Lin Wei-Hsiang, Giachello Carlo N G, Baines Richard A
Division of Neuroscience and Experimental Psychology, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, UK.
Division of Neuroscience and Experimental Psychology, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, UK
Dis Model Mech. 2017 Feb 1;10(2):141-150. doi: 10.1242/dmm.027045. Epub 2016 Dec 14.
Epilepsy is a significant disorder for which approximately one-third of patients do not respond to drug treatments. Next-generation drugs, which interact with novel targets, are required to provide a better clinical outcome for these individuals. To identify potential novel targets for antiepileptic drug (AED) design, we used RNA sequencing to identify changes in gene transcription in two seizure models of the fruit fly Drosophila melanogaster The first model compared gene transcription between wild type (WT) and bangsenseless (para), a gain-of-function mutant in the sole fly voltage-gated sodium channel (paralytic). The second model compared WT with WT fed the proconvulsant picrotoxin (PTX). We identified 743 genes (FDR≤1%) with significant altered expression levels that are common to both seizure models. Of these, 339 are consistently upregulated and 397 downregulated. We identify pumilio (pum) to be downregulated in both seizure models. Pum is a known homeostatic regulator of action potential firing in both flies and mammals, achieving control of neuronal firing through binding to, and regulating translation of, the mRNA transcripts of voltage-gated sodium channels (Na). We show that maintaining expression of pum in the CNS of para flies is potently anticonvulsive, whereas its reduction through RNAi-mediated knockdown is proconvulsive. Using a cell-based luciferase reporter screen, we screened a repurposed chemical library and identified 12 compounds sufficient to increase activity of pum Of these compounds, we focus on avobenzone, which significantly rescues seizure behaviour in para flies. The mode of action of avobenzone includes potentiation of pum expression and mirrors the ability of this homeostatic regulator to reduce the persistent voltage-gated Na current (I) in an identified neuron. This study reports a novel approach to suppress seizure and highlights the mechanisms of neuronal homeostasis as potential targets for next-generation AEDs.
癫痫是一种严重的疾病，约三分之一的患者对药物治疗无反应。需要与新靶点相互作用的新一代药物，为这些患者提供更好的临床疗效。为了确定抗癫痫药物（AED）设计的潜在新靶点，我们使用RNA测序来鉴定果蝇两种癫痫模型中基因转录的变化。第一个模型比较了野生型（WT）和无触角（para）之间的基因转录，无触角是果蝇唯一的电压门控钠通道（麻痹）功能获得性突变体。第二个模型比较了喂食惊厥剂印防己毒素（PTX）的WT和WT之间的情况。我们鉴定出743个基因（FDR≤1%），其表达水平在两种癫痫模型中均有显著改变。其中，339个基因持续上调，397个基因下调。我们发现pumilio（pum）在两种癫痫模型中均下调。Pum是果蝇和哺乳动物中已知的动作电位发放稳态调节因子，通过与电压门控钠通道（Na）的mRNA转录本结合并调节其翻译来控制神经元发放。我们表明，在para果蝇的中枢神经系统中维持pum的表达具有显著的抗惊厥作用，而通过RNAi介导的敲低降低其表达则会诱发惊厥。使用基于细胞的荧光素酶报告基因筛选，我们筛选了一个重新利用的化学文库，并鉴定出12种足以增加pum活性的化合物。在这些化合物中，我们重点研究阿伏苯宗，它能显著挽救para果蝇的癫痫行为。阿伏苯宗的作用方式包括增强pum的表达，反映了这种稳态调节因子降低已鉴定神经元中持续性电压门控钠电流（I）的能力。本研究报告了一种抑制癫痫的新方法，并强调神经元稳态机制作为新一代AEDs潜在靶点的重要性。
