Progesterone Receptors and Seizure Susceptibility

描述(申请人提供)：孕酮(P)在月经性癫痫的病理生理学中起关键作用。月经性癫痫是一种与月经周期有关的癫痫，影响许多患有癫痫的女性。虽然P的内分泌作用是由孕激素受体(PR)介导的，但P在癫痫发作活动中的分子作用机制尚不清楚。P部分通过转化为别孕酮来调节癫痫发作活动，从而增强GABAA受体的功能。最近，我们有了一个新的发现，即PR在癫痫易感性中起着关键作用。我们的初步结果表明，PR基因敲除(PRKO)小鼠表现出较高的癫痫阈值和对点燃癫痫发生的抵抗，表明PR参与了癫痫易感性的调节。在这一应用中，我们建议利用遗传学、分子和电生理方法来阐明PR增加海马区癫痫敏感性的分子机制。我们推测，孕酮对癫痫的作用部分是由于PR调节了海马区GABA受体亚单位的表达和功能，从而减少了对癫痫的抑制，从而降低了癫痫的敏感性。PR介导的“亚单位转换”导致GABAA受体亚单位组成和功能特性的改变是P控制癫痫敏感性的分子机制。我们建议使用我们实验室建立的三个模型系统来关键地检验这一假说：1)PRKO转基因小鼠模型；2)野生型小鼠的反义PR抑制；3)RU-486阻断野生型小鼠的PR功能。该方案的具体目的是：1)利用癫痫点燃模型，确定PR是否增加癫痫敏感性；2)确定PR是否参与了P引起的GABAA受体亚单位表达和通道功能的改变。目的1研究WT和PRKO小鼠在使用和不使用P和/或非那雄胺治疗的情况下，海马区点燃的比率，作为癫痫发生的指标。目的研究P，P+非那雄胺对WT和PRKO小鼠海马区GABAA受体亚单位表达的影响，并在停药后24小时检测GABAA受体亚单位的表达水平。我们将通过记录急性分离的海马神经元的GABA门控CI电流来验证P诱导的GABAA受体亚单位表达的功能重要性。意义。拟议的研究将提供关于P机制调控癫痫易感性的新信息，这将有助于更好地理解月经性癫痫。相关性。月经性癫痫的女性癫痫发作围绕其月周期聚集。然而，目前这种情况的病理生理学还不完全清楚。这一应用中提出的研究将有助于更好地了解疾病机制，从而为月经性癫痫和其他与孕酮水平波动相关的疾病开发特定的治疗方法。

DESCRIPTION (provided by applicant): Progesterone (P) plays a key role in the pathophysiology of "catamenial epilepsy", a menstrual cyclerelated seizure disorder that affects many women with epilepsy. While P's endocrine actions are mediated by the progesterone receptor (PR), P's molecular mechanism of action in seizure activity is not clearly understood. P modulates seizure activity partly through conversion to allopregnanolone, which enhances GABAA receptor function. Recently, we have made the novel discovery that PRs play a key role in seizure susceptibility. Our preliminary results underscore that PR knockout (PRKO) mice exhibit elevated seizure threshold and resistance to kindling epileptogenesis, suggesting that PRs mediate seizure susceptibility. In this application, we propose to elucidate the molecular mechanisms whereby PRs increase seizure susceptibility in the hippocampus, utilizing genetic, molecular and electrophysiological approaches. We hypothesize that progesterone's actions on seizure activity are due in part to PR regulation of hippocampal GABA^ receptor subunit expression and function that results in reduced inhibition and thereby seizure susceptibility. PR-mediated "subunit switching" leading to alterations of GABAA receptor subunit composition and functional properties are the molecular mechanism by which P controls seizure susceptibility. We propose to test this hypothesis critically utilizing 3 model systems that are well established in our lab: 1) PRKO transgenic mouse model; 2) Antisense PR inhibition in wild-type mice; and 3) RU-486 blockade of PR function in wild-type mice. The Specific Aims of this proposal are: 1) to determine whether PRs increase seizure susceptibility using the kindling model of epilepsy; and 2) to determine whether PRs are involved in P induced alterations in GABAA receptor subunit expression and channel function. Aim 1 will examine the rate of hippocampus kindling in WT and PRKO mice, with and without P and/or finasteride treatment, as an indicator of epileptogenesis. Aim 2 will assess mRNA (real-time PCR) and protein levels (Westerns blots) of GABAA receptor subunits in hippocampal subfields in WT and PRKO mice treated with P, P+finasteride, and 24 hrs after neurosteroid withdrawal. We will verify the functional importance of P induced changes in GABAA receptor subunit expression by recording GABA-gated CI" currents in acutely isolated hippocampal neurons using patch-clamp electrophysiology. Significance. The proposed study will provide novel information on the P mechanisms governing regulation of seizure susceptibility, which could lead to improved understanding of catamenial epilepsy. Relevance. Women with catamenial epilepsy have seizures clustered around their monthly cycle. However, currently the pathophysiology of this condition is not fully understood. The studies proposed in this application will help better understand the disease mechanism and thereby allow development of specific therapies for catamenial epilepsy and other conditions associated with fluctuations in progesterone levels.