Changing social norm compliance with noninvasive brain stimulation.

All known human societies have maintained social order by enforcing compliance with social norms. The biological mechanisms underlying norm compliance are, however, hardly understood. We show that the right lateral prefrontal cortex (rLPFC) is involved in both voluntary and sanction-induced norm compliance. Both types of compliance could be changed by varying the neural excitability of this brain region with transcranial direct current stimulation, but they were affected in opposite ways, suggesting that the stimulated region plays a fundamentally different role in voluntary and sanction-based compliance. Brain stimulation had a particularly strong effect on compliance in the context of socially constituted sanctions, whereas it left beliefs about what the norm prescribes and about subjectively expected sanctions unaffected. Our findings suggest that rLPFC activity is a key biological prerequisite for an evolutionarily and socially important aspect of human behavior.

In vitro tests to evaluate potential biological activity in natural substances.

The biological activity of polysaccharides from the mycelia of 40 Basidiomycetes was studied using an uncommon toxicity test technique, the planaria bioassay, and the better known potato disk bioassay. The results showed the utility of this duo of 'in vitro' tests as a preliminary screening of the toxicity of substances that are present in aqueous fungal extracts.

Effects of the ionophore antibiotic monensin on hepatic biotransformations and target organ morphology in rats.

As a preliminary in vivo approach in order to study the mechanism of toxicity of the veterinary anticoccidial monensin, male Wistar rats were orally administered 0, 2 and 12 mg kg-1 body wt. day-1 of monensin for 7 days. At the end of the experiment, effects of the ionophore on serum creatine kinase, lactic dehydrogenase and selected drug metabolising enzyme activities were investigated. Furthermore, liver, heart and quadriceps femoris muscle samples were submitted to morphological investigations. Clinical signs or increasing levels of enzymic markers of muscle injury attributable to monensin toxicosis have never been observed in treated animals. As a matter of fact all drug metabolising enzymes activities checked have not shown significant changes, except for a significant decrease of ethoxyresorufin O-deethylase (up to 31%) and aminopyrine N-demethylase (17%) activities. Morphologically, mitochondrial cristae fragmentation and initial formation of 'myelinic sheaths-like' structures have been noticed in heart and muscle fibres. As far as rat study is concerned, these results confirm heart and muscle as target organs of monensin toxicity. In addition, these findings suggest that the inhibition of hepatic biotransformation processes following the i.p. administration of the ionophore, as reported previously by other authors, might reflect unspecific cellular toxic effects rather than a specific enzyme damage.

Involvement of capsaicin-sensitive nerves in paraquat-induced mortality.

Paraquat (PQ), a broad spectrum herbicide, produces severe lung inflammation and necrosis resulting in pulmonary fibrosis and respiratory failure. Tachykinins are peptides released by sensory C fibers and have the ability of influencing respiratory functions and cellular proliferation. To examine whether the damage caused by PQ involves tachykinins, rats were depleted in their content of tachykinins by systemic treatment with capsaicin prior to PQ exposure. The animal subjected to this treatment showed a 3-fold higher viability compared to those treated with PQ alone (75 vs 27%). Depletion of reduced glutathione (GSH) is associated with oxidative stress produced by reactive oxygen intermediates during PQ metabolism. This is considered to be critical in the pathogenesis of lung damage by PQ. PQ treatment induced a significant depletion of GSH during the first days and a similar effect was also observed in the group of capsaicin-pretreated rats. Four weeks after PQ treatment the levels of GSH were similar to controls in rat pretreated or not with capsaicin plus PQ. This may indicate that the reduced levels of GSH may be associated to the toxicity observed in the acute phase, but not of importance in the final PQ-induced mortality. Neutral endopeptidase (NEP) is an enzyme considered to be critical in controlling the levels of tachykinins. Exposure of crude membrane preparations of rat lung to PQ resulted in a dose-dependent inhibition of NEP activity. Since NEP inactivation may occur in lung following a PQ exposure in vivo, the results indicate that during PQ intoxication a more sustained activity of tachykinins may be present, producing effects such as cell proliferation, fluid extravasation and bronchoconstriction. In conclusion, this finding supports the hypothesis that neuropeptides released from capsaicin-sensitive nerves could be involved in the modulation of PQ-induced lung damage.

Zinc ethylene-bis-dithiocarbamate (Zineb)-mediated inhibition of monooxygenases and lipid peroxidation in bovine liver microsomes.

Hepatic bovine microsomes were incubated with Zineb concentrations ranging from 2.5 mM to 2.5 microM. Only the higher concentrations of the fungicide (2.5 and 0.25 mM) elicited a sharp decline in cytochrome P450, cytochrome b5 and total sulphydryl groups content as well as in the activities of NADPH cytochrome c reductase, aminopyrine N-demethylase and aniline 4-hydroxylase. The loss of cytochrome P450 was matched by a concomitant increase in the amount of cytochrome P420, which represents a catalytically inactive form of cytochrome P450. The same concentrations of the fungicide, either alone or in the presence of NADPH 1 mM, failed to increase the amount of thiobarbituric reactive substances with respect to control incubations, thereby excluding the possibility of lipid peroxidation as a contributing factor in the loss of cytochrome P450 and in the inhibition of cytochrome P450-mediated metabolism. It is concluded that Zineb can depress monooxygenase activity in bovine hepatic microsomes mainly through the denaturation of cytochrome P450 and the impaired transfer of reducing equivalents to the complex cytochrome P450-substrate. These mechanisms might also account for the inhibition in lipid peroxidation brought about by the fungicide.

[Eco-toxicological study conducted with a battery of biological and phytological tests on sediments carried out on a series of 24 tributaries of the Po in 1994 and 1995]. / Ricerche ecotossicologiche eseguite con batterie di tests biologici e fitologici sui sedimenti prelevati in sequenza da 24 fiumi affluenti del Po, negli anni 1994 e 1995.

In spring and autumn 1994 and 1995 affluent water and bed sediment were sampled from 24 tributaries of the Po river, always at the same site and at the nearest place to the confluence. In the laboratory the pore water was separated from the particle fraction of the sediment. The organic compounds bound to the latter component were extracted with solvents and brought to water solution by means of dimethyl sulfoxide. The observed animal species, along with the seeds of Lepidium, were exposed to effluent water, to pore water and to water solutions of the organic compounds extracted from bed sediment. Toxicity was evaluated on the basis of 1) direct lethality, 2) the delay of embryo development, 3) the impairment of regeneration, in the animal species, while the germination index was used for the Lepidium susceptibility. The results of these investigations demonstrate that 1) the challenged species cover a broad range of sensitivities toward environmental toxins, 2) toxicity found in river samples appears almost exclusively bound to the sediment, 3) the noxious effects found in the tributaries of the Po river increase moving downstream, and 4) likewise the sediment-bound toxicity varies among the different samplings, both as a consequence of changes in rain-dependent river flow, and because of the man-made interventions on the river sides and bed.

[Toxicity of atrazine and its metabolite deethylatrazine in Thamnocephalus platyurus and Dugesia gonocephala]. / Tossicità dell'atrazina e del suo metabolita desetilatrazina per il Thamnocephalus platyurus e per la Dugesia gonocephala.

The toxicity of Atrazine and that of its Desethylatrazine metabolite has been defined employing two organisms already tested in our labs the Dugesia gonocephala, belonging to a scissiparous strain coming from the island of Tavolara (Sardinia) and the Thamnocephalus platyurus, crustacean anostracan produced under form of quiescent cysts from Creasel Ltd. (Deinze, Belgium). It has been defined the lethal concentrations at 50% of Atrazine and of Desethylatrazine, of which it has been studied also the report dose-effect in the comparisons of the rectilinear motility. The results highlight that in the comparisons of the T. platyurus Atrazine expounds a toxicity of around three times that of its metabolite; this is not seen with the Planarians that are equally sensitive to both the compounds. The use of these two experimental models for the evaluation of the contamination of bodies of water is shown to be particularly useful given their great sensitivity to pollutants.

Gender differences in ethanol oxidation and cytochrome P4502E1 content and functions in hepatic microsomes from alcohol-preferring and non-preferring rats.

1. We have studied the hepatic microsomal metabolism of ethanol (MEOS), CYP2E1 expression and catalytic activity, and the response to phenobarbital (PB) induction or CCl4 challenge in rats of either sex genetically selected for their preference (P) or aversion (NP) for ethanol. 2. In P versus NP females, the amount of both total cytochrome P450 and P450 binding to metyrapone was lower, whereas the activities of MEOS, aniline 4-hydroxylase (4-AOH), and 4-nitrophenol hydroxylase (PNP-OH) as well as the level of immunodetectable CYP2E1 content were consistently higher. By contrast, no substantial differences were observed between P and NP males. 3. Despite an apparent down-regulation of CYP2E1 expression occurring in all rats as a result of PB induction, P females maintained higher 2E1 levels and showed enhanced MEOS, 4-AOH and PNP-OH activities with respect to NP females. No such changes were detected in the male counterparts. 4. No sex-related differences in CCl4-mediated inhibition of monooxygenase or MEOS activities were evident between P and NP animals. 5. These results indicate that, in females only, the behavioural trait of ethanol preference is apparently associated not only with higher constitutive levels of CYP2E1 and rate of microsomal metabolism of ethanol but also with altered susceptibility to PB induction.

Effects of the subchronic administration of zinc ethylene-bis-dithiocarbamate (zineb) to rabbits.

The effects of subchronic administration (90 d) of zineb were studied in male New Zealand White rabbits. Rabbits were allotted to 3 groups of 8 animals each and offered diets containing 0, 0.3 or 0.6% zineb. A marked decline in weight gain, hemoglobin concentration, hematocrit, and erythrocyte and leucocyte counts occurred at the highest zineb dosage. There was a dose-related depression in circulating thyroid hormones, whereas serum lipid concentration, particularly that of cholesterol and triglycerides, increased. Hepatic lipid concentration was considerably reduced in rabbits exposed to 0.6% zineb. Neither serum testosterone nor the activities of selected testicular enzymes showed changes suggestive of testicular involvement. Pathological changes were in agreement with biochemical findings; there was a marked dose-related enlargement of the thyroid showing histological colloid struma. An increase in relative weight and moderate glycogenosis were detected in liver, whereas no lesions occurred in testes. It was concluded that thyroid and liver are the main targets for zineb toxicity in the rabbit. Unlike the results from previous studies conducted on other food-producing species, repeated exposure of rabbits to zineb failed to cause testicular damage. This might be related to the inability of zineb to significantly accumulate in the testes.

Experimental model of cirrhosis in rabbits exposed to carbon tetrachloride by inhalation.

Orthotopic hepatic transplantation is the most diffused surgical treatment used when attempting to substitute an irreversibly damaged liver. However, this practice faces two main problems. Firstly, surgeons need to explant the organ from the donor who is a just deceased human being, with all the implications involved. Secondly, the host patient must be rendered immunologically tolerant to the graft, through pharmacological suppression. Some hepatic alterations allow a structural and functional compensation by means of an autotransplant, but liver cirrhosis, which is widely diffused in humans and easily produced in experimental animals, has not yet been treated with this technique. This research was undertaken with the aim of procuring hepatic cirrhosis in a medium-sized animal--the rabbit--exposed for some months to CCl4 vapours. Preliminary experiments were performed in order to shorten the challenge period by means of liver induction. This approach was unsuccessful because of the natural susceptibility of this species, which was heightened by the barbiturate administration (0.05% sodium phenobarbital in drinking water, for one week). However, induction, rendered impossible the survival after carbon tetrachloride given orally (125 microliters/kg body weight) or by inhalation (1000 ppm for 2 hours). Finally, CCl4 was administered to normal rabbits by inhalation at the initial concentration of 100 ppm, for 2 hours twice a week. The level of the hepatotoxin was subsequently raised stepwise, according to the body weight curve, up to the maximum of 600 ppm by week 23. At the fourth week of intoxication, the metabolic activity of liver microsomes appeared severely depressed, as shown by the 300% increase of hexobarbital sleeping time. The observation of the surface of liver in situ, performed through an explorative laparatomy, showed initial but clear signs of hepatic fibrosis. At sacrifice, after six or ten months of cirrhogenic treatment, the histopathology examination of the liver demonstrated severe and massive cirrhosis, respectively for the two different steps. This experimental schedule appears to be suitable for producing hepatic cirrhosis in a medium-sized animal, which could be used as a model for pioneering attempts of liver autotransplantation. Furthermore, we point out two important aspects of these results. First, the poison has been inhaled by the experimental animals as it generally happens for humans in the environment. Second, the ratio between the length of exposure versus the life span of rabbits--6 months and 8 years, respectively--parallels that reported for human cirrhosis due to halogenated hydrocarbons--5 or 10 years versus 60 or 80 years.

[The goal-oriented project "pathology of environmental pollutants" (M.U.R.S.T. 40%)]. / Il progetto finalizzato "patologia da tossici ambientali" (M.U.R.S.T. 40%).

The purpose of this study was to estimate to toxicity bound to effluent water, pore water and bed sediment sampled twice during 1994 (spring and autumn) from 16 tributaries of Po river. Toxicity was determined by using a battery of bioassays (Artemia salina, Dugesiagonocephala and Thamnocephalus platyurus) and a battery of phytoassays, including Oryza sativa and Triticum sativum. Results show that practically no toxicity is found in effluent water, mild adverse effects are obtained with sediment pore water, while the most harmful compounds are those bound to sediment particles. These data strengthen the hypothesis that sediment is the true historical memory of the ecotoxicological conditions of a water body. Finally, the map of our results illustrates that pollution severity is progressively increasing from western to eastern tributaries in Po river valley.

[Toxicologic effects of cadmium on gametes and embryonal kinetics of paracentrotus lividus]. / Effetti tossicologici da cadmio sui gameti e sulla cinetica embrionale del paracentrotus lividus.

The adverse effects of cadmium chloride or cadmium sulphate on both fertilization of sea urchin eggs by spermatozoa and cadmium treated embryos have been studied. Cd treated ova can be fertilized by control spermatozoa. On the contrary, the preliminary treatment of spermatozoa with CdCl2 or CdSO4 succeeds in reducing the fertilization rate of control eggs at concentrations as low as 2 micrograms/ml. Cd sulphate appears to be more noxious than Cd chloride. The study of the residual motility of sea urchin embryos exposed to either CdCl2 or CdSO4 has evidenced a fairly effect at 21 hours after fertilization. On the contrary, concentrations of 25 micrograms/ml Cd chloride or Cd sulphate significantly affect the motility of 48 hours embryos. Higher levels of cadmium, for example, 150 or 200 micrograms/ml, completely block the translation of developing organisms. In this instance, Cd chloride is more effective than Cd sulphate. In addition, a few embryos are affected by malformations and are underdeveloped. These results are interpreted on the basis of the inhibitory effect of cadmium on the CaCO3 uptake by tissues and structures which physiologically need to be calcified in order to develop and function. Further, they give support to the predictive significance of the toxicological tests on the sea urchin plutei in monitoring environmental hazards.

[Protective effect on nephropathy and on cataract in the streptozotocin-diabetic rat of the vanadium-lazaroid combination]. / Azione protettiva sulla nefropatia e sulla cataratta nel ratto diabetico da streptozotocina da parte della combinazione Vanadio-Lazaroide.

Experimental work from our laboratory has confirmed the protective power of vanadium compounds on hyperglycemia and glycosuria in streptozotocin (STZ) diabetes. Furthermore, the diabetic cataract too has been partially prevented. The protection slightly increased, when vanadium was administered in combination with vitamin E. This investigation has introduced a combination of Na3VO4 plus the lazaroid U-83836E, a liposoluble antioxidant much more efficacious than tocopherol, in order to improve the insufficient protection when vitamin E was used. Male Wistar rats, rendered diabetic with STZ, were treated for 12 weeks with Na3VO4 in drinking water, U-83836E carried by the food, or both. The most significant metabolic parameters (food and fluid intake, diuresis and excreted feces) were studied monthly by means of metabolic cages. Body weight, glycemia, glycosuria and proteinuria were also recorded. At week 6 and 12 of the treatment, the opaqueness of the eye lenses was controlled. Circulation glycosylated hemoglobin (HbA1c), fructosamine, N-acetyl-beta-D-glucosaminidase (NAG) and fluorescent peroxides were evaluated at the end of the experiment. After the first month of treatment U-83836E improved significantly the protective effect of vanadate alone on polydipsia and polyuria, but more efficiently on hyperglycemia and glycosuria. The further ameliorating effect of the lazaroid was observed also on HbA1c, NAG and, most important, on the cataract. In conclusion, these findings demonstrate that the lazaroid U-83836E succeeds in further protecting the most important symptoms of diabetes treated with vanadate, and that this antioxidant acts effectively even when it is administered per os, in a non invasive manner.

Amelioration of diabetes and cataract by Na3VO4 plus U-83836E in streptozotocin treated rats.

Experimental work in our laboratory has confirmed the protective activity of vanadium compounds on hyperglycemia and glycosuria in streptozotocin (STZ) diabetes. Furthermore, diabetic cataract has also been partially prevented. Nevertheless, the combination of a natural antioxidant, vitamin E, with Na3 VO4 has not further enhanced this ameliorating effect. Our experimental approach has been an attempt to block the prooxidant activity of both STZ and vanadate, with the purpose of eliciting the best possible antidiabetic protection. More recently, a lipid soluble synthetic antioxidant U-78517F, a 2-methylaminochroman, has been reported to have a significant protective effect against brain injury and ischemia. This compound inhibits the iron-dependent lipid peroxidation 100 times more effectively than vitamin E. This investigation has introduced a combination of the vanadium compound plus the aforesaid lazaroid, as its (-) enantiomer, U-83836E, in order to improve the insufficient protection when vitamin E was used. For twelve weeks, male Wistar rats, rendered diabetic with STZ, were administered Na3VO4 in drinking water along with the lazaroid carried by the food. Four, eight and twelve weeks after the beginning of the protective treatment, fluid and food intake, diuresis and excreted feces, glycosuria and proteinuria were determined on biological samples obtained in metabolic cages; body weight and glycemia were also recorded. At weeks 6 and 12 of the treatment, the opaqueness of the eye lenses was controlled and registered. At the end of the experiment, circulating glycosylated hemoglobin (HbA1c), fructosamine, N-acetyl-beta-D-glucosaminidase (NAG), and fluorescent peroxides were evaluated. Within the first month of treatment, protection by the combination paralleled that elicited by vanadate alone. At subsequent steps, U-83836E significantly improved the protective effect of vanadate alone on polydipsia and polyuria, but especially on hyperglycemia and glycosuria. The further ameliorating effect of the lazaroid was also observed on HbA1c and NAG, and, most important, on the cataract. In conclusion, these findings demonstrate that the lazaroid U-83836E succeeds in further protecting the most important symptoms of diabetes treated with vanadate, and that this antioxidant acts effectively even when it is administered orally in food, in a non invasive manner.

Induction of hepatic drug metabolizing enzymes and interaction with carbon tetrachloride in rats after a single oral exposure to atrazine.

A single oral dose (430 mg/kg) of atrazine, a widely employed s-triazine herbicide, was administered to young male rats. There was a significant increase of the in vivo elimination of hexobarbital and a significant induction of the activity of 7-pentoxyresorufin-O-dealkylase, while cytochrome P-450 content and other mixed function oxidase activities remained unaltered. The administration of carbon tetrachloride (CCl4) to atrazine pretreated rats did not substantially augment the impairment of drug metabolizing enzymes brought about by CCl4 alone. Results suggest that atrazine behaves like a relatively weak inducer of phenobarbital-inducible families of cytochrome P-450.

In vivo studies on halogen compound interactions. III. Effect of carbon tetrachloride plus 1,2-dichloroethane on liver necrosis and fatty accumulation.

The effect of a single dose of carbon tetrachloride (CT), 1,2-dichloroethane (DCE) and the mixture on liver toxicity was investigated. The co-presence of both toxins exerts a more than additive effect on liver necrosis and on TBA-reactive substances produced by liver homogenates incubated at 37 degrees C. Both these effects are prevented in animals treated with vitamin E. The liver GSH is not involved in the synergistic action. The liver triglyceride levels of rats treated with the mixture are lower than in those treated with CT alone. This finding cannot be explained either by an improvement in the lipoprotein secretion or by the fact that the liver receives less NEFA. It is thus likely that the apparent protection against liver steatosis is the sign of more severe damage to the liver cell that partially blocks triglyceride synthesis. The mixture composed of CT+DCE exerts a potentiating action on liver toxicity with CT+DCE having the same characteristics as those, as previously reported, exerted by the co-presence of CT+DBE.

In vivo studies on halogen compound interactions. IV. Interaction among different halogen derivatives with and without synergistic action on liver toxicity.

The liver toxicity of several halogen compound mixtures have been tested. The compounds were selected on the basis of their metabolic pathways: carbon tetrachloride (CT) and trichlorobromomethane (TCBM) undergo a dehalogenation via P450-dependent enzyme system, 1,2-dichloroethane (DCE) and 1,2-dibromoethane (DBE) are mainly conjugated with the cytosolic glutathione (GSH) by means of the GSH-S-transferase. The mixture TCBM+DBE shows a more than additive action on lipid peroxidation and liver necrosis. TCBM, like CT, reduces the hepatic level of GSH-S-transferase, increasing the amount of DBE available for cytochrome P450-dependent metabolism, with the production of toxic metabolites. Thus, the behavior of the mixture TCBM+DBE is very similar to that of the mixture CT+DBE, previously reported. Mixtures composed of CT+TCBM and DCE+DBE do not show any synergistic effect on liver toxicity. The results allow one to conclude that the toxicity of mixtures of halogen compounds can be partly predicted on the basis of their metabolic pathways. When the metabolism is quite different, a synergistic toxicity can occur if one pathway interferes with a detoxification mechanism of the other compound. If the two metabolisms are very similar they produce, at most, an additive toxicity.

T-cell subpopulations in pediatric healthy children: age-normal values.

Assessment of the percentage and absolute number of T cells as well as of their main subpopulations is presently a routine procedure for the diagnosis and follow-up of a wide array of pediatric immunologic disorders. For several clinical applications (severe immunodeficiencies or leukaemias) the diagnostic usefulness of their enumeration does not require close comparison with age normal values, while in other circumstances such as follow-up of immunomodulating or immunosuppressive treatments or detection of minor immune defects, the expected changes of T cell subsets are more subtile and they are likely to be detected only by comparison with well-defined age normal values. In the present study CD3, CD4 and CD8 positive cells were enumerated in a group of 410 healthy children of age ranging from 30 days to 9 years. No significant changes in percentage or absolute number were observed during infancy and childhood. Furthermore the sum of CD4 and CD8 positive cells was close to the percentage of CD3 positive cells, suggesting a phenotype maturity of T cells from infancy.

Lethality, hexobarbital narcosis and behavior in rats exposed to atrazine, bentazon or molinate.

Previous findings from our laboratory suggested a possible interaction of atrazine, bentazon and molinate with other environmental and/or occupational poisons. The aim of this research was to obtain further toxicological information by using phenobarbital-induced rats and to characterize the effects of these herbicides on the hepatic microsomal metabolism of xenobiotics. Acute experiments have shown that the LD50 is augmented by the barbiturate pretreatment when atrazine is used, remains unchanged in the case of bentazon, but is lowered when molinate is given. Recrystallized atrazine, in the absence of the wetting compounds, elicits the same acute toxicity found when animals are challenged with a commercial preparation. No significant sex-related differences have been observed. In long-term treatment with these toxicants, atrazine shortened the hexobarbital narcosis, but no effect was observed after administration of either bentazon or molinate. Further studies on hexobarbital sleeping time demonstrated that females are more susceptible than males to the narcotic effect of this compound. The induction-like effect of atrazine exposure has been confirmed, mainly in young animals. At the end of the sleeping time, the actual serum concentration of hexobarbital is practically the same, and is not related to the length of the sleeping time. The absence of behavioral alterations in the open field tests exclude possible neurological effects of the triazine herbicide. In conclusion, these data demonstrate that atrazine by itself induces the hepatic pharmacometabolic system, while its metabolites result less toxic than the parent compound. On the contrary, metabolic transformations render the toxic effects of bentazon more severe.

Age- and sex-related effects on hepatic drug metabolism in rats chronically exposed to dietary atrazine.

Male and female Wistar rats were administered a diet containing 450 ppm atrazine as early as 60 days prior to the cohabitation period and the same diet was offered to their offspring. Hexobarbital sleeping time and further in vitro assays pointed to a monooxygenase induction which appeared to be more marked in males vs females and most significant in the offspring at weaning. At this age, induction involved also the cytosolic glutathione S-transferase, a phase II enzyme. Results would suggest that the inducing properties of the herbicide can be transferred to the offspring via the placental and/or the mammary route.