Pharmacological blockage and P2X7 deletion hinder aversive memories: reversion in an enriched environment.

Adenosine triphosphate (ATP) plays a role in cell signaling. It was soon proposed that ATP activates ionotropic P2X receptors, exerting an influence on neurons as well as on glial cells. In addition to the fact that the activation of P2X and P2Y receptors can stimulate or inhibit the release of glutamate from rat hippocampal neurons, the release of ATP has been implicated in hippocampal long-term potentiation (LTP). Through different behavioral paradigms, this study aimed to investigate the participation of P2X7R in genetically modified (knockout (KO)) mice with the suppressed expression of this receptor and in the pharmacological blockage of this receptor in rats, as well as to evaluate the effect of environmental enrichment on potential mnemonic deficits. The results suggest that P2X7R participates in aversive memory processes: pharmacological blockage with the selective P2X7R antagonist, A-740003, in different time frames elicited dose-dependent impairments in memory acquisition, consolidation and retrieval in rats that were submitted to the contextual fear-conditioning (FC) task, and the deletion of P2X7R hampered the aversive memory processes of mice that were subjected to the FC paradigm. Experiments using mice that were subjected to environmental enrichment suggest that this form of stimulation reverses mnemonic impairments that are ascribed to the absence of the P2X7R, suggesting that these receptors do not participate on such a reversal. Finally, no alterations were observed in the habituation memory of P2X7KO mice.

Enzymatic GST levels and overall health of mullets from contaminated Brazilian Lagoons.

Glutathione S-transferase (GST) assays in non-mammalian organisms are usually conducted inappropriately, since no previous standardization of the optimal concentrations of proteins and substrates and adequate pH is conducted. Standardization is a key task to adjust enzyme assays at their kinetically correct maximal initial velocities, if one wants these velocities to indicate the amount of enzyme in a sample. In this paper GST assays were standardized in liver cytosol to compare seasonal GST levels in liver of mullet from two contaminated lagoons in the Rio de Janeiro to those from a reference bay. GST potential as a biomarker of sublethal intoxication in this species was also evaluated. Mullet liver GST levels assayed with substrates that corresponded to three different GST isoenzymes varied throughout the year. The differences indicated that mullets are suffering from sublethal intoxication from contaminants in these lagoons. Seasonal variations of activity were relevant, since these could indicate differences in xenobiotic input into the areas. An analysis of overall mullet health condition using a morphological index (the Fulton Condition Factor) and macroscopic abnormalities corroborated the differences in GST levels, with fish from one of the sites in worse overall health condition showing lower and significantly different FCF when compared to the reference site. Therefore, GST standardized activity levels are useful biomarkers of environmental contamination for mullet.

Metals in water, sediments, and biota of an offshore oil exploration area in the Potiguar Basin, Northeastern Brazil.

Metal concentrations were evaluated in water, bottom sediments, and biota in four field campaigns from 2002 to 2004 in the Potiguar Basin, northeastern Brazil, where offshore oil exploration occurs. Analyses were performed by inductively coupled plasma mass spectrometry and inductively coupled plasma optical emission spectrometry. Total metal concentrations in water (dissolved + particulate) and sediments were in the range expected for coastal and oceanic areas. Abnormally high concentrations in waters were only found for Ba (80 µg l(-1)) and Mn (12 µg l(-1)) at the releasing point of one of the outfalls, and for the other metals, concentrations in water were found in stations closer to shore, suggesting continental inputs. In bottom sediments, only Fe and Mn showed abnormal concentrations closer to the effluent releasing point. Metal spatial distribution in shelf sediments showed the influence of the silt-clay fraction distribution, with deeper stations at the edge of the continental shelf, which are much richer in silt-clay fraction showing higher concentrations than shallower sediments typically dominated by carbonates. Metal concentrations in estuarine (mollusks and crustaceans) and marine (fish) organisms showed highest concentrations in oysters (Crassostrea rhizophorae). Fish tissues metal concentrations were similar between the continental shelf influenced by the oil exploration area and a control site. The results were within the range of concentrations reported for pristine environments without metals contamination. The global results suggest small, if any, alteration in metal concentrations due to the oil exploration activity in the Potiguar Basin. For monitoring purposes, the continental inputs and the distribution of the clay-silt fraction need to be taken into consideration for interpreting environmental monitoring results.

Direct determination of tin in whole blood and urine by GF AAS.

OBJECTIVE: The aim of this work was to develop a procedure for the determination of tin in whole blood and urine by GF AAS with a minimum sample pre-treatment, using Pd/Mg as chemical modifier. DESIGN AND METHODS: The analyses of tin were conducted using an atomic absorption spectrometer with Zeeman background correction. The laboratory staff volunteered blood and urine samples for the experimental studies and application of the methodology. RESULTS: Samples were just diluted with 0.2% v/v Triton X-100, and pyrolysis and atomization temperatures of 1300 and 2200 °C were used. External calibration was performed with matrix matched calibration solutions. Limits of detection of 2.7 and 0.8 µgL(-1) were reached for blood and urine, respectively. The method was applied to the determination of Sn in blood and urine of eleven subjects not occupationally exposed, working in a laboratory of toxicology in a large Brazilian city, and the results ranged from 7.4 to 11.2 µgL(-1) and ≤0.8 to 2.2 µgL(-1), for blood and urine, respectively. Accuracy was assessed by analysis of standard reference materials for tin in blood (Contox I, lot TM144-1097, Kaulson Laboratories, USA) and urine (Seronorm, lot 0511545, Sero AS, Norway). CONCLUSIONS: Results showed good agreement between experimental and reference values according to the Student's t test at 95% of confidence.

Alterations in morphometric and organosomatic indices and histopathological analyses indicative of environmental contamination in Mullet, Mugil liza, from Southeastern Brazil.

Mullet (Mugil liza) were sampled in five different areas along the Guanabara Bay, southeastern Brazil, classified as non-contaminated, moderately contaminated and contaminated. Morphometric (Fulton condition factor, relative condition factor and weight to length scaling coefficient) and organosomatic (hepatosomatic index) indices of environmental stress were analysed. Fish from the differentially contaminated areas show statistically different Fulton and relative condition factors and hepatosomatic indices, but not the weight to length scaling coefficient. The Kn and the FCF followed the same trend, with fish from São Gonçalo (1.07 ± 0.04 and 0.89 ± 0.03), Itaipu (0.84 ± 0.01 and 0.86 ± 0.01) and the Rodrigo de Freitas Lagoon (1.03 ± 0.01 and 0.87 ± 0.20) showing higher FCFs than fish from Magé (0.96 ± 0.01 and 0.81 ± 0.01). Fish from Itaipu showed significantly higher HSI values than the other sampling sites (1.68 ± 0.07), with fish from Olaria and Ipiranga showing the lowest (1.56 ± 0.12 and 1.60 ± 0.07, respectively).

First-time report of metalloproteinases in fish bile and their potential as bioindicators regarding environmental contamination.

Gallbladder bile from 2 fish species, mullet (Mugil liza) and tilapias (Tilapia rendalli), contain substantial matrix metalloproteinases (MMPs). Extensive purification studies were conducted in order to obtain workable samples for SDS-PAGE and zymography analysis. Proteinase activities were assayed by gelatin substrate zymography. Several protein bands were observed, corresponding to molecular weights of 200, 136, 43, 36, 34, 29, 23 and 14 kDa in mullet bile and 179, 97, 79, 61, 54, 45, 36, 33 and 21 kDa in tilapia bile. Specific inhibitor studies were conducted, in which MMPS were inhibited by EDTA and 1,10 phenanthroline, but not by serine and cysteine protease inhibitors, such as phenylmethylsulfonyl fluoride (PMSF) and transepoxysuccinyl-l-leucylamido-l-guanidino butane (E-64), confirming the proteinase identities as MMPs. Differences in proteinase expression were observed in fish from a contaminated and reference site. Some studies regarding MMPs in different fish tissues exist, however this is the first study conducted in fish bile, and their involvement in detoxification processes and organism protection against the effects of aquatic contaminants may be a possibility.

Mercury and methyl mercury ratios in caimans (Caiman crocodilus yacare) from the Pantanal area, Brazil.

The Pantanal region is the largest floodplain area in the world and of great biological importance due to its unique flora and fauna. This area is continuously undergoing increasing anthropogenic threats, and has also experienced mercury contamination associated with gold mining and other anthropogenic activities. Pantanal caimans are top-level predators, and, as such, show great potential to accumulate mercury (Hg) by biomagnification. In this study 79 specimens from four locations in the Pantanal were analyzed for total Hg and methyl mercury (MeHg) by cold vapor atomic absorption spectrometry. Total Hg contents ranged from 0.02 to 0.36 µg g(-1) (ww), and most specimens presented MeHg ratios above 70%. One of the sites, impacted by anthropogenic activities, presented significantly higher total Hg in comparison to three less impacted sites, supporting the hypothesis that caimans can, in fact, be considered effective bioindicators of ecosystem health.

Total-Hg and organic-Hg in Cephalopholis fulva (Linnaeus, 1758) from inshore and offshore waters of NE Brazil.

To investigate whether source proximity or bioavailability is the major factor controlling both Hg concentration and Hg speciation in marine fishes, total- and organic-Hg content in muscle and liver tissues from different populations of Cephalopholis fulva (piraúna) from inshore and offshore waters of the Brazilian northeastern coast were analyzed. Average total-Hg in muscle (104 ng x g(-1) w.w.) and liver (2,865 ng x g(-1) w.w.) tissues, as well as organic-Hg concentrations in muscle (169 ng x g(-1) w.w.) and liver (1,038 ng x g(-1) w.w.), were much higher in the offshore population of C. fulva than in the inshore ones. In the inshore population total-Hg and organic-Hg average concentrations in muscle tissue were similar and reached only 49 ng x g(-1) w.w., while in liver they reached 412 ng x g(-1) w.w. for total-Hg and 180 ng x g(-1) w.w., for organic-Hg. Concentrations of both Hg species in the two populations were higher in liver than in muscle. The average percentage contribution of organic-Hg to the total Hg content was higher in muscle (98-100%) than in liver (42-53%), but similar between the two populations. Total-Hg and organic-Hg concentrations in muscle and liver from the offshore population showed significant (P < 0.05) positive correlation with fish length. However, in the inshore population only the total-Hg and organic-Hg in muscle tissues correlate significantly with fish size. Although the coastal environments are enriched in total Hg relative to open waters, the significant higher Hg concentrations in the offshore population of C. fulva and the significant correlation found between organic-Hg in liver with fish size suggest higher bioavailability of Hg in offshore waters relative to inshore ones.

Total-Hg and organic-Hg in Cephalopholis fulva (Linnaeus, 1758) from inshore and offshore waters of NE Brazil

To investigate whether source proximity or bioavailability is the major factor controlling both Hg concentration and Hg speciation in marine fishes, total- and organic-Hg content in muscle and liver tissues from different populations of Cephalopholis fulva (piraúna) from inshore and offshore waters of the Brazilian northeastern coast were analyzed. Average total-Hg in muscle (104 ng.g-1 w.w.) and liver (2,865 ng.g-1 w.w.) tissues, as well as organic-Hg concentrations in muscle (169 ng.g-1 w.w.) and liver (1,038 ng.g-1 w.w.), were much higher in the offshore population of C. fulva than in the inshore ones. In the inshore population total-Hg and organic-Hg average concentrations in muscle tissue were similar and reached only 49 ng.g-1 w.w., while in liver they reached 412 ng.g-1 w.w. for total-Hg and 180 ng.g-1 w.w., for organic-Hg. Concentrations of both Hg species in the two populations were higher in liver than in muscle. The average percentage contribution of organic-Hg to the total Hg content was higher in muscle (98-100 percent) than in liver (42-53 percent), but similar between the two populations. Total-Hg and organic-Hg concentrations in muscle and liver from the offshore population showed significant (P < 0.05) positive correlation with fish length. However, in the inshore population only the total-Hg and organic-Hg in muscle tissues correlate significantly with fish size. Although the coastal environments are enriched in total Hg relative to open waters, the significant higher Hg concentrations in the offshore population of C. fulva and the significant correlation found between organic-Hg in liver with fish size suggest higher bioavailability of Hg in offshore waters relative to inshore ones.

As concentrações de Hg-total e Hg-orgânico foram determinadas em diferentes populações de Cephalopholis fulva (piraúna) capturadas em águas costeiras e em bancos oceânicos do litoral nordeste do Brasil. A comparação entre as duas populações permitiu investigar o efeito da proximidade de fontes sobre as concentrações, e a especiação de Hg em músculo e fígado desta espécie. As concentrações médias de Hg-total em músculo (104 ng.g-1 w.w.) e fígado (2,865 ng.g-1 w.w.), assim como as concentrações de Hg-orgânico em músculo (169 ng.g-1 w.w.) e fígado (1,038 ng.g¹ w.w.) foram muito maiores na população capturada nos bancos oceânicos do que na população costeira. Nesta, as concentrações médias de Hg-total e Hg-orgânico na musculatura de C. fulva foram similares e baixas (49 ng.g-1 w.w.), enquanto que atingiram 412 ng.g-1 w.w. de Hg-total e 180 ng.g-1 w.w. de Hg-orgânico no fígado destes animais. As concentrações das duas espécies de Hg foram significativamente maiores no fígado do que na musculatura. A contribuição percentual média de Hg-orgânico para a concentração total de Hg nos peixes foi maior para músculo (98-100 por cento) que para fígado (42-53 por cento), mas semelhante entre as duas populações. As concentrações de Hg-total e Hg-orgânico na musculatura e no fígado de C. fulva mostraram-se positivamente correlacionadas com o tamanho do animal (P < 0,05). Entretanto, na população costeira somente as concentrações destas espécies de Hg na musculatura apresentaram correlações significativas com o tamanho do animal. Embora o ambiente costeiro seja relativamente enriquecido em Hg, em relação aos bancos oceânicos, as maiores concentrações de Hg foram verificadas na população oceânica de C. fulva. A correlação significativa entre Hg-orgânico no fígado e tamanho do animal no fígado sugerem uma maior biodisponibilidade do Hg em águas oceânicas quando comparada às águas costeiras.

Mercury contamination in farmed fish setup on former garimpo mining areas in the Northern Mato Grosso State, Amazonian region, Brazil.

The garimpo gold mining activity has released about 2.500 tons of mercury in the Brazilian Amazonian environment in the 1980-1995 period. The northern region of Mato Grosso State, an important gold mining and trading area during the Amazonian gold rush is now at a turning point regarding its economic future. Nowadays, the activities related to gold mining have only a low relevance on its economy. Thus, the local communities are looking for economic alternatives for the development of the region. Cooperative fish farming is one of such alternatives. However, some projects are directly implemented on areas degraded by the former garimpo activity and the mercury left behind still poses risks, especially by its potential accumulation in fish. The objective of the present study was to evaluate the levels of mercury contamination in two fish farming areas, Paranaíta and Alta Floresta, with and without records of past gold-washing activity, respectively. Data such as mercury concentration in fish of different trophic level, size, and weight as well as the water physical and chemical parameters were measured and considered. These preliminary data have shown no significant difference between these two fish farming areas, relatively to mercury levels in fish.

On-line mercury and methylmercury pre-concentration by adsorption of their dithiophosphoric acid diacyl ester chelates on a C18 column and cold-vapor atomic-absorption detection.

The pre-concentration of mercury(II) and methylmercury by adsorption of their dithiophosphoric acid diacyl ester (DDTP) chelates on a C18 column, then detection with cold-vapor atomic-absorption spectrometry was investigated. Conditions such as sample pH, reductant and chelating agent flow and concentration, and eluent and carrier gas flow were optimized. Optimization was performed by use of evolutionary operation with a proper factorial design. At a sample flow of 5.3 mL min(-1) and a loading time of 4.5 min, column adsorption efficiency ranged from 88 to 93% for both species. Detection limits down to 10 ng L(-1) were obtained at a sample throughput of 12 h(-1). There was good agreement between found and certified values in the analysis of certified reference materials after their microwave-assisted mineralization with HNO3 and H2O2.

Fourier transform infrared spectrometric determination of Ziram.

A procedure has been developed for vapour-phase Fourier transform infrared determination of Ziram, a dithiocarbamate pesticide. The method is based on the evolution of CS(2), after decomposition of the dithiocarbamate with diluted H(2)SO(4) at 50 degrees C. The CS(2) evolved was swept by a carrier flow of nitrogen to a laboratory-made infrared gas cell of 39 mm pathlength and 490 mul volume. The signals were registered as a function of time. The area of peaks obtained from absorbance measurement in the wavenumber range between 1600 and 1450 cm(-1) were interpolated in a calibration line established from Ziram standards treated in the same way as samples. The method provided an absolute limit of detection of 0.055 mg, a variation coefficient of the order of 6% for an analyte mass of 50 mg, and an analysis time of 3.5 min.

Exposure to mercury in pregnant women from Alta Floresta-Amazon basin, Brazil.

The region of Alta Floresta in the South of the Amazon basin, close to the Teles Pires River, was one of the main prospecting gold areas in the Amazon Basin until the beginning of the 1990s. The economic growth was accompanied by a considerable increase in the population, due to a massive influx of migrants from the southern region of the country. Women had an important role during that process. They worked in the "garimpos" in different activities, such as cooking or managing, where they were exposed mainly to indoor elemental mercury during burning of amalgam. They also worked in gold dealers' shops, where a great amount of amalgam was burned daily. Fish consumption was an important dietary protein source and also a possible exposure pathway, due to the high Hg concentrations reported in carnivorous species. The present study evaluates the mercury uptake and consequent risks involved for pregnant women from Alta Floresta at the end of the gold rush period. The survey included women at different pregnancy stages and it was supported by clinical exams, followed by an interview with a specific questionnaire. Mercury hair concentrations ranging from 0.05 to 8.2 microg/g were found among women, with 13% of them showing concentrations above 2 microg/g. According to the results, mean differences in hair mercury concentration were statistically higher for pregnant women if they had worked in gold mining areas, they had consumed alcohol, their husbands had worked as "garimpeiros," they ate fish, and/or they had malaria before and/or during pregnancy

Flow injection-FTIR determination of dithiocarbamate pesticides.

A flow injection Fourier transform infrared spectrometric procedure has been developed for the determination the dithiocarbamate pesticides Ziram and Thiram in solid samples. All the operations involved, such as extraction, filtration and measurement, were integrated in the experimental set-up in order to avoid excessive manipulation of samples and standards. Ultrasonic assisted and mechanical extraction were evaluated for the solubilization of the analytes and, additionally, the effect of carrier flow rate, sample loop volume and the ratio between sample mass and volume of solvent employed were studied. Quantitative extractions with chloroform were obtained for both Ziram and Thiram, after 5 and 2 min, respectively, of mechanical shaking of sample slurries. Absorbance measurement, in the wavenumber range of 1600-1460 cm-1 for Ziram and 1400-1315 cm-1 for Thiram, was carried out, and the area values of the peaks obtained, as a function of time, were interpolated in external calibration lines prepared from standard solutions of Ziram and Thiram in chloroform. Analyses of commercial formulations and spiked soil samples incubated two weeks were in a good agreement with values found by other methodologies. Absolute detection limits of 400 micrograms for Ziram and 785 micrograms for Thiram and variation coefficients of 6.4% and 2.5% were obtained by use of the aforementioned methodology.

Fourier transform infrared determination of CO2 evolved from carbonate in carbonated apatites.

A quantitative method based on FTIR has been developed to determine carbonate in synthetic apatites. The method measures the evolved CO2 after reaction of 50 mg apatite with 2 mL of hydrochloric acid (0.5 M) in a reaction vessel, heated to 40 degrees C. The CO2 evolved was swept by a carrier of nitrogen to a laboratory-made infrared gas cell of 39 mm pathlength and 490 microL volume. The signals were recorded as a function of time and the areas of the chemigram peaks obtained from the measurements in the wavenumber range of 2,500-2,150 cm(-1), were interpolated using a calibration curve. The method can be used to study apatites with carbonate contents below 0.2% with a sampling frequency of 8 h(-1).

Determination of cadmium in whole blood and urine electrothermal atomic absorption spectrometry using palladium-based modifiers and in situ decontamination.

The determination of Cd in whole blood and urine by electrothermal atomic absorption spectrometry using Pd-based modifiers and in situ decontamination was studied. The performance of Pd, Pd + NH4NO3, and Pd + Mg(NO3)2 as modifiers were compared with respect to maximum pyrolysis and atomization temperatures, background attenuation, linear calibration range and matrix interference. Samples were diluted (1 + 4) with 0.1% Triton X-100 (blood) or 0.2% HNO3 (urine), and introduced onto the graphite tube platform already containing the in situ dried and decontaminated modifier. The influence of modifier volume and concentration, as well as that of the dilution ratio was investigated. The proposed procedure involves the use of Pd(NO3)2 + (Mg(NO3)2 as modifier, a pyrolysis temperature of 700 degree C and an in situ decontamination step of 1200 degrees C. The atomization temperature was 1700 degrees C. The detection limit (n = 10, k = 3) was 0.22 micrograms l-1 for both samples. Typical relative standard deviation values (n = 3) were below 3%. Using matrix-matched standards, good agreement was observed between experimental and reference values in the analysis of certified reference materials. Interlaboratory comparison data have confirmed the validity of the proposed analytical procedure.

Determination of Sb, Ni and V in slurry from airborne particulate material collected on filter by graphite furnace atomic absorption spectrometry.

A microdigestion procedure performed directly in the autosampler cup is proposed. Small quartz filter pieces loaded with the particulate material are transferred to the cup. The sample is digested by a mixture of nitric, sulfuric and hydrofluoric acid (1:1:1) under sonication. After the addition of a boric acid solution the elements are determined by graphite furnace atomic absorption spectrometry using the autosampler to deliver the slurry into the furnace. A mixture of palladium and magnesium nitrates was used as a modifier for Sb. Spiking studies showed recoveries close to 100% using aqueous analytical curves for Ni and Sb. For V, an analytical curve in the blank slurry, obtained by submitting an unloaded filter to the same procedure, was used. The method was applied to two standard reference materials, Coal Fly Ash (NIST 1633a) and Urban Particulate (NIST 1648) and the concentrations showed good agreement with the certified or recommended values using aqueous analytical solutions.