, the mechanisms underlying α2-subunit plasticity remain unclear. Here, we utilized the neurosteroid synthesis inhibitor finasteride and PR knockout mice to investigate the role of PRs in α2-subunit in the hippocampus. α2-Subunit expression was significantly upregulated during the high-P state of diestrous stage and with P treatment in wildtype and PR knockout mice. In contrast, there was no change in α2-subunit expression when metabolism of P into neurosteroids was blocked by finasteride in both genotypes. These findings suggest that ovarian cycle-related P and neurosteroids regulate α2-GABA-A receptor expression in the hippocampus via a non-PR pathway, which may be relevant to menstrual-cycle related brain conditions.

Keywords: Epilepsy; GABA receptor; Neurosteroid; Perimenstrual; Progesterone; α2-Subunit.

Fig. 1. Changes in GABA-A receptor α2-subunit mRNA expression in the hippocampus subfields CA1, CA3, and dentate gyrus (DG) during various phases of the estrous cycle

Total RNA was extracted from the microdisected hippocampus tissue and cDNA was prepared for TaqMan PCR analysis. The TaqMan PCR data was normalized in every assay using GAPDH and expressed as percent over the control. The data represents the mean ±SEM (n=8 mice per group). *p<0.05 vs. estrus group.

Fig.2. Changes in GABA-A receptor α2-subunit mRNA expression in the hippocampus during neurosteroid exposure in wildtype (A) and PRKO (B) mice

The α2-subunit mRNA expression was quantified in the hippocampus samples collected from female mice following 7-day treatment with vehicle, or neurosteroid exposure by progesterone (P) treatment. The α2-subunit mRNA expression was normalized with GAPDH and expressed as percent over the control. The data represents the mean ±SEM (n=8 mice per group). *p<0.05 vs. vehicle group.

Fig. 3. Changes in GABA-A receptor α2-subunit mRNA expression in the hippocampus during neurosteroid inhibition in WT (A) and PRKO (B) mice

The GABA-AR α2-subunit mRNA expression was quantified in the hippocampus samples collected from wild-type (WT) mice following 7-day treatment with vehicle, progesterone (P) or progesterone and finasteride (P+F). The α2-subunit mRNA expression was normalized with GAPDH and expressed as percent over the control. The data represents the mean ±SEM (n=8 mice per group). *p<0.05 vs. vehicle group; ^#p<0.05 vs. P treatment group.