中华按蚊拟除虫菊酯杀虫剂击倒抗性突变的研究进展

主要阐述了钠离子通道的结构和功能、拟除虫菊酯杀虫剂的作用机理,总结了中华按蚊钠离子通道中与拟除虫菊酯杀虫剂抗性相关的kdr突变及其抗性机理,为后续中华按蚊抗性机制的深入研究提供了理论依据。     

氯虫酰胺对甜菜夜蛾神经元钠离子通道电流的影响

为了研究新型杀虫剂氯虫酰胺对甜菜夜蛾（Laphygma exigua）的杀虫机制,应用全细胞膜片钳技术记录了氯虫酰胺对急性分离的甜菜夜蛾幼虫中枢神经细胞电压门控钠离子通道的影响。结果显示,在电压钳模式下,0.1 g/L药物作用1 min、5 min、15 min后平均钠离子通道电流峰值分别是（-9.627±0.114）nA、（-11.668±0.131）nA、（-8.726±0.398）nA。在电流钳模式下,0.4 g/L药物作用1 min、3 min、5 min后动作电位峰值分别为（94.366±4.596）m V、（90.363±2.258） m V、（83.15±2.959）m V ,时程分别为（5.025±0.884）m s、（6.032±0.073） m s、（6.387±0.376） ms ,药物作用10 min后只能产生电紧张,无法发放动作电位。以上结果表明,氯虫酰胺对钠电流的作用明显具有时间依赖性,0.1g/L药物初始可增加钠离子通道电流峰值,但随用药时间延长效果减弱。高质量浓度氯虫酰胺可时间依赖性地降低动作电位峰值和延长时程,降低细胞膜兴奋性。甜菜夜蛾中枢神经细胞钠离子通道是氯虫酰胺药物的作用靶标之一。     

电压门控钠离子通道scn1Laa参与斑马鱼脑神经发育和运动行为调节的研究

电压门控钠离子通道对于脊椎动物脑神经起始、传播动作电位具有重要作用。为了解斑马鱼电压门控钠离子通道基因scn1Laa在脑神经中的作用,通过CRISPR/Cas9基因编辑技术,首次构建了可稳定遗传的生长没有受明显影响的scn1Laa缺陷型（scn1Laa-/-）斑马鱼家系。相比野生型,5 dpf（days post-fertilization,受精后5天）scn1Laa缺陷型斑马鱼兴奋抑制性神经元（氨基丁酸类神经元）表达相对增加,兴奋类神经元（谷氨酸能类神经元）和成熟神经元显著减少,脑部细胞增殖也显著减少。受精后5天和90 天的 scn1Laa缺陷型斑马鱼的运动较同时期野生型斑马鱼更为活跃,受精后90天的 scn1Laa缺陷型斑马鱼的运动具有明显的爆发性。以上结果表明,scn1laa缺失导致兴奋类神经元（谷氨酸能类神经元）以及神经细胞增殖减少,影响脑周围神经放电,导致运动神经调节障碍,出现运动行为异常活跃。即电压门控钠离子通道基因scn1Laa参与斑马鱼脑神经发育和生长,间接参与运动行为调节。同时本文也为进一步探究电压门控钠离子通道在脑神经中的作用奠定基础。     

氰氟虫腙的作用机制及抗药性研究进展

氰氟虫腙是一种新型的钠离子通道抑制剂,作用机制独特,未发现氰氟虫腙与现有杀虫剂存在交互抗性。对氰氟虫腙的研发历史、杀虫谱、作用机制、结合位点、抗性情况进行综合评估,以期阐明氰氟虫腙在害虫抗药性治理以及农药安全使用方面的重要性。     

桔小实蝇电压门控钠离子通道基因cDNA克隆及其生物信息学分析

【目的】克隆获得桔小实蝇（Bactrocera dorsalis）电压门控钠离子通道基因cDNA序列,明确其典型特征,为研究桔小实蝇抗性分子机理奠定基础。【方法】采用RT-PCR和PCR技术,克隆桔小实蝇钠离子通道基因cDNA序列,利用相关软件对其序列进行生物信息学分析。【结果】克隆得到1条长为6 446 bp的cDNA序列,包含1个6 405 bp的完整开放阅读框,共编码2 134个氨基酸。同源比对发现,桔小实蝇与果蝇（Drosophila melanogaster,NP_001188635）和家蝇（Musca domestica,AAB47604）钠离子通道基因相似度分别高达91.7%和86.9%,而与人的钠离子通道Nav1.2基因（Homo sapiens,NP_066287）相似度为42.3%。所克隆序列包含昆虫钠离子通道所有典型特征。【结论】成功地从桔小实蝇中克隆得到钠离子通道基因完整开放阅读框。该钠离子通道基因存在丰富的选择性剪接,发现了3个可能与抗性相关的碱基取代。钠离子通道基因有可能发展成为一种昆虫系统发育研究的分子标记。     

柑橘全爪螨钠离子通道基因差异性分析

为弄清钠离子通道基因与柑橘全爪螨Panonychus citri噻螨酮抗性的关系,通过柑橘全爪螨转录组分析鉴定了钠离子通道基因;采用序列比对和Sanger测序,分别对柑橘全爪螨敏感品系和噻螨酮抗性品系钠离子通道基因序列进行单核苷酸多态性分析;进一步采用RPKM法和荧光定量PCR对抗性品系和敏感品系钠离子通道基因进行表达差异分析.从柑橘全爪螨转录组中获得了39条钠离子通道基因,相对于敏感品系,抗性品系中有21条表达上调,17条表达下调,Nay1.8.1和Nay1.6.4分别为上调倍数最高的2个钠离子通道基因[log2 Ratio(RS/SS)分别为10.59,10.30],进一步荧光定量PCR分析发现,Nav1.8.1和Nav1.6.4上调倍数分别为3.92和1.68;基因序列比较和Sanger测序发现,抗性品系中,Nay1.9.4在344,441,468位和Nav1.7.3在第786分别有3个和1个SNP位点,进一步分析编码的氨基酸序列发现,Nav1.9.4只有第468位G突变为A导致蛋氨酸突变为异亮氨酸,Nav1.7.3在第786位T突变为C导致亮氨酸突变为色氨酸.钠离子通道基因差异性分析及验证为进一步研究柑橘全爪螨抗性分子机理提供了有价值的基因资源.     

3种稻飞虱抗药性基因的生物信息学分析

目前褐飞虱、白背飞虱和灰飞虱的转录组数据均已获得,本研究利用生物信息学方法从3种稻飞虱转录组中搜索获得8类与抗药性相关的基因,包括解毒代谢酶基因和杀虫剂靶标基因.其中,羧酸酯酶基因26条,细胞色素p450基因293条,谷胱甘肽转移酶基因39条.在靶标基因中,乙酰胆碱酯酶基因10条,乙酰胆碱受体33条,钠离子通道24条,γ-氨基丁酸受体21条,鱼尼丁受体59条.在褐飞虱中找到更多的基因序列,分析认为这可能与褐飞虱转录组测序深度有关.对这3种同翅目害虫抗药性相关的基因做了多序列比对和进化分析.     

新烟碱类杀虫剂对白符跳虫毒性的研究

目前新烟碱类杀虫剂已经成为全球使用最广泛的一类杀虫剂。白符跳虫(Folsomia candida)是多年来国内外土壤生态安全评估中普遍使用物种之一。国内有关新烟碱类杀虫剂对白符跳虫毒性的研究报道甚少,国外已有部分关于新烟碱类杀虫剂对白符跳虫毒性的研究报道,从新烟碱类杀虫剂对白符跳虫的急性毒性、慢性毒性、繁殖毒性等方面进行阐述,为新烟碱类杀虫剂污染的生态风险评价提供理论依据。     

新烟碱类杀虫剂及使用

新烟碱类是全新结构的超高效杀虫剂,也是自菊酯类以来销售量增长最快的一类杀虫剂。除农民朋友较熟悉的吡虫啉以外,市场上可见到的新烟碱类杀虫剂还有啶虫脒和噻虫嗪等。其中吡虫啉和啶虫脒称为第一代新烟碱类杀虫剂,噻虫嗪等称为第二代新烟碱类杀虫剂。新烟碱类杀虫剂虽然也作用于害虫的神经系统,但与传统的有机磷、氨基甲酸酯和菊酯类农药不同,不存在交互抗性。     

杀虫剂的分类

按照目前较习惯的化学结构分类方法,杀虫剂可分为有机磷类杀虫剂,如甲胺磷、敌敌畏、氧化乐果等;氨基甲酸脂类杀虫剂,如克百威、灭多威等;拟除虫菊酯类杀虫剂,如氯氰菊酯、杀灭菊酯等;有机氯杀虫剂,如硫丹、甲氧滴滴涕等;沙蚕毒素类杀虫剂,如杀螟丹、杀虫双等;苯甲酰脲类杀虫剂,如除虫脲、噻嗪酮等。此外,还有根据有效成分物质来源划分的植物源类杀虫剂,如鱼藤酮、烟碱等;微生物源类杀虫剂,如BT、多角体病毒等。以及含有至少两种有效成分的混合杀虫剂。杀虫剂的分类     

The molecular basis of acid insensitivity in the African naked mole-rat

Acid evokes pain by exciting nociceptors; the acid sensors are proton-gated ion channels that depolarize neurons. The naked mole-rat (Heterocephalus glaber) is exceptional in its acid insensitivity, but acid sensors (acid-sensing ion channels and the transient receptor potential vanilloid-1 ion channel) in naked mole-rat nociceptors are similar to those in other vertebrates. Acid inhibition of voltage-gated sodium currents is more profound in naked mole-rat nociceptors than in mouse nociceptors, however, which effectively prevents acid-induced action potential initiation. We describe a species-specific variant of the nociceptor sodium channel Na(V)1.7, which is potently blocked by protons and can account for acid insensitivity in this species. Thus, evolutionary pressure has selected for an Na(V)1.7 gene variant that tips the balance from proton-induced excitation to inhibition of action potential initiation to abolish acid nociception.     

Calmodulin activation of calcium-dependent sodium channels in excised membrane patches of Paramecium

Calmodulin is a calcium-binding protein that participates in the transduction of calcium signals. The electric phenotypes of calmodulin mutants of Paramecium have suggested that the protein may regulate some calcium-dependent ion channels. Calcium-dependent sodium single channels in excised patches of the plasma membrane from Paramecium were identified, and their activity was shown to decrease after brief exposure to submicromolar concentrations of calcium. Channel activity was restored to these inactivated patches by adding calmodulin that was isolated from Paramecium to the cytoplasmic surface. This restoration of channel activity did not require adenosine triphosphate and therefore, probably resulted from direct binding of calmodulin, either to the sodium channel itself or to a channel regulator that was associated with the patch membrane.     

Mechanism of voltage gating in potassium channels

The mechanism of ion channel voltage gating-how channels open and close in response to voltage changes-has been debated since Hodgkin and Huxley's seminal discovery that the crux of nerve conduction is ion flow across cellular membranes. Using all-atom molecular dynamics simulations, we show how a voltage-gated potassium channel (KV) switches between activated and deactivated states. On deactivation, pore hydrophobic collapse rapidly halts ion flow. Subsequent voltage-sensing domain (VSD) relaxation, including inward, 15-angstrom S4-helix motion, completes the transition. On activation, outward S4 motion tightens the VSD-pore linker, perturbing linker-S6-helix packing. Fluctuations allow water, then potassium ions, to reenter the pore; linker-S6 repacking stabilizes the open pore. We propose a mechanistic model for the sodium/potassium/calcium voltage-gated ion channel superfamily that reconciles apparently conflicting experimental data.     

Genomic organization and deduced amino acid sequence of a putative sodium channel gene in Drosophila

The deduced amino acid sequence of a Drosophila gene isolated with a vertebrate sodium channel complementary DNA probe revealed an organization virtually identical to the vertebrate sodium channel protein; four homologous domains containing all putative membrane-spanning regions are repeated in tandem with connecting linkers of various sizes. All areas of the protein presumed to be critical for channel function show high evolutionary conservation. These include those proposed to function in voltage-sensitive gating, inactivation, and ion selectivity. All 24 putative gating charges of the vertebrate protein are in identical positions in the Drosophila gene. Ten introns interrupt the coding regions of the four homology units; introns with positions conserved among homology units bracket a region hypothesized to be the selectivity filter for the channel. The Drosophila gene maps to the right arm of the second chromosome in region 60D-E. This position does not coincide with any known mutations that confer behavioral phenotypes, but is close to the seizure locus (60A-B), which has been hypothesized to code for a voltage-sensitive sodium channel.     

昆虫离子通道及抗性的研究进展

对钠、氯、钾和钙离子通道的结构、药理、分子抗性及基因调控机理的研究进展进行了综述，认为研究昆虫离子通道对于害虫抗性治理和开发新农药具有重要意义。     

Changes in sodium channel gating produced by point mutations in a cytoplasmic linker

Voltage-gated sodium channels are transmembrane proteins of approximately 2000 amino acids and consist of four homologous domains (I through IV). In current topographical models, domains III and IV are linked by a highly conserved cytoplasmic sequence of amino acids. Disruptions of the III-IV linker by cleavage or antibody binding slow inactivation, the depolarization-induced closed state characteristic of sodium channels. This linker might be the positively charged "ball" that is thought to cause inactivation by occluding the open channel. Therefore, groups of two or three contiguous lysines were neutralized or a glutamate was substituted for an arginine in the III-IV linker of type III rat brain sodium channels. In all cases, inactivation occurred more rapidly rather than more slowly, contrary to predictions. Furthermore, activation was delayed in the arginine to glutamate mutation. Hence, the III-IV linker does not simply act as a charged blocker of the channel but instead influences all aspects of sodium channel gating.     

DNA甲基化抑制剂对水稻幼苗生长及耐盐性的影响

DNA甲基化抑制剂能够通过改变植物基因组的DNA甲基化水平从而对植物的开花、生长发育和抗逆性产生影响。以耐盐和盐敏感水稻品种为材料,分别在水稻萌发期和幼苗期用DNA甲基化抑制剂进行处理,并对处理后的幼苗进行盐胁迫。结果表明:萌发期抑制剂处理导致种子萌发率显著降低,幼苗出现矮化、成簇、死亡等症状,株高、根长和干重也受到了显著的抑制;同时,DNA甲基化抑制剂处理可以降低盐胁迫时地上部的钠离子浓度、提高抗坏血酸过氧化物酶(APX)的活性。上述结果表明DNA甲基化抑制剂通过改变基因组DNA的甲基化水平来影响水稻的生长发育,并通过影响离子吸收和抗氧化酶类的活性来影响水稻的耐盐性。     

The molecular basis of neuronal excitability

Neurons process and transmit information in the form of electrical signals. Their electrical excitability is due to the presence of voltage-sensitive ion channels in the neuronal plasma membrane. In recent years, the voltage-sensitive sodium channel of mammalian brain has become the first of these important neuronal components to be studied at the molecular level. This article describes the distribution of sodium channels among the functional compartments of the neuron and reviews work leading to the identification, purification, and characterization of this membrane glycoprotein.     

海葵多肽类神经毒素的结构与药理学功能

海葵以其触手刺细胞中的毒液行使捕食和防御功能,其毒液中富含各种多肽类神经毒素,分子量为3—7kDa之间,分子序列中含多对二硫键以稳定其结构。海葵神经毒素以钠离子通道毒素和钾离子通道毒素为其主要成分,此外还发现有作用于其他离子通道的成分,此外,还有部分海葵毒素目前尚不清楚其分子靶标。不同类型的海葵毒素具有不同的空间结构。海葵毒素多肽的分子多样性使其成为动物毒素研究的一个重要分支,同时海葵多肽毒素对不同离子通道的特异性和高亲和性,使得它们成为神经生理学和药理学研究的一种重要工具。     

Ion channels in yeast

Voltage-dependent ion channels have been found in the plasma membrane of the yeast Saccharomyces cerevisiae. Ion channel activities were recorded from spheroplasts or patches of plasma membrane with the patch-clamp technique. The most prominent activities came from a set of potassium channels with the properties of activation by positive but not negative voltages, high selectivity for potassium over sodium ion, unit conductance of 20 picosiemens, inhibition by tetraethylammonium or barium ions, and bursting kinetics.