摘要:
杀菌剂氟啶胺是哺乳动物细胞中一种典型的线粒体氧化磷酸化解偶联试剂,极易在鱼体内富集,且对鱼类具有极高的毒性。本文测定了氟啶胺对斑马鱼(Danio rerio)胚胎的96 h半数致死浓度LC50值,并研究了该浓度下氟啶胺对斑马鱼胚胎线粒体呼吸耗氧速率和多巴胺神经通路中关键基因表达的干扰效应。氟啶胺对斑马鱼胚胎的96 h-LC50值为0.5 μmol·L-1,该浓度下氟啶胺能够导致胚胎发生中轴骨骼发育不全和心脏水肿等畸形现象,这说明该杀菌剂对斑马鱼胚胎具有极高的毒性。在该浓度下,胚胎的基础呼吸速率和ATP产量均受到了显著抑制,这说明氟啶胺在斑马鱼细胞内同样具有解偶联活性,该杀菌剂能够干扰斑马鱼的线粒体氧化磷酸化过程。另外,LC50浓度氟啶胺还能够激活线粒体内锰超氧化物歧化酶基因(sod2)的表达,这说明氟啶胺能够引发线粒体内氧化自由基的产生,从而导致线粒体功能异常。由于多巴胺系统对线粒体能量产生和氧气消耗具有较高的敏感性,因此本文检测了斑马鱼体内多巴胺系统关键基因的转录水平。LC50浓度的氟啶胺能够显著抑制与多巴胺合成有关的酪氨酸羟化酶基因(th)和与多巴胺接收有关的多巴胺受体基因(drd2a)的转录水平,这说明斑马鱼在发育中多巴胺系统是氟啶胺引发神经疾病的有效靶位。一直以来氟啶胺在水环境中的残留是备受关注的环境问题,因此研究氟啶胺对斑马鱼胚胎的毒性作用机制具有重要意义。
Abstract:
Fluazinam, a widely used agricultural fungicide, is one typical mitochondrial uncoupler in mammalian cells. This fungicide is considered to be very highly accumulative and toxic to fish. In the present study, 96-h median lethal concentration (LC50) of fluazinam to zebrafish (Danio rerio) embryos was determined, and the toxic effect of LC50 fluazinam on mitochondrial oxidative respiration and dopamine system was then determined to better understand molecular mechanisms underlying fluazinam-induced toxicity. The 96-h LC50 of fluazinam to zebrafish was calculated to be 0.5 µmol·L-1. Fluazinam of LC50 level casued axial skeleton defects and cardiac edemas in fish embryos. This suggested that the fungicide was highly toxic to zebrafish embryos. At LC50 level, fluazinam was determined to significantly inhibit basal respiratory rate and ATP production of zebrafish embryos, indicating that fluazinam also has the uncoupling activity in fish cells, as that in mammalian cells, and can also disrupt mitochondrial oxidative phosphorylation in zebrafish embryos. Additionally, fluazinam of LC50 level can activate the transcription of mitochondrial manganese superoxide dismutase (sod2), indicating that fluazinam can induce oxyradical production in mitochondria, thus leading to mitochondrial dysfunction. As dopamine system is highly sensitive to mitochondrial oxygen and energy homeostasis, the transcripts of genes related to dopamine system were determined in the present study. Our results showed that fluazinam of LC50 level significantly inhibited the expressions of tyrosine hydroxylase (th) and dopamine receptor 2a (drd2a). This indicated that the dopamine system is an effective target for fluazinam- induced nervous diseases in fish embryos. Nowadays, fluazinam residue in aquatic environment is of great concern due to high risk to aquatic organisms, and the investigation of fluazinam toxicity to fish larvae therefore is of great significance.
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