慢性亚硝酸盐暴露引起雄性斑马鱼脑神经递质紊乱及行为异常
Chronic Nitrite Exposure Causes Brain Neurotransmitter Disorders and Behavioral Abnormalities in Male Zebrafish
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摘要: 为了探究亚硝酸盐慢性暴露致斑马鱼神经毒性影响及其可能的作用机制，本研究将成年雄性斑马鱼（Danio rerio）浸泡在0、0.2、2和20 mg·L-1的亚硝酸盐养殖环境中30 d，暴露期结束后观察并记录斑马鱼的神经行为学变化和脑组织病理学变化，并测定脑神经递质系统相关指标以及氧化-抗氧化系统参数变化。研究结果显示：水环境中亚硝酸盐的慢性暴露导致鱼体血浆中高铁血红蛋白的累积，进而诱发组织缺氧。而斑马鱼脑部丙二醛（malondialdehyde，MDA）含量的剂量依赖性增加和总抗氧化能力（total antioxidant capacity，T-AOC）的下降，表明亚硝酸盐的暴露破坏了脑组织抗氧化防御系统与活性氧之间的平衡，造成了氧化应激。超氧化物歧化酶（superoxide dismutase，SOD）活性的降低以及相关基因Cu/Znsod和Mnsod mRNA表达水平的下降进一步证实，亚硝酸盐暴露降低了斑马鱼脑的抗氧化能力。同时，检测到亚硝酸盐暴露诱导斑马鱼的行为出现异常，包括运动能力下降、对同类鱼群聚集的倾向性降低以及学习和记忆能力减弱。脑组织病理学结果显示，随着亚硝酸盐暴露浓度的升高，中脑小球周灰质带的神经细胞数量减少，并在20 mg·L-1亚硝酸盐浓度组中出现血栓。在测定神经递质系统相关指标时发现，多巴胺（dopamine，DA）含量下降，DA代谢相关基因（dat、drd1b和drd4a）、胆碱能系统相关基因（ache、chrna4b和chrna7）、γ-氨基丁酸（γ-aminobutyric acid，GABA）系统中的gabra1和gat1以及5-羟色胺受体家族基因（htr1aa、htr1ab和htr2a）的mRNA转录水平均有不同程度的下调，表明亚硝酸盐通过抑制神经递质的合成、转运及受体的功能导致信息传递受阻，最终导致运动、学习和记忆能力受损。综上所述，亚硝酸盐慢性暴露诱导氧化应激发生，使机体抗氧化能力下降、脑组织结构损伤，进而造成神经递质系统功能紊乱及行为异常。Abstract: To investigate the neurotoxic effects of nitrite on zebrafish and its possible mechanisms, adult male zebrafish (Danio rerio) were exposed to 0, 0.2, 2 and 20 mg·L-1 nitrite for 30 d. At the end of the experiment, we recorded zebrafish behavior changes and examined alterations in brain histopathology, the neurotransmitter system-related indicators and oxidative-antioxidative system parameters. The result that plasma methemoglobin contents were increased implied that nitrite caused hypoxia. The concentration-dependent increase of malondialdehyde (MDA) content in the brain indicated that zebrafish suffered from nitrite-induced oxidative stress. Coincident with the decrease of total antioxidant capacity (T-AOC), superoxide dismutase (SOD) activity and Cu/Znsod as well as Mnsod mRNA expression levels were depressed in zebrafish brain. Meantime, chronic exposure to nitrite significantly decreased locomotor activity, the tendency to gather in similar fish groups, and learning as well as memory abilities. Brain histopathology results displayed a decrease in the number of neuronal cells in periglomerular gray zone of mesencephalon and the formation of thrombus. The detection of indicators related to the neurotransmitter system showed that dopamine (DA) levels were decreased along with the mRNA levels of genes related to DA metabolism (dat, drd1b and drd4a), cholinergic system (ache, chrna4b and chrna7), the γ-aminobutyric acid (GABA) system (gabra1 and gat1), and 5-hydroxytryptamine receptor family (htr1aa, htr1ab and htr2a). In summary, chronic exposure to nitrite caused oxidative stress and affected zebrafish brain structure, which further leads to behavioral abnormalities and dysfunction of the neurotransmitter system.
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Key words:
- nitrite /
- zebrafish /
- neurotoxicity /
- neurotransmitters /
- oxidative stress
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