Linking mountaintop removal mining to water quality for imperiled species using satellite data.

Environmental laws need sound data to protect species and ecosystems. In 1996, a proliferation of mountaintop removal coal mines in a region home to over 50 federally protected species was approved under the Endangered Species Act. Although this type of mining can degrade terrestrial and aquatic habitats, the available data and tools limited the ability to analyze spatially extensive, aggregate effects of such a program. We used two large, public datasets to quantify the relationship between mountaintop removal coal mining and water quality measures important to the survival of imperiled species at a landscape scale across Kentucky, Tennessee, Virginia, and West Virginia. We combined an annual map of the extent of surface mines in this region from 1985 to 2015 generated from Landsat satellite imagery with public water quality data collected over the same time period from 4,260 monitoring stations within the same area. The water quality data show that chronic and acute thresholds for levels of aluminum, arsenic, cadmium, conductivity, copper, lead, manganese, mercury, pH, selenium, and zinc safe for aquatic life were exceeded thousands of times between 1985 and 2015 in streams that are important to the survival and recovery of species on the Endangered Species List. Linear mixed models showed that levels of manganese, sulfate, sulfur, total dissolved solids, total suspended solids, and zinc increased by 6.73E+01 to 6.87E+05 µg/L and conductivity by 3.30E+06 µS /cm for one percent increase in the mined proportion of the area draining into a monitoring station. The proportion of a drainage area that was mined also increased the likelihood that chronic thresholds for copper, lead, and zinc required to sustain aquatic life were exceeded. Finally, the proportion of a watershed that was mined was positively related to the likelihood that a waterway would be designated as impaired under the Clean Water Act. Together these results demonstrate that the extent of mountaintop removal mining, which can be derived from public satellite data, is predictive of water quality measures important to imperiled species-effects that must be considered under environmental law. These findings and the public data used in our analyses are pertinent to ongoing re-evaluations of the effects of current mine permitting regulations to the recovery and survival of federally protected species.

Effects of bisphenol A (BPA) on brain-specific expression of cyp19a1b gene in swim-up fry of Labeo rohita.

Estrogen regulates numerous developmental and physiological processes and effects are mediated mainly by estrogenic receptors (ERs), which function as ligand-regulated transcription factor. ERs can be activated by many different types endocrine disrupting chemicals (EDCs) and interfere with behaviour and reproductive potential of living organism. Estrogenic regulation of membrane associated G protein-coupled estrogen receptor, GPER activity has also been reported. Bisphenol A (BPA), a ubiquitous endocrine disruptor is present in many household products, has been linked to many adverse effect on sexual development and reproductive potential of wild life species. The present work is aimed to elucidate how an environmentally pervasive chemical BPA affects in vivo expression of a known estrogen target gene, cyp19a1b in the brain, and a known estrogenic biomarker, vitellogenin (Vg) in the whole body homogenate of 30 days post fertilization (dpf) swim-up fry of Labeo rohita. We confirm that, like estrogen, the xenoestrogen BPA exposure for 5-15 days induces strong overexpression of cyp19a1b, but not cyp19a1a mRNA in the brain and increase concentration of vitellogenin in swim-up fry. BPA also induces strong overexpression of aromatase B protein and aromatase activity in brain. Experiments using selective modulators of classical ERs and GPER argue that this induction is largely through nuclear ERs, not through GPER. Thus, BPA has the potential to elevate the levels of aromatase and thereby, levels of endogenous estrogen in developing brain. These results indicate that L. rohita swim-up fry can be used to detect environmental endocrine disruptors either using cyp19a1b gene expression or vitellogenin induction.

Effect of Tunceli garlic on some immunological parameters in Cyprinus carpio exposed to chlorpyrifos.

In this study, the effect of Tunceli garlic (Allium tuncelianium) on hemoglobin (Hb) level, oxidative radical production of neutrophils (Nitoblue tetrazolium assay-NBT activity) and total immunoglobulin (TI) content in carp (Cyprinus carpio) exposed to chlorpyrifos (CPF)  was investigated. The 96 hour LC50 value of CPF on C. carpio was calculated to as 0.230 mg/L. The fishes were exposed to sublethal concentration of chlorpyrifos (1/8 of LC50 value: 0.029 mg/L), and Tunceli garlic (20 and 40 g/kg diet) was simultaneously administered for 21 days. Blood samples were taken from the fishes at 7, 14 and 21 days and analysed to determine the Hb levels, the NBT activity and the TI content. There was a significant decrease in the Hb level, the NBT activity and the TI content of CPF-treated fish. However, Tunceli garlic reversed the Hb level, the NBT activity and the TI content. In conclusion, this study demonstrated that CPF had a negative effect on the immunological values of the fish. The simultaneous administration of Tunceli garlic was neutralised CPF-induced toxicity.

Constraining the Teratogenicity of Pesticide Pollution by a Synthetic Nanoreceptor.

The teratogenicity of the pesticide nereistoxin (NTX) and its derivative thiocyclam (THI) towards aquatic life was dramatically constrained by a synthetic nanoreceptor, cucurbit[7]uril, through selective encapsulation of the pesticides (KCB[7]-NTX of 3.24(±0.31)×106 m-1 and KCB[7]-THI of 7.46(±0.10)×105 m-1 ), as evidenced by the rate of hatchability, morphology development, and tyrosinase activity of zebrafish larvae incubated with the pesticides (3-300 µm) in the absence and in the presence of 300 µm cucurbit[7]uril, demonstrating the significant potential of the nanoreceptor in managing ecological pollution of these pesticides.

Effects of caffeine on behavioral and inflammatory changes elicited by copper in zebrafish larvae: Role of adenosine receptors.

This study investigated the effects of caffeine in the behavioral and inflammatory alterations caused by copper in zebrafish larvae, attempting to correlate these changes with the modulation of adenosine receptors. To perform a survival curve, 7dpf larvae were exposed to 10µM CuSO4, combined to different concentrations of caffeine (100µM, 500µM and 1mM) for up to 24h. The treatment with copper showed lower survival rates only when combined with 500µM and 1mM of caffeine. We selected 4 and 24h as treatment time-points. The behavior evaluation was done by analyzing the traveled distance, the number of entries in the center, and the length of permanence in the center and the periphery of the well. The exposure to 10µM CuSO4 plus 500µM caffeine at 4 and 24h changed the behavioral parameters. To study the inflammatory effects of caffeine, we assessed the PGE2 levels by using UHPLC-MS/MS, and TNF, COX-2, IL-6 and IL-10 gene expression by RT-qPCR. The expression of adenosine receptors was also evaluated with RT-qPCR. When combined to copper, caffeine altered inflammatory markers depending on the time of exposure. Adenosine receptors expression was significantly increased, especially after 4h exposure to copper and caffeine together or separately. Our results demonstrated that caffeine enhances the inflammation induced by copper by decreasing animal survival, altering inflammatory markers and promoting behavioral changes in zebrafish larvae. We also conclude that alterations in adenosine receptors are related to those effects.

Inhibitory effect of cadmium on estrogen signaling in zebrafish brain and protection by zinc.

The present study was conducted to assess the effects of Cd exposure on estrogen signaling in the zebrafish brain, as well as the potential protective role of Zn against Cd-induced toxicity. For this purpose, the effects on transcriptional activation of the estrogen receptors (ERs), aromatase B (Aro-B) protein expression and molecular expression of related genes were examined in vivo using wild-type and transgenic zebrafish embryos. For in vitro studies, an ER-negative glial cell line (U251MG) transfected with different zebrafish ER subtypes (ERα, ERß1 and ERß2) was also used. Embryos were exposed either to estradiol (E2 ), Cd, E2 +Cd or E2 +Cd+Zn for 72 h and cells were exposed to the same treatments for 30 h. Our results show that E2 treatment promoted the transcriptional activation of ERs and increased Aro-B expression, at both the protein and mRNA levels. Although exposure to Cd, does not affect the studied parameters when administered alone, it significantly abolished the E2 -stimulated transcriptional response of the reporter gene for the three ER subtypes in U251-MG cells, and clearly inhibited the E2 induction of Aro-B in radial glial cells of zebrafish embryos. These inhibitory effects were accompanied by a significant downregulation of the expression of esr1, esr2a, esr2b and cyp19a1b genes compared to the E2 -treated group used as a positive control. Zn administration during simultaneous exposure to E2 and Cd strongly stimulated zebrafish ERs transactivation and increased Aro-B protein expression, whereas mRNA levels of the three ERs as well as the cyp19a1b remained unchanged in comparison with Cd-treated embryos. In conclusion, our results clearly demonstrate that Cd acts as a potent anti-estrogen in vivo and in vitro, and that Cd-induced E2 antagonism can be reversed, at the protein level, by Zn supplement. Copyright © 2016 John Wiley & Sons, Ltd.

Green tea and vitamin C ameliorate some neuro-functional and biochemical signs of arsenic toxicity in rats.

BACKGROUND/OBJECTIVES: Nervous system damage is one of the consequences of oral exposure to waterborne inorganic arsenic. In this work, the role of oxidative status in the neurotoxicity of arsenic and the possible role of two foodborne antioxidants in ameliorating arsenic-related oxidative stress were investigated. METHODS: Male Wistar rats were given 10 mg/kg b.w. of trivalent inorganic arsenic (in the form of NaAsO2), 5 day/week for 6 weeks by gavage, combined with vitamin C solution (1 g/l) or green tea infusion (2.5 g in 500 ml boiled water) as antioxidants given in the drinking fluid. RESULTS: Body weight gain was reduced by arsenic from the second week and the antioxidants had no effect on that. Cortical evoked potentials had increased latency, tail nerve conduction velocity was reduced, and this latter effect was counteracted by the antioxidants. The effect of green tea was stronger than that of vitamin C, and green tea also diminished lipid peroxidation induced by As. There was fair correlation between brain As levels, electrophysiological changes, and lipid peroxidation, suggesting a causal relationship. DISCUSSION: Natural antioxidants might be useful in the protection of the central nervous system against the toxicity of oral As.

[Experimental evaluation of the impact of low doses of the herbicide 2,4-D in drinking water on some indices of lipid and immune status].

There were studied features of the manifestation of incoming with potable water nontoxic doses of the herbicide 2,4-D on the body mass index, some indices of lipid metabolism and immune system in different types of diets in the experiment in rats. There was shown a significant gain in body weight of animals, and MDA and leptin level in the serum under the action of a herbicide in conjunction with a high-calorie diet. In all experimental groups there was noted the increased level of total cholesterol, tendency to the increase of LDL cholesterol. Under the action of the herbicide there was noted an increase of IL-6, TNF-a and the numbers of leukocytes and, on the contrary, the reduction of the number of thymocytes and kariocytes in thymus and spleen.

Mangiferin ameliorates aluminium chloride-induced cognitive dysfunction via alleviation of hippocampal oxido-nitrosative stress, proinflammatory cytokines and acetylcholinesterase level.

Mangiferin is a phytochemical primarily present in the stem, leaves and bark of Mangifera indica. It offers neuroprotection mainly through inhibition of oxidative stress, and decreasing proinflammatory cytokines level in the brain. Aluminium has been reported to cause oxidative stress-associated damage in the brain. In the present investigation, protective effect of mangiferin against aluminium chloride (AlCl3)-induced neurotoxicity and cognitive impairment was studied in male Swiss albino mice. AlCl3 (100 mg/kg) was administered once daily through oral gavage for 42 days. Mangiferin (20 and 40 mg/kg, p.o.) was given to mice for last 21 days of the study. We found cognitive dysfunction in AlCl3-treated group, which was assessed by Morris water maze test, and novel object recognition test. AlCl3-treated group showed elevated level of oxidative stress markers, proinflammatory cytokines level and lowered hippocampal brain-derived neurotrophic factor (BDNF) content. Mangiferin (40 mg/kg) prevented the cognitive deficits, hippocampal BDNF depletion, and biochemical anomalies induced by AlCl3-treatment. In conclusion, our data demonstrated that mangiferin offers neuroprotection in AlCl3-induced neurotoxicity and it may be a potential therapeutic approach in the treatment of oxido-nitrosative stress and inflammation-associated neurotoxicity.

Arsenic inhibits hedgehog signaling during P19 cell differentiation.

Arsenic is a toxicant found in ground water around the world, and human exposure mainly comes from drinking water or from crops grown in areas containing arsenic in soils or water. Epidemiological studies have shown that arsenic exposure during development decreased intellectual function, reduced birth weight, and altered locomotor activity, while in vitro studies have shown that arsenite decreased muscle and neuronal cell differentiation. The sonic hedgehog (Shh) signaling pathway plays an important role during the differentiation of both neurons and skeletal muscle. The purpose of this study was to investigate whether arsenic can disrupt Shh signaling in P19 mouse embryonic stem cells, leading to changes muscle and neuronal cell differentiation. P19 embryonic stem cells were exposed to 0, 0.25, or 0.5 µM of sodium arsenite for up to 9 days during cell differentiation. We found that arsenite exposure significantly reduced transcript levels of genes in the Shh pathway in both a time and dose-dependent manner. This included the Shh ligand, which was decreased 2- to 3-fold, the Gli2 transcription factor, which was decreased 2- to 3-fold, and its downstream target gene Ascl1, which was decreased 5-fold. GLI2 protein levels and transcriptional activity were also reduced. However, arsenic did not alter GLI2 primary cilium accumulation or nuclear translocation. Moreover, additional extracellular SHH rescued the inhibitory effects of arsenic on cellular differentiation due to an increase in GLI binding activity. Taken together, we conclude that arsenic exposure affected Shh signaling, ultimately decreasing the expression of the Gli2 transcription factor. These results suggest a mechanism by which arsenic disrupts cell differentiation.

Reduction in acute ecotoxicity of paper mill effluent by sequential application of xylanase and laccase.

In order to reduce the ecotoxicity of paper mill, four different enzymatic pretreatment strategies were investigated in comparison to conventional chemical based processes. In strategy I, xylanase-aided pretreatment of pulp was carried out, and in strategy II, xylanase and laccase-mediator systems were used sequentially. Moreover, to compare the efficiency of Bacillus stearothermophilus xylanase and Ceriporiopsis subvermispora laccase in the reduction of ecotoxicity and pollution, parallel strategies (III and IV) were implemented using commercial enzymes. Conventional C(D)E(OP)D(1)D(2) (C(D), Cl(2) with ClO2; EOP, H2O2 extraction; D1 and D2, ClO2) and X/XLC(D)E(OP)D(1)D(2) (X, xylanase; L, laccase) sequences were employed with non-enzymatic and enzymatic strategies, respectively. Acute toxicity was determined by the extent of inhibition of bioluminescence of Vibrio fischeri with different dilutions of the effluent. Two-fold increase was observed in EC50 values for strategy I compared to the control process. On the other hand, sequential application of commercial enzymes resulted in higher acute toxicity compared to lab enzymes. In comparison to the control process, strategy II was the most efficient and successfully reduced 60.1 and 25.8% of biological oxygen demand (BOD) and color of effluents, respectively. We report for the first time the comparative analysis of the ecotoxicity of industrial effluents.

Edaravone mitigates hexavalent chromium-induced oxidative stress and depletion of antioxidant enzymes while estrogen restores antioxidant enzymes in the rat ovary in F1 offspring.

Environmental contamination of drinking water with chromium (Cr) has been increasing in more than 30 cities in the United States. Previous studies from our group have shown that Cr affects reproductive functions in female Sprague Dawley rats. Although it is impossible to completely remove Cr from the drinking water, it is imperative to develop effective intervention strategies to inhibit Cr-induced deleterious health effects. Edaravone (EDA), a potential inhibitor of free radicals, has been clinically used to treat cancer and cardiac ischemia. This study evaluated the efficacy of EDA against Cr-induced ovarian toxicity. Results showed that maternal exposure to CrVI in rats increased follicular atresia, decreased steroidogenesis, and delayed puberty in F1 offspring. CrVI increased oxidative stress and decreased antioxidant (AOX) enzyme levels in the ovary. CrVI increased follicle atresia by increased expression of cleaved caspase 3, and decreased expression of Bcl2 and Bcl2l1 in the ovary. EDA mitigated or inhibited the effects of CrVI on follicle atresia, pubertal onset, steroid hormone levels, and AOX enzyme activity, as well as the expression of Bcl2 and Bcl2l1 in the ovary. In a second study, CrVI treatment was withdrawn, and F1 rats were injected with estradiol (E2) (10 µg in PBS/ethanol per 100 g body weight) for a period of 2 wk to evaluate whether E2 treatment will restore Cr-induced depletion of AOX enzymes. E2 restored CrVI-induced depletion of glutathione peroxidase 1, catalase, thioredoxin 2, and peroxiredoxin 3 in the ovary. This is the first study to demonstrate the protective effects of EDA against any toxicant in the ovary.

Isolation of a new butenolide from the South China Sea gorgonian coral Subergorgia suberosa.

Chemical investigation on the South China Sea gorgonian coral Subergorgia suberosa has led to the isolation of one new butenolide (5R)-5-(1-ethoxypropyl)-5-hydroxy-3,4-dimethylfuran-2(5H)-one (1) as a pair of inseparable epimers, along with two known butenolides (S)-5-hydroxy-3,4-dimethyl-5-propylfuran-2(5H)-one (2) and (S)-5-hydroxy-3,4-dimethyl-5-pentylfuran-2(5H)-one (3). Their structures including absolute configurations were determined unambiguously by detailed analyses of spectroscopic data and by comparison with the available literature. Compounds 1-3 exhibited moderate antifouling activities against the settlement of Balanus amphitrite larvae, whereas compound 4, the acetyl derivative of 3, showed no antifouling activity at the concentrations up to 25 µg/mL.

Effect of cadmium-polluted water on plasma levels of tumor necrosis factor-α, interleukin-6 and oxidative status biomarkers in rats: protective effect of curcumin.

UNLABELLED: The present study was designed to investigate the effect of CdCl2-polluted drinking water (40 mg CdCl2/L) on the level of TNF-α and IL-6, as well as oxidative status biomarkers in plasma of rats. The possible protective effect of oral administration of curcumin (50 mg/kg body weight/day) was assessed. Results illustrated that Cd exposure significantly elevated the plasma levels of TNF-α and IL-6 (p<0.001) as compared to normal rats. Also, Cd administration resulted in a significant elevation in the lipid peroxidation and markedly reduction in the activities of SOD and catalase as well as the level of glutathione and total antioxidant capacity in plasma. The co-treatment of Cd with curcumin significantly reduced the levels of TNF-α and IL-6 and ameliorated the alteration in oxidative status biomarkers induced by Cd. Negative correlation between IL-6 or TNF-α was and the plasma activities of catalase, SOD and the level of total antioxidant capacity were found in rats exposed to Cd. CONCLUSION: Cadmium toxicity induced the release of TNF-α and IL-6 which is associated with systemic oxidative stress. This may be involved in the mechanism of the Cd toxicity. On the other hand, the findings suggest the curative action of curcumin against Cd toxicity.

Efficacy of lycopene against fluoride toxicity in rats.

CONTEXT: Oxidative damage to cellular components such as lipids and cell membranes by free radicals and reactive oxygen species (ROS) is thought to be associated with the development of degenerative diseases. Fluoride intoxication is associated with oxidative stress and altered anti-oxidant defense mechanism. Lycopene is a lipid-soluble powerful anti-oxidant that scavenges free radicals and ROS. OBJECTIVE: This study was extended to investigate lycopene anti-oxidant efficacy in different organs of fluoride-intoxicated rats. METHODS: Twenty-four adult rats were randomly divided into four groups of six animals each. Rats in group I received daily doses of vehicle. Group II rats were given lycopene (10 mg/kg body weight/day), by tubes, dissolved in 0.5 ml of corn oil for 5 weeks. Group III rats were given sodium fluoride (NaF) (10.3 mg/kg body weight/day), by tubes, for 5 weeks. In group IV rats, lycopene was administered 1 h later and NaF was administered for 5 weeks. RESULTS: NaF administration induced oxidative stress as evidenced by elevated levels of lipid peroxidation (51.3, 65.9 and 67.6%) measured as malondialdehyde and total nitrate/nitrite (61.0, 59.7 and 68.9%) in red blood cells, heart and brain tissues. Moreover, significantly decreased reduced glutathione level, total anti-oxidant capacity and superoxide dismutase activity were observed in the examined tissues. The induced oxidative stress and the alterations in anti-oxidant system were normalized by the oral administration of lycopene treatment. CONCLUSION: Lycopene administration could minimize the toxic effects of fluoride indicating its free-radical scavenging and powerful anti-oxidant activities.

Antagonistic interaction of mercury and selenium in a marine fish is dependent on their chemical species.

It is well-known that selenium (Se) shows protective effects against mercury (Hg) bioaccumulation and toxicity, but the underlying effects of Se chemical species, concentration, and administration method are poorly known. In this study, we conducted laboratory studies on a marine fish Terapon jurbua to explain why Hg accumulation is reduced in the presence of Se observed in field studies. When Se and Hg were administrated concurrently in the fish diets, different Se species including selenite, selenate, seleno-dl-cystine (SeCys), and seleno-dl-methionine (SeMet) affected Hg bioaccumulation differently. At high concentration in fish diet (20 µg g(-1) normally), selenate and SeCys significantly reduced the dietary Hg(II) assimilation efficiency (AE) from 38% to 26%. After the fish were pre-exposed to dietary selenite or SeMet (7 µg g(-1) normally) for 22 days with significantly elevated Se body concentrations, the Hg(II) AEs were pronouncedly reduced (from 41% to 15-26%), whereas the dissolved uptake rate constant and elimination rate constant were less affected. In contrast to Hg(II), all the MeHg biokinetic parameters remained relatively constant whether Se was administrated simultaneously with the fish diet or when the fish were pre-exposed to Se with elevated body concentrations. Basic biokinetic measurements thus revealed that Se had direct interaction with Hg(II) during dietary assimilation rather than with MeHg and that different Se species had variable effects on Hg assimilation.

Tributyltin-binding protein type 1, a lipocalin, prevents inhibition of osteoblastic activity by tributyltin in fish scales.

Tributyltin-binding protein type 1 (TBT-bp1) is a member of the lipocalin family of proteins which bind to small hydrophobic molecules. In this study, we expressed a recombinant TBT-bp1 (rTBT-bp1, ca. 35kDa) in a baculovirus expression system and purified the protein from the hemolymph of silkworm larvae injected with recombinant baculovirus. After incubation of a mixture of rTBT-bp1 and TBT and its fractionation by means of gel filtration chromatography, TBT was detected in the elution peak of rTBT-bp1, confirming the binding potential of rTBT-bp1 for TBT. An assay of the ability of rTBT-bp1 or native TBT-bp1 (nTBT-bp1) to restore osteoblastic activity inhibited by TBT showed that co-treatment of the scales with rTBT-bp1 or nTBT-bp1 in combination with TBT restored osteoblastic activity in goldfish scales, whereas treatment with TBT alone significantly inhibited osteoblastic activity. These results suggest that TBT-bp1 as a lipocalin member might function to decrease the toxicity of TBT by binding to TBT.

Gene expression profiling in the fetal cardiac tissue after folate and low-dose trichloroethylene exposure.

BACKGROUND: Previous studies show gene expression alterations in rat embryo hearts and cell lines that correspond to the cardio-teratogenic effects of trichloroethylene (TCE) in animal models. One potential mechanism of TCE teratogenicity may be through altered regulation of calcium homeostatic genes with a corresponding inhibition of cardiac function. It has been suggested that TCE may interfere with the folic acid/methylation pathway in liver and kidney and alter gene regulation by epigenetic mechanisms. According to this hypothesis, folate supplementation in the maternal diet should counteract TCE effects on gene expression in the embryonic heart. APPROACH: To identify transcriptional targets altered in the embryonic heart after exposure to TCE, and possible protective effects of folate, we used DNA microarray technology to profile gene expression in embryonic mouse hearts with maternal TCE exposure and dietary changes in maternal folate. RESULTS: Exposure to low doses of TCE (10 ppb) caused extensive alterations in transcripts encoding proteins involved in transport, ion channel, transcription, differentiation, cytoskeleton, cell cycle, and apoptosis. Exogenous folate did not offset the effects of TCE exposure on normal gene expression, and both high and low levels of folate produced additional significant changes in gene expression. CONCLUSIONS: A mechanism by which TCE induces a folate deficiency does not explain altered gene expression patterns in the embryonic mouse heart. The data further suggest that use of folate supplementation, in the presence of this toxin, may be detrimental and not protective of the developing embryo.

Toxicity and characterization of cholinesterase-inhibition induced by diisopropyl fluorophosphate in Artemia salina larvae.

The acute toxicity of diisopropyl fluorophosphate (DFP) on three age classes of Artemia salina was evaluated. An increase in toxicity of this organophosphorous (OP) compound was found following longer development of A. salina larvae. The effects of pretreatment with the non-selective muscarinic antagonist atropine, the two reversible acetylcholinesterase inhibitors physostigmine and pyridostigmine, and the cholinesterase-reactivating oxime 2-pyridine aldoxime methoiodide (2-PAM), as individual and combined pretreatments, on DFP-induced lethality in 24h Artemia were also investigated. The lethal action of DFP was not prevented by pretreatment of 24h Artemia with atropine, physostigmine, and pyridostigmine, while 2-PAM proved effective against intoxication with this OP compound. The inhibitory effects of combinations of atropine (10(-5)M) plus 2-PAM or physostigmine were greater than those elicited by either drug alone, with the maximum protection afforded being 100%. Pretreatment with 2-PAM (10(-6)M) plus physostigmine or pyridostigmine was ineffective. These results suggest that the combinations of atropine plus 2-PAM or physostigmine are effective in the prevention of the lethal effects induced by DFP in A. salina larvae.

Is Cl- protection against silver toxicity due to chemical speciation?

In freshwater teleosts, the primary mechanism of acute silver toxicity is inhibition of Na(+)/K(+) ATPase and carbonic anhydrase at the gill, leading to net Na(+) and Cl(-) loss due to the continued diffusion of these ions into the hypoosmotic external environment. External Cl(-) has been shown to protect rainbow trout (Oncorhychus mykiss) against silver toxicity presumably by complexation to form AgCl. However, Cl(-) does not appear to greatly influence silver toxicity to at least two other species, the European eel (Anguilla Anguilla) and the fathead minnow (Pimephales promelas). We hypothesized that differences in protective effects of Cl(-) at the gill were due to differing requirements or mechanisms for Cl(-) uptake among fish species. To test this hypothesis, we exposed Fundulus heteroclitus, which does not take up Cl(-) across the gills, and Danio rerio and P. promelas, which do rely on Cl(-) uptake across the gills, to Ag(+) in waters of varying Cl(-) concentration. The 96-h LC50s of F. heteroclitus exposed to Ag(+) in soft water with 10 microM Cl(-), 1mM KCl, and 0.5mM MgCl(2) were 3.88, 1.20, and 3.20 microg/L, respectively, and not significantly different. The 96-h LC50s for D. rerio exposed to Ag(+) in soft water with 10 microM Cl(-) and 1mM KCl were 10.3 and 11.3 microg/L, respectively and P. promelas exposed under the same conditions were 2.32 and 2.67 microg/L, respectively. Based on these results, increasing external Cl(-) concentration by as much as 1mM (35.5mg/L) did not offer protection against Ag(+) toxicity to any fish species tested. Although previous results in our laboratory have demonstrated that P. promelas do take up Cl(-) at the gill, a mechanism of uptake has not been identified. Additional experiments, investigating the mechanisms of Na(+) and Cl(-) influx at the gill of P. promelas and the influence of silver, demonstrated that Cl(-) uptake in P. promelas acclimated to soft water occurs through both a Na(+):K(+):2Cl(-) co-transporter and a Cl(-)/HCO(3)(-) exchanger, but is not dependent on carbonic anhydrase. Further, acclimation water chemistry was found to greatly influence subsequent branchial silver accumulation, but Cl(-) uptake was not sensitive to 10 microg/L Ag(+).