3β-Methyl-Neurosteroid Analogs Are Preferential Positive Allosteric Modulators and Direct Activators of Extrasynaptic δ-Subunit γ-Aminobutyric Acid Type A Receptors in the Hippocampus Dentate Gyrus Subfield
3β-Methyl-Neurosteroid Analogs Are Preferential Positive Allosteric Modulators and Direct Activators of Extrasynaptic δ-Subunit γ-Aminobutyric Acid Type A Receptors in the Hippocampus Dentate Gyrus Subfield
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DOI：
10.1124/jpet.117.246660
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发表时间：
2018-06-01
影响因子：
3.5
通讯作者：
Reddy, Doodipala Samba
中科院分区：
文献类型：
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作者：
Chuang, Shu-Hui;Reddy, Doodipala Samba
神经类固醇是γ -氨基丁酸(GABA)-A受体的强力调节剂。Ganaxolone （3 α -羟基-3 β -甲基-5 α -孕酮-20- 1，GX）和神经类固醇异孕酮（AP）的合成类似物被设计用于治疗癫痫和相关疾病。然而，它们在天然神经元中的确切作用机制尚不清楚。在这里，我们试图通过分析突触外受体介导的张力电流和突触受体介导的相电流来确定GX及其类似物对天然海马神经元GABA-A受体的作用方式。GX在两种细胞类型（含δ的齿状回颗粒细胞（DGGCs）和含γ 2的CA1锥体细胞（CA1 PCs））中的浓度-响应谱。GX在DGGCs中产生的GABA-A受体激活的氯离子电流的增强（500%）明显大于CAIPCs（200%）。在缺乏GABA的情况下，GX在DGGCs中引起的内向电流是CA1PCs的2倍，而CA1PCs在DGGCs中引起的内向电流是AP的2倍。在海马切片中，GX增强并直接激活DGGCs的强直电流。这些反应在δ亚基敲除（δ KO）小鼠的DGGCs中显著减弱，证实了GX对δ GABA-A受体的选择性。与AP一样，抑制蛋白激酶C可阻止GX强直电流的增强。此外，GX对海马点燃癫痫发作的保护作用在delta KO小鼠中显著减弱。GX类似物在GABA-A受体介导的强直抑制中表现出比GX更强的效力和疗效。综上所述，这些结果提供了强有力的证据，证明GX及其类似物是突触外δ GABA-A受体的优先变构调节剂和直接激活剂，调节齿状回的网络抑制和癫痫发作。因此，这些发现为临床使用合成神经类固醇治疗癫痫和发作性疾病提供了机制依据。
Neurosteroids are powerful modulators of gamma-aminobutyric acid (GABA)-A receptors. Ganaxolone (3 alpha-hydroxy-3 beta-methyl-5 alpha-pregnan-20-one, GX) and synthetic analogs of the neurosteroid allopregnanolone (AP) are designed to treat epilepsy and related conditions. However, their precise mechanism of action in native neurons remains unclear. Here, we sought to determine the mode of action of GX and its analogs at GABA-A receptors in native hippocampal neurons by analyzing extrasynaptic receptor-mediated tonic currents and synaptic receptor-mediated phasic currents. Concentration-response profiles of GX were determined in two cell types: delta-containing dentate gyrus granule cells (DGGCs) and gamma 2-containing CA1 pyramidal cells (CA1 PCs). GX produced significantly greater potentiation of the GABA-A receptor-activated chloride currents in DGGCs (500%) than CAIPCs (200%). In 1he absence of GABA, GX evoked 2-fold greater inward currents in DGGCs than CA1PCs, which were 2-fold greater than AP within DGGCs. In hippocampus slices, GX potentiated and directly activated tonic currents in DGGCs. These responses were significantly diminished in DGGCs from delta-subunit knockout (delta KO) mice, confirming GX's selectivity for delta GABA-A receptors. Like AP, GX potentiation of tonic currents was prevented by protein kinase C inhibition. Furthermore, GX's protection against hippocampus-kindled seizures was significantly diminished in delta KO mice. GX analogs exhibited greater potency and efficacy than GX on delta GABA-A receptor-mediated tonic inhibition. In summary, these results provide strong evidence that GX and its analogs are preferential allosteric modulators and direct activators of extrasynaptic delta GABA-A receptors regulating network inhibition and seizures in the dentate gyrus. Therefore, these findings provide a mechanistic rationale for the clinical use of synthetic neurosteroids in epilepsy and seizure disorders.