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Perinatal polychlorinated biphenyl (PCB) exposures still remain a serious health concern because offspring receive PCB burden from mother during vulnerable processes of development. Since cytochrome P450 (CYP) represents a toxicological endpoint, in the present study, representing an extended investigation of a previous multitasked one, we explored the long-term responsiveness of CYP1A and CYP2B isoforms by Western blot analysis in liver and whole brain of lactating (PN12), weaning (PN21), and adult offspring (PN60) rats prenatally and lactationally exposed to a reconstituted PCB mixture (RM) of noncoplanar PCB138, 153, 180, and coplanar PCB126 congeners. We chose highly chlorinated PCBs instead of lower chlorinated one, because their recalcitrance to biotransformation makes easy their accumulation/persistence in tissues and breast milk. Dioxin-like congener PCB126 binding aryl hydrocarbon receptor (AHR) is responsible of many toxic effects. Pregnant Sprague-Dawley dams with high affinity AHR received subcutaneous injection of RM (10 mg/kg body weight) daily during gestation (days 15-19) and twice a week during breast-feeding. The results evidenced a transfer of PCBs to neonates through milk and a significant responsiveness of hepatic CYP in both mothers and offspring. In liver of exposed progeny, CYP isoforms exhibited a significant increment at PN12 (70% over control) and at PN21 (270% over control). Contrary to dams, in adult PCB offspring CYP levels showed a decline up to values similar to those of control. This transient developmental responsiveness of CYP isoforms in offspring liver reflects roughly the time course of hepatic PCB levels previously reported. Even if congeners were detected in brain, we failed in evidencing a responsiveness of CYP isoforms probably because of region-specific CYP expression in this organ. In conclusion, induction of offspring hepatic CYP is index of liver PCB burden, and despite the insensitivity of whole brain CYP we cannot exclude brain vulnerability toward PCB. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 856-866, 2014.

Napoleon Bonaparte (1769 to 1821) is one of the most studied historical figures in European history. Not surprisingly, amongst the many mysteries still surrounding his person is the cause of his death, and particularly the suspicion that he was poisoned, continue to intrigue medical historians. After the defeat of the Napoleonic Army at the battle of Waterloo in 1815, Napoleon was exiled to the small island of Saint Helena in the South Atlantic, where he died 6 years later. Although his personal physician, Dr François Carlo Antommarchi, stated in his autopsy report that stomach cancer was the cause of death, this diagnosis was challenged in 1961 by the finding of an elevated arsenic concentration in one of Napoleon's hair samples. At that time it was suggested that Napoleon had been poisoned by one of his companions in exile who was allegedly supported by the British Government. Since then Napoleon's cause of death continues to be a topic of debate. The aim of this review is to use a multidisciplinary approach to provide a systematic and critical assessment of Napoleon's cause of death. Copyright © 2011 by Lippincott Williams & Wilkins.

The influence of a high quality compost amendment on two soils contaminated with diesel oil and polycyclic aromatic hydrocarbons, PAHs, respectively, was evaluated, with respect to contaminant biodegradation, microbial community composition and soil toxicity and genotoxicity. For each of the two soils, two 20-kg biopiles were set up, one without amendments and one compost-amended. GC/FID analyses revealed that compost was effective in enhancing biodegradation of diesel oil and of four-ring PAHs. It also influenced microbial community composition, as inferred by ARDRA analyses and partial sequencing of 16S rDNA of clones from libraries constructed from each soil sample. Microtox® analyses on soil aqueous saline extracts and Solid Phase Tests showed some toxicity reduction due to compost addition, while the Comet assay, performed on coelomocytes of earthworms exposed to contaminated soils, did not show genotoxicity reduction. In general, the use of compost amendment to hydrocarbon-contaminated soils in a bioremediation process proved to be effective for depletion of contaminants and reduction of toxicity.

The gender-specific expression pattern of aromatase and 5alpha-reductases (5alpha-R) during brain development provides neurons the right amount of estradiol and DHT to induce a dimorphic organization of the structure. Polychlorinated biphenyls (PCBs) are endocrine disruptive pollutants; exposure to PCBs through placental transfer and breast-feeding may adversely affect the organizational action of sex steroid, resulting in long-term alteration of reproductive neuroendocrinology. The study was aimed at: a) evaluating the hypothalamic expression of aromatase, 5alpha-R1 and 5alpha-R2 in fetuses (GD20), infant (PN12), weaning (PN21) and young adult (PN60) male and female rats exposed to PCBs during development; b) correlating these parameters with the time of testicular descent, puberty onset, estrous cyclicity and copulatory behavior; c) evaluating possible alterations of some non reproductive behaviors (locomotion, learning and memory, depression/anxiety behavior). A reconstituted mixture of four indicator congeners (PCB 126, 138, 153 and 180) was injected subcutaneously to dams at the dose of 10 mg/kg daily from GD15 to GD19 and then twice a week till weanling. The results indicated that developmental PCB exposure produced important changes in the dimorphic hypothalamic expression of both aromatase and the 5alpha-Rs, which were still evident in adult animals. We observed that female puberty onset occurs earlier than in control animals without cycle irregularity, while testicular descent in males was delayed. A slight but significant impairment of sexual behavior and an important alteration in memory retention were also noted specifically in males. We conclude that PCBs might affect the dimorphic neuroendocrine control of reproductive system and of other neurobiological processes.

The effect of arbuscular mycorrhiza on heavy metal uptake and translocation was investigated in Cannabis sativa. Hemp was grown in the presence and absence of 100 microg g-1 Cd and Ni and 300 microg g-1 Cr(VI), and inoculated or not with the arbuscular mycorrhizal fungus Glomus mosseae. In our experimental condition, hemp growth was reduced in inoculated plants and the reduction was related to the degree of mycorrhization. The percentage of mycorrhizal colonisation was 42% and 9% in plants grown in non-contaminated and contaminated soil, suggesting a significant negative effect of high metal concentrations on plant infection by G. mosseae. Soil pH, metal bioavailability and plant metal uptake were not influenced by mycorrhization. The organ metal concentrations were not statistically different between inoculated and non-inoculated plants, apart from Ni which concentration was significantly higher in stem and leaf of inoculated plants grown in contaminated soil. The distribution of absorbed metals inside plant was related to the soil heavy metal concentrations: in plant grown in non-contaminated soil the greater part of absorbed Cr and Ni was found in shoots and no significant difference was determined between inoculated and non-inoculated plants. On the contrary, plants grown in artificially contaminated soil accumulated most metal in root organ. In this soil, mycorrhization significantly enhanced the translocation of all the three metals from root to shoot. The possibility to increase metal accumulation in shoot is very interesting for phytoextraction purpose, since most high producing biomass plants, such as non-mycorrhized hemp, retain most heavy metals in roots, limiting their application.

The present study was to assess the effect of heavy metal stress on the DNA methylation of a metal-sensitive plant, Trifolium repens L. and of a metal-tolerant plant, Cannabis sativa L. The changes in the level of 5-methylcytosine (5mC) in the root DNA of plants grown on soils contaminated with different concentrations of Ni 2+ , Cd 2+ and Cr 6+ compared with that of untreated plants, were determined by immunolabelling with a monoclonal antibody, using the Slot-Blot technique. Results showed that DNA of hemp control plants was about three times more methylated than clover DNA, for the same amount of root DNA. Heavy metal treatments induced a global dose-dependent decrease of 5mC content, both in hemp and clover, ranging from 20 to 40%. Changes in methylation pattern of 5'-CCGG-3' containing sequences were investigated by methylation-sensitive amplification polymorphism (MSAP) technique. Control plants of the same species showed a very similar pattern, suggesting that, in normal condition, methylation involves precise sites. Heavy metals induced DNA methylation changes mainly related to hypomethylation events. These variations were not randomly directed but involved specific DNA sequences, since the detected polymorphisms were the same in all the plants analysed for each treatment.

Experiments in semi-natural conditions were undertaken to assess hemp metal tolerance and its ability to accumulate cadmium, nickel and chromium. Cannabis sativa was grown in two soils, S1 and S2, containing 27, 74, 126 and 82, 115, 139 μg g−1 of Cd, Ni and Cr, respectively. After two months from germination and at ripeness, no significant alteration in plant growth or morphology was detected. On the contrary, a high hemp reactivity to heavy metal stress with an increase in phytochelatin and DNA content was observed during development, suggesting the Cannabis sativa ability to avoid cell damage by activating different molecular mechanisms. Metals were preferentially accumulated in the roots and only partially translocated to the above-ground tissues. The mean shoot Cd content was 14 and 66 μg g−1 for S1 and S2 soil, respectively. Although not negligible concentrations they were about 100 times lower than those calculated for the hyperaccumulator Thlaspi caerulescens. Similarly Ni uptake was limited if compared with that of the Ni-hyperaccumulator Alyssum murale. Chromium uptake was negligible. As expected on the base of the metal concentration detected in ripe plants, no statistically significant variation in soil metal content was detected after one crop of hemp. Nevertheless, a consistent amount (g) of Cd and Ni is expected to be extracted by 1 ha biomass of hemp (about 10 t) per year and along the time a slow restoration of deeper soil portions can be obtained by its wide root system (at least 0,5 m deep). In addition, the possibilities of growing hemp easily in different climates and using its biomass in non-food industries can make heavy metal contaminated soils productive. This means economical advantage along with a better quality of soil.

Perinatal polychlorinated biphenyl (PCB) exposures still remain a serious health concern because offspring receive PCB burden from mother during vulnerable processes of development. Since cytochrome P450 (CYP) represents a toxicological endpoint, in the present study, representing an extended investigation of a previous multitasked one, we explored the long-term responsiveness of CYP1A and CYP2B isoforms by Western blot analysis in liver and whole brain of lactating (PN12), weaning (PN21), and adult offspring (PN60) rats prenatally and lactationally exposed to a reconstituted PCB mixture (RM) of noncoplanar PCB138, 153, 180, and coplanar PCB126 congeners. We chose highly chlorinated PCBs instead of lower chlorinated one, because their recalcitrance to biotransformation makes easy their accumulation/persistence in tissues and breast milk. Dioxin-like congener PCB126 binding aryl hydrocarbon receptor (AHR) is responsible of many toxic effects. Pregnant Sprague-Dawley dams with high affinity AHR received subcutaneous injection of RM (10 mg/kg body weight) daily during gestation (days 15-19) and twice a week during breast-feeding. The results evidenced a transfer of PCBs to neonates through milk and a significant responsiveness of hepatic CYP in both mothers and offspring. In liver of exposed progeny, CYP isoforms exhibited a significant increment at PN12 (70% over control) and at PN21 (270% over control). Contrary to dams, in adult PCB offspring CYP levels showed a decline up to values similar to those of control. This transient developmental responsiveness of CYP isoforms in offspring liver reflects roughly the time course of hepatic PCB levels previously reported. Even if congeners were detected in brain, we failed in evidencing a responsiveness of CYP isoforms probably because of region-specific CYP expression in this organ. In conclusion, induction of offspring hepatic CYP is index of liver PCB burden, and despite the insensitivity of whole brain CYP we cannot exclude brain vulnerability toward PCB. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 856-866, 2014.

The present findings provide experimental evidence for the hypothesis that an impairment of mitochondrial function may be involved in manganese neurotoxicity. Specifically, the treatment of dopaminergic neuronal-derived cell line (PC12) with MnCl2 produced a significant inhibition of some mitochondrial complexes of the respiratory chain, while in the glial-derived cell line (C6) this effect was not observed. In PC12 the decrease in complex I activity was more pronounced than in other mitochondrial complexes. However treatment of cells with ZnSO4 exerted no significant variations in enzymatic activities. A direct exposure of mitochondrial fraction to MnCl2 reduced enzymatic activities of mitochondria in both cell lines adding further support to the proposed theory that the different sensitivity of the cells to manganese may be explained by a difference in uptake or intracellular storage. These data indicate that manganese neurotoxicity could be the result of a direct effect just on complex I activity or due to a secondary effect of oxidative stress induced by an excess of this transition metal.

Rat primary hepatocyte cultures have been used to study the effect of Benomyl alone or in combination with Pirimiphos-methyl. The results presented demonstrate that Benomyl alone is responsible for the microtubular disorganization in both a time- and dose-dependent manner, that the effect is reversible after the agent is removed, and that Benomyl is a potent glutathione-depleting agent. Pirimiphos-methyl, alone or combined with Benomyl had no effect on microtubule organization, but reinforced the decrease in glutathione.

This study explored the comet assay as a possible tool to assess genotoxicity in erythrocytes of Columba livia to detect genotoxic effects induced by exposure to urban air pollution. Fieldwork was conducted between June 2004 and June 2005 in the city of Milan, Italy, by sampling pigeons in different areas almost twice a week. Six air contaminants-CO, PM10, NO(2), O(3) (ozone), SO(2), and C6H6-plus polycyclic aromatic hydrocarbons (PAHs) in fine particles, temperature, and ultraviolet index, were considered. Genotoxicity levels, expressed as %DNA migrated, tail moment, and damage index (DI), were always higher in wild pigeons than in pigeons living indoors (controls). Animals exposed to urban air showed significant differences from season to season, and the genotoxic parameters presented the highest values in summer (45.30% +/- 1.40% %DNA migrated, 12.73 +/- 0.80 tail moment, and 22.30 +/- 0.15 x DI x 10(-1)); regression analyses showed a positive relation between DI and O(3) concentrations (P < 0.001). The use of the comet assay DI parameter as a rapid assessment of incipient genotoxic risk by pollution, as measured in C. livia living in urban areas, is also discussed.

The study has explored two conservative biomarkers, porphyrin pattern in guano and methemoglobin levels in blood of Columba livia, in order to assay their potential use for monitoring toxic effects induced by exposure to urban air pollution. Fieldwork was conducted between October 2003 and June 2005 in the city of Milan, Italy, by sampling the pigeons in different areas almost twice a week. Six air contaminants, CO, PM10, NO(2), O(3), SO(2), and C(6)H(6), plus polycyclic aromatic hydrocarbons (PAHs) in fine particles, temperature, and UV index, were considered. Protoporphyrins from pigeon excreta sampled outdoors were always higher than in indoor animals, particularly in winter. A positive correlation (p < 0.001) of protoporphyrin with PAHs was observed. In terms of the values of methemoglobinemia, animals exposed to urban air showed significant differences from season to season but only in summer and autumn did the outdoor pigeons show higher values than indoor animals. In summer, a regression model with a positive correlation between O(3) and benzene was determined (p < 0.01). The use of Columba livia as a sentinel for urban air pollution as well as for detecting complex interactions between contaminants in the urban air pollutant mixture was discussed.

The aim of this study was to improve knowledge about transplacental transfer of an environmentally relevant PCB mixture by evaluating congener levels in livers and brains of rat dams and fetuses after maternal exposure, and correlating them to the levels of CYP450 and P-glycoprotein, involved in biotransformation and xenobiotics export, respectively. Pregnant dams were injected daily from gestation day (GD) 15 to 19 with 10 mg kg-1 of a reconstituted mixture (RM) composed of PCB138, 153, 180 and 126. Our data indicate that at GD20 RM is partitioned among maternal tissues, and that fetuses are not excluded from this distribution, evidencing a placental transfer of PCBs. Considering the ratio of maternal and fetal PCB concentrations based on lipid-weight, the amounts of congeners were 7-fold lower in fetal livers than in maternal livers and 25-30-fold higher in fetal brains than in maternal ones. Moreover, in dams the congeners were able to induce hepatic CYP450 response (total CYP450, CYP1A and CYP2B), but failed to increase P-170 expression, while in fetuses the constitutive expression of CYP450 and P-170 was not induced by treatment. Pearson Product-Moment Correlation applied to treated group data suggests that PCB accumulation in fetal livers, but not in brains, depended principally on their mothers' intoxication pattern. On the whole, these results emphasize the maternal liver and the fetal brain as depot organs for PCB sequestration and their susceptibility towards PCB toxicological risk. Moreover they highlight the lack of a coordinated response between the investigated defence mechanism. © 2009 Elsevier Ltd. All rights reserved.

Mixed air pollutants are considered a major cause of DNA damage in living species. In this study Trifolium repens L. cv Regal was used as a bioindicator to assess the genotoxicity of air stressors in the Italian province of Novara. Two on-site biomonitoring experiments were performed during the spring and autumn of 2004. Test plants were exposed at 19 monitoring sites distributed homogeneously throughout the province, and each experiment lasted for a period of 6 weeks. Genotoxicity was evaluated with Amplified Fragment Length Polymorphism (AFLP) molecular markers. The results show the predominantly rural central-west region of the Novara Province to have the worst air quality with regard to genotoxicity. Analyses of geomorphology, land use and climatic factors suggest that the compromised air quality in the region could be attributed to wind strength and direction, transporting pollution from vehicular traffic on the A4 highway and from the urban/industrialized centres of Novara and Vercelli. Plant growth, changes in plant photochemical efficiency and the presence of ozone related leaf injuries were also measured to better interpret the results of genotoxicity. Statistical analyses show that although climatic factors such as light intensity and temperature influence plant growth, they do not contribute to atmospheric stressor-induced DNA damage. Further analyses indicated that, as expected, a mixture of genotoxic and non-genotoxic pollutants coexist in the Novara Province troposphere, and that the elevated ozone concentrations experienced during the study may have contributed to the DNA damage in the tested plants by enhancing genotoxicity via interaction with other air stressors.

The setting up of efficient early warning systems is a challenge to research for preventing environmental alteration and human disease. In this paper, we report the development and the field application of a new biomonitoring methodology for assessing soil genotoxicity. In the first part, the use of amplified fragment length polymorphism and flow cytometry techniques to detect DNA damage induced by soils artificially contaminated with heavy metals as potentially genotoxic compounds is explained. Results show that the combination of the two techniques leads to efficient detection of the sublethal genotoxic effect induced in the plant bioindicator by contaminated soil. By contrast, the classic mortality, root, and shoot growth vegetative endpoints prove inappropriate for assessing soil genotoxicity because, although they cause genotoxic damage, some heavy metals do not affect sentinel plant development negatively. The statistical elaboration of the data obtained led to the development of a statistical predictive model which differentiates four different levels of soil genotoxic pollution and can be used everywhere. The second part deals with the application of the biomonitoring protocol in the genotoxic assessment of two areas surrounding a steelworks in northern Italy and the effectiveness of this methodology. In this particular case, in these areas, the predictive model reveals a pollution level strictly correlated to the heavy metal concentrations revealed by traditional chemical analysis.

Levels of 12 trace elements were measured in samples of the bryophyte Hypnum cupressiforme Hedw. and in soil collected in the surroundings of an oil-fired power plant in Northern Italy. Metal bioaccumulation in moss was estimated after soil correction in order to obtain deposition patterns and individuate potentially toxic metals emitted from the plant. V and Ni, occurring together in fuel oil, showed highest bioaccumulation values near the stacks. Mean contamination of the study area for these elements is 5.5 (V) and 3.3 (Ni) times the background levels of the reference site. Other elements showed only limited alterations of bioaccumulation values, in relation to agricultural and industrial activity in the study area.

Midazolam, a water soluble benzodiazepine used as a preanaesthetic and hypnotic drug, showed a concentration-related (0.1-0.75 mM) depressant effect on both Adenosine 5'-diphosphate (ADP)-induced oxygen consumption and oxidative phosphorylation of rat liver mitochondria if the substrate was oxidized at different steps in the oxidation chain, but not when the substrate was ascorbate plus tetramethyl-p-phenylenediamine (complex IV). Furthermore, midazolam did not affect citrate synthase activity, but inhibited the 2,4 dinitrophenol (DNP)-uncoupled mitochondrial respiration. This result shows that midazolam primarily acts as a mitochondrial electron transport inhibitor. This inhibition is mainly due to the fact that midazolam decreases NADH ubiquinone reductase (complex I) and ubiquinol cytochrome c reductase (complex III) activities, but it also inhibits complex II activity. Spectrophotometric measurements of redox states of rat skeletal muscle mitochondria cytochromes show a decrease in the reduction of aa3 and c+c1 cytochromes in the presence of the benzodiazepine. Midazolam significantly decreased the reduced ubiquinone/total ubiquinone ratio (evaluated by means of HPLC and electrochemical detection) in rat liver mitochondria in both beta-hydroxybutyrate and succinate. Ubisemiquinone may be the redox component affected by midazolam, whether or not bound to the iron-sulfur proteins present in all three mitochondrial complexes. These effects of midazolam, not necessarily related to the preanaesthetic and hypnotic action are probably mediated via mitochondrial benzodiazepine receptors.

Organization of the actin cytoskeleton, the cytosolic free-calcium concentrations and ATP levels were analyzed in 3T3 mouse fibroblasts treated with 0.75 or 1.5 mM MPTP. In the presence of the drug actin filaments were time- and dose-dependently presently disorganized, ATP level was unaffected and intracellular calcium increased within 5 s. The correlation between MPTP cytotoxicity and [Ca 2+ ] i level emerging from these results, suggests that the primary effect of the molecule itself is on the plasma membrane's integrity for calcium ion regulation.

Chick embryos were treated on day 0 of incubation with two phenoxy herbicides, 2-methyl-4-chlorophenoxyacetic acid (MCPA) (0.4, 2 mg/egg) and 2,4-dichlorophenoxyacetic acid (2, 4-D) (1, 2, 4 mg/egg). Both herbicides seemed to exert toxic effects mainly on the liver of 19-day-old embryos. Specific histological analysis indicated biliary stasis. Ethoxycoumarin O-deethylase was depressed by MCPA but raised by 2, 4-D. Other hepatic monooxygenase activities were unaffected by the herbicides and no significant changes were found in cytochromes. The higher dose of MCPA increased NADPH-cytochrome P450 reductase. 2,4-D treatment increased by activity of glutathione-S-transferases in the hepatic post-microsomal fraction while MCPA increased them at the lower dose and significantly reduced them at the higher. The phenoxyacetic herbicides appear thus to have some effects on hepatic drug metabolizing enzymes of the chick embryo which cannot be easily interpreted. Biliary retention, produced in particular by MCPA, could be partly responsible for these effects.

The aim of our research was the in vitro evaluation of the neurotoxic effects of three EBDCs (Nabam, Zineb and Maneb) and ETU on SK-N-BE human neuroblastoma cells as a model for neurotoxicity in humans. The EC50 value was used as an index of the toxicities of these compounds. Since Zineb and Maneb contain zinc and manganese as cations, respectively, in order to determine the contributions of these metals, the EC50s of zinc chloride and manganese chloride were also evaluated. Nabam, Zineb and Maneb had EC50 values ranging from 1μM to 30μM; the EC50s of manganese and zinc in this human cell line were found to be of the same order of magnitude as those of the EBDC fungicides. These in vitro effects are discussed in relation to the possible use of neuronal cell lines for detecting the neurotoxicities of these compounds.

Activities of catalase, glutathione peroxidase (GSH-Px) and glutathione-S-transferase (GST) were measured in livers of the crested newt (Urodele Amphibian) after 16 days of percutaneous exposure to the herbicide Agroxone 3, a commercial formulation of 2-methyl-4-chlorophenoxyacetic acid (MCPA) containing 25% MCPA as sodium salt and 75% wetting and dispersing ingredients. Animals of both sexes were exposed to 800 p.p.m. of the active ingredient. There was a three-fold increase in catalase and a clear enhancement of GSH-Px in the MCPA-exposed animals of both sexes. A sex-dependent effect of MCPA was observed on GST activity, which increased significantly in female newts only. Similar liver enzyme changes were observed in female newts, and not in males, given the hypolipidaemic drug clofibrate (13 mg/animal/day, by gavage, for 8 days), which is structurally related to phenoxyherbicides. These findings indicate that the crested newt is sensitive to the hepatic effects of the peroxisome proliferators.

The toxicity of the herbicide Erbitox E30, a commercial formulation of 2-methyl-4-chlorophenoxyacetic acid (MCPA) containing 28% MCPA as sodium-potassium salt and 72% of unknown ingredients, was tested on chick embryos. Sterile aqueous solutions of MCPA were injected into the air chamber at doses of 0, 1.5, 3.0, 6.0, 9.0, or 10.5 mg/egg on day 0 or on day 4 of incubation. The mortality rate for the embryos treated on day 0 of incubation was high in the first 5 days, low from 5-12 days and again increased by 15 days. The 15-day LD50 was 4.4 mg/egg (95% C.I. 3.7-5.3 mg/egg). HPLC analysis of albumen and yolk showed that concentrations of MCPA in the albumen were detectable at 5 min, highest at 7 days and markedly diminished by 14 days of incubation; a significantly lower concentration of MCPA was found in the yolk throughout the incubation period, except at 14 days when the yolk concentration was 4 times higher than the albumen concentration. At 15 days of incubation, MCPA was evenly distributed in the tissues of the embryo. MCPA was more toxic to 4-day embryos; concentrations above 6.0 mg/egg were lethal to all embryos within the first week of incubation. The 15-day LD50 for treatment on day 4 of incubation was 2.8 mg/egg (95% C.I. 2.5-3.2 mg/egg). The liver was affected by treatment with MCPA, being green in treated embryos. However, histological examination revealed few changes in the liver parenchyma.

Camel synthetic beta-endorphin focused in three bands by isoelectrofocusing on 1 mm polyacrylamide thin gel. All the bands have opioid activity, measured on guinea pig ileum, and radioimmunologically react with beta-endorphin antiserum. Since beta-endorphin from rat pituitary gland, particularly from the neurointermediate lobe, also focused in several bands, we hypothesized that the camel peptide occurs in different conformations. A quick, simple technique based on histoelectrofocusing is proposed as a good approach to separating and measuring beta-endorphin from rat pituitary lobes. The method gives very high recovery.

The influence of some cytostatic agents (the antibiotic daunomycin, the polypeptide peptikemio and the nitrosourea derivative BCNU) on the antinociceptive effect of morphine is here investigated. It is demonstrated that daunomycin and peptikemio significantly enhance morphine-induced analgesia, while on the other hand, BCNU is ineffective. Daunomycin neither prevents morphine binding to blood proteins nor facilitates its access to the brain. A role of calcium ions in daunomycin potentiating effect is suggested.

Pentazocine, like amobarbital, enhances at low concentrations, but depresses at higher concentrations the QO2 of brain cortical slices from normal rats; under this condition morphine was in

The administration of prostaglandin E1 intracerebrally through a cannula implanted in a lateral ventricle, antagonizes morphine analgesia in rats.

Summary The adrenal ascorbic acid depletion, elicited in rats by conjunctival phlogosis provoked by application of croton oil, has been assumed as a test for quantitative evaluation of antiphlogistic effect of the following topically applied steroids: hydrocortisone acetate, triamcinolone acetonide, dexamethasone valerate. It is shown that the ascorbic acid depletion is reduced by such steroids and that such a reduction is linearly related to the log of their increasing concentrations. The parallelism of the calculated regression lines enables the evaluation of their antiphlogistic potency in reference to hydrocortisone acetate, which is 3.80 for triamcinolone and 4.27 for dexamethasone. The results obtained are compared with those obtained by the other experimental methods devised in order to evaluate the topical antiphlogistic effect of corticosteroids.

Tolerance and cross tolerance to the antinociceptive effect of equipotent doses of morphine (10 mug/kg) and mescaline (100 mug/kg) are shown in the rabbit after their repeated intracerebroventricular administration. The recording of the electrical activity of different brain areas indicates that a partial tolerance also develops to the EEG effects in animals undergoing chronic treatment with mescaline. The comparison of certain of the mescaline-induced effects with those of morphine suggests that some biochemical and neural patterns are common to the 2 drugs.

The isoelectric point of the camel and the human β-endorphin, of the α-endorphin and the enkephalins were determined by analytical isoelectric focusing on 1 mm thin polyacrylamide gel slab. The difficulty of staining peptides as short as β-endorphin or smaller was overcomed using a modification of Bibring and Baxandall's or Faupel and Von Arx's staining method. The camel β-endorphin gives two bands having isoelectric point of 10.3 and 10.4, the human β-endorphin focus at pH 9.9, while α-endorphin, leu and met-enkephalin at pH 5.9, 5.5 and 5.45 respectively. The staining method described coupled with the isoelectric focusing seems to be fit for discriminating β-endorphin in a crude rat pituitary extract.

Periaqueductal gray matter (PAG) has been shown to be one of the sites in the central nervous system where microinjections of morphine strongly inhibit intestinal transit. To investigate the nature of opioid receptor populations involved in this central effect, selective opioid agonists, FK 33824 for mu, DALA for delta, dynorphin for kappa and tentatively beta-endorphin for epsilon, were microinjected in all PAG areas previously identified as morphine-sensitive for intestinal inhibition. The PAG-induced inhibition of intestinal transit appears to be mediated mainly by mu receptors and possibly by epsilon receptors. kappa and delta receptors seem not to be involved.

Acute administration of Morphine (20 mg/kg/s.c.) in the rat results in a rise of liver tyrosine aminotransferase (TAT) expressed as μmoles of p-hy-droxyphenylpyruvate/100 mg/h. With chronic administration, a tolerance develops to this enzymatic effect. TAT induction is not evident in pregnant rats, given the narcotic, in which enzyme levels are already initially high. Afer delivery TAT returns to normal levels and it is possible to show both induction and tolerance developing to morphine. Enzyme activity in fetal livers is much lower than that of adult animals: after maternal administration of morphine only a modest TAT increase is seen which is not, however, statistically significant. TAT activity is fully evident in livers of offspring, with much higher mean levels in newborn rats from morphine-treated animals, as a possible consequence of morphine deprivation. In this latter group of newborn rats narcotic administration causes TAT activity to return to levels as high as those of naive animals. On the other hand, morphine administration to the prole of naive rats results in an induction of liver TAT.

Summary Endorphin and ACTH-like materials levels in rat plasma and pituitary were measured by radioimmunoassay under baseline and cold stress conditions. Cold stress significantly increased plasma β-endorphin and ACTH immuno-reactivity. A rise in these two peptides was also found in the neurointer-mediate lobe of the pituitary, while in the anterior lobe their levels were unaffected. These findings suggest that the rise of β-endorphin and ACTH content in the neurointermediate lobe occurs as a compensatory biosynthetic mechanism for the peptides released from the adenohypophysis.

Calcitonin (CT) injected into the brain ventricles (ICV) of conscious rabbits induced an analgesia not reversable by naloxone which could be repeatedly elicited (for 5 days), while tolerance to morphine developed. CT and morphine synergized in vivo when administered ICV in combination. CT did not alter electrically-induced contractions of guinea-pig myenteric plexus-longitudinal muscle and no displace of 3 H-dihydromorphine by CT was observed in brain opiate receptor preparations. We have concluded that the mechanism of centrally induced CT analgesia may be opiate-independent.

The endogenous opioid met-enkephalin intraventricularly adminstered to the rat at the dose of 100 microgram raised rectal temperature, whereas 400 microgram of the pentapeptide caused a diphasic effect, i.e., hypothermia followed by hyperthermia. Met-enkephalin was ineffective when administered i.p. The effects on temperature were substantially similar to those elicited, for both routes of administration, by morphine, which may either raise or lower rat temperature depending on the dose. More naloxone was required to antagonize thermic effects of met-enkephalin than morphine. Finally, there was a lack of effects on temperature for met-enkephalin centrally administered to morphine-tolerant animals, thus providing further evidence, in vivo, of cross tolerance between opiates and naturally occurring ligands of opiate receptors.

A l t h o u g h the h e r b i c i d e 4 c h l o r o 2 m e t h y l p h e n o x y a c e t i c a c i d (MCPA) i s used w i d e l y as an a g r i c u l t u r a l c h e m i c a l , few s t u d i e s have been made of i t s b i o l o g i c a l e f f e c t s in a n i m a l s . E p i d e m i o l o g i c a l r e s e a r c h has s u g g e s t e d i t may be c a r c i n o g e n i c ( H a r d e l l and Sands t rom 1979) and r e c e n t s t u d i e s have s u b s t a n t i a t e d the h y p o t h e s i s t h a t phenoxyacetic herbicides exert their carcinogenic action indirectly via peroxisome proliferation (Vainio et al. 1983; Hietanen et al. 1985).

Some effects of intraventricular injection of mescaline are examined in conscious rabbits. By means of electrical stimulation of the tooth pulp it is shown that an acute treatment with 70, 100, 150 mug/kg of mescaline elicits analgesia, the intensity of which is dose-dependent: with daily administration of 100 mug/kg for 5 days a complete tolerance develops to the antinociceptive effect. A tolerance also develops to the behavioral effects of mescaline after repeated administrations, with the exception of the stuporous state, a symptom which, on the contrary, is accentuated as the treatment proceeds. An EEG arousal is induced in the rabbit by acutely administered mescaline; the chronic treatment (100 mug/kg) makes the return of voltage to original levels progressively slower. Finally, the confrontation of certain of the mescaline-induced effects with those of morphine suggests some biochemical and neural patterns common to the 2 drugs.

The effect of Leu5-enkephalin on growth hormone (GH) and prolactin (PRL) release was studied in vivo in the infant rat and compared to that of morphine. In 10 day-old pups, intracerebroventricular injection of Leu5-enkephalin (50, 75 and 100 μg) resulted in a dose-related increase in plasma GH; morphine was active as GH releaser at the dose of 5 and 10 μg, but not at 2.5 μg. Pretreatment with naloxone (2 mg/kg ip) suppressed the GH-releasing effect of either Leu5-enkephalin (100 μg) or morphine (10 μg). Leu5-enkephalin (75 and 100 μg) induced a rise in plasma PRL which was neither dose-related nor antagonized by naloxone; morphine (5 and 10 μg) was active as PRL releaser and its effect was antagonized by naloxone. These results indicate that: 1) Leu5-enkephalin stimulates both GH and PRL release; 2) the release of GH by Leu5-enkephalin but likely not that of PRL involves specific opiate receptors; 3) morphine releases GH and PRL through specific opiate receptors.

Chronic exposure to manganese-laden dusts induces, in humans and lower primates, neurological disorders with clinicopathological features that resemble idiopathic Parkinson's disease. As many authors have suggested, manganese neurotoxicity could be related to the capability of this metal to increase catechol autoxidation in catecholaminergic neurons, therefore increasing the formation of toxic compounds such as peroxides, superoxides, free radicals, and semi-orthoquinones. Oxidative stresses and consequent neuronal damage could then occur if physiological scavenger mechanisms fail in their detoxifying action. We here report that manganese chloride weakly inhibits, in a dose-dependent way by a reversible competitive mechanism, human brain glutathione-S-transferases possibly suggesting that manganese intoxication could cause intraneuronal accumulation of cytotoxic compounds. We also report that both 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, a neurotoxin known to induce in man Parkinson-like syndromes, and one of its metabolites 1-methyl-4-phenylpyridinium failed to decrease glutathione-S-transferase activity.