A Caenorhabditis elegans assay of seizure-like activity optimised for identifying antiepileptic drugs and their mechanisms of action.
A Caenorhabditis elegans assay of seizure-like activity optimised for identifying antiepileptic drugs and their mechanisms of action.
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DOI：
10.1016/j.jneumeth.2018.09.004
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发表时间：
2018-11-01
影响因子：
3
通讯作者：
Morgan A
中科院分区：
文献类型：
--
作者：
Wong SQ;Jones A;Dodd S;Grimes D;Barclay JW;Marson AG;Cunliffe VT;Burgoyne RD;Sills GJ;Morgan A
带有突变的GABAA受体的蠕虫在暴露于戊四氮后表现出惊厥。经批准的抗癫痫药物乙琥胺可预防抽搐。线虫模型是一种比啮齿动物发作模型更高的吞吐量和道德选择。癫痫影响约1%的人，但现有的抗癫痫药物(AED)仅能缓解症状，对约30%的患者无效。因此，迫切需要新的AED。然而，一个主要的瓶颈是传统啮齿动物模型中早期AED筛查的低通量性质。通过使用更简单的无脊椎动物系统，例如线虫秀丽线虫，这一过程可能会加快。此前有报道称，在编码GABAA受体的UNC-49功能丧失突变的线虫中，戊四氮(PTZ)可诱发头部摆动惊厥。鉴于人类GABAA受体中与癫痫相关的突变已被很好地记录下来，这可能代表着一种临床相关的系统，用于早期AED筛查。然而，最初的琼脂平板法不适合大规模筛查，也没有被验证用于AEDs的鉴定。因此，我们建立了一种替代的流线型、高通量的方法，通过液体孵育的方式用PTZ和AEDs处理突变株。在UNC-49突变体中，PTZ暴露后几分钟内诱发的惊厥可被已建立的AED乙琥胺强烈抑制。这种保护作用不依赖于乙琥胺所建议的靶点--T-型钙通道，因为蠕虫CCA-1同源基因的零突变不影响乙琥胺的抗惊厥作用。我们简化的分析是AED验证的，对于更高吞吐量的化合物筛选是可行的，并且可以促进对AED作用机制的深入了解。基于癫痫相关的遗传背景，这种线虫UNC-49癫痫样活动模型提供了一种道德上的、更高吞吐量的替代传统啮齿动物癫痫发作模型的初始AED筛查。
Worms with mutant GABAA receptors exhibit convulsions upon exposure to pentylenetetrazol. Convulsions are prevented by the approved anti-epileptic drug, ethosuximide. C. elegans model is a higher throughput, ethical alternative to rodent seizure models. Epilepsy affects around 1% of people, but existing antiepileptic drugs (AEDs) only offer symptomatic relief and are ineffective in approximately 30% of patients. Hence, new AEDs are sorely needed. However, a major bottleneck is the low-throughput nature of early-stage AED screens in conventional rodent models. This process could potentially be expedited by using simpler invertebrate systems, such as the nematode Caenorhabditis elegans. Head-bobbing convulsions were previously reported to be inducible by pentylenetetrazol (PTZ) in C. elegans with loss-of-function mutations in unc-49, which encodes a GABAA receptor. Given that epilepsy-linked mutations in human GABAA receptors are well documented, this could represent a clinically-relevant system for early-stage AED screens. However, the original agar plate-based assay is unsuited to large-scale screening and has not been validated for identifying AEDs. Therefore, we established an alternative streamlined, higher-throughput approach whereby mutants were treated with PTZ and AEDs via liquid-based incubation. Convulsions induced within minutes of PTZ exposure in unc-49 mutants were strongly inhibited by the established AED ethosuximide. This protective activity was independent of ethosuximide’s suggested target, the T-type calcium channel, as a null mutation in the worm cca-1 ortholog did not affect ethosuximide’s anticonvulsant action. Our streamlined assay is AED-validated, feasible for higher throughput compound screens, and can facilitate insights into AED mechanisms of action. Based on an epilepsy-associated genetic background, this C. elegans unc-49 model of seizure-like activity presents an ethical, higher throughput alternative to conventional rodent seizure models for initial AED screens.
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影响因子：
3.8
作者：
Boyd WA;McBride SJ;Rice JR;Snyder DW;Freedman JH
通讯作者：
Freedman JH
影响因子：
11.2
作者：
COULTER, DA;HUGUENARD, JR;PRINCE, DA
通讯作者：
PRINCE, DA
影响因子：
--
作者：
Chen X;Barclay JW;Burgoyne RD;Morgan A
通讯作者：
Morgan A
影响因子：
2.5
作者：
COULTER, DA;HUGUENARD, JR;PRINCE, DA
通讯作者：
PRINCE, DA
DOI：
10.1016/b978-0-12-804066-9.00024-9
发表时间：
2017-01-01
期刊：
MODELS OF SEIZURES AND EPILEPSY, 2ND EDITION
影响因子：
--
作者：
Baines, Richard A.;Giachello, Carlo N. G.;Lin, Wei-Hsiang
通讯作者：
Lin, Wei-Hsiang