Effect of subchronic administration of ethanol and methylmercury in combination on the tissue distribution of mercury in rats

The effect of oral administration for 14 weeks of 8 g.kg-1.day-1 ethanol and 0.5 mg.kg-1.day-1 methylmercuric chloride in combination to rats fed isocaloric diets has been investigated. Ethanol, in contrast to published studies, failed to influence the tissue distribution of methylmercury and its inorganic mercury metabolite in brain and kidney, and did not inhibit the increase in kidney weight induced by methylmercury. Ethanol and methylmercury, in combination and individually, reduced the renal but not the hepatic activity of gamma-glutamyltransferase, but did not affect the renal and biliary concentration of reduced glutathione. Further study is required to determine the circumstances under which ethanol can influence the tissue distribution of methylmercury and its inorganic mercury metabolite.     

Steroid sulfatase and sulfuryl transferase activity in monkey brain tissue

Dehydroepiandrosterone and its sulfated form are commonly known as modulators of gamma-aminobutyrate A and N-methyl-D-aspartate receptors. In spite of poor permeability of the blood-brain barrier for sulfated steroids, high concentrations of dehydroepiandrosterone and also its sulfate have been found in brain tissue. Physiological concentrations of these neuromodulators are maintained by two enzymes present in the blood and many peripheral tissues, including the brain, namely, steroid sulfatase and neurosteroid sulfuryl transferase (NSST). This prompted us to investigate activities of these enzymes in primate brain tissue. Rather low neurosteroid sulfuryl transferase activity was detectable in in vitro incubations of cytosol fractions from male and female Macaca mulatta brains, dissected to cerebral cortex, subcortex, and cerebellum. In male monkeys, the highest activity was found in the cerebellum followed by cortex and subcortex. On the other hand, in female monkeys, the highest activity was determined in the cortex followed by subcortex and cerebellum. Steroid sulfatase activity was determined in in vitro microsomal samples from each of the above-mentioned brain regions. Specific activities in female cerebral regions declined in the order: cerebellum, cortex, and subcortex. In male monkeys, no significant difference among the studied regions was observed. Using dehydroepiandrosterone sulfate as a substrate, the apparent kinetic characteristics of steroid sulfatase were determined as follows: K(M) 36.10 +/- 8.33 microM, V(max) 8.38 +/- 1.68 nmol/h/mg protein. These results will serve as a basis for further studies concerning the pathophysiology of human brain tumors.     

汞在新建水库食物网中生物累积与风险评价研究进展

汞是唯一参与全球循环的液态重金属。1974年,自美国学者Smith首次报道水库中鱼类总汞含量高于邻近自然湖泊以来,水库中鱼类汞升高的风险成为新建水库环境影响评价中的重要内容之一。汞在水库生态系统生物组分和非生物组分中含量升高的现象先后在世界各国报道,包括加拿大、美国、芬兰、泰国和巴西等。通过对系列的野外研究进行回顾,表明了水库形成后生态系统中汞的甲基化过程发生了变化。水库形成对汞在食物网中的鱼类、底栖生物、浮游生物的累积产生影响。水库中汞的生物累积、迁移转化主要与被淹没土壤和植物腐解过程有着直接或间接的关系。水库形成后,总汞、甲基汞和甲基汞比例在生态系统食物网各组分中的变化并不一致。蓄水后,水体中总汞变化较小,甲基汞和甲基汞比例上升明显;浮游生物尤其是浮游动物中总汞升高,但甲基汞和甲基汞比例升高更为明显;与浮游动物类似,底栖水生昆虫中总汞升高,甲基汞和甲基汞比例升高也更为明显;鱼类作为食物网顶级消费者,甲基汞比例一般在80%以上,在水库形成后鱼类总汞和甲基汞均明显升高,但甲基汞比例变化已经不大。这些变化揭示了水库形成后甲基汞在食物网传递的两个主要可能途径,一是微型生物食物网。通过悬浮颗粒物、浮游植物、浮游动物这一环节,甲基汞和甲基汞比例有明显的增加。第二个途径是底层生物食物网。通过悬浮颗粒物、细菌、碎屑食性底栖水生昆虫、肉食型底栖水生昆虫环节,甲基汞和甲基汞比例明显增加。这两种途径均能导致以水生昆虫、小鱼、甲壳类等为食的肉食性鱼类汞含量增加。水库形成后,生态系统中汞的甲基化发生了明显的"加速"过程。这种"加速"过程最直接的因素是成库后大量土壤淹没使得汞的甲基化平衡被打破。这个过程主要有两方面的影响。一方面是直接影响,被淹没土壤和植被在腐解过程中主动或被动地将甲基汞释放到水库生态系统中;另一方面是间接影响,被淹没土壤和植被的腐解使水库底部形成厌氧环境,有利于无机汞从被淹没土壤和植被中溶出,为甲基化反应提供充裕的、可供甲基化的无机汞,同时腐解产生的大量营养物质为微生物提供丰富食物来源,使硫酸盐还原菌大量繁殖,促进无机汞的甲基化。在我国,有关汞在新建水库食物网中生物累积和风险评价的研究有待进一步加强。     

COMPARISON OF THE BINDING’ OF β-ALANINE AND y-AMINOBUTYRIC ACID IN SYNAPTOSOMAL-MITOCHONDRIAL FRACTIONS OF RAT BRAIN1

Abstract— Na⁺-dependent ‘binding’ of β-alanine and GABA was examined with synaptosomal-mitochondrial fractions of rat brain incubated for 10 min at 0°C. GABA was bound to a much greater extent than β-alanine to particles of cerebral cortex, whole cerebellum and brain stem. For cerebral cortex, the binding capacity (B_max) for GABA was about 18 limes greater than that for β-alanine. and the affinity of the particles for GABA was about 2′ times greater than for β-alanine. The order of potency of GABA binding to brain regions was cerebral cortex > cerebellum > brain stem, whereas that for β-alanine was the reverse. If the binding of β-alanine is taken to indicate the glial component of the Na⁺-dependent binding process for GABA, then most of the GABA was bound to neuronal elements under the conditions employed.     

Schleim aus der Braunalge Nemacystus decipiens,Mozuku 1. Mitt. Bestandteile des Schleims und ihre Eigenschaften

We examined the distribution of mercury and selenium in fifteen tissues of striped dolphins (Stenella coeruleoalbd). The total mercury level in the mature dolphins showed differences among the tissues and was highest in the liver. The total mercury concentration in most tissues increased with age, and reached a constant level at 20 to 25 years of age. The total mercury level in the tissues was not appreciably different among mature males, pregnant females, lactating females and resting females. In the muscle of mature individuals, the total mercury level of striped dolphins collected in 1977 and 1980 was appreciably higher than that of those collected 1978 and 1979. Methylmercury showed less variation in concentration among the tissues. The ratio of methylmercury to total mercury in muscle decreased with growth after about 10 years of age when the increase of methylmercury stopped. Selenium levels in the dolphins increased with age as total mercury levels did. High correlation coefficients were found between the total mercury and selenium levels in spleen, muscle, pancreas and liver. The concentrations of total mercury in the various tissues of immature dolphins were much lower than those of mature ones.     

Activity of pyruvate- and ketoglutarate dehydrogenase complexes on various regions of the rat brain

The activity of pyruvate dehydrogenase and ketoglutarate dehydrogenase complexes (PDC; EC 1.2.4.1 and KDC; EC 1.2.4.2, respectively) was studied in extracts and lysates of mitochondria isolated from the cortex, cerebellum and stem of the rat brain. In all the mentioned cerebral areas the PDC activity calculating per 1 mg protein noticeably increased that for KDC. Under conditions of solubilization the activity of KDC lowered to a greater extent than that of PDC. The studied brain areas are arranged according to the activity of PDC and KDC in lysates in the following order: stem much much greater than cortex greater than cerebellum, however, the highest stock of the PDC activity manifesting with activation of endogenous phosphatase and dephosphorylation of the complex is observed in cerebellum. When calculating per mitochondria isolated from 1 g of tissue, the value of the PDC activity/KDC activity ratio in all areas of the brain is more than 1, and in cerebellum allowing for the complete PDC activity it exceeds 3. The data obtained emphasize the biochemical originality of the cerebellum and the presence of specific peculiarities in regulation of the pyruvate dehydrogenase complex activity in this cerebral area.     

Dose and duration related methylmercury deposition, glycosidases inhibition, myelin degeneration and chelation therapy

Methylmercury accumulation in different parts of the CNS (olfactory bulbs, cerebral hemispheres, cerebellum, medulla oblongata and spinal cord) in relation to the cytoarchitectural changes in myelin sheath as well as in glycosidases levels have been reported. Male albino rats were treated with low and high doses of methylmercury chloride (1 mg/kg and 10 mg/kg) N-acetyl-DL-homocysteine thiolactone (40 mg/kg and 80 mg/kg) and glutathione (100 mg/kg and 150 mg/kg) for varied time periods. The study shows a dose and duration dependent accumulation of mercury in all the CNS areas coinciding with a progressive myelin degeneration and inhibition of the glycosidases. A casual relationship between the amount of mercury accumulation and the extent of enzymes inhibition, in any particular area of CNS, could not be established. Similarly none of the antagonists is (though has been successful in recovering the enzymes and lessening the mercury burden in a few isolated cases) able to bring an absolute control value in any group.     

Quantification of glial fibrillary acidic protein in developing sheep brain cortex, cerebellum and stem

1. The glial fibrillary acidic protein (GFAP) content of foetal, young (lamb) and adult sheep brain white (stem and cerebellum) and grey (cortex) matter-enriched regions has been determined by means of an improved ELISA using one layer of anti-human GFAP monoclonal antibody. 2. The order of GFAP concentration in brain regions was as follows: brain stem greater than cerebellum greater than cortex. 3. Postnatal brain development accounts for an increase of GFAP in all the regions. The most important increase in GFAP was observed in the adult brain and was proportionally more significant in the grey matter-enriched cortex.     

Effects of mercury compounds on the spontaneous and potassium-evoked release of [3H]dopamine from mouse striatal slices

The effects of mercury compounds on the spontaneous and potassium-evoked release of [3H]dopamine from mouse striatal slices have been examined. All mercury compounds examined produced concentration-dependent increases in the spontaneous release of [3H]dopamine, with an order of potency of methylmercury greater than mercuric (Hg2+) mercury greater than p-choloromercuribenzene sulfonic acid. Methylmercury had no effect on the 25 mM potassium evoked release of [3H]dopamine in the presence of 1.3 mM calcium. However, in calcium-free conditions, methylmercury significantly increased the potassium-evoked release of [3H]dopamine. Mercuric mercury significantly reduced the 25 mM potassium evoked release of [3H]dopamine in the presence of 1.3 mM calcium, and this response was not reversible with brief washing of the tissue. In calcium-free conditions, mercuric mercury significantly elevated the evoked release of [3H]dopamine, similar to the result obtained with methylmercury. It is suggested that mercury compounds alter dopaminergic synaptic function, possibly by disrupting calcium homeostasis or calcium-dependent processes, and that methylmercury and mercuric mercury can have differential effects to alter dopaminergic neurotransmission.     

Accumulation of mercury in estuarine food chains

To understand the accumulation of inorganic mercury and methylmercury at the base of the estuarine food chain, phytoplankton (Thalassiosira weissflogii) uptake and mercury speciation experiments were conducted. Complexation of methylmercury as methylmercury-bisulfide decreased the phytoplankton uptake rate while the uptake rate of the methylmercury-cysteine and -thiourea complexes increased with increasing complexation by these ligands. Furthermore, our results indicated that while different ligands influenced inorganic mercury/methylmercury uptake by phytoplankton cells, the ligand complex had no major influence on either where the mercury was sequestered within the phytoplankton cell nor the assimilation efficiency of the mercury by copepods. The assimilation efficiency of inorganic mercury/methylmercury by copepods and amphipods feeding on algal cells was compared and both organisms assimilated methylmercury much more efficiently; the relative assimilation efficiency of methylmercury to inorganic mercury was 2.0 for copepods and 2.8 for amphipods. The relative assimilation is somewhat concentration dependent as experiments showed that as exposure concentration increased, a greater percentage of methylmercury was found in the cytoplasm of phytoplankton cells, resulting in a higher concentration in the copepods feeding on these cells. Additionally, food quality influenced assimilation by invertebrates. During decay of a T. weissflogii culture, which served as food for the invertebrates, copepods were increasingly less able to assimilate the methylmercury from the food, while even at advanced stages of decay, amphipods were able to assimilate mercury from their food to a high degree. Finally, fish feeding on copepods assimilated methylmercury more efficiently than inorganic mercury owing to the larger fraction of methylmercury found in the soft tissues of the copepods.     

Regional distribution and relative amounts of glutamate decarboxylase isoforms in rat and mouse brain.

The levels of the two isoforms of glutamate decarboxylase (GAD) were measured in 12 regions of adult rat brain and three regions of mouse brain by sodium dodecylsulfate-polyacrylamide gel electrophoresis and immunoblotting with an antiserum that recognizes the identical C-terminal sequence in both isoforms from both species. In rat brain the amount of smaller isoform, GAD65, was greater than that of the larger isoform, GAD67, in all twelve regions. GAD65 ranged from 77-89% of total GAD in frontal cortex, hippocampus, hypothalamus, midbrain, olfactory bulb, periaqueductal gray matter, substantia nigra, striatum, thalamus and the ventral tegmental area. The proportion of GAD65 was lower in amygdala and cerebellum but still greater than half of the total. There was a strong correlation between total GAD protein and GAD activity. In the three mouse brain regions analysed (cerebellum, cerebral cortex and hippocampus) the proportion of GAD65 (35,47, and 51% of total GAD) was significantly lower than in the corresponding rat-brain regions. The amount of GAD67 was greater than the amount of GAD65 in mouse cerebellum and was approximately equal to the amount of GAD65 in mouse cerebral cortex and hippocampus.     

Regional levels of glucose,amino acids,high energy phosphates,and cyclic nucleotides in the central nervous system during hypoglycemic stupor and behavioral recovery

The effects of insulin-induced hypoglycemic stupor and subsequent treatment with glucose on mouse cerebral cortical, cerebellar and brain stem levels of glucose, glycogen, ATP, phosphocreatine, glutamate, aspartate and GABA and on cerebral cortical and cerebellar levels of cyclic AMP and cyclic GMP have been measured. Hypoglycemia decreased glucose, glycogen and glutamate levels and had no effect on ATP levels in all three regions of brain. GABA levels were decreased only in cerebellum. Aspartate levels rose in cerebral cortex and brain stem, and creatine phosphate increased in cerebral cortex and cerebellum. In the hypoglycemic stuporous animals, cyclic GMP levels were elevated in cerebral cortex and depressed in cerebellum whereas cyclic AMP levels were unchanged from control values. Intravenous administration of 2.5-3.5 mmol/kg of glucose to the hypoglycemic stuporous animals produced recovery of near normal neurological function within 45 s. Only brain glucose and aspartate levels returned to normal prior to behavioral recovery. These results suggest that of the several substances examined in this study, only glucose and perhaps aspartate have important roles in the biochemical mechanisms producing neurological abnormalities in hypoglycemic animals.     

Postnatal ontogeny of uridine kinase in the cerebellum,hypothalamus, and cerebral cortex of the rat

Postnatal developmental patterns of uridine kinase were determined in crude subcellular fractions of the rat cerebellum, hypothalamus and cerebral cortex at ages 3 through 60 days. The highest specific activity and predominant distribution of enzyme was in the 105,000g supernatant of the 3 brain regions. Enzyme activity in hypothalamus and cerebral cortex was maximum at 3 days and decreased with age; in cerebellum it increased through 13 days and decreased thereafter. Thus, the pattern of activity in hypothalamus and cerebral cortex paralleled changes in DNA and RNA synthesis through age 60 days; in cerebellum, it more closely approximated changes in DNA synthesis during early development. Changes inK _m with aging suggest that the brain regions contain more than one form of enzyme. The highest particulate activity was in the microsomal fraction of the cerebellum and hypothalamus at all ages and in the cortex at 35 and 60 days. Relative specific activity for microsomal fractions of the brain regions at 60 days indicate a concentration of the enzyme which may be relevant in the maintenance of RNA activity in adult brain.     

Immuno-detection of aluminium and aluminium induced conformational changes in calmodulin - implications in Alzheimer's disease

Studies were conducted to ascertain any involvement of free radical mediated prooxidative processes in different brain regions following diazopam administration. A significant decrease in TBA reactive substance formation was observed in cerebral cortex, cerebellum and brain stem regions after single doses of 1.5, 3 and 6 mg/kg b.wt. For further studies rats were given diazepam (i.p.) at 3 mg/kg body weight dose and sacrificed after 1 h to follow changes in the pro/antioxidant status. An enhancement in the TBARS formation was found in the mitochondrial fractions from cerebral cortex and brain stem. This effect was highest in brain stem being 107% as compared to controls. In the post mitochondrial fraction, cerebellum showed 49% enhancement whereas decreased formation of thiobarbituric acid reactive substances was observed in cerebral cortex and brain stem. Isozymes of superoxide dismutase showed a decrease in activity which was region dependent. Even though, total thiols were not significantly altered, free thiols showed depletion in cerebellum (39.8%) and brain stem (50%). Glutathione reductase activity was also decreased in cerebellum and brain stem. The results indicate that a single dose of diazepam causes free radical mediated changes and the modulatory response of antioxidant defences appears to be region specific.     

Preventive Effects of Dextromethorphan on Methylmercury-Induced Glutamate Dyshomeostasis and Oxidative Damage in Rat Cerebral Cortex

Methylmercury (MeHg) is a well-known environmental pollutant leading to neurotoxicant associated with aberrant central nervous system (CNS) functions, but its toxic mechanisms have not yet been fully recognized. In the present study, we tested the hypothesis that MeHg induces neuronal injury via glutamate (Glu) dyshomeostasis and oxidative damage mechanisms and that these effects are attenuated by dextromethorphan (DM), a low-affinity and noncompetitive N-methyl-d-aspartate receptor (NMDAR) antagonist. Seventy-two rats were randomly divided into four groups of 18 animals in each group: control group, MeHg-treated group (4 and 12 μmol/kg), and DM-pretreated group. After the 4-week treatment, we observed that the administration of MeHg at a dose of 12 μmol/kg significantly increased in total mercury (Hg) levels, disrupted Glu metabolism, overexcited NMDARs, and led to intracellular calcium overload in the cerebral cortex. We also found that MeHg reduced nonenzymatic and enzymatic antioxidants, enhanced neurocyte apoptosis, induced reactive oxygen species (ROS), and caused lipid, protein, and DNA peroxidative damage in the cerebral cortex. Moreover, glutamate/aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1) appeared to be inhibited by MeHg exposure. These alterations were significantly prevented by the pretreatment with DM at a dose of 13.5 μmol/kg. In conclusion, these findings strongly implicate that DM has potential to protect the brain from Glu dyshomeostasis and oxidative damage resulting from MeHg-induced neurotoxicity in rat.     

Accumulation of methylmercury and inorganic mercury in the brain

Differences in metabolism between different mercury species are well recognized. Conclusions that only a minor demethylation of methylmercury takes place in the brain are based primarily on results from short term studies. Results from a number of studies on humans exposed for many years to methylmercury have shown high concentrations of inorganic mercury in the brain in relation to total mercury. Similar evidence is available from studies on monkeys exposed for several years to methylmercury. The results indicate that a significant accumulation of inorganic mercury takes place with time despite the fact that the demethylation rate is slow. Differences in biological halftimes between different mercury species will explain the results. Some data do still need confirmation using different analytical methods. There is reason to believe that the one-compartment model for methyl mercury cannot be used without reservations. Inorganic mercury has a complicated metabolism. After exposure to metallic mercury vapor, inorganic mercury, probably bound to selenium, accumulates in the brain. A fraction of the mercury is excreted, with a long biological halftime. Studies on rats and monkeys indicate that inorganic mercury penetrates the blood-brain barrier only to a very limited-extent.     

Elevated mercury bound to serum proteins in methylmercury poisoned rats after selenium treatment

Methylmercury is a toxic pollutant and is generated by microbial methylation of elemental or inorganic mercury in the environment. Previous study found decreased hepatic MDA levels and urinary mercury levels in methylmercury poisoned rats after sodium selenite treatment. This study further found increased mercury levels in serum samples from methylmercury poisoned rats after selenium treatment. By using size exclusion chromatography coupled to inductively coupled plasma mass spectrometry, three Hg- binding protein fractions and two Se-binding protein fractions were identified with the molecular weight of approximately 21, 40, and 75 kDa and of 40 and 75 kDa, respectively. Elevated mercury level in the 75 kDa protein fraction was found binding with both Hg and Se, which may explain the decreased urinary Hg excretion in MeHg poisoned rats after Se treatment. MALDI-TOF-MS analysis of the serum found that the 75 kDa protein fractions were albumin binding with both Hg and Se and the 21 kDa fraction was Hg- binding metallothionein.     

Effect of Acute Administration of 3-Butyl-1-Phenyl-2-(Phenyltelluro)Oct-En-1-One on Oxidative Stress in Cerebral Cortex,Hippocampus, and Cerebellum of Rats

Organotellurium compounds have been synthesized since 1840, but pharmacological and toxicological studies about them are still incipient. Therefore, the objective of this study was to verify the effect of acute administration of the organochalcogen 3-butyl-1-phenyl-2-(phenyltelluro)oct-en-1-one on some parameters of oxidative stress in the brain of 30-day-old rats. Animals were treated intraperitoneally with a single dose of the organotellurium (125, 250, or 500 μg/kg body weight) and sacrificed 60 min after the injection. The cerebral cortex, the hippocampus, and the cerebellum were dissected and homogenized in KCl. Afterward, thiobarbituric acid reactive substances (TBARS), carbonyl, sulfhydryl, catalase (CAT), superoxide dismutase (SOD), nitric oxide (NO) formation, and hydroxyl radical production were measured in the brain. The organotellurium enhanced TBARS in the cerebral cortex and the hippocampus, and increased protein damage (carbonyl) in the cerebral cortex and the cerebellum. In contrast, the compound provoked a reduced loss of thiol groups measured by the sulfhydryl assay in all the tissues studied. Furthermore, the activity of the antioxidant enzyme CAT was reduced by the organochalcogen in the cerebral cortex and the cerebellum, and the activity of SOD was inhibited in all the brain tissues. Moreover, NO production was increased in the cerebral cortex and the cerebellum by this organochalcogen, and hydroxyl radical formation was also enhanced in the cerebral cortex. Our findings indicate that this organotellurium compound induces oxidative stress in the brain of rats, corroborating that this tissue is a potential target for organochalcogen action.     

Phytoremediation of Mercury- and Methylmercury-Polluted Soils Using Genetically Engineered Plants

Inorganic mercury in contaminated soils and sediments is relatively immobile, though biological and chemical processes can transform it to more toxic and bioavailable methylmercury. Methylmercury is neurotoxic to vertebrates and is biomagnified in animal tissues as it is passed from prey to predator. Traditional remediation strategies for mercury contaminated soils are expensive and site-destructive. As an alternative we propose the use of transgenic aquatic, salt marsh, and upland plants to remove available inorganic mercury and methylmercury from contaminated soils and sediments. Plants engineered with a modified bacterial mercuric reductase gene, merA, are capable of converting Hg(II) taken up by roots to the much less toxic Hg(0), which is volatilized from the plant. Plants engineered to express the bacterial organo-mercurial lyase gene, merB, are capable of converting methylmercury taken up by plant roots into sulfhydryl-bound Hg(II). Plants expressing both genes are capable of converting ionic mercury and methylmercury to volatile Hg(0) which is released into an enormous global atmospheric Hg(0) pool. To assess the phytoremediation capability of plants containing the merA gene, a variety of assays were carried out with the model plants Arabidopsis thaliana, and tobacco (Nicotiana tabacum).