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Short fragments and fragment analogues of beta-amyloid 1-42 peptide (Abeta1-42) display a protective effect against Abeta-mediated neurotoxicity. After consideration of our earlier results with in vitro bioassay of synthetic Abeta-recognition peptides and toxic fibrillar amyloids, five pentapeptides were selected as putative neuroprotective agents: Phe-Arg-His-Asp-Ser amide (Abeta4-8) and Gly-Arg-His-Asp-Ser amide (an analogue of Abeta4-8), Leu-Pro-Tyr-Phe-Asp amide (an analogue of Abeta17-21), Arg-Ile-Ile-Gly-Leu amide (an analogue of Abeta30-34), and Arg-Val-Val-Ile-Ala amide (an analogue of Abeta38-42). In vitro electrophysiological experiments on rat brain slices demonstrated that four of these peptides counteracted with the field excitatory postsynaptic potential-attenuating effect of Abeta1-42; only Arg-Val-Val-Ile-Ala amide proved inactive. In in vivo experiments using extracellular single-unit recordings combined with iontophoresis, all these pentapeptides except Arg-Val-Val-Ile-Ala amide protected neurons from the NMDA response-enhancing effect of Abeta1-42 in the hippocampal CA1 region. These results suggest that Abeta recognition sequences may serve as leads for the design of novel neuroprotective compounds.

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Beta-amyloid peptides (Abetas) bind to several G-protein coupled receptor proteins and stimulate GTPase activity in neurons. In this study we determined the effects of Abeta(1-42), Abeta(1-40), Abeta(25-35) and their mixtures on [(35)S]GTP binding in rat brain cortical membranes in the absence and presence of zinc. We found that the Abetas alone induced a concentration-dependent activation of G-proteins (IC50 approximately 10(-6) m), while aggregated Abeta fibrils only affected GTP binding at concentrations above 10(-5) m. Mixing Abeta(25-35) with Abeta(1-42) or Abeta(1-40) induced a several-fold increase in GTP-binding. This potentiation followed a bell shaped curve with a maximum at 50 : 50 ratios. No potentiating effect could be seen by mixing Abeta(1-40) and Abeta(1-42) or highly aggregated Abetas. Zinc had no effect on Abeta(1-40/42) but strongly potentiated the Abeta(25-35) or the mixed peptides-induced GTP-binding. Changes in secondary structure accompanied the mixed peptides or the peptide/zinc complexes induced potentiation, revealing that structural alterations are behind the increased biological action. These concentration dependent potentiating effects of zinc and the peptide mixtures could be physiologically important at brain regions where peptide fragments and/or zinc are present at elevated concentrations.

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Whereas a cardinal role for beta-amyloid protein (Abeta) has been postulated as a major trigger of neuronal injury in Alzheimer's disease, the pathogenic mechanism by which Abeta deranges nerve cells remains largely elusive. Here we report correlative in vitro and in vivo evidence that an excitotoxic cascade mediates Abeta neurotoxicity in the rat magnocellular nucleus basalis (MBN). In vitro application of Abeta to astrocytes elicits rapid depolarization of astroglial membranes with a concomitant inhibition of glutamate uptake. In vivo Abeta infusion by way of microdialysis in the MBN revealed peak extracellular concentrations of excitatory amino acid neurotransmitters within 20-30 min. Abeta-triggered extracellular elevation of excitatory amino acids coincided with a significantly enhanced intracellular accumulation of Ca2+ in the Abeta injection area, as was demonstrated by 45Ca2+ autoradiography. In consequence of these acute processes delayed cell death in the MBN and persistent loss of cholinergic fibre projections to the neocortex appear as early as 3 days following the Abeta-induced toxic insult. Such a sequence of Abeta toxicity was effectively antagonized by the N-methyl-D-aspartate (NMDA) receptor ligand dizocilpine maleate (MK-801). Moreover, Abeta toxicity in the MBN decreases with advancing age that may be associated with the age-related loss of NMDA receptor expression in rats. In summary, the present results indicate that Abeta compromises neurons of the rat MBN via an excitotoxic pathway including astroglial depolarization, extracellular glutamate accumulation, NMDA receptor activation and an intracellular Ca2+ overload leading to cell death.

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Organic pollution of water and soil has various harmful effects on biological systems (1). Chlorine substituted benzol compounds are one these xenobiotic substances, which are toxic to the environment (2). They can also accumulate in plant and animal tissues (3), which provides ample reason to study the effects of sublethal doses of chloro-benzols on various cell cultures. In this study the toxic effects of chloro-benzols were investigated on avian fibroblast and mammalian hepatocyte cultures. The fibroblast cultures were prepared from eggs preadapted to chloro-benzol during a fourteen-day-long incubation period. The Wistar rat hepatocyte monolayer cultures were exposed to a direct treatment of 1,2,4-tri-chloro-benzol (0.01 microgram/ml-1 microgram/ml) for 3 hours. Following the treatment with chloro-benzol, the viability of the cells was measured, together with lactic dehydrogenase activity, in both kinds of cultures. The effect of tri-chloro-benzol treatment on chicken eggs was not significant. The cells of chicken embryos were not damaged after the 1,2,4-tri-chloro-benzol treatment. The hepatocyte cultures showed the toxic effects of 1,2,4-tri-chloro-benzol after the direct and acute treatment. The cell viability decreased and the LDH activity increased significantly. These results show that the primary cell cultures are suitable for studying the effects of organochlorine compounds.

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Analytical methods were introduced for the determination of residues of ectoparasiticides containing pyrethroid and organophosphate active ingredients in foods. Milk and edible tissues of cows treated with three experimental ectoparasiticides (containing cypermethrin + diazinon, deltamethrin + diazinon and alphamethrin + diazinon, respectively) were assayed for the presence of active ingredient residues. Synthetic pyrethroid residues were not detected in any of the samples processed. Diazinon residues could only be detected in milk samples taken on the first day after treatment (0.005-0.025 mg/kg) and in liver and fat tissue samples taken on the day of slaughtering (0.12 and 0.01 mg/kg, respectively). Permethrin and propetamphos residues were determined in the skin, meat and liver of chickens kept on 'Blotic-B' treated litter and in eggs collected at different times after the treatment of layer houses. Permethrin residues could not be detected in any of the samples (< 0.01 mg/kg). Meat and fat tissues of chickens slaughtered on the day after treatment contained small amounts of propetamphos (0.003 and 0.02 mg/kg, respectively). In the case of chickens kept on the treated litter and slaughtered after one week, active ingredient was not detected in meat, but 0.006 mg/kg propetamphos was present in the fat. The residue content of other samples (liver, egg) was below the detection limit of the applied method at all sampling times. From the food toxicological point of view these pesticide combinations can be used safely if the recommended withdrawal period is observed between ectoparasiticide administration and slaughter.

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The aggregation of beta-amyloid peptides is very important for their neurotoxic effect; standardization of the aggregation grade is necessary for biological experiments. Measurement of aggregation with physicochemical methods is a difficult task. The present work revealed that FT-IR can be used for studying the aggregation properties of beta-amyloid peptides and the effects of environmental variables (solvent, pH, ions, and temperature) on aggregation. In dimethyl sulfoxide or hexafluoroisopropanol, amyloid peptides are in a monomeric state; on dilution with phosphate buffer just before measurement is made, aggregation begins. A detailed two-dimensional FT-IR correlation spectroscopic study was made of the conformational transitions that occur during the aggregation of beta-amyloid peptides. Two processes (random/helix-to-beta-sheet and aggregation of beta-sheets) and multiple conformational states were observed before the most stable form was attained. beta-Amyloid peptides undergo decomposition in basic buffers containing Ca(2+); this process should be avoided during aging experiments.

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Long-term behavioral effects, changes in learning and memory functions and aberrations of cholinergic fibers projecting to the parietal cortex were investigated after bilateral injections of beta-amyloid(Phe(SO3H)24)25-35 peptide in rat nucleus basalis magnocellularis (nbm). The beta-amyloid peptide used in these experiments contained the original beta-amyloid 25-35 sequence which was coupled to a phenylalanine-sulphonate group at position 24. This additional residue serves as a protective cap on the molecule without influencing its neurotoxic properties and results in water-solubility, stability and low rates of peptide metabolism. In this paper, home cage, locomotor and open-field activities, passive shock-avoidance and 'Morris' water maze learning abilities were assessed throughout a 35-day survival period. Subsequently, acetylcholinesterase (AChE) histochemistry was used to visualize alterations of parietal cortical cholinergic innervation. In response to the neurotoxic action of beta-amyloid(Phe(SO3H)24)25-35, a progressive hyperactivity developed in the rats in their home cages which were maintained throughout the 5-week post-injection period. This was accompanied by a significant hypoactivity in the novel environment of a locomotor arena. Beta-amyloid(Phe(SO3H)24)25-35-treated animals showed greatly impaired cortical memory functions in the step-through passive shock-avoidance paradigm, while spatial learning processes remained unaffected. Moreover, beta-amyloid(Phe(SO3H)24)25-35 injections in the nucleus basalis suppressed explorative behavior in rats and inhibited conditioned stress responses 28 days after surgery. Reductions of cortical cholinergic (AChE-positive) projections provided anatomical substrate for the behavioral changes. This indicated extensive, long-lasting neurodegenerative processes as a result of beta-amyloid(Phe(SO3H)24)25-35 infusion.

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The effect exerted by a biogenic performance enhancer of high nucleotide content on meat quality in 20 Norwegian Landrace pigs (90 to 95 kg) was examined. The diet of the treated group was supplemented with the performance enhancer for the last 30 days of fattening. The stress effect was transport to the slaughterhouse and slaughter itself. Plasma creatine kinase (CK), lactate dehydrogenase (LDH), and aspartate aminotransferase (AST) activities, glucose and cortisol concentrations, and muscle pH were determined. Serious stress damage was found in the cardiac and skeletal muscle, as indicated by the high CK (980 U/L), LDH (> 1600 U/L) and AST (67 U/L) activities in the untreated group; values were significantly lower in the experimental pigs (458, 468 and 17 U/L, respectively). There were no significant differences in glucose and cortisol concentrations between the two groups. In the control group the pH values were significantly lower and more muscle samples showed PSE character than in the treated group (75 and 30%, respectively).

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The effect of Ca2+ and Al3+ binding on the conformation of the neurotoxic amyloid fragment H-Ile-Ile-Gly-Leu-Met-NH2 [betaA(31-35)NH2] was studied in trifluoroethanol solutions and in the presence of liposomes. Comparative circular dichroism and Fourier-transform infrared spectroscopic studies revealed that the peptide forms a specific 1:1 complex with Ca2+ which coordinates the polar amide carbonyl groups of the peptide backbone. The results suggest the importance of a folded structure in the complexation of Ca2+. On the contrary, the increasing Al3+ concentration causes a gradual shift of the conformational equilibrium toward beta-sheet structure reflecting no specific binding site for Al3+. In the presence of liposomes the peptide adopts a conformation similar to that of the Ca(2+)-peptide complex. The relevance of the stabilization of peptide conformation by Ca2+ and liposome binding to the bioactive conformation of betaA(31-35)NH2 is also discussed.

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beta-Amyloid(1-42) peptide (beta AP(1-42) was injected into the medial septum of rats. After a 14-day survival time, neuronal alterations in the septal cholinergic and GABAergic systems were visualized by means of histo- and immunocytochemical methods. Neurons insulted by the peptide were primary choline acetyltransferase-immunoreactive (ir), while only minor effects of beta AP(1-42) were observed on parvalbumin-ir interneurons. These results indicate that the changes in intracellular Ca2+ level elicited by beta AP(1-42) may contribute to beta-amyloid neurotoxicity, and Ca(2+)-binding proteins may play an important role in the protection against the neurotoxic effects of beta AP(1-42).

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Beta-amyloid(1-42) peptide (betaAP) was injected into the right nucleus basalis magnocellularis (nbm) of rats. After a 14-day survival time, the acetylcholinesterase and choline acetyltransferase activities and the number of muscarinic receptors were found biochemically to be significantly reduced in the ipsilateral frontal cortices. Confirmation of these data with silver staining also revealed degeneration of the projective fibers of the nbm to the frontal cortex. These results demonstrate the cholinotoxicity of betaAP in an in vivo animal model.

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The secondary structure of amyloid beta A(25-35) and its deletion analogues was studied by circular dichroism (CD), Fourier transform infrared (FTIR) spectroscopy and molecular dynamics calculation. Data of our comparative CD and FTIR measurements in trifluoroethanol suggest that beta A(25-35)NH2 has a preferred beta-sheet conformation. Contrary to this beta A(31-35)NH2 tends to adopt a beta-turn conformation. Based on the comparable neurotoxic effect of beta A(25-35)NH2 and beta A(31-35)NH2 the neurotoxicity likely involves the same 31-35 core sequence and the "biologically active conformation" is a beta-turn rather than a beta-sheet structure.

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A subacute toxicological study of the effects of the ethylene-bisdithiocarbamate-containing fungicide Dithane M-45 (80% mancozeb) was carried out in male Wistar rats. The rats were given Dithane mixed in the feed at doses of 0, 10, 50, 75, 113, 169, 253 or 379 mg/kg body weight for 12 wk. One-third of the rats given 379 mg Dithane/kg body weight died, and doses of greater than or equal to 169 mg/kg decreased the growth of the rats as well as the nutrient utilization. The relative weights of the liver and thyroid were significantly increased in rats given greater than or equal to 75 mg Dithane/kg body weight, and those of the kidneys, adrenals and testes were significantly increased in the two highest dose groups. The serum cholesterol level was increased significantly in groups given greater than or equal to 75 mg/kg, and doses of 113, 169 or 253 mg/kg caused the elevation of the triglyceride content of the liver. The two highest doses of Dithane decreased the detoxicating capacity of the liver. The function of the thyroids was impaired even by very small doses of Dithane: at a dose of 10 mg/kg body weight the quantity of iodine stored in the thyroids was decreased by 20%. At doses of greater than or equal to 50 mg/kg statistically significant decreases in thyroid iodine content were observed. Histological examination of the thyroids showed dose-dependent hyperplasia in rats treated with Dithane.

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Acute (LD50) and short-term (14 days) toxicological examinations were performed in animal experiments on the interaction of a synthetic pyrethroid Decis 2,5 EC (25 g deltamethrin/l) and of ethylene-bisdithiocarbamate/Dithane M-45 (80% mancozeb), using a 1:5 deltamethrin/mancozeb mixture. LD50 value of the mixture was similar to that of the more toxic Decis. In the short-term examination, some pathologically high AST and ALT values were observed in the treated groups and the deltamethrin content of fatty tissue samples increased parallel with the increase of Decis consumption. The chymotrypsin and lipase activities in the small intestinal mucosa and gamma-GT and LAP activities in the content of the bowels were reduced in several treated groups. The administration of Dithane in a dose in accordance with 20% of the LD50 value (3125 mg/kg b.m.) proved to be more toxic than expected and caused the death of the animals.

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In the GDR the benzo[a]pyrene (BaP) contents of vegetables grown in areas far from industrial plants range from 0.01 to 2.3 microgram/kg of fresh weight with a mean of 0.43 microgram/kg; and in the Hungarian People's Republic the BaP contents of such vegetables lie between 0.05 and 3.85 microgram/kg with a mean of 0.6 microgram/kg. The BaP contents of vegetables from areas near industrial plants or in the immediate proximity of emitters range from 0.15 to 12.4 microgram/kg (in the GDR) and from 0.05 to 19.6 microgram/kg (in the Hungarian People's Republic), being markedly superior to the values obtained with comparable vegetables from areas far from industrial plants. The BaP contents of samples from the Hungarian People's Republic collected near a relatively new petrochemical plant were 3.5 times as high as those of samples collected in areas far from industrial plants; and the BaP contents obtained with samples collected in the proximity of an old petrochemical plant were 8 times as high as those observed with samples collected in areas far from industrial plants. The contamination depends strongly upon the type of emitter. Samples collected in the GDR in the immediate proximity of coal-preparation plants (such as tar extraction, coking or carbon black plants) contained BaP contents 10 to 100 times as high as those found in samples collected in areas far from industrial plants. Targeted measures aiming at diminishing air pollution will widely reduce the contamination of harvested crops with cancerogenic hydrocarbons.

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Smoking is a well-known source of food contaminated caused by carcinogenic polycyclic aromatic hydrocarbons. Epidemiological studies indicates a statistical correlation between the increased occurrence of cancer of the intestinal tract and the frequent intake of smoked foods. As observed during the last 10 years in a certain district of Hungary with a Slovenian population, the percentage of stomach cancer among all types of cancer is nearly twice as high (47-50%) as in Hungary altogether (29.9%). In this special district, predominantly home-smoked meat products are consumed. Using identical techniques, the authors investigated the contamination of smoked foodstuffs by carcinogenic, cocarcinogenic and other polyaromatic hydrocarbons in the German Democratic Republic and in Hungary. No significant differences have been found either in the average values or in the ranges of the benzo(a)pyrene (BaP) contents in meats smoked by industrial processes used in the GDR and in Hungary. In the GDR, industrially smoked foods contain on average 0.43 micrograms/kg, products smoked in handicraft workshops 0.76 micrograms/kg, home-smoked products 0.74 micrograms/kg. The mean BaP content of all smoked meat and sausage products amounts of 0.55 micrograms/kg. In Hungary, the following average values have been found: 0.6 micrograms/kg for industrially smoked products, 0.74 micrograms/kg for home-smoked products, the total being 0.7 micrograms/kg. The average BaP value of home-smoked (softwood) products of the Slovenian population in Hungary is as high as 4.16 micrograms/kg. Apart from this particular case, the techniques used in both countries permit the production of smoked meat and sausages with a BaP content of less than 1 microgram/kg.

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The quantity of phycyclic aromatic hydrocarbons (PAHs) in Hungarian foods and consumer goods were measured in more than 300 samples of food and other consumables. In more than 60% of the samples, pyrene, fluoranthene, benz(a)anthracene and benzo(g.h.i)perylene were detectable besides the highly carcinogenic benzo(a)pyrene (BaP). Investigational results and consumption data show that the average per man-year BaP intake with food in Hungary is 300-600 microgram.

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In 1975, in cooperation with 8 County Hygienic Stations, total mercury content in fish from rivers and lakes in Hungary was measured. The mercury content in fish muscle-tissue from 164 fish samples averaged 0.36 mg/kg. Average levels in different fish species ranged from 0.10 to 0.57 mg/kg. The amount of mercury in fish from rivers and lakes, with the exception of the Danube, was, in general, lower than the tolerance level (0.50 mg/kg) as adopted in many countries. Mercury levels in fish from Lake Balaton did not amount to more than 0.30 mg/kg. The circulating cold vapour atomic absorption method was used to determined the mercury content in the fish samples.

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In the GDR, industrially smoked foods contain on an average 0.43 microgram Benzo(a)pyrene (BaP)/kg; products smoked according to trade rules, 0.76 microgram/kg; home-smoked products, 0.74 microgram/kg. The mean BaP content of smoked meat and sausage products in the GDR is 0.55 microgram/kg. Smoked fishes (without skin) contained on an average 1.39 microgram BaP/kg. The following average values were found in the Hungarian People's Republic: 0.6 microgram/kg (industrially smoked), 0.74 microgram/kg (home-smoked), 0.7 microgram/kg (total mean). Smoked cheese contained on an average 0.85 microgram/kg. The average BaP value of home-smoked (softwood) products of a Slovene ethnic group in the Hungarian People's Republic is 4.16 microgram/kg; the population of this region shows a considerably increased rate of stomach cancer. Apart from this particular case, the techniques used in both countries permit to obtain smoked meat and sausage products and smoked cheese with BaP contents less than 1 microgram/kg.

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