Effect of Bacopa monniera Extract on Methylmercury-Induced Behavioral and Histopathological Changes in Rats

Methylmercury (MeHg) is a well-recognized environmental contaminant with established health risk to human beings by fish and marine mammal consumption. Bacopa monniera (BM) is a perennial herb and is used as a nerve tonic in Ayurveda, a traditional medicine system in India. This study was aimed to evaluate the effect of B. monniera extract (BME) on MeHg-induced toxicity in rat cerebellum. Male Wistar rats were administered with MeHg orally at a dose of 5 mg/kg b.w. for 21 days. Experimental rats were given MeHg and also administered with BME (40 mg/kg, orally) 1 h prior to the administration of MeHg for 21 days. After treatment period, MeHg exposure significantly decreases the body weight and also caused the following behavioral changes. Decrease tail flick response, longer immobility time, significant decrease in motor activity, and spatial short-term memory. BME pretreatment reverted the behavioral changes to normal. MeHg exposure decreases the DNA and RNA content in cerebellum and also caused some pathological changes in cerebellum. Pretreatment with BME restored all the changes to near normal. These findings suggest that BME has a potent efficacy to alleviate MeHg-induced toxicity in rat cerebellum.     

Effects of methylmercury on epigenetic markers in three model species: Mink,chicken and yellow perch

We previously reported that methylmercury (MeHg) exposure is associated with DNA hypomethylation in the brain stem of male polar bears. Here, we conveniently use archived tissues obtained from controlled laboratory exposure studies to look for evidence that MeHg can disrupt DNA methylation across taxa. Brain (cerebrum) tissues from MeHg-exposed mink (Neovison vison), chicken (Gallus gallus) and yellow perch (Perca flavescens) were analyzed for total Hg levels and global DNA methylation. Tissues from chicken and mink, but not perch, were also analyzed for DNA methyltransferase (DNMT) activity. In mink we observed significant reductions in global DNA methylation in an environmentally-relevant dietary exposure group (1 ppm MeHg), but not in a higher group (2 ppm MeHg). DNMT activity was significantly reduced in all treatment groups. In chicken or yellow perch, no statistically significant effects of MeHg were observed. Dose-dependent trends were observed in the chicken data but the direction of the change was not consistent between the two endpoints. Our results suggest that MeHg can be epigenetically active in that it has the capacity to affect DNA methylation in mammals. The variability in results across species may suggest inter-taxa differences in epigenetic responses to MeHg, or may be related to differences among the exposure scenarios used as animals were exposed to MeHg through different routes (dietary, egg injection), for different periods of time (19–89 days) and at different life stages (embryonic, juvenile, adult).     

Experimental and Natural Warming Elevates Mercury Concentrations in Estuarine Fish

Marine food webs are the most important link between the global contaminant, methylmercury (MeHg), and human exposure through consumption of seafood. Warming temperatures may increase human exposure to MeHg, a potent neurotoxin, by increasing MeHg production as well as bioaccumulation and trophic transfer through marine food webs. Studies of the effects of temperature on MeHg bioaccumulation are rare and no study has specifically related temperature to MeHg fate by linking laboratory experiments with natural field manipulations in coastal ecosystems. We performed laboratory and field experiments on MeHg accumulation under varying temperature regimes using the killifish, Fundulus heteroclitus. Temperature treatments were established in salt pools on a coastal salt marsh using a natural temperature gradient where killifish fed on natural food sources. Temperatures were manipulated across a wider range in laboratory experiments with killifish exposed to MeHg enriched food. In both laboratory microcosms and field mesocosms, MeHg concentrations in killifish significantly increased at elevated temperatures. Moreover, in field experiments, other ancillary variables (salinity, MeHg in sediment, etc.) did not relate to MeHg bioaccumulation. Modeling of laboratory experimental results suggested increases in metabolic rate as a driving factor. The elevated temperatures we tested are consistent with predicted trends in climate warming, and indicate that in the absence of confounding factors, warmer sea surface temperatures could result in greater in bioaccumulation of MeHg in fish, and consequently, increased human exposure.     

Genome-Wide Association Analysis of Tolerance to Methylmercury Toxicity in Drosophila Implicates Myogenic and Neuromuscular Developmental Pathways

Methylmercury (MeHg) is a persistent environmental toxin present in seafood that can compromise the developing nervous system in humans. The effects of MeHg toxicity varies among individuals, despite similar levels of exposure, indicating that genetic differences contribute to MeHg susceptibility. To examine how genetic variation impacts MeHg tolerance, we assessed developmental tolerance to MeHg using the sequenced, inbred lines of the Drosophila melanogaster Genetic Reference Panel (DGRP). We found significant genetic variation in the effects of MeHg on development, measured by eclosion rate, giving a broad sense heritability of 0.86. To investigate the influence of dietary factors, we measured MeHg toxicity with caffeine supplementation in the DGRP lines. We found that caffeine counteracts the deleterious effects of MeHg in the majority of lines, and there is significant genetic variance in the magnitude of this effect, with a broad sense heritability of 0.80. We performed genome-wide association (GWA) analysis for both traits, and identified candidate genes that fall into several gene ontology categories, with enrichment for genes involved in muscle and neuromuscular development. Overexpression of glutamate-cysteine ligase, a MeHg protective enzyme, in a muscle-specific manner leads to a robust rescue of eclosion of flies reared on MeHg food. Conversely, mutations in kirre, a pivotal myogenic gene identified in our GWA analyses, modulate tolerance to MeHg during development in accordance with kirre expression levels. Finally, we observe disruptions of indirect flight muscle morphogenesis in MeHg-exposed pupae. Since the pathways for muscle development are evolutionarily conserved, it is likely that the effects of MeHg observed in Drosophila can be generalized across phyla, implicating muscle as an additional hitherto unrecognized target for MeHg toxicity. Furthermore, our observations that caffeine can ameliorate the toxic effects of MeHg show that nutritional factors and dietary manipulations may offer protection against the deleterious effects of MeHg exposure.     

Methylmercury and brain development: A review of recent literature

Methylmercury (MeHg) is a potent environmental pollutant, which elicits significant toxicity in humans. The central nervous system (CNS) is the primary target of toxicity, and is particularly vulnerable during development. Maternal exposure to MeHg via consumption of fish and seafood can have irreversible effects on the neurobehavioral development of children, even in the absence of symptoms in the mother. It is well documented that developmental MeHg exposure may lead to neurological alterations, including cognitive and motor dysfunction. The neurotoxic effects of MeHg on the developing brain have been extensively studied. The mechanism of toxicity, however, is not fully understood. No single process can explain the multitude of effects observed in MeHg-induced neurotoxicity. This review summarizes the most current knowledge on the effects of MeHg during nervous system development considering both, in vitro and in vivo experimental models. Considerable attention was directed towards the role of glutamate and calcium dyshomeostasis, mitochondrial dysfunction, as well as the effects of MeHg on cytoskeletal components/regulators.     

Proteome modifications of juvenile beluga (Huso huso) brain as an effect of dietary methylmercury

Methylmercury (MeHg) is the most toxic form of mercury which is bioaccumulated in the aquatic food chain. It has been shown that one of the main targets of MeHg toxicity is the brain, but there is little knowledge of the molecular mechanisms of its toxic effects. In this work we used a proteomics analysis to determine the changes in the brain proteome of juvenile beluga (Huso huso) exposed to dietary MeHg. The juvenile beluga were fed the diet containing 0.8 ppm MeHg for 70 days. Proteins of the brain tissue were analyzed using two-dimensional electrophoresis and MALDI-TOF/TOF mass spectrometry. We found eight proteins with significant altered expression level in the fish brain exposed to MeHg. These proteins are involved in different cell functions including cell metabolism, protein folding, cell division, and signal transduction. Our results support the idea that MeHg exerts its toxicity through oxidative stress induction and apoptotic effects. They also suggest that chronic MeHg exposure would induce an important metabolic deficiency in the brain. These findings provide basic information to understand possible mechanisms of MeHg toxicity in aquatic ecosystems.     

Dietary Exposure of the Red-Crowned Crane (Grus japonensis) to Total and Methyl Mercury in Zhalong Wetland,Northeastern China

To determine the dietary exposure of the migratory red-crowned crane to mercury (Hg), this study analyzed the concentrations of total mercury (T-Hg) and methyl mercury (MeHg) in its prey, i.e., reeds and three aquatic animal families (Perccottus glenni Dybowski, Cybister japonicus Sharp, and Viviparidae) in northeastern China. Results indicated that the Hg concentration in Zhalong Wetland was elevated through the food chain, and the prey of the red-crowned crane contained measurable levels of T-Hg and MeHg. In prey tissues, MeHg was the main form of the Hg element and accounted for 61 % of total Hg concentration in Viviparidae, 58 % in C. japonicus Sharp, and 85 % in P. glenni Dybowski. The highest T-Hg and MeHg concentrations ranged from 1.66 to 3.89 ppm and from 1.12 to 2.67 ppm, respectively, and they were detected in the feathers of the red-crowned cranes. The lowest T-Hg concentration was determined in the excretions of wild red-crowned cranes at 0.21 ppm; furthermore, the content of MeHg was below the detection limit. In Zhalong Wetland, the level of dietary exposure of the population of red-crowned cranes to Hg is below the threshold of Hg toxicity. Moreover, eggshells are suitable indicators of Hg risk levels to the red-crowned crane.     

Salivary parameters alterations after early exposure to environmental methylmercury: A preclinical study in offspring rats

BACKGROUNDMethylmercury (MeHg) is still considered a global pollutant of major concern; thus, it becomes relevant to investigate and validate alternative diagnostic methods to track early-life human exposure. This study aimed to evaluate the salivary parameters and to characterize potential mechanisms of oxidative damage on the salivary glands (SG) of offspring rats after pre- and postnatal environmental-experimental MeHg exposure.METHODSPregnant Wistar rats were daily exposed to 40 μg/kg MeHg during both gestational and lactation periods. Then, the saliva of offspring rats was analyzed in terms of flow rate, amylase activity, and total protein concentration. The SG of the offspring rats were dissected to perform the oxidative biochemistry analyses of antioxidant capacity against peroxyl radicals (ACAP), lipid peroxidation (LPO), and nitrite levels.RESULTSExposure to MeHg significantly decreased the ACAP, increased LPO and nitrite levels, decreased salivary flow rate, amylase activity, and total protein concentration.CONCLUSIONSaliva analyses can predict damages induced by early-life MeHg exposure and may be used as an auxiliary diagnostic method.     

Rapidly increasing methyl mercury in endangered ivory gull (Pagophila eburnea) feathers over a 130 year record

Mercury (Hg) is increasing in marine food webs, especially at high latitudes. The bioaccumulation and biomagnification of methyl mercury (MeHg) has serious effects on wildlife, and is most evident in apex predators. The MeHg body burden in birds is the balance of ingestion and excretion, and MeHg in feathers is an effective indicator of overall MeHg burden. Ivory gulls (Pagophila eburnea), which consume ice-associated prey and scavenge marine mammal carcasses, have the highest egg Hg concentrations of any Arctic bird, and the species has declined by more than 80% since the 1980s in Canada. We used feathers from museum specimens from the Canadian Arctic and western Greenland to assess whether exposure to MeHg by ivory gulls increased from 1877 to 2007. Based on constant feather stable-isotope (δ¹³C, δ¹⁵N) values, there was no significant change in ivory gulls'' diet over this period, but feather MeHg concentrations increased 45× (from 0.09 to 4.11 µg g⁻¹ in adults). This dramatic change in the absence of a dietary shift is clear evidence of the impact of anthropogenic Hg on this high-latitude threatened species. Bioavailable Hg is expected to increase in the Arctic, raising concern for continued population declines in high-latitude species that are far from sources of environmental contaminants.     

The Nicotine-Evoked Locomotor Response: A Behavioral Paradigm for Toxicity Screening in Zebrafish (Danio rerio) Embryos and Eleutheroembryos Exposed to Methylmercury

This study is an adaptation of the nicotine-evoked locomotor response (NLR) assay, which was originally utilized for phenotype-based neurotoxicity screening in zebrafish embryos. Zebrafish embryos do not exhibit spontaneous swimming until roughly 4 days post-fertilization (dpf), however, a robust swimming response can be induced as early as 36 hours post-fertilization (hpf) by means of acute nicotine exposure (30–240μM). Here, the NLR was tested as a tool for early detection of locomotor phenotypes in 36, 48 and 72 hpf mutant zebrafish embryos of the non-touch-responsive maco strain; this assay successfully discriminated mutant embryos from their non-mutant siblings. Then, methylmercury (MeHg) was used as a proof-of-concept neurotoxicant to test the effectiveness of the NLR assay as a screening tool in toxicology. The locomotor effects of MeHg were evaluated in 6 dpf wild type eleutheroembryos exposed to waterborne MeHg (0, 0.01, 0.03 and 0.1μM). Afterwards, the NLR assay was tested in 48 hpf embryos subjected to the same MeHg exposure regimes. Embryos exposed to 0.01 and 0.03μM of MeHg exhibited significant increases in locomotion in both scenarios. These findings suggest that similar locomotor phenotypes observed in free swimming fish can be detected as early as 48 hpf, when locomotion is induced with nicotine.     

Postnatal Methylmercury Exposure Induces Hyperlocomotor Activity and Cerebellar Oxidative Stress in Mice: Dependence on the Neurodevelopmental Period

During the early postnatal period the central nervous system (CNS) is extremely sensitive to external agents. The present study aims at the investigation of critical phases where methylmercury (MeHg) induces cerebellar toxicity during the suckling period in mice. Animals were treated with daily subcutaneous injections of MeHg (7 mg/kg of body weight) during four different periods (5 days each) at the early postnatal period: postnatal day (PND) 1–5, PND 6–10, PND 11–15, or PND 16–20. A control group was treated with daily subcutaneous injections of a 150 mM NaCl solution (10 ml/kg of body weight). Subjects exposed to MeHg at different postnatal periods were littermate. At PND 35, behavioral tests were performed to evaluate spontaneous locomotor activity in the open field and motor performance in the rotarod task. Biochemical parameters related to oxidative stress (levels of glutathione and thiobarbituric acid reactive substances, as well as glutathione peroxidase and glutathione reductase activity) were evaluated in cerebellum. Hyperlocomotor activity and high levels of cerebellar thiobarbituric acid reactive substances were observed in animals exposed to MeHg during the PND 11–15 or PND 16–20 periods. Cerebellar glutathione reductase activity decreased in MeHg-exposed animals. Cerebellar glutathione peroxidase activity was also decreased after MeHg exposure and the lowest enzymatic activity was found in animals exposed to MeHg during the later days of the suckling period. In addition, low levels of cerebellar glutathione were found in animals exposed to MeHg during the PND 16–20 period. The present results show that the postnatal exposure to MeHg during the second half of the suckling period causes hyperlocomotor activity in mice and point to this phase as a critical developmental stage where mouse cerebellum is a vulnerable target for the neurotoxic and pro-oxidative effects of MeHg.     

Linking fish consumption patterns and health risk assessment of mercury exposure in a coastal community of NW Mexico

We aimed at characterizing the methylmercury (MeHg) exposure through fish consumption in two populations (common general and fishing-related population (FRP)) using a probabilistic health risk assessment in children, women of childbearing age, and adults in Mazatlán. The hazard quotients (HQs) were obtained from fish consumption, defined through a survey, and the levels of mercury in fishery products, obtained from published information. The average fish ingestion rate (IR_food) was higher in the FRP (167.85 g d^?1) than in the general population (GP) (140.9 g d^?1). However, HQs were significantly (p < 0.05) higher in the GP (ranging from 0.18 to 10.91) compared to the FRP (0.20 to 2.48); significant differences were also found among groups of both populations. Remarkably, children in both populations exhibited the highest proportions of risk, reaching up to 97% in GP. For all populations, fish consumption was the most important variable influencing MeHg exposure. Overall, for MeHg exposure, there is no safe level of fish consumption without risk, and actions should be taken to mitigate possible risk; further research with current data is needed to assess potential health risks associated with MeHg exposure, particularly in children.     

Methylmercury Effects On Cell Cycle Kinetics

Abstract. Methylmercury (MeHg) effects on cell cycle kinetics were investigated to help identify its mechanisms of action. Flow cytometric analysis of normal human fibroblasts grown in vitro in the presence of BrdU allowed quantitation of the proportion of cells in G₁, S, G₂ and the next G₁ phase. This technique provides a rapid and easily performed method of characterizing phase lengths and transition rates for the complete cell cycle. After first exposure to MeHg the cell cycle time was lengthened due to a prolonged G₁. At 3, μm MeHg the G₁ phase length was 25% longer than the control. the G₁/S transition rate was also decreased in a dose-related manner. Confluent cells exposed to MeHg and replated with MeHg respond in the same way as cells which have not been exposed to MeHg before replating. Cells exposed for long times to MeHg lost a detectable G₁ effect, and instead showed an increase in the G₂ percentage, which was directly related to MeHg concentration and length of exposure. After 8 days at 5 μM MeHg, 45% of the population was in G₂. the G₂ accumulation was reversible up to 3 days, but at 6 days the cells remained in G₂ when the MeHg was removed. Cell counts and viability indicated that there was not a selective loss of cells from the MeHg. MeHg has multiple effects on the cell cycle which include a lengthened G₁ and decreased transition probability after short term exposure of cycling cells, and a G₂ accumulation after a longer term exposure. There were no detectable S phase effects. It appears that mitosis (the G₂ accumulation) and probably synthesis of some macromolecules in G₁ (the lengthened G₁ and lowered transition probability) are particularly susceptible to MeHg.     

Methylmercury Alters the Activities of Hsp90 Client Proteins,Prostaglandin E Synthase/p23 (PGES/23) and nNOS

Methylmercury (MeHg) is a persistent pollutant with known neurotoxic effects. We have previously shown that astrocytes accumulate MeHg and play a prominent role in mediating MeHg toxicity in the central nervous system (CNS) by altering glutamate signaling, generating oxidative stress, depleting glutathione (GSH) and initiating lipid peroxidation. Interestingly, all of these pathways can be regulated by the constitutively expressed, 90-kDa heat shock protein, Hsp90. As Hsp90 function is regulated by oxidative stress, we hypothesized that MeHg disrupts Hsp90-client protein functions. Astrocytes were treated with MeHg and expression of Hsp90, as well as the abundance of complexes of Hsp90-neuronal nitric oxide synthase (nNOS) and Hsp90-prostaglandin E synthase/p23 (PGES/p23) were assessed. MeHg exposure decreased Hsp90 protein expression following 12 h of treatment while shorter exposures had no effect on Hsp90 protein expression. Interestingly, following 1 or 6 h of MeHg exposure, Hsp90 binding to PGES/p23 or nNOS was significantly increased, resulting in increased prostaglandin E₂ (PGE₂) synthesis from MeHg-treated astrocytes. These effects were attenuated by the Hsp90 antagonist, geldanmycin. NOS activity was increased following MeHg treatment while cGMP formation was decreased. This was accompanied by an increase in •O₂ ⁻ and H₂O₂ levels, suggesting that MeHg uncouples NO formation from NO-dependent signaling and increases oxidative stress. Altogether, our data demonstrates that Hsp90 interactions with client proteins are increased following MeHg exposure, but over time Hsp90 levels decline, contributing to oxidative stress and MeHg-dependent excitotoxicity.     

Selenium Health Benefit Values as Seafood Safety Criteria

Selenium (Se) is absolutely required for activity of 25–30 genetically unique enzymes (selenoenzymes). All forms of life that have nervous systems possess selenoenzymes to protect their brains from oxidative damage. Homeostatic mechanisms normally maintain optimal selenoenzyme activities in brain tissues, but high methylmercury (MeHg) exposures sequester Se and irreversibly inhibit selenoenzyme activities. However, nutritionally relevant amounts of Se can replace the Se sequestered by MeHg and maintain normal selenoenzyme activities, thus preventing oxidative brain damage and other adverse consequences of MeHg toxicity. Findings of studies that seem contradictory from MeHg exposure perspectives are entirely consistent from MeHg:Se molar ratio perspectives. Studies that have reported dose-dependent consequences of maternal MeHg exposures on child development uniformly involved seafoods that contained much more Hg than Se. Meanwhile more typical varieties of ocean fish contain much more Se than Hg. This may explain why maternal MeHg exposure from eating ocean fish is associated with major IQ benefits in children instead of harm. Therefore, instead of being avoided, ocean fish consumption should be encouraged during pregnancy. However, the safety of freshwater fish consumption is less certain. In freshwater fish, MeHg bioaccumulation and toxicity are both inversely related to Se bioavailability. Their Se can be far lower than their MeHg contents, potentially making them more dangerous than pilot whale meats. Therefore, to provide accurate and appropriate regulatory advice regarding maternal consumption of seafoods and freshwater fish, Hg:Se molar ratios need to be incorporated in food safety criteria.     

The effect of aqueous speciation and cellular ligand binding on the biotransformation and bioavailability of methylmercury in mercury-resistant bacteria

Mercury resistant bacteria play a critical role in mercury biogeochemical cycling in that they convert methylmercury (MeHg) and inorganic mercury to elemental mercury, Hg(0). To date there are very few studies on the effects of speciation and bioavailability of MeHg in these organisms, and even fewer studies on the role that binding to cellular ligands plays on MeHg uptake. The objective of this study was to investigate the effects of thiol complexation on the uptake of MeHg by measuring the intracellular demethylation-reduction (transformation) of MeHg to Hg(0) in Hg-resistant bacteria. Short-term intracellular transformation of MeHg was quantified by monitoring the loss of volatile Hg(0) generated during incubations of bacteria containing the complete mer operon (including genes from putative mercury transporters) exposed to MeHg in minimal media compared to negative controls with non-mer or heat-killed cells. The results indicate that the complexes MeHgOH, MeHg-cysteine, and MeHg-glutathione are all bioavailable in these bacteria, and without the mer operon there is very little biological degradation of MeHg. In both Pseudomonas stutzeri and Escherichia coli, there was a pool of MeHg that was not transformed to elemental Hg(0), which was likely rendered unavailable to Mer enzymes by non-specific binding to cellular ligands. Since the rates of MeHg accumulation and transformation varied more between the two species of bacteria examined than among MeHg complexes, microbial bioavailability, and therefore microbial demethylation, of MeHg in aquatic systems likely depends more on the species of microorganism than on the types and relative concentrations of thiols or other MeHg ligands present.     

Comparison of dietary mercury exposure in two sympatric top predator fishes, largemouth bass and northern pike: a bioenergetics modeling approach

Physical and ecologicalfactors, including lake temperature, fishphysiology, and diet, influence methylmercury(MeHg) exposure in fish. We employedbioenergetics modeling to compare dietary MeHgexposure in sympatric top predators, largemouthbass (Micropterus salmoides) and northernpike (Esox lucius). We comparedsimulations using field data to hypotheticalsimulations with (1) ± 25% change in meandaily lake temperature for juvenile and adultbass and pike; (2) ± 25% change inlong-term growth rate of pike; (3) adult bassdiet shift from generalist predator to strictpiscivore. Bass and pike MeHg exposures weresimilar in baseline simulations and reflectedpatterns in field tissue concentrations. Thisoccurred despite the fact that bass consumedhighly contaminated benthic invertebrates,while pike exclusively consumed lesscontaminated fish prey. Higher temperaturesincreased adult bass and pike MeHg exposures by35% and 27%, respectively. Shifting adultbass diets to 100% fish resulted in a 54%decrease in exposure, while increasing pikegrowth rates resulted in a 24% decrease. Bioenergetics modeling proved useful inunderstanding the influence of temperature,prey-base, and predator growth on differencesin Hg exposure across fish species.