Tissue distribution and urinary excretion of inorganic arsenic and its methylated metabolites in mice following acute oral administration of arsenate.

The relationship of exposure dose and tissue concentration of parent chemical and metabolites is a critical issue in cases where toxicity may be mediated by a metabolite or by parent chemical and metabolite acting together. This has emerged as an issue for inorganic arsenic (iAs), because both its trivalent and pentavalent methylated metabolites have unique toxicities; the methylated trivalent metabolites also exhibit greater potency than trivalent inorganic arsenic (arsenite, As(III)) for some endpoints. In this study, the time-course tissue distributions for iAs and its methylated metabolites were determined in blood, liver, lung, and kidney of female B6C3F1 mice given a single oral dose of 0, 10, or 100 micromol As/kg (sodium arsenate, As(V)). Compared to other organs, blood concentrations of iAs, mono- (MMA), and dimethylated arsenic (DMA) were uniformly lower across both dose levels and time points. Liver and kidney concentrations of iAs were similar at both dose levels and peaked at 1 h post dosing. Inorganic As was the predominant arsenical in liver and kidney up to 1 and 2 h post dosing, with 10 and 100 micromol As/kg, respectively. At later times, DMA was the predominant metabolite in liver and kidney. By 1 h post dosing, concentrations of MMA in kidney were 3- to 4-fold higher compared to other tissues. Peak concentrations of DMA in kidney were achieved at 2 h post dosing for both dose levels. Notably, DMA was the predominant metabolite in lung at all time points following dosing with 10 micromol As/kg. DMA concentration in lung equaled or exceeded that of other tissues from 4 h post dosing onward for both dose levels. These data demonstrate distinct organ-specific differences in the distribution and methylation of iAs and its methylated metabolites after exposure to As(V) that should be considered when investigating mechanisms of arsenic-induced toxicity and carcinogenicity.     

Arsenic metabolites in urine and feces of hamsters pretreated with PCB

High pressure liquid chromatography and graphite tube atomizing atomic absorption spectrometry were used to quantify monomethyl arsonic acid (MMA), dimethyl arsinic acid (DMA), and inorganic arsenic (IA: arsenite plus arsenate) in the urine and feces of male and female hamsters pretreated with a single ip injection of PCB (100 mg/kg) and 4 days later given a single po dose of arsenite (10 mg As/kg). Approximately 17 to 23% and 35 to 63% of the arsenic given was eliminated in the urine and feces, respectively, during the 5 days after the administration of arsenic. Both DMA and MMA were found in the urine but only MMA was detected in the feces, as methylated metabolite. Fecal excretion of arsenic was significantly larger in female than in males. PCB influenced the metabolism of arsenic by significantly increasing the proportion of DMA excreted into the urine of female hamsters during the 5 days after the arsenic administration, but did not alter the total amount of arsenic metabolites in any group of male or female hamsters. PCB did not affect the cumulative amounts of fecal arsenic in any group, although the excretion in the PCB-treated group of females reached the maximum level 1 day earlier than in the controls. These results suggest that the metabolism of arsenic may be regulated by certain sex-relating factors which are influenced by PCB.     

Metabolism and Excretion of Gallium Arsenide and Arsenic Oxides by Hamsters following Intratracheal Instillation

The increasing use of gallium arsenide (GaAs) in the electronicsindustry has produced the need for pharmacokinetic and toxicologicdata on GaAs. The disposition in male Syrian golden hamsters(n = 4) following intratracheal instillation of GaAs (mean volumediameter 5.8 µm), arsenic(III) oxide (arsenite), and arsenic(V)oxide (arsenate) at a dose of 5 mg/kg body weight was examined.Blood, kidney, liver, and lung samples were collected at 1,2, and 4 days after administration. Excreta were collected daily.Urinary metabolite profiles were determined after separationon a mixed anion–cation-exchange column. Total As contentwas analyzed by direct hydride flame atomic absorption spectrophotometryafter digestion. Arsenic blood levels after GaAs, arsenite,and arsenate administration were 0.185 ± 0.041, 0.596± 0.117, and 0.3 10 ± 0.045 ppm, respectively,after Day 1. Arsenic blood levels after GaAs administrationincreased to 0.279 ± 0.021 ppm on Day 2 indicating continuedabsorption while levels decreased for the arsenite and arsenategroups. At Day 1 the liver contained 0.565 ± 0.036, 2.62± 0.26, and 0.579 ± 0.144% of the arsenic doseof GaAs, arsenite, and arsenate, respectively. The" arseniteand arsenate were rapidly excreted in the urine with almosthalf the dose appearing after 4 days; in contrast, only about5% of the GaAs was found at the corresponding time. Total recoveries,as arsenic equivalents, for the three compounds were between75 and 80%. Ratios of the two major urinary metabolites (dimethylarsinicacid/total Inorganic As species) were 1.41, 1.71, and 0.983for GaAs, arsenite, and arsenate, respectively. GaAs is metabolizedto the same compounds as arsenite and arsenate, and shows ametabolic profile most similar to that observed for sodium arsenite.     

Monomethylarsonic acid reductase and monomethylarsonous acid in hamster tissue

The formation of monomethylarsonous acid (MMA(III)) by tissue homogenates of brain, bladder, spleen, liver, lung, heart, skin, kidney, or testis of male Golden Syrian hamsters was assessed using [(14)C]monomethylarsonic acid (MMA(V)) as the substrate for MMA(V) reductase. The mean +/- SEM of MMA(V) reductase specific activities (nanomoles of MMA(III) per milligram of protein per hour) were as follows: brain, 91.4 +/- 3.0; bladder, 61.8 +/- 3.7; spleen, 30.2 +/- 5.4; liver, 29.8 +/- 1.4; lung, 21.5 +/- 0.8; heart, 19.4 +/- 1.5; skin, 14.7 +/- 1.6; kidney, 10.6 +/- 0.4; and testis, 9.8 +/- 0.6. The concentrations of MMA(III) in male Golden Syrian hamster livers were determined 15 h after administration of a single intraperitoneal dose of 145 microCi of [(73)As]arsenate (2 mg of As/kg of body weight). Trivalent arsenic species (arsenite, MMA(III), and dimethylarsinous acid, DMA(III)) were extracted from liver homogenates using carbon tetrachloride (CCl(4)) and 20 mM diethylammonium salt of diethyldithiocarbamic acid (DDDC). Pentavalent arsenicals (arsenate, MMA(V), and dimethylarsinic acid, DMA(V)) remained in the aqueous phase. The organic and the aqueous phases then were analyzed by HPLC. Metabolites of inorganic arsenate present in hamster liver after 15 h were observed in the following concentrations (nanograms per gram of liver +/- SEM): MMA(III), 38.5 +/- 2.9; DMA(III), 49.9 +/- 10.2; arsenite, 35.5 +/- 3.0; arsenate, 118.2 +/- 8.7; MMA(V), 31.4 +/- 2.8; and DMA(V), 83.5 +/- 6.7. This first-time identification of MMA(III) and DMA(III) in liver after arsenate exposure indicates that the significance of arsenic species in mammalian tissue needs to be re-examined and re-evaluated with respect to their role in the toxicity and carcinogenicity of inorganic arsenic.     

Altered arsenic disposition in experimental nonalcoholic Fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) is represented by a spectrum of liver pathologies ranging from simple steatosis to nonalcoholic steatohepatitis (NASH). Liver damage sustained in the progressive stages of NAFLD may alter the ability of the liver to properly metabolize and eliminate xenobiotics. The purpose of the current study was to determine whether NAFLD alters the disposition of the environmental toxicant arsenic. C57BL/6 mice were fed either a high-fat or a methionine-choline-deficient diet to model simple steatosis and NASH, respectively. At the conclusion of the dietary regimen, all mice were given a single oral dose of either sodium arsenate or arsenic trioxide. Mice with NASH excreted significantly higher levels of total arsenic in urine (24 h) compared with controls. Total arsenic in the liver and kidneys of NASH mice was not altered; however, NASH liver retained significantly higher levels of the monomethyl arsenic metabolite, whereas dimethyl arsenic was retained significantly less in the kidneys of NASH mice. NASH mice had significantly higher levels of the more toxic trivalent form in their urine, whereas the pentavalent form was preferentially retained in the liver of NASH mice. Moreover, hepatic protein expression of the arsenic biotransformation enzyme arsenic (3+ oxidation state) methyltransferase was not altered in NASH animals, whereas protein expression of the membrane transporter multidrug resistance-associated protein 1 was increased, implicating cellular transport rather than biotransformation as a possible mechanism. These results suggest that NASH alters the disposition of arsenical species, which may have significant implications on the overall toxicity associated with arsenic in NASH.     

Accumulation and metabolism of arsenic in mice after repeated oral administration of arsenate

Exposure to the human carcinogen inorganic arsenic (iAs) occurs daily. However, the disposition of arsenic after repeated exposure is not well known. This study examined the disposition of arsenic after repeated po administration of arsenate. Whole-body radioassay of adult female B6C3F1 mice was used to estimate the terminal elimination half-life of arsenic after a single po dose of [(73)As]arsenate (0.5 mg As/kg). From these data, it was estimated that steady-state levels of whole-body arsenic could be attained after nine repeated daily doses of [(73)As]arsenate (0.5 mg As/kg). The mice were whole-body radioassayed immediately before and after the repeated dosing. Excreta were collected daily and analyzed for arsenic-derived radioactivity and arsenicals. Whole-body radioactivity was determined 24 h after the last repeated dose, and five mice were then euthanized and tissues analyzed for radioactivity. The remaining mice were whole-body radioassayed for 8 more days, and then their tissues were analyzed for radioactivity. Other mice were administered either a single or nine repeated po doses of non-radioactive arsenate (0.5 mg As/kg). Twenty-four hours after the last dose, the mice were euthanized, and tissues were analyzed for arsenic by atomic absorption spectrometry (AAS). Whole-body radioactivity was rapidly eliminated from mice after repeated [(73)As]arsenate exposure, primarily by urinary excretion in the form of dimethylarsinic acid (DMA(V)). Accumulation of radioactivity was highest in bladder, kidney, and skin. Loss of radioactivity was most rapid in the lung and slowest in the skin. There was an organ-specific distribution of arsenic as determined by AAS. Monomethylarsonic acid was detected in all tissues except the bladder. Bladder and lung had the highest percentage of DMA(V) after a single exposure to arsenate, and it increased with repeated exposure. In kidney, iAs was predominant. There was a higher percentage of DMA(V) in the liver than the other arsenicals after a single exposure to arsenate. The percentage of hepatic DMA(V) decreased and that of iAs increased with repeated exposure. A trimethylated metabolite was also detected in the liver. Tissue accumulation of arsenic after repeated po exposure to arsenate in the mouse corresponds to the known human target organs for iAs-induced carcinogenicity.     

Biliary and urinary excretion of inorganic arsenic: monomethylarsonous acid as a major biliary metabolite in rats.

In rats exposed to arsenite (AsIII) or arsenate (AsV), the biliary excretion of arsenic depends completely on availability of hepatic glutathione, suggesting that both AsIII and AsV are transported into bile in thiol-reactive trivalent forms (Gyurasics et al. [1991], Biochem. Pharmacol. 42, 465-468). To test this hypothesis, the bile and urine of bile duct-cannulated rats injected with AsIII or AsV (50 micromol/kg, iv) were collected periodically for 2 h and analyzed for arsenic metabolites by HPLC-hydride generation-atomic fluorescence spectrometry. Arsenic was excreted predominantly into bile in AsIII-injected rats, but the urine was the main route of excretion in AsV-exposed rats. Injected AsIII was excreted in urine practically unchanged, whereas both AsV and AsIII appeared in urine after administration of AsV. Irrespective of the arsenical administered, the bile contained 2 main arsenic species, namely AsIII and a hitherto unidentified metabolite. Formation of this metabolite could be prevented by pretreatment of the rats with the methylation inhibitor periodate-oxidized adenosine, indicating that it is a methylated arsenic compound. This metabolite could be converted in vitro into monomethylarsonic acid (MMAsV) by oxidation, whereas synthetic MMAsV could be converted into the unknown metabolite by reduction. Consequently, this biliary metabolite of both AsIII and AsV is monomethylarsonous acid (MMAsIII), a long-hypothesized, but never identified, intermediate in the biotransformation of AsIII and AsV. Although MMAsIII is thought to be formed from an oxidized precursor, rats injected with MMAsV did not excrete MMAsIII. In summary, the inorganic arsenicals investigated are transported into bile exclusively in trivalent forms, namely as AsIII and MMAsIII, but are excreted in urine in both tri- and pentavalent forms. Identification of MMAsIII is signified by the fact that this metabolite is more toxic than AsIII and AsV and thus formation of MMAsIII represents toxification of inorganic arsenic.     

Arsenic urinary speciation in Mthfr deficient mice injected with sodium arsenate

In most mammalian species, arsenic biotransformation occurs primarily by biomethylation and reduction reactions, with dimethylarsinic acid being the predominant metabolite excreted in the urine. Methylenetetrahydrofolate reductase (Mthfr) plays a key role in folate metabolism by channeling one-carbon units between nucleotide synthesis and methylation reactions. In the study on transgenic Mtfhr knockout mice we investigated: (1) whether Mthfr is an important determinant in arsenic biotransformation by performing urinary arsenic speciation, and (2) whether dietary folate deficiency alters arsenic biotransformation in these mice. The Mthfr mice fed folate replete or folate deficient diet were injected with sodium arsenate 1 mg/kg, and placed in metabolic cages for a urine collection. The urine was analyzed for arsenic species. Additionally, folate and homocysteine plasma level was analyzed in Mthfr mice. When fed a folate control diet, the Mthfr^−/− mice excreted significantly less of the total arsenic in urine than did the Mthfr^+/+ and Mthfr^+/− mice. The Mthfr^−/− had significantly lower levels of pentavalent arsenic in their urine than did the Mthfr^+/+mice. The wild type mice excreted significantly less pentavalent arsenic when they were fed folate deficient diet comparing to control diet. The current data suggest that both the Mthfr status and food folate level modulate in a significant manner excretion of arsenic in mice, following intraperitoneal administration of sodium arsenate.     

Forced uptake of trivalent and pentavalent methylated and inorganic arsenic and its cyto-/genotoxicity in fibroblasts and hepatoma cells.

Mammals are able to convert inorganic arsenic to mono-, di-, and trimethylated metabolites. In previous studies we have shown that the trivalent organoarsenic compounds are more toxic than their inorganic counterparts and that the toxicity is associated with the cellular uptake of the arsenicals. In the present study, we investigated cyto-/genotoxic effects of the arsenic compounds arsenate [As(i)(V)], arsenite [As(i)(III)], monomethylarsonic acid [MMA(V)], monomethylarsonous acid [MMA(III)], dimethylarsinic acid [DMA(V)], dimethylarsinous acid [DMA(III)], and trimethylarsine oxide [TMAO(V)] after an extended exposure time (24 h) and compared the uptake capabilities of fibroblasts (CHO-9 cells: Chinese hamster ovary) used for genotoxicity studies, with those of hepatic cells (Hep G2: hepatoma cell-line). To find out whether the arsenic compounds are bound to membranes or if they are present in the cytosol, the amount of arsenic was measured in whole-cell extracts and in membrane-removed cell extracts by inductively coupled plasma-mass spectrometry (ICP-MS). In addition, we forced the cellular uptake of the arsenic compounds into CHO-9 cells by electroporation and measured the intracellular arsenic concentrations before and after this procedure. Our results show that organic and inorganic arsenicals are taken up to a higher degree by fibroblasts compared to hepatoma cells. The arsenic metabolite DMA(III) was the most membrane permeable species in both cell lines and induced strong genotoxic effects in CHO-9 cells after an exposure time of 24 h. The uptake of all other arsenic species was relatively low (<1% by Hep G2 and <4% by CHO cells), but was dose-dependent. Electroporation increased the intracellular arsenic levels as well as the number of induced MN in CHO-9 cells. With the exception of As(i)(III) and DMA(III) in CHO-9 cells, the tested arsenic compounds were not bound to cell membranes, but were present in the cytosol. This may indicate the existence of DMA(III)-specific exporter proteins as are known for As(i)(III). Our results indicate that the uptake capabilities of arsenic compounds are highly dependent upon the cell type. It may be hypothesized that the arsenic-induced genotoxic effects observed in fibroblasts are due to the high uptake of arsenicals into this cell type. This may explain the high susceptibility of skin fibroblasts to arsenic exposure.     

The Metabolism of Arsenite and Arsenate by the Rat

The Metabolism of Arsenite and Arsenate by the Rat. Lerman,S.A. and Clarkson, T.W. (1983). Fundam. App. Toxicol. 3: 309–314.Differences in distribution and metabolism of arsenite and arsenatewere studied in rats and in rat liver and kidney slices andhepatocytes. Five minutes after i.v. administration of 4.8 nmolarsenite or arsenate to male Sprague-Dawley rats, blood levelsof arsenic were only 10 percent of the initial dose. Blood arseniclevels then rose: by 4 hours about 67 percent of the initialdose of arsenite and 28 percent of the initial dose of arsenatewere in the blood compartment. The predominant form of arsenicin the RBC was dimethylarsinic acid (DMA). Arsenite was rapidlydistributed to both liver and kidney; arsenate was rapidly distributedto kidney only. After 4 hours of exposure to arsenite, liverslices had taken up six times more arsenic and kidney slicestwo times more arsenic than after exposure to arsenate. Isolatedhepatocytes took up as much as 20 times more arsenic after arseniteexposure. DMA was found in the medium of the liver slices andhepatocytes exposed to arsenite, but very little DMA was foundin the medium of the arsenate-exposed liver slices and hepatocytes.However, five times more DMA was found in the medium of thekidney slices exposed to arsenate than in the medium of theliver slices. Phosphate inhibited uptake and metabolism of arsenateby kidney slices. These studies indicate that inorganic arsenicis rapidly taken up by liver and kidney and methylated. Arseniteis methylated by both organs, whereas arsenate may be methylatedby kidney only.     

Pharmacokinetics and pharmacodynamics of norfluoxetine in rats: Increasing extracellular serotonin level in the frontal cortex

Norfluoxetine is the most important active metabolite of the widely used antidepressant fluoxetine. Although the pharmacokinetics/pharmacodynamics (PK/PD) relationship and neurochemical profile of fluoxetine is well characterized in human and in animals, little is known about the effect of its metabolite. The aim of this study was to characterize extracellular level of serotonin (5-hydroxytryptamine, 5-HT)-time profile of norfluoxetine after acute administration over 18 h post dose and to establish the relationship between this pharmacodynamic (PD) profile and its pharmacokinetic (PK) properties. Following subcutaneous administration of fluoxetine in rats, plasma and brain PK of fluoxetine and norfluoxetine were monitored respectively by liquid chromatography/tandem mass spectrometry (LC/MS/MS). The extracellular level of 5-HT in the frontal cortex was measured by microdialysis as a PD endpoint. Norfluoxetine when directly administrated to rats caused a significant increase in extracellular level of 5-HT in the frontal cortex and maintained for 18 h. This result is correlated well with higher plasma and brain concentration and longer plasma and brain retention time of norfluoxetine. Our results showed that norfluoxetine contributes to 5-HT transporter inhibition and extends fluoxetine efficacy.     

Effect of α-tocopherol on carbon tetrachloride intoxication in the rat liver

Carbon tetrachloride (1 ml/kg body weight as a 1:1 mixture of CCl₄ and mineral oil) was orally administered to rats. After 12 h, the activity of plasma ALT (alanine aminotransferase) was significantly higher than that of the control group, and plasma ALT and AST (aspartate aminotransferase) activities significantly increased 24 h after CCl₄ administration. These results indicated that the necrotic process had initiated at about 12 h and developed thereafter. After 6–24 h of CCl₄ administration, the hepatic level of vitamin C, the most sensitive indicator of oxidative stress, decreased significantly, indicating that oxidative stress was significantly enhanced 6 h after CCl₄ intoxication and thereafter. Oral administration of vitamin E (1 ml/kg body weight as a 1:1 mixture of α-tocopherol and mineral oil) 12 h before CCl₄ administration caused a significant elevation of liver vitamin E level and ameliorated liver necrosis 24 h after CCl₄ intoxication based on plasma AST and ALT. Vitamin E also significantly restored the hepatic vitamin C concentration 12 and 24 h after CCl₄ intoxication, demonstrating that vitamin E functioned as an antioxidant. The liver vitamin E concentration was not changed by vitamin E supplementation to rats that did not receive CCl₄. This result indicated that vitamin E accumulated in the damaged liver. The activation of JNK, ERK1/2 and p38 MAPK took place 1.5 h after CCl₄ administration. Co-administration of α-tocopherol with CCl₄ did not affect these early changes in MAPKs.     

Effects of exogenous glutathione on arsenic burden and NO metabolism in brain of mice exposed to arsenite through drinking water

Chronic exposure to inorganic arsenic (iAs) is associated with neurotoxicity. Studies to date have disclosed that methylation of ingested iAs is the main metabolic pathway, and it is a process relying on reduced glutathione (GSH). The aim of this study was to explore the effects of exogenous GSH on arsenic burden and metabolism of nitric oxide (NO) in the brain of mice exposed to arsenite via drinking water. Mice were exposed to sodium arsenite through drinking water contaminated with 50 mg/L arsenic for 4 weeks and treated intraperitoneally with saline solution, 200 mg/kg body weight (b.w), 400 mg/kg b.w, or 800 mg/kg b.w GSH, respectively, at the 4th week. Levels of iAs, monomethylarsenic acid, and dimethylarsenic acid (DMAs) in the liver, blood, and brain were determined by method of hydride generation coupled with atomic absorption spectrophotometry. Activities of nitric oxide synthase (NOS) and contents of NO in the brain were determined by colorimetric method. Compared with mice exposed to arsenite alone, administration of GSH increased dose-dependently the primary and secondary methylation ratio in the liver, which caused the decrease in percent iAs and increase in percent DMAs in the liver, as a consequence, resulted in significant decrease in iAs levels in the blood and total arsenic levels in both blood and brain. NOS activities and NO levels in the brain of mice in iAs group were significantly lower than those in control; however, administration of GSH could increase significantly activities of NOS and contents of NO. Findings from this study suggested that exogenous GSH could promote both primary and secondary arsenic methylation capacity in the liver, which might facilitate excretion of arsenicals, and consequently reduce arsenic burden in both blood and brain and furthermore ameliorate the effects of arsenicals on NO metabolism in the brain.     

Paraoxonase 1 activity in subchronic low‐level inorganic arsenic exposure through drinking water

Epidemiological evidences indicate close association between inorganic arsenic exposure via drinking water and cardiovascular diseases. While the exact mechanism of this arsenic‐mediated increase in cardiovascular risk factors remains enigmatic, epidemiological studies indicate a role for paraoxonase 1 (PON1) in cardiovascular diseases. To investigate the association between inorganic arsenic exposure and cardiovascular diseases, rats were exposed to sodium arsenite (trivalent; 50, 100, and 150 ppm As) and sodium arsenate (pentavalent; 100, 150, and 200 ppm As) in their drinking water for 12 weeks. PON1 activity towards paraoxon (PONase) and phenylacetate (AREase) in plasma, lipoproteins, hepatic, and brain microsomal fractions were determined. Inhibition of PONase and AREase in plasma and HDL characterized the effects of the two arsenicals. While the trivalent arsenite inhibited PONase by 33% (plasma) and 46% (HDL), respectively, the pentavalent arsenate inhibited the enzyme by 41 and 34%, respectively. AREase activity was inhibited by 52 and 48% by arsenite, whereas the inhibition amounted to 72 and 67%, respectively by arsenate. The pattern of inhibition in plasma and HDL indicates that arsenite induced a dose‐dependent inhibition of PONase whereas arsenate induced a dose‐dependent inhibition of AREase. In the VLDL + LDL, arsenate inhibited PONase and AREase while arsenite inhibited PONase. In the hepatic and brain microsomal fractions, only the PONase enzyme was inhibited by the two arsenicals. The inhibition was more pronounced in the hepatic microsomes where a 70% inhibition was observed at the highest dose of pentavalent arsenic. Microsomal cholesterol was increased by the two arsenicals resulting in increased cholesterol/phospholipid ratios. Our findings indicate that decreased PON1 activity observed in arsenic exposure may be an incipient biochemical event in the cardiovascular effects of arsenic. Modulation of PON1 activity by arsenic may also be mediated through changes in membrane fluidity brought about by changes in the concentration of cholesterol in the microsomes. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 154–162, 2016.     

Systemic indicators of inorganic arsenic toxicity in four animal species

The effect of arsenic compounds depends on the chemical form and is specific for certain organs. The lack of specific biological indicators for the effects of each arsenic species makes it difficult to differentiate their toxicity. Five prospective biological indicators of systemic toxicity were examined at time points ranging from 15 min to 24 h using male Sprague-Dawley rats, B6C3F1 mice, Golden-Syrian hamsters, and Hartley guinea pigs, following intraperitoneal dosing with 0.1 and 1 mg/kg sodium arsenite. Rats and mice were also dosed with 1 mg/kg sodium arsenate. Total blood arsenic levels were determined in all animal species to show that exposure occurred and as an index of the severity of the change is an indicator of toxicity. Total blood arsenic levels were increased in all animal species. This increase was dose, arsenic species, and animal dependent. Renal pyruvate dehydrogenase activity was significantly decreased at early time points in mice, hamsters, and guinea pigs, and at later time points in rats dosed with arsenite. Rats and mice dosed with arsenate also exhibited PDH decrease at early time points. Blood hematocrit and glucose were increased in the rat and guinea pig, respectively, after arsenite administration. Creatinine and urea nitrogen were found to be unresponsive to arsenic in most animal species. Data suggested that the mouse and secondly the hamster appear to be the most appropriate animal models for the study of acute arsenic toxicity.     

太白楤木皂苷对砷诱导的肝细胞线粒体途径凋亡的保护作用

目的探讨太白楤木皂苷对砷暴露损伤的保护作用及分子机制。方法人正常肝细胞HL-7702进行砷暴露及太白楤木皂苷保护处理。砷暴露组用10μmol L-1终浓度的NaAsO2处理,太白楤木皂苷保护组用不同浓度太白楤木皂苷预处理12 h后,再进行砷暴露处理。24 h后MTT法检测细胞活力;12 h后收集细胞样品,分别检测细胞的凋亡、活性氧（ROS）、还原型谷胱甘肽（GSH）,氧化损伤产物丙二醛（MDA）水平以及线粒体凋亡途径的重要因子细胞色素C（Cyto C）释放和Caspase-3的激活情况。结果砷暴露可以引起肝细胞活力显著降低,给予不同浓度的太白楤木皂苷具有一定的保护作用,其中5μg mL-1太白楤木皂苷保护效果最佳。砷暴露可以引起细胞凋亡比例显著增加,细胞ROS水平升高,GSH水平降低,氧化损伤产物MDA水平增加,线粒体凋亡途径因子Cyto C和Caspase-3的激活。太白楤木皂苷（5μg mL-1）预处理能够显著拮抗砷暴露引起的细胞上述指标变化。结论砷暴露可以通过诱导细胞氧化应激,激活线粒体途径细胞凋亡,发挥其损伤毒性作用。太白楤木皂苷可以通过抑制细胞氧化应激,抑制线粒体途径的细胞凋亡,从而发挥其抗砷暴露损伤的保护作用。     

Comparison of the effects of acute and subchronic administration of atipamezole on reaction to novelty and active avoidance learning in rats

The effects of an α₂-adrenoceptor antagonist, atipamezole, on exploratory behaviour in a novel environment, spontaneous motor activity and active avoidance learning were studied after acute injection and continuous infusion (0.1 mg/kg h) for 24 h and 6–9 days in rats. The effects of atipamezole on biogenic amines and their main metabolites in brain were studied after an acute injection (0.3 mg/kg s.c.) and continuous infusion (0.1 mg/kg h) for 24 h and 10 days. The level of central α₂-adrenoceptor antagonism and the drug concentration in blood and in the brain were measured after continuous infusion for 24 h and 10 days. In behavioural tests, atipamezole had no effect on spontaneous motor activity at any of the doses studied. However, after both acute administration and continuous 24-h infusion, atipamezole decreased exploratory behaviour in a staircase test, but no longer after 6 days of continuous infusion. Acute administration of atipamezole impaired performance in active avoidance learning tests causing a learned helplessness-like behaviour. When the training was started after 7 days of continuous infusion, atipamezole significantly improved active avoidance learning. There was a significant increase in the metabolite of noradrenaline (NA), 3-methoxy-4-hydroxyphenylethyleneglycol sulphate (MHPG-SO₄), after 24 h but not any longer after 10 days of continuous atipamezole infusion, although the extent of central α₂-adrenoceptor antagonism was unchanged and the atipamezole concentration present in brain was even elevated at 10 days compared to levels after 24-h infusion. In conclusion, these results reveal that acute and sub-chronic atipamezole treatments have different and even opposite effects on behaviour in novel, stressful situations. After acute treatment, atipamezole potentiates reaction to novelty and stress, causing a decrease in exploratory activity and impairment in shock avoidance learning. After subchronic treatment, there was no longer any effect on exploratory behaviour and, in fact, there was an improvement in the learning of a mildly stressful active avoidance test. The changes in behaviour occurred in parallel with attenuation in the MHPG-SO₄-increasing effect, thus the suppressed behaviour in the present test conditions after acute atipamezole injection is associated with a major increase in central NA release. The results support the role of α₂-adrenoceptors and noradrenergic system in reactions both to novelty and stress and have possible implications in cognitive functions as well as in depression. Received: 6 August 1998 / Accepted: 4 January 1999     

Effects of exogenous methionine on arsenic burden and NO metabolism in brain of mice exposed to arsenite through drinking water

The aim of this study was to explore the effects of exogenous methionine (Met) on arsenic burden and metabolism of nitric oxide (NO) in the brain of mice exposed to arsenite via drinking water. Mice were exposed to sodium arsenite through drinking water contaminated with 50 mg/L arsenic for four consecutive weeks, and treated intraperitoneally with saline solution, 100 mg/kg body weight (b.w), 200 mg/kg b.w or 400 mg/kg b.w of Met, respectively at the fourth week. Levels of inorganic arsenic (iAs), monomethylarsenic acid (MMAs), and dimethylarsenic acid (DMAs) in the liver, blood and brain were determined by method of hydride generation coupled with atomic absorption spectrophotometry. Nitric oxide synthase (NOS) activities and NO levels in the brain were determined by colorimetric method. Compared with mice exposed to arsenite alone, administration of Met increased significantly the primary methylation ratio in the liver, which resulted in decrease of percent iAs and increase of percent DMAs in the liver, and decrease of iAs, MMAs and total arsenic levels (TAs) in the blood and DMAs and TAs in the brain. NOS activities and NO levels in the brain of mice exposed to arsenite alone were significantly lower than those in control, however administration of Met could increase significantly NO levels. Findings from this study suggested that exogenous Met could benefit the primary arsenic methylation in the liver, which might increase the production of methylated arsenicals and facilitate arsenic excretion. As a consequence, arsenic burden in both blood and brain was reduced, and toxic effects on NO metabolism in the brain were ameliorated. © 2011 Wiley Periodicals, Inc. Environ Toxicol 2012.