Kainate Receptors as a Target for the Anticonvulsant Perampanel

摘要 潘生丁(PMP)是第三代抗惊厥药物，起到非竞争性变构调节剂的作用。 AMPA受体中的离子亲性谷氨酸受体(IGluRs)是AMPA受体的主要介体。 中枢神经系统的兴奋性神经传递。它的抗惊厥活性被认为是由于润湿而产生的 AMPA受体抑制在癫痫脑中的过度兴奋性。最近的结构性决议 PMP在GluA2 AMPA受体亚基上的结合位点揭示了其NAM决定因素的精细细节 但也强调了AMPA和海人藻酸受体(KAR)关键结合残基的保守性 亚单位，一个不同但相关的iGluR家族。KARs在中枢神经系统中起着各种作用，通常 以调节兴奋和抑制音之间的平衡为特征的。 我们假设PMP在KARS上起到NAM的作用，这种活动在一定程度上有助于其治疗 功效。我们的初步结果初步支持了这一假说，并进一步表明 PMP的抑制依赖于特定的KAR亚基GluK5与受体复合体的结合。在……里面 在这个项目中，我们建议在三个相关的目标上进一步探索这一观察结果。在具体目标1中，我们将测试 PMP是KARS亚单位选择性非竞争性拮抗剂的假设及其重要性探讨 PMP结合区的关键氨基酸残基控制KARs对调节剂的敏感性。在……里面 具体目标2，我们将测试PMP是否抑制CA3上海马苔藓纤维突触的神经元KARs 锥体神经元和背根神经节神经元。这两种类型的神经元受体都是已知的 含有GluK5亚基。我们将比较野生型和GluK5-/-小鼠受体的相对抑制情况，以进行测试 GluK5在中枢神经系统中形成抑制PMP的关键底物的假说。在具体目标3中，我们将 比较PMP对AMPA和海人藻酸受体的调节活性 野生型和GluK5/-基因敲除小鼠的动物癫痫、焦虑和疼痛模型，PMP 显示出功效。 这些目标意义重大，因为它们可能改变我们对 第三代抗惊厥药物帕姆帕特。有效区分两种类型的抑制剂的优化 AMPA和海人藻酸受体可能会带来治疗指数更大的新一代药物。

SUMMARY Perampanel (PMP) is a third-generation anticonvulsant that acts as a noncompetitive allosteric modulator (NAM) of AMPA receptors, the ionotropic glutamate receptors (iGluRs) that serve as principle mediators of excitatory neurotransmission in the CNS. Its anticonvulsant activity is thought to result from dampening hyperexcitability in an epileptic brain through AMPA receptor inhibition. The recent structural resolution of the binding site for PMP on the GluA2 AMPA receptor subunit revealed fine details into determinants of its NAM activity but also underscored the conservation of critical binding residues in AMPA and kainate receptor (KAR) subunits, a distinct but related family of iGluRs. KARs serve a variety of functions in the CNS that are generally characterized as modulating a balance between excitation and inhibition tone. We hypothesize that PMP acts as a NAM on KARs and that this activity in part contributes to its therapeutic efficacy. Our preliminary results provide initial tentative support for this hypothesis and additionally reveal that PMP inhibition depends on the incorporation of a specific KAR subunit, GluK5, into the receptor complex. In this project, we propose to explore this observation further in three related aims. In Specific Aim 1, we will test the hypothesis that PMP is a subunit-selective noncompetitive antagonist of KARs and explore the importance of key amino acid residues in the PMP binding domains that control sensitivity of KARs to the modulator. In Specific Aim 2, we will test if PMP inhibits neuronal KARs at hippocampal mossy fiber synapses on CA3 pyramidal neurons and in dorsal root ganglion neurons. Both of these types of neuronal receptors are known to contain the GluK5 subunit. We will compare relative inhibition of receptors in wildtype and GluK5-/- mice to test the hypothesis that GluK5 forms a key substrate for PMP inhibition in the CNS. In Specific Aim 3, we will discriminate between modulatory activity on AMPA vs. kainate receptors by comparing potency of PMP in wildtype and GluK5-/- knockout mice in animal seizure, anxiety, and pain models, indications in which PMP shows efficacy. These objectives are significant because they could change our understanding of the mechanism of action of the third-generation anticonvulsant perampanel. Optimization of inhibitors that effectively discriminate between AMPA and kainate receptors could lead to a new generation of drugs with larger therapeutic indices.