氨基脲诱导的大鼠神经行为毒性及其生化机制
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摘要
为探究氨基脲(SEM)染毒对SD大鼠的神经行为毒性及其作用机制,将44只SPF级SD雄性大鼠随机分为4组:对照组,SEM低、中、高剂量组,每组11只,分别以0,7.5,15,30mg/(kg·bw)SEM的剂量连续灌胃染毒28d.染毒前后分别通过旷场实验和高架十字迷宫实验测试神经行为.采用液相色谱法测定γ-氨基丁酸(GABA)和谷氨酸(GLU),ELISA法测定5-羟色胺(5-HT)、去甲肾上腺素(NE)、多巴胺(DA)、单胺氧化酶(MAO)以及N-甲基-D-天氡氨酸受体(NMDAR)含量.结果显示,高剂量染毒组大鼠在旷场实验中运动总距离和中央区域距离显著低于对照组(P<0.05),各剂量组大鼠在高架十字迷宫实验中进入开臂时间百分比和进入开臂次数百分比均显著低于对照组(P<0.05或P<0.01).与对照组相比,各染毒组大鼠脑组织中GABA含量和MAO活性均有不同程度降低,而GLU,NMDAR,5-HT,NE和DA水平均有不同程度上升.SEM诱导大鼠神经行为毒性的机制与破坏GABA和GLU的相互转化、增加NMDAR含量以及抑制MAO活性导致单胺类神经递质水平上升3个途径有关.
To study the neurobehavioral toxicity of semicarbazide(SEM) in rats and its biochemical mechanism. A total of 44 male SD rats were randomly divided into 4 groups(n=11 per group): control group, low-dose group, medium-dose group, and high-dose group. SEM was intragastrically administrates at the dosage of 0, 7.5, 15, 30 mg/kg(body weight) for 28 days. The open field test(OFT) and elevated plus maze(EPM) were used to evaluated the neurobehavior in rats before and after exposure to SEM. The gamma-aminobutyric acid(GABA) and glutamate acid(GLU) of hippocampus were determined by liquid chromatography. ELISA method was used to mesure the contents of 5-hydroxytryptamine(5-HT), norepinephrine(NE), dopamine(DA), monoamine oxidase(MAO) and N-methyl-d-aspartic acid receptor(NMDAR). The results showed that the total distance traveled and the distance traveled in the center were significantly lower in the high-dose SEM group compared with the control group(P<0.05). The open arm time percentage and open arm entries percentage were significantly lower in the SEM groups than in the control group(P<0.05 or P<0.01). Compared with the control group, the concentrations of GABA and the activity of MAO were decreased, whereas the levels of GLU, NMDAR, 5-HT, NE and DA were increased in different exposed groups. SEM might affect the nervous system of rats by damaging the mutual transformation of GABA and GLU, increasing the concentration of NMDAR and inhibiting the activity of MAO that directly enhance the levels of monoamine neurotransmitters.
To study the neurobehavioral toxicity of semicarbazide(SEM) in rats and its biochemical mechanism. A total of 44 male SD rats were randomly divided into 4 groups(n=11 per group): control group, low-dose group, medium-dose group, and high-dose group. SEM was intragastrically administrates at the dosage of 0, 7.5, 15, 30 mg/kg(body weight) for 28 days. The open field test(OFT) and elevated plus maze(EPM) were used to evaluated the neurobehavior in rats before and after exposure to SEM. The gamma-aminobutyric acid(GABA) and glutamate acid(GLU) of hippocampus were determined by liquid chromatography. ELISA method was used to mesure the contents of 5-hydroxytryptamine(5-HT), norepinephrine(NE), dopamine(DA), monoamine oxidase(MAO) and N-methyl-d-aspartic acid receptor(NMDAR). The results showed that the total distance traveled and the distance traveled in the center were significantly lower in the high-dose SEM group compared with the control group(P<0.05). The open arm time percentage and open arm entries percentage were significantly lower in the SEM groups than in the control group(P<0.05 or P<0.01). Compared with the control group, the concentrations of GABA and the activity of MAO were decreased, whereas the levels of GLU, NMDAR, 5-HT, NE and DA were increased in different exposed groups. SEM might affect the nervous system of rats by damaging the mutual transformation of GABA and GLU, increasing the concentration of NMDAR and inhibiting the activity of MAO that directly enhance the levels of monoamine neurotransmitters.
引文
[1]高素,汝少国.氨基脲的毒性效应研究进展[J].环境科学研究,2013,(6):637-644.
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[4]Pereira A S,Donato J L,De Nucci G.Implications of the use of semicarbazide as a metabolic target of nitrofurazone contamination in coated products[J].Food Additives&Contaminants,2004,21(1):63-69.
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[7]谭志军,翟毓秀,冷凯良,等.呋喃西林和呋喃唑酮代谢物在大菱鲆组织中的消除规律[J].中山大学学报(自然科学版),2008,47(S1):63-69.
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[9]Ye J,Wang X H,Sang Y X,et al.Assessment of the Determination of Azodicarbonamide and Its Decomposition Product Semicarbazide:Investigation of Variation in Flour and Flour Products[J].Journal of Agricultural&Food Chemistry,2011,59(17):9313-9318.
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[11]朱乐玫,袁萍,张贝贝,等.原花青素对氨基脲致雄性小鼠生殖毒性的拮抗作用[J].实用预防医学,2012,19(2):165-168.
[12]周一平.用Excel软件进行药物毒理实验的随机分组[J].药学进展,2005,29(9):425-427.
[13]高素.氨基脲对斑马鱼(Danio rerio)抗雌激素效应研究[D].山东:中国海洋大学,2013.
[14]GB15193-2014食品安全性毒理学评价程序和方法[S].
[15]俞军龄,陈再兴,毛小元,等.HPLC法测定tremor大鼠脑组织中谷氨酸和GABA含量[J].中国药理学通报,2009,25(11):1530-1533.
[16]王维刚.旷场实验在小鼠行为分析中的应用[J].中国细胞生物学学报,2016,33(11):1191-1196.
[17]范兴文,贯士阔,吴开良.脑部照射对大鼠情绪和记忆的影响[J].中国癌症杂志,2014,24(11):814-819.
[18]王维刚,吴文婷,周嘉斌,等.小鼠动物实验方法系列专题(五)应用高架十字迷宫分析小鼠焦虑行为[J].中国细胞生物学学报,2011,5:466-472.
[19]刘友平,刘田,毕航,等.品系、性别和生物节律对小鼠旷场实验的影响[J].西安交通大学学报(医学版),2014,35(5):634-638.
[20]孙世光,李自发,孙鹏,等.昆明小鼠旷场实验评价方法的三维结构[J].中华行为医学与脑科学杂志,2011,20(10):875-877.
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[22]雷娜,鲁亚平.γ-氨基丁酸生理机理研究进展[J].清远职业技术学院学报,2011,4(3):9-11.
[23]Kah O,Trudeau V L,Sloley B D,et al.Influence of GABA on gonadotrophin release in the goldfish[J].Neuroendocrinology,1992,55(4):396-404.
[24]Trudeau V L,Spanswick D,Fraser E J,et al.The role of amino acid neurotransmitters in the regulation of pituitary gonadotropin release in fish[J].Biochemistry and cell biology=Biochimie et biologie cellulaire,2000,78(3):241-259.
[25]李连娥,唐涛,李亚国,等.阿尔茨海默病和衰老中γ-氨基丁酸系统与学习记忆关系的研究进展[J].昆明理工大学学报(自然科学版),2017,6:81-85.
[26]Santos J M,Macedo C E,Brand?o M L.Gabaergic mechanisms of hypothalamic nuclei in the expression of conditioned fear[J].Neurobiology of Learning&Memory,2008,90(3):560-568.
[27]Maranghi F,Tassinari R,Lagatta V,et al.Effects of the food contaminant semicarbazide following oral administration in juvenile Sprague-Dawley rats.[J].Food&Chemical Toxicology,2009,47(2):472-479.
[28]Gore A C.Gonadotropin-releasing hormone neurons,NMDA receptors,and their regulation by steroid hormones across the reproductive life cycle[J].Brain Research Reviews,2001,37(1-3):235-248.
[29]李欢欢,林文娟.中枢N-甲基-D-天冬氨酸受体在应激所致行为改变中的作用[J].心理科学进展,2005,13(3):320-326.
[30]区硕俊,罗展远,曾广丰,等.UPLC-Q-TOF-MS法测定小鼠大脑中4种单胺类神经递质的含量[J].分析测试学报,2016,35(12):1569-1574.
[31]Sardella R,Scorzoni S,Conte C,et al.Novel orthogonal liquid chromatography methods to dose neurotransmitters involved in Parkinson's disease[J].Journal of Pharmaceutical&Biomedical Analysis,2014,98:253-259.
[32]赖玉婷,欧阳俊彦,郭佑廷,等.壬基酚对大鼠5-羟色胺分解代谢通路的影响[J].中国环境科学,2014,34(9):2408-2412.
[33]刘焕,杨婕,黄少文,等.桑葚粗提液对壬基酚诱导大鼠焦虑行为的干预作用及其机制[J].食品工业科技,2017,38(14):294-298.
[34]喻东山,梅安昌.惊恐障碍的发生机制[J].四川精神卫生,2009,22(2):122-124.
[2]李嘉.食品添加剂副产物氨基脲的毒理学研究[D].吉林:吉林农业大学,2008.
[3]Cooper K M,Samsonova J V,Plumpton L,et al.Enzyme immunoassay for semicarbazide-The nitrofuran metabolite and food contaminant[J].Analytica Chimica Acta,2007,592(1):64.
[4]Pereira A S,Donato J L,De Nucci G.Implications of the use of semicarbazide as a metabolic target of nitrofurazone contamination in coated products[J].Food Additives&Contaminants,2004,21(1):63-69.
[5]Diblikova I,Cooper K M,Kennedy D G,et al.Monoclonal antibody-based ELISA for the quantification of nitrofuran metabolite3-amino-2-oxazolidinone in tissues using a simplified sample preparation[J].Analytica Chimica Acta,2005,540(2):285-292.
[6]Cooper K M,Caddell A,Elliott C T,et al.Production and characterisation of polyclonal antibodies to a derivative of3-amino-2-oxazolidinone,a metabolite of the nitrofuran furazolidone[J].Analytica Chimica Acta,2004,520(1):79-86.
[7]谭志军,翟毓秀,冷凯良,等.呋喃西林和呋喃唑酮代谢物在大菱鲆组织中的消除规律[J].中山大学学报(自然科学版),2008,47(S1):63-69.
[8]Khong S P,Gremaud E,Richoz J,et al.Analysis of matrix-bound nitrofuran residues in worldwide-originated honeys by isotope dilution high-performance liquid chromatography-tandem mass spectrometry[J].Journal of Agricultural&Food Chemistry,2004,52(17):5309-5315.
[9]Ye J,Wang X H,Sang Y X,et al.Assessment of the Determination of Azodicarbonamide and Its Decomposition Product Semicarbazide:Investigation of Variation in Flour and Flour Products[J].Journal of Agricultural&Food Chemistry,2011,59(17):9313-9318.
[10]Maranghi F,Tassinari R,Marcoccia D,et al.The food contaminant semicarbazide acts as an endocrine disrupter:Evidence from an integrated in vivo/in vitro approach.[J].Chemico-biological interactions,2010,183(1):40-48.
[11]朱乐玫,袁萍,张贝贝,等.原花青素对氨基脲致雄性小鼠生殖毒性的拮抗作用[J].实用预防医学,2012,19(2):165-168.
[12]周一平.用Excel软件进行药物毒理实验的随机分组[J].药学进展,2005,29(9):425-427.
[13]高素.氨基脲对斑马鱼(Danio rerio)抗雌激素效应研究[D].山东:中国海洋大学,2013.
[14]GB15193-2014食品安全性毒理学评价程序和方法[S].
[15]俞军龄,陈再兴,毛小元,等.HPLC法测定tremor大鼠脑组织中谷氨酸和GABA含量[J].中国药理学通报,2009,25(11):1530-1533.
[16]王维刚.旷场实验在小鼠行为分析中的应用[J].中国细胞生物学学报,2016,33(11):1191-1196.
[17]范兴文,贯士阔,吴开良.脑部照射对大鼠情绪和记忆的影响[J].中国癌症杂志,2014,24(11):814-819.
[18]王维刚,吴文婷,周嘉斌,等.小鼠动物实验方法系列专题(五)应用高架十字迷宫分析小鼠焦虑行为[J].中国细胞生物学学报,2011,5:466-472.
[19]刘友平,刘田,毕航,等.品系、性别和生物节律对小鼠旷场实验的影响[J].西安交通大学学报(医学版),2014,35(5):634-638.
[20]孙世光,李自发,孙鹏,等.昆明小鼠旷场实验评价方法的三维结构[J].中华行为医学与脑科学杂志,2011,20(10):875-877.
[21]Moraes C L,Bertoglio L J,Carobrez A P.Interplay between glutamate and serotonin within the dorsal periaqueductal gray modulates anxiety-related behavior of rats exposed to the elevated plus-maze[J].Behavioural Brain Research,2008,194(2):181-186.
[22]雷娜,鲁亚平.γ-氨基丁酸生理机理研究进展[J].清远职业技术学院学报,2011,4(3):9-11.
[23]Kah O,Trudeau V L,Sloley B D,et al.Influence of GABA on gonadotrophin release in the goldfish[J].Neuroendocrinology,1992,55(4):396-404.
[24]Trudeau V L,Spanswick D,Fraser E J,et al.The role of amino acid neurotransmitters in the regulation of pituitary gonadotropin release in fish[J].Biochemistry and cell biology=Biochimie et biologie cellulaire,2000,78(3):241-259.
[25]李连娥,唐涛,李亚国,等.阿尔茨海默病和衰老中γ-氨基丁酸系统与学习记忆关系的研究进展[J].昆明理工大学学报(自然科学版),2017,6:81-85.
[26]Santos J M,Macedo C E,Brand?o M L.Gabaergic mechanisms of hypothalamic nuclei in the expression of conditioned fear[J].Neurobiology of Learning&Memory,2008,90(3):560-568.
[27]Maranghi F,Tassinari R,Lagatta V,et al.Effects of the food contaminant semicarbazide following oral administration in juvenile Sprague-Dawley rats.[J].Food&Chemical Toxicology,2009,47(2):472-479.
[28]Gore A C.Gonadotropin-releasing hormone neurons,NMDA receptors,and their regulation by steroid hormones across the reproductive life cycle[J].Brain Research Reviews,2001,37(1-3):235-248.
[29]李欢欢,林文娟.中枢N-甲基-D-天冬氨酸受体在应激所致行为改变中的作用[J].心理科学进展,2005,13(3):320-326.
[30]区硕俊,罗展远,曾广丰,等.UPLC-Q-TOF-MS法测定小鼠大脑中4种单胺类神经递质的含量[J].分析测试学报,2016,35(12):1569-1574.
[31]Sardella R,Scorzoni S,Conte C,et al.Novel orthogonal liquid chromatography methods to dose neurotransmitters involved in Parkinson's disease[J].Journal of Pharmaceutical&Biomedical Analysis,2014,98:253-259.
[32]赖玉婷,欧阳俊彦,郭佑廷,等.壬基酚对大鼠5-羟色胺分解代谢通路的影响[J].中国环境科学,2014,34(9):2408-2412.
[33]刘焕,杨婕,黄少文,等.桑葚粗提液对壬基酚诱导大鼠焦虑行为的干预作用及其机制[J].食品工业科技,2017,38(14):294-298.
[34]喻东山,梅安昌.惊恐障碍的发生机制[J].四川精神卫生,2009,22(2):122-124.