有机磷类杀虫剂对脑突触体烟碱型自主受体功能的影响

目的　探讨有机磷杀虫剂 (OPs)对脑突触体烟碱型自主受体功能 (NAF)的影响 ,揭示OPs可能存在的其他作用途径。方法　 (1)体外实验 :以放射性核素氚标记的胆碱与大鼠脑突触体共同温育后用生理缓冲液灌流 ,根据不同有机磷刺激时的放射性胆碱释放量计算NAF值 ,分析氧毒死蜱和对氧磷分别加入灌流系统后NAF的变化 ;(2 )体内实验 :用毒死蜱给大鼠染毒 ,96h后取其大脑皮质制备突触体 ,测定其NAF和乙酰胆碱酯酶 (AChE)活力。结果　对氧磷在体外对NAF影响较小(平均抑制率 <2 3% ) ,但氧毒死蜱却彻底抑制NAF(平均抑制率 >10 0 % )。体内实验亦显示 ,毒死蜱在抑制AChE活力 (抑制率为 91% )的同时 ,对NAF有较强的阻断作用 (抑制率为 6 6 % )。结论　不同的OPs对脑突触体NAF的作用不同 ,毒死蜱可阻断成年大鼠脑突触体的NAF。     

甲胺磷和乙酰甲胺磷对大鼠脑组织内游离钙及环腺苷酸的影响

目的　研究高毒有机磷化合物甲胺磷与其替代品乙酰甲胺磷对大鼠脑组织内游离钙[Ca2 ]i及环腺苷酸 (cAMP)的影响。方法　SD大鼠随机分为 2个染毒组与对照组 ,染毒组连续 5d分别腹腔注射 1 2 0LD50 甲胺磷和乙酰甲胺磷 ;以Fura - 2 AM为荧光指示剂 ,RF - 5 30 1PC荧光分光光度计测定大鼠神经细胞 [Ca2 ]i浓度的变化 ;FJ - 2 0 0 3 2 0 0γ免疫计数器测定大鼠脑组织cAMP含量。结果　甲胺磷染毒组大鼠皮层及海马细胞 [Ca2 ]i分别为 (32 3.86± 2 3.6 0 )、(2 80 .79± 2 1.92 )nmol L ;乙酰甲胺磷染毒组大鼠皮层及海马细胞 [Ca2 ]i分别为 (2 2 5 .2 4± 19.86 )、(2 17.84± 11.0 3)nmol L。甲胺磷组大鼠皮层及海马 [Ca2 ]i高于对照组 [(2 0 9.82± 10 .73)、(199.0 1± 17.17)nmol L],差异有显著性 (P <0 .0 1)。甲胺磷染毒组大鼠皮层及海马cAMP含量为 (35 .2 4± 7.6 3)、(75 .36± 11.31)pmol mg;乙酰甲胺磷染毒组大鼠皮层及海马cAMP含量为 (30 .75± 5 .76 )、(6 9.5 7± 12 .5 9)pmol mg,两者与对照组 [(2 9.4 3± 4 .73)、(6 0 .99± 7.4 1)pmol mg]的差异均无显著性 (P >0 .0 5 )。结论　甲胺磷和乙酰甲胺磷对第二信使影响存在差异 ,可能是两者最终神经毒性作用不同的原因之一。     

拟除虫菊酯对大鼠脑组织γ-氨基丁酸转氨酶活力的影响

目的　研究拟除虫菊酯对大鼠脑组织γ 氨基丁酸转氨酶 (GABAT)活力的影响。方法　应用偶联酶学紫外分光光度法 ,在体外和整体实验中 ,观察溴氰菊酯 (DM)和氯菊酯 (PM)对大鼠大脑皮层、海马、纹状体和小脑GABAT活力的影响。结果　在体外 ,终浓度为 10 -9～ 10 -4mol/L的DM或PM对大鼠大脑皮层、海马、纹状体和小脑的GABAT活力无明显影响 ;一次性经口给予 37.5mg/kg的DM组大鼠大脑皮层、海马和小脑的GABAT活力分别为 (2 .96± 0 .4 3)、(2 .13± 0 .4 4 )、(5 .12± 1.36 )nmol·mgpro-1·min-1,低于对照组 [(3.4 3± 0 .4 1)、(2 .6 8± 0 .4 7)、(6 .74± 1.6 4 )nmol·mgpro-1·min-1],差异有显著性 (P <0 .0 5 ) ,6 0 0mg/kgPM一次性染毒组大鼠大脑皮层和海马的GABAT活力分别为(4.5 7± 0 .30 )、(4.18± 0 .6 3)nmol·mgpro-1·min-1,高于对照组 ,差异有显著性 (P <0 .0 5 ) ;连续 5d每天 1次经口给予 12 .5mg/kg的DM或 2 0 0mg/kg的PM ,均对大鼠大脑皮层、海马、纹状体和小脑的GABAT活力无明显影响。结论　在体外 ,DM和PM均对大鼠脑组织GABAT活力无明显影响 ;在整体实验中 ,DM和PM对大鼠脑组织GABAT活力的影响可能存在差异。     

抗虫灵对大 鼠全血和人红细胞膜乙酰胆碱酯酶的抑制作用

目的应用Ellman法或羟胺法检测乙酰胆碱酯酶(AChE)活性.方法研究有机磷杀虫剂抗虫灵对大鼠全血和人红细胞膜AchE活性的影响.结果雄性大鼠每天经口给以抗虫灵6.81mg/kg连续5d可引起全血AChE活性明显下降;动物亚慢性90d染毒后,雄性中剂量(0.25mg/kg)和高剂量(1.00mg/kg)组动物与雌性低剂量(0.05mg/kg)和高剂量(1.00mg/kg)组动物全血AChE活性明显下降;体外试验发现,抗虫灵在所试验的5×10-8～5×10-8mol/L的浓度范围内对人红细胞膜AChE活性都有明显抑制作用,并有剂量和时间依赖性,而且这种抑制为不可逆抑制.结论抗虫灵对AChE活性的抑制是其毒作用的重要机理,符合有机磷杀虫剂毒作用的一般规律.     

固相萃取-气相色谱-质谱联用法测定人血清中甲胺磷和乙酰甲胺磷

目的建立一种人血清中乙酰甲胺磷及其代谢物甲胺磷的测定方法,即固相萃取-气相色谱-质谱联用法。方法取0.3ml待测血清,用去离子水稀释至3ml,经固相萃取小柱(Carb柱,400mg)提取,目标化合物经丙酮洗脱,氮气吹干后,加入10μl的内标物(苯酚),再用丙酮定容至1ml,取1μl进样。结果甲胺磷和乙酰甲胺磷的线性范围分别为20～100mg/L和10～80mg/L;方法的加标回收率范围为:92.1%～114%;RSD为:3.5%-10.4%。当取样量为0.3ml时,甲胺磷和乙酰甲胺磷的最低检出限分别为10 mg/L和5 mg/L。结论该方法分析快速,定性、定量准确,完全能满足突发性中毒事件的确证分析。     

氯解磷定对不同有机磷杀虫剂中毒后乙酰胆碱酯酶活力恢复的影响

目的比较氯解磷定（PAM-Cl）对急性甲胺磷、敌敌畏和氧乐果中毒后乙酰胆碱酯酶（AChE）活力恢复的影响，为临床上合理治疗提供科学依据。方法收集4省1市7家县、市级医院收治的101例急性有机磷杀虫剂中毒患者资料，按接触杀虫剂品种不同分为急性甲胺磷中毒组59例、急性敌敌畏（或敌百虫）中毒组32例、急性氧乐果中毒组10例，比较3组患者红细胞AChE活力抑制程度和PAM-Cl对AChE的复能作用。结果入院时甲胺磷组、敌敌畏组、氧乐果组红细胞AChE活力分别为（9．12±7．99）、（7．32±4．62）和（12．01±9．53）U／gHb，均受抑制，但差异无统计学意义（P〉O．05）。经PAM-Cl治疗后3组患者中毒症状减轻或消失。甲胺磷组红细胞AChE活力在治疗后12、24、48、72h及出院时分别为（11．37±8．67）、（12．51±6．98）、（15．90±7-31）、（18．33±4．78）及（18．91±7．00）U／gHb，均较入院时明显升高并且呈不断回升趋势，差异有统计学意义（P〈O．01或P〈0．05）。敌敌畏组患者AChE活力在治疗后3、12、24、48、72h及出院时均较入院时明显增高，差异有统计学意义（P〈0．05）；12h后各时间点AChE活力则无进一步上升。氧乐果组患者AChE活力只有出院时较入院时明显增高，差异有统计学意义（P〈0．05）；其余各时间点AChE活力较入院时始终无明显恢复，差异均无统计学意义（P〉0.05）。结论PAM-Cl对甲胺磷、敌敌畏、氧乐果中毒均有治疗作用，对甲胺磷抑制的AChE活力有明显的重活化作用，对敌敌畏（敌百虫）抑制的AChE活力有一定重活化作用，而对氧乐果（乐果）抑制的AChE活力无重活化作用。     

雌二醇对溴氰菊酯染毒大鼠皮层突触体神经毒性的影响

目的观察雌激素对溴氰菊酯染毒动物皮层突触体膜兴奋性氨基酸递质释放和ATPase活力的影响.方法用高效液相色谱(HPLC)检测不同水平17β-雌二醇(10-5、10-8、10-111mol/L)对2×10-5mol/L溴氰菊酯染毒的去卵巢大鼠皮层突触体氨基酸释放的影响;用化学比色法检测17β-雌二醇(10-5、10-8、10-11mol/L)对2×10-4mol/L溴氰菊酯染毒的去卵巢大鼠皮层突触体Na+-K+ATPase、Mg2+-ATPase、Ca2+-ATPase、Ca2+-Mg2+-ATPage活力的影响,并观察雌激素受体拮抗剂Tamoxifen对雌二醇作用的影响.结果2×10-5mol/L氰菊酯处理可增加高钾去极化状态下大鼠皮层突触体天冬氨酸(Asp)和谷氨酸(Glu)的释放;10-8、10-11mol/L的17β-雌二醇可抑制溴氰菊酯所致高钾去极化状态下突触体Asp和Glu的释放,对Asp释放抑制率为28.42%、24.36%,对Glu释放抑制率为21.52%、14.57%;Tamoxifen对17β-雌二醇的抑制作用未见拮抗.2×10-4mol/L溴氰菊酯可抑制突触体4种ATPage活力;10-5mol/L雌二醇可拮抗溴氰菊酯对4种ATPase活力的抑制;l0-8、10-11mol/L雌二醇可增加Ca2+-ATPase活力;Tamoxifen对雌二醇的拮抗作用仍未见明显影响.结论 17β-雌二醇对溴氰菊酯染毒所致皮层突触体兴奋性氨基酸递质释放增加和ATPase活力抑制有一定的保护作用,该作用可能是雌二醇非基因学机制的表现.     

溴氰菊酯对脑代谢型谷氨酸受体结合功能的影响

为探讨溴氰菊酯对大鼠脑组织代谢型谷氨酸受体 (m Glu R)的影响 ,采用放射性配基受体结合试验研究了溴氰菊酯对大鼠大脑皮层和海马 m Glu R结合功能的影响。结果表明 :溴氰菊酯分别在 2× 10 - 8～ 2× 10 - 4 mol/ L和 2× 10 - 1 2 ～2× 10 - 4 mol/ L剂量范围内能增加大脑皮层和海马突触膜 m Glu R与 3H - Glu的特异结合量 ,并呈现剂量效应关系。与对照组相比 ,溴氰菊酯分别在 2× 10 - 6 ～ 2× 10 - 4 mol/ L和 2× 10 - 1 0 ～ 2× 10 - 4 m ol/ L剂量范围内显著增加大脑皮层和海马突触膜m Glu R与 3H- Glu的特异结合量。这一体外试验结果说明溴氰菊酯能增强大脑皮层和海马突触膜 m Glu R对 Glu的亲和力     

混配农药施药员的生物监测

目的 探讨混配农药中的拟除虫菊酯对有机磷抑制红细胞乙酰胆碱酯酶（AChE）作用的影响。方法 在江苏和山东省随机选择1周内未接触任何农药的健康施药员作为生物监测对象,其中,混配农药组（简称混配组）65人,使用甲胺磷与溴氰菊酯（江苏省）或甲基对硫磷与氯氰菊酯（山东省）混配的农药;单一农药组（简称单一组）67人,使用甲胺磷（江苏省）或甲基对硫磷（山东省）。喷药时间为2h,其个人防护等外部条件均与本地平常情况一致。同时在当地乡镇卫生院随机选择未接触农药的健康医务人员2人为对照组。使用硫代乙酰胆碱二硫代双硝基苯甲酸（DTNB）分光光度法,分别测定喷药前、喷药后即刻和喷药后1h的红细胞AChE活力。结果 喷药后混配组和单一组的红细胞AChE活力均受到抑制,与喷药前相比差异有高度显著性（P＜0．01）;红细胞AChE活力的喷药后即刻抑制率和喷药后1h恢复率,混配组均高于单一组,但差异均无显著性（t=－0．9270,df=64.1,P=0.3574;t=-1.3413,df=64.3,P=0.1845)。结论 本次监测未观察到混配农药中的拟除虫菊酯对有机磷抑制红细胞AChE活力的作用有明显影响。     

溴氰菊酯对大鼠脑中苄氧基试卤灵O-脱烷基化酶活力及CYP2B1/2B2蛋白水平的影响

目的　研究溴氰菊酯 (deltamethrin ,DM)对大鼠脑组织中苄氧基试卤灵O 脱烷基化酶(benzyloxyresorufinO dealkylatase,BROD)活力及CYP2B1 / 2B2蛋白水平的影响。方法　采用荧光分光光度法 ,在体内、体外研究了DM对大鼠脑组织中BROD活力的影响 ,并用Western blot法检测了DM对CYP2B1 / 2B2蛋白水平的影响。结果　大鼠连续 5d腹腔注射DM 1 2 .5mg·kg- 1 ·d- 1 染毒后 ,其全脑、大脑皮层及小脑BROD活力明显被抑制 ,其抑制率分别是 2 6 .7%、2 3 .8%和 33 .3 % ,差异均有显著性(P <0 .0 5) ;在体外 ,DM浓度在 2× 1 0 - 8～ 2× 1 0 - 4mol/L范围内对BROD活力无明显影响 ;DM在体内明显降低小脑中CYP2B1 / 2B2蛋白水平 ,其酶蛋白合成抑制率为 42 .6 %。结论　DM在体内能明显抑制脑内BROD活力 ,其机制与抑制该酶蛋白合成有关     

Molecular properties and inhibition kinetics of acetylcholinesterase obtained from rat brain and cockroach ganglion.

1. The molecular composition of acetylcholinesterase (AChE) obtained from cockroach neural, and rat brain tissues was different. Vertebrate enzyme exhibited a higher degree of polymerization than insect enzyme. 2. Acephate was a potent inhibitor of cockroach AChE, but a poor inhibitor of rat AChE. Unlike acephate, methamidophos was a potent inhibitor of both cockroach and rat enzymes. Acephate exhibited greater affinity for the cockroach-AChE than for the rat-AChE, and acephate phosphorylated the cockroach-AChE several times faster than the rat enzyme. The rate of phosphorylation of insect and rat AChE was similar in the presence of methamidophos. Solubilization of AChE by Triton X-100 altered the kinetics of inhibition of rat AChE by acephate. However, solubilization did not alter the kinetics of inhibition of rat AChE by methamidophos or the kinetics of inhibition of cockroach AChE by acephate or methamidophos. 3. The mechanism of acephate-cockroach AChE interaction was different than the mechanism of acephate-rat AChE interaction. It is proposed that both the rat and cockroach enzyme may contain, along with the anionic and esteratic sites, an "electron deficient" (ED) binding site which may exhibit selectivity for acephate and nefopam. The ED site in rat-AChE has allosteric properties, whereas the cockroach-AChE does not. It is also proposed that the ED site in cockroach-AChE may be situated in or adjacent to the active site and, therefore, acephate may be bound to the ED site such that the phosphate moiety of acephate interacts with the enzyme's "esteratic" site. Although nefopam also bound to the ED site in cockroach AChE, it did not inhibit the enzyme. This study also indicated that the ED site in rat-AChE may be peripheral to the active site, and that the binding of acephate to this site prevented the phosphorylation by methamidophos of the rat-AChE. Unlike acephate, methamidophos specifically bound to the active site in both the rat- and cockroach-AChE.     

Biological monitoring of human exposure to acephate

Acephate is a water-soluble organophosphate insecticide whose action on insects has been related to its conversion to methamidophos, a very potent anticholinesterase agent which has caused delayed neuropathy in man.Inhalation and skin exposure to acephate was evaluated in four workers engaged in 8-day campaigns with the formulation of the 97%-pure technical product. Before, during, and after exposure, the workers were monitored for the urine content of acephate and methamidophos, and for erythrocyte (AChE) and plasma (PChE) cholinesterase levels. Median air concentrations (8-hr TWA) ranged from 0.278 to 2.170 mg/m³; median total-body skin deposition ranged from 26.1 to 41.9 mg/day. Based on these values, daily workers' absorption of acephate was estimated to be in the order of 10–20 mg. Urinary excretion of unchanged acephate followed a pattern consistent with exposure, showing peak values of excretion during the workshift or in the eight hr after the end of the workshift. The urine levels of unchanged acephate were found to vary from 1 to 10 mg/L. Methamidophos was not detected in any urine sample (detection limit: 30 u.g/L). High correlation (r=0.78) was found between skin exposure level and urine acephate elimination. No changes in AChE or PChE were observed for the workers whose urinary concentrations of acephate were 1 or 2 mg/L. One subject who had urinary acephate excretion between 3 and 8 mg/L, showed slightly decreased values of PChE during exposure and of AChE after exposure.     

Clonal variation in acetylcholinesterase biomarkers and life history traits following OP exposure in Daphnia magna

Two clones of Daphnia magna (Standard and Ruth) were exposed for 7 days to sub-lethal concentrations of acephate (5.0 and 10.0 mg/L). Survivorship, individual growth, reproduction and the population growth rate (lambda) were evaluated over three weeks. Acetylcholinesterase (AChE) activity was measured on days 2, 7 and 21. Acephate exposure inhibited AChE activity but had no direct effect on life history (LH) traits. There was also no effect of clone on AChE activity, LH and lambda. However, a significant interaction between clone and acephate concentration was found on both fecundity and lambda. AChE inhibition at 48 h was associated with a decrease in lambda in the Standard clone and an increase in lambda in clone Ruth. Therefore, our findings show that genotypic variation will influence the link between AChE activity and toxic effects at higher levels of biological organisation in D. magna.     

Potential of the insecticides acephate and methamidophos to contaminate groundwater

The possible contamination of groundwater by the insecticides acephate and methamidophos was assessed using the behavior assessment model (BAM) and the groundwater pollution-potential model (GWP). The dissipation coefficients of the two insecticides in two soils (Annei silt loam and Pingchen silt clay loam) at different moisture contents (50 and 100% field capacity) and soil temperatures (20 and 30 degrees C) were studied by determining the degradation and adsorption of each insecticide in the soil. The movement of acephate and methamidophos was studied by leaching each insecticide in a soil column in the laboratory. The absorption coefficient of methamidophos was much higher than that of acephate in both types of soil. In the leaching test, methamidophos more easily leached out from the Pingchen soil column than from the Annei soil column. Methamidophos was rapidly degraded, with a half-life of 1.11 to 1.61 days in the Annei soil and 7.50 to 13.20 days in the Pingchen soil at different temperatures and soil water contents. Acephate was found to have a longer half-life than methamidophos in soil; however, the mobility of methamidophos in both soils was slower than that of acephate. The mobility of acephate in soil is somewhat faster than that of methamidophos, and thus acephate may lead to the contamination of groundwater much more easily than methamidophos under normal conditions.     

Toxicity of acephate and methamidophos to dark-eyed juncos

The calculated, acute oral LD₅₀ of acephate and methamidophos to dark-eyed juncos (Junco hyemalis) was 106 mg/kg and 8 mg/kg, respectively. Brain cholinesterase (ChE) activity in birds that died after acephate poisoning was depressed 80% below that of control birds. Birds that died of acute methamidophos poisoning had brain ChE depression of 60%. The birds killed by acephate had brain acephate residue concentrations greater than 2 mg/kg and methamidophos concentrations usually greater than 0.25 mg/kg. Eighty percent of the birds killed with methamidophos had brain methamidophos concentrations greater than 0.1 mg/kg. The five-day feeding LC₅₀ for acephate was 1485 mg/kg. Brain ChE activities of birds which died early in the study were less depressed (51.5%) than those which died at a later date (69.6%). Brain residues of acephate and methamidophos were lower in these birds than in the birds of the acute oral LD₅₀ studies. Brain ChE activity returned to normal within three days after the birds received a single sublethal dose of acephate. These studies indicate that the amount of acephate needed to produce the ChE depression found in other investigations in most dark-eyed juncos exposed to forest applications of insecticide is about one-fifth of the LD₅₀; however, in a few birds the ChE activity may be depressed to near lethal levels.Formerly, Pacific Southwest Forest and Range Experiment Station, Forest Service, U.S. Department of Agriculture, Berkeley, CA 94701.     

Neurotoxic effects of low-level chronic acephate exposure in rats

Rats were administered the organophosphorus insecticide acephate at 1.0 or 10.0 mg/kg.day for 15 weeks. Blood and brain samples were collected at the end of the treatment and analyzed for cholinesterase, acetylcholinesterase, and glutamic acid decarboxylase activities and catecholamine and amino acid levels. No significant inhibition in the activity of brain AChE was noted at doses of 1.0 or 10.0 mg/kg.day. Low levels of acephate exposure (1.0 mg/kg.day), which did not alter plasma cholinesterase or RBC acetylcholinesterase activity levels, resulted in a significant elevation of plasma epinephrine and norepinephrine levels. Decreased GABA, dopamine, and tyrosine levels and glutamic acid decarboxylase activity were observed in brains of these rats. Similar changes occurred in rats exposed to 10 mg of acephate/kg.day; however, plasma cholinesterase and RBC acetylcholinesterase activities were inhibited. These observations suggest that chronic exposure to acephate altered the activity of the noncholinergic system without altering the cholinergic activity, and that low-level chronic exposure to organophosphorous compounds cannot be predicted by measuring the ChE or AChE enzyme activities.     

Concentration effects of selected insecticides on brain acetylcholinesterase in the common carp (Cyprinus carpio L.)

The differential inhibition of acetylcholinesterase (AChE) by organophosphate (OP) and carbamate (C) is followed by the distinct duration of exposure effect on common carp AChE. Hence, in the present study in vivo exposure period effect and in vitro concentration-response of chlorfenvinphos, chlorpyrifos diazinon, and carbofuran were investigated on Cyprinus carpio L. AChE. Individuals of 1-year-old carp were exposed for 96 h to different concentrations of insecticides, after which AChE activity was measured in the brain. The highest concentrations of carbofuran (2.44 mg x L(-1)), chlorfenvinphos (2.9 mg x L(-1)), and diazinon (2.5 mg x L(-1)) killed all the test animals after only 4 h, although there was no statistically significant difference from the control group's brain AChE activity. The lowest concentration significantly inhibited brain AChE after 96 h. Chlorfenvinphos was the most potent inhibitor in vivo and chlorpyrifos the least active inhibitor after 96 h of exposure time. In vitro experimentation with the same pesticide indicated that several concentrations inhibited 50% of the AChE activity (I50), ranging from 4.1x10(-7) to 8.12x10(-4) M in both single inhibitory action and joint inhibitory effect. The results suggest that in biomonitoring programs carp brain AChE can be a good diagnostic tool for chronic OP nd C pollution.     

拟除虫菊酯对大鼠脑组织及肝脏生物转化酶的影响

目的 实验研究溴氰菊酯（ＤＭ）和氯菊酯（ＰＭ）对大鼠脑组织及肝脏某些生物转化酶的影响。方法 应用荧光分光光度法、二硝基苯肼比色法和Ｈａｂｉｇ法检测７－乙氧基试卤灵－Ｏ－脱乙基酶９ＥＲＯＤ０、Ｂ型单胺氧化酶９ＭＡＯＢ）和谷胱甘肽Ｓ－转移酶（ＧＳＴ）活力。结果 整体实验中，ＤＭ染毒动物肝脏ＥＲＯＤ、ＭＡＯＢ活力分别下降了４７．９４％和２２．７６％；脑组织和肝脏ＧＳＴ活力分别增加了２２．２０％和３９．５