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Mutagenicity of methylazoxymethanol acetate in the presence of alcohol dehydrogenase, aldehyde dehydrogenase, and rat liver microsomes in Salmonella typhimurium His G46.

Methylazoxymethanol (MAM) is the short-lived toxic and carcinogenic aglycone of cycasin, a natural component of the cycad plant. In the present study, the stable acetate ester of MAM, MAM acetate, was tested in combination with porcine liver esterase and Salmonella typhimurium His G46 to study the comparative mutagenicity of this compound in the presence of rat hepatic alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), and rat liver microsomes. In the presence of rat liver microsomes and an NADPH-generating system, mutagenicity of MAM acetate was not significantly altered. However, addition of rat liver 105,000g supernatant fraction and/or NAD+ significantly increased the number of his+ revertants above control. A concentration-dependent increase in mutagenicity of MAM acetate was observed for NAD+ from 50 to 200 microM, while NADP+ caused a decrease in mutagenicity of MAM acetate in this same concentration range. Pyrazole (100-500 microM) had no significant effect on mutagenicity of MAM acetate in the presence of rat liver 105,000g supernatant, while disulfiram at 500 microM resulted in a significant decrease in mutagenicity of MAM acetate. The results of this study implicate ALDH as essential in activation of MAM acetate to a mutagenic species in this system, while the role of ADH and microsomes appears to be minimal.

Alcohol Dehydrogenase↗

Rapid decrease of cytochrome P-450IIE1 in primary hepatocyte culture and its maintenance by added 4-methylpyrazole.

Studies were conducted to evaluate the possible induction or the maintenance of cytochrome P-450IIE1 in primary hepatocyte cultures by the inducing agent 4-methylpyrazole. Hepatocytes were isolated from control (noninduced) rats and from rats treated in vivo with either pyrazole or 4-methylpyrazole to induce P-450IIE1. The content of P-450IIE1 was determined by Western blots with antipyrazole P-450 IgG, and catalytic activity was assessed by assays of dimethylnitrosamine demethylase activity. The treatment with 4-methylpyrazole in vivo increased the content of P-450IIE1 and dimethylnitrosamine demethylase activity sevenfold and fourfold, respectively. In cultures prepared from noninduced hepatocytes, P-450IIE1 levels fell to values of 76%, 65%, 31% and 1% of freshly isolated hepatocytes after 1, 3, 6 and 9 days in culture. A similar decrease in dimethylnitrosamine demethylase was observed during this time. In cultures prepared from induced hepatocytes, the decline in P-450IIE1 was more rapid as levels fell to 77%, 31%, 3% and 3% of initial values after 1, 3, 6 and 9 days in culture. Again, the fall in dimethylnitrosamine demethylase activity paralleled the decline in content of P-450IIE1 and was more rapid with the induced hepatocytes. With cultures prepared from noninduced or induced hepatocytes, the addition of 4-methylpyrazole in vitro did not increase the content of P-450IIE1 or the activity of dimethylnitrosamine demethylase over the initial values. However, 4-methylpyrazole appeared to stabilize the P-450IIE1 and to decrease its rate of decline in culture. In noninduced cultures, the percent remaining content of P-450IIE1 after 6 days was 31% in the absence of and 52% in the presence of 5 mol/L 4-methylpyrazole. In cultures from 4-methylpyrazole-induced hepatocytes, the percent remaining P-450IIE1 after 3 days was 31% in the absence of inducer and 59% with 4-methylpyrazole added in vitro. Similarly 4-methylpyrazole helped to prevent the rapid decline of dimethylnitrosamine demethylase activity in induced and noninduced cultures. Viability of the induced and noninduced cultures in the absence or presence of added 4-methylpyrazole was similar. Levels of mRNA for P-450IIE1 were similar for livers from control rats and from rats treated in vivo with 4-methylpyrazole. The mRNA levels rapidly declined in induced and noninduced cultures, and this decline, unlike the fall in P-450IIE1 or dimethylnitrosamine demethylase activity, could not be prevented by the addition of 4-methylpyrazole in vitro to the cultures.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

Induction of cytochrome P450 2E1 increases hepatotoxicity caused by Fas agonistic Jo2 antibody in mice.

Cytochrome P450 2E1 (CYP2E1) may be a central pathway in generating oxidative stress, reactive oxygen species, and causing hepatotoxic injury by alcohol and various hepatotoxins. This study evaluated the ability of CYP2E1 to potentiate or synergize the hepatotoxicity of Fas in vivo. C57BL/6 mice were injected intraperitoneally with pyrazole (Pyr) to induce CYP2E1. Then, 16-hour fasted mice were administered agonistic Jo2 anti-Fas antibody ip. Other mice were treated with Pyr or Jo2 alone. Levels of serum aminotransferase were 8.3- and 6.3-fold higher in the Pyr/Jo2 group compared with Jo2 alone, respectively. Histological evaluation of liver showed more extensive acidophilic necrosis and severe pathological changes in the Pyr/Jo2-treated mice. DNA fragmentation and caspase-8 and -3 activities were more elevated in the Pyr/Jo2 group compared with Jo2 alone. CYP2E1 activity and protein levels were higher in the Pyr/Jo2 group than in Jo2 alone. Levels of inducible nitric oxide synthase, 3-nitrotyrosine protein adducts, malondialdehyde, and protein carbonyls were also higher in the Pyr/Jo2 group compared with Jo2 alone. Glutathione and activities of catalase and Cu-Zn superoxide dismutase were decreased in the Pyr/Jo2 group. Administration of chlormethiazole, an inhibitor of CYP2E1, to the Pyr/Jo2-treated mice caused a significant decrease of alanine aminotransferase and liver pathological changes in association with a decrease in CYP2E1 protein and activity. In conclusion, enhanced hepatotoxicity of Fas was found in mice with elevated levels of CYP2E1. We speculate that overexpression of CYP2E1 might synergize and increase the susceptibility to Fas induced-liver injury.

Animals↗

Nrf2 is increased by CYP2E1 in rodent liver and HepG2 cells and protects against oxidative stress caused by CYP2E1.

Induction of CYP2E1 by ethanol is one pathway through which ethanol generates oxidative stress. Nrf2 is a transcription factor that regulates important antioxidant and phase II detoxification genes. Nrf2 induction by CYP2E1 and its importance in the adaptive response to increased oxidative stress caused by CYP2E1 was studied. Increases in Nrf2 protein and mRNA were observed in livers or hepatocytes of chronic alcohol-fed mice or rats and of pyrazole-treated rats or mice, conditions known to elevate CYP2E1. HepG2 cells expressing CYP2E1 (E47 cells) showed increased Nrf2 mRNA and protein expression compared with control HepG2 C34 cells. Nrf2 is activated in E47 cells as shown by an increase in nuclear Nrf2 levels and Nrf2-antioxidant-responsive element binding activity, and upregulation of Nrf2-regultated genes, glutamate cysteine ligase catalytic subunit (GCLC), and heme oxygenase 1 (HO-1). Increases in Nrf2 protein and mRNA are blocked by inhibitors of CYP2E1 activity and a reactive oxygen species (ROS) scavenger, N-acetylcysteine, which decrease ROS levels as well as Nrf2 mRNA induction. Upregulation of GCLC and HO-1 in E47 cells is dependent on Nrf2 and is prevented by siRNA-Nrf2. Blocking Nrf2 by siRNA-Nrf2 decreases glutathione and increases ROS and lipid peroxidation, resulting in decreased mitochondrial membrane potential and loss of cell viability of E47 cells but not C34 cells. These results suggest that Nrf2 is activated and that levels of protein and mRNA are increased when CYP2E1 is elevated. In conclusion, Nrf2 plays a key role in the adaptive response against increased oxidative stress caused by CYP2E1.

Animals↗

Comparative effects of chronic ethanol and acetaldehyde exposure on myocardial function in rats.

This study was conducted to compare separately the chronic effects of high blood levels of ethanol and acetaldehyde on the metabolism of the heart. Levels of ethanol and acetaldehyde were altered by administration of either 4-methylpyrazole (4-MP), a potent alcohol dehydrogenase inhibitor, or pargyline (PAR), a monoamine oxidase inhibitor that markedly increases acetaldehyde levels in the blood following ethanol administration. Measurements were made in rats consuming ethanol for three to four weeks. Mitochondrial respiration, in vitro contractility of glycerinated heart muscle fibers, and myocardial protein synthesis were determined. As compared to animals receiving only ethanol, administration of either-4-methyl-pyrazole or pargyline plus ethanol resulted in more severe damage to mitochondrial respiration and myocardial protein synthesis. The data illustrate that both acetaldehyde and ethanol in high concentrations can cause severe damage to myocardial metabolism.

Acetaldehyde↗

Extensive protein carbonylation precedes acrolein-mediated cell death in mouse hepatocytes.

Allyl alcohol hepatotoxicity is mediated by an alcohol dehydrogenase-derived biotranformation product, acrolein. This highly reactive alpha,beta-unsaturated aldehyde readily alkylates model proteins in vitro, forming, among other products, Michael addition adducts that possess a free carbonyl group. Whether such damage accompanies acrolein-mediated toxicity in cells is unknown. In this work we established that allyl alcohol toxicity in mouse hepatocytes involves extensive carbonylation of a wide range of proteins, and that the severity of such damage to a subset of 18-50 kDa proteins closely correlated with the degree of cell death. In addition to abolishing cytotoxicity and glutathione depletion, the alcohol dehydrogenase inhibitor 4-methyl pyrazole strongly attenuated protein carbonylation. Conversely, cyanamide, an aldehyde dehydrogenase inhibitor, enhanced cytotoxicity and protein carbonylation. Since protein carbonylation clearly preceded the loss of membrane integrity, it may be associated with the toxic process leading to cell death.

Acrolein↗

Alteration of liver microsomal monooxygenases and substrate competition with aniline hydroxylase from rats chronically fed low-fat and high-fat-containing alcohol diets.

Male Sprague-Dawley rats fed ethanol (EtOH) 36% of total calories for four weeks in a liquid diet containing either 34% (HF) or 12% (LF) of calories as fat were studied with respect to induction of microsomal monooxygenases (MFO) and substrate competition with EtOH-inducible aniline hydroxylase. The specific activity and turnover of aniline hydroxylase were induced to similar extents by HF-EtOH and LF-EtOH diets. Whereas, both LF-EtOH and HF-EtOH caused a decrease in the turnover of arylhydrocarbon (benzo[a]pyrene) hydroxylase (AHH) and aldrin epoxidase compared to pair-fed (PF) controls, LF-EtOH but not HF-EtOH increased the turnover of ethoxycoumarin and ethoxyresorufin O-deethylase (ECOD and EROD). The increase in ECOD and EROD and the decrease in AHH by EtOH is contrary to the parallel induction of these activities by 3-methylcholanthrene (3-MC) and Aroclor 1254 (Aroclor). Benzo(a)pyrene (BaP) stimulated aniline hydroxylase in the HF-EtOH and PF systems, whereas with LF diet, stimulation was seen only in the EtOH group. Ethoxycoumarin (EC) inhibited aniline hydroxylase by microsomes from EtOH- and pyrazole-treated rats, whereas it stimulated aniline hydroxylase by control microsomes, suggesting that the EC effects were associated with EtOH-inducible cytochrome P-450. Ethoxyresorufin (ER) inhibited aniline hydroxylase in EtOH and PF groups, thus the differential effects of EC were not nonspecific O-deethylase effects. The effects of EtOH feeding on ECOD, EROD, and AHH (ie, substrates for 3-MC-inducible cytochrome P-450) displayed a greater differential between the experimental and control group with the LF- than with the HF-containing diet. The findings suggest that the alteration of certain MFO activities by chronic EtOH ingestion can be modified by the content of dietary fat. Moreover, the competition dynamics of MFO substrates toward EtOH-inducible aniline hydroxylase are altered by EtOH feeding and, in turn, modified by dietary fat.

Aniline Hydroxylase↗

Unambiguous assignment of trifluoromethylpyrazole regioisomers by 19F-15N correlation spectroscopy.

Analysis of the 19F chemical shifts of trifluoromethylpyrazole regioisomers has shown that while chemical shift is in general a reliable predictor of regiochemistry in this series, there is a narrow chemical shift range in which the two isomers overlap and the regiochemistry cannot be assigned with certainty. We have examined the usage of 19F--15N correlation spectroscopy as a method to provide a second unambiguous confirmation of regiochemistry of 3- and 5-trifluoromethylpyrazole regioisomers. In the case of 3-trifluoromethyl analogs, one expects a three-bond coupling to the pyridine type nitrogen (N-2). In the case of the 5-trifluoromethyl pyrazole the situation is exactly reversed, with the fluorines of the trifluoromethyl moiety being 3-bonds from the pyrrole type nitrogen (N-1). We have observed that 3-trifluromethyl analogs follow the expected pattern of readily observable 3-bond coupling. However, 19F--15N couplings in 5-trifluromethyl analogs do not follow the normal pattern of 3-bond coupling. Using this information we have been able to develop unambiguous methods to distinguish the isomers.

Fluorine Radioisotopes↗

Effect of ethanol and methanol on the motility of Saccostrea commercialis sperm and sperm models.

The organic solvents methanol and ethanol at concentrations of 2.5% and 5% (v/v), respectively, were found to significantly (P less than 0.001) decrease the radius of curvature and track velocity of S. commercialis sperm. To observe the effects of the solvent directly on the axoneme, S. commercialis sperm models were prepared by extraction with Triton X-100 and reactivation with ATP in media containing acetate anions, DTT, magnesium, and cAMP. Concentrations of 0.1% Triton X-100 demembranated sperm while 0.01% and 0.05% Triton X-100 permeabilized sperm. Sperm models were successfully produced after reactivation with 1 mM ATP. At pH 8.25, 1% (v/v) ethanol or methanol was observed to increase waveform asymmetry and significantly (P less than 0.001) decrease track velocity of 0.1% Triton X-100 demembranated sperm models. Similarly 1% (v/v) ethanol increased tail-wave asymmetry and decreased track velocity of 0.01% and 0.05% Triton X-100 permeabilized sperm models. Reactivated motility of 0.05% Triton X-100 permeabilized sperm models prepared at pH 7.8 were poor and improved after treatment with 7% (v/v) ethanol, which increased waveform asymmetry and doubled the track velocity of sperm. This stimulatory effect of ethanol was unchanged in the presence of the alcohol dehydrogenase inhibitor pyrazole. Concerning the precise mechanism of action of ethanol on the axoneme, we conclude that a stimulatory or inhibitory effect of ethanol is dependent on the pH of the sperm model system used.

Alcohol Dehydrogenase↗

Synthesis and herbicidal activity of phenyl-substituted benzoylpyrazoles.

A novel series of substituted 3-phenyl benzoylpyrazoles were prepared and tested as potential grass herbicides. The targeted materials were prepared by three newly developed synthetic routes, which allowed a comprehensive study of the SAR (structure-activity relationships) of this series. The best combination of grass weed activity (Avena fatua L, Setaria viridis (L) Beauv and Alopecurus myosuroides Huds) and wheat selectivity was obtained with an alkoxy group in the 4-position of the phenyl ring. Activity was further enhanced by the presence of tert-butyl on the pyrazole and a methyl group at the C-2 position of the benzoyl moiety. The alkoxy-substituted 3-phenylbenzoylpyrazoles are a novel class of herbicides with potential utility for control of important grass weeds in cereals.

4-Hydroxyphenylpyruvate Dioxygenase↗

Ethanol increases content and activity of human cytochrome P4502E1 in a transduced HepG2 cell line.

Using recombinant retroviral expression, a HepG2 cell line which stably and constitutively expresses the coding sequences of the human cytochrome P4502E1 was previously established. Addition of ethanol (2 to 100 mM) to the culture medium of this cell line for two days resulted in an increase in the content of P4502E1 as determined by immunoblotting and an increase in HepG2 microsomal oxidation of p-nitrophenol, aniline, and N,N-dimethylnitrosamine. The ethanol-induced increase in microsomal oxidation of these substrates was prevented by ligands and inhibitors of P4502E1 as well as anti-human P4502E1 IgG and corresponded to the increase in P4502E1 content. Several other agents including pyrazole, 4-methylpyrazole, isoniazid, pyridine, and DMSO also increased the content of P4502E1 in this cell line but not oxidation of substrates, presumably a reflection of remaining tightly bound to the active site of P4502E1. Slot blot analysis indicated that ethanol addition did not increase P4502E1 mRNA levels. These results indicate that ethanol can increase the content of P4502E1 as well as catalytic oxidation of substrates dependent on P4502E1 in this experimental model, perhaps by stabilization of the protein against degradation.

Blotting, Western↗

Evidence that cytochrome P450 2E1 is involved in the (omega-1)-hydroxylation of lauric acid in rat liver microsomes.

The present study examined changes in hepatic CYP2E1 content and (omega-1)-hydroxylation of lauric acid in rats treated with pyridine, pyrazole, acetone, ethanol and 3-methylcholanthrene. The (omega-1)-hydroxylase activity was strongly correlated with chlorzoxazone 6-hydroxylation (r = 0.76) and 4-nitrophenol-hydroxylase (r = 0.91). Both these activities are carried out by CYP2E1. (omega-1) hydroxylase activity was inhibited by ethanol (Ki = 3.5 mM), dimethylsulfoxide and diethyldithiocarbamate. Furthermore, polyclonal antibody directed against rat CYP2E1 inhibited (omega-1)-hydroxylation by more than 90% while it had no effect on the omega-hydroxylation. These results suggest that the (omega-1)-hydroxylation of lauric acid is mediated principally by the CYP2E1 enzyme in rat liver microsomes.

Acetone↗

Preclinical toxicity evaluation of tepoxalin, a dual inhibitor of cyclooxygenase and 5-lipoxygenase, in Sprague-Dawley rats and beagle dogs.

Tepoxalin [5- (4-chlorophenyl)-N-hydroxy-1-(4-methoxyphenyl)-N-methyl-1H-pyrazole -3-propanamide] is an orally active anti-inflammatory agent, which inhibits both cyclooxygenase and 5-lipoxygenase activities. The oral toxicity of tepoxalin was evaluated in 1- and 6-month rat (up to 50 mg/kg/day) and dog (up to 150 mg/kg bid) studies. In rats, increased liver weight, centrilobular hypertrophy, and hepatic necrosis were observed at dosages >/=20 mg/kg/day. Renal changes indicative of analgesic nephropathy syndrome (i.e., papillary edema or necrosis, cortical tubular dilatation) were seen at >/=15 mg/kg. In rats treated for 1 month, these hepatic and renal effects were largely reversible after a 1-month recovery period. Gastrointestinal erosions and ulcers were seen in female rats given 40 mg/kg/day for 6 months. Changes in clinical pathology parameters included decreases in red blood cell count, hemoglobin, and hematocrit mean values; elevation in platelet counts; and an increase in prothrombin and activated partial thromboplastin times. Mild increases in alanine aminotransferase, aspartate aminotransferase, and cholesterol were also noted in rats. Decreased erythrocyte parameters, increased leukocyte counts, and decreased total protein, albumin, and/or calcium were noted in some dogs in the 300 mg/kg/day group following 6 months of dosing. Small pyloric ulcerations were seen at 100 and 300 mg/kg/day dosages for up to 6 months. In both rats and dogs, no accumulation of tepoxalin or its carboxylic acid metabolite was detected in plasma following multiple dosing over a range of 5 to 50 mg/kg/day for rats and 20 to 300 mg/kg/day for dogs. Plasma concentrations of the carboxylic acid metabolite were severalfold higher than those of the parent compound. The no-effect dosages in rats (5 mg/kg/day) and dogs (20 mg/kg/day) were approximately one and six times the ED50 (3.5 mg/kg), respectively, for inhibition of inflammatory effects in the adjuvant arthritic rat without gastric mucosal damage. In terms of severity, the relative lack of gastrointestinal side effects, within the estimated therapeutic dose range, distinguishes tepoxalin from most marketed anti-inflammatory drugs.

Absorption↗

The catalytic triad in Drosophila alcohol dehydrogenase: pH, temperature and molecular modelling studies.

Drosophila alcohol dehydrogenase belongs to the short chain dehydrogenase/reductase (SDR) family which lack metal ions in their active site. In this family, it appears that the three amino acid residues, Ser138, Tyr151 and Lys155 have a similar function as the catalytic zinc in medium chain dehydrogenases. The present work has been performed in order to obtain information about the function of these residues. To obtain this goal, the pH and temperature dependence of various kinetic coefficients of the alcohol dehydrogenase from Drosophila lebanonensis was studied and three-dimensional models of the ternary enzyme-coenzyme-substrate complexes were created from the X-ray crystal coordinates of the D. lebanonensis ADH complexed with either NAD(+) or the NAD(+)-3-pentanone adduct. The kon velocity for ethanol and the ethanol competitive inhibitor pyrazole increased with pH and was regulated through the ionization of a single group in the binary enzyme-NAD(+) complex, with a DeltaHion value of 74(+/-4) kJ/mol (18(+/-1) kcal/mol). Based on this result and the constructed three-dimensional models of the enzyme, the most likely candidate for this catalytic residue is Ser138. The present kinetic study indicates that the role of Lys155 is to lower the pKa values of both Tyr151 and Ser138 already in the free enzyme. In the binary enzyme-NAD(+) complex, the positive charge of the nicotinamide ring in the coenzyme further lowers the pKa values and generates a strong base in the two negatively charged residues Ser138 and Tyr151. With the OH group of an alcohol close to the Ser138 residue, an alcoholate anion is formed in the ternary enzyme NAD(+) alcohol transition state complex. In the catalytic triad, along with their effect on Ser138, both Lys155 and Tyr151 also appear to bind and orient the oxidized coenzyme.

Alcohol Dehydrogenase↗

The effect of chronic ethanol consumption on pathways of ethanol metabolism.

The contribution of the known hepatic pathways for the disposal of ethanol was studied utilizing two different experimental designs. In vitro studies were carried out in hepatocytes isolated from rats fed Purina chow. Rates of ethanol oxidation in these preparations increased with increasing levels of ethanol (10-50 mM). After inhibition of alcohol dehydrogenase (ADH) by pyrazole (2 mM) and the catalase by azide (1 mM) approximately 25% of the ethanol oxidizing activity remained. The residual activity was also dependent upon ethanol concentration and the apparent Km was 13 mM. Hepatocytes from ethanol-fed rats exhibited rates of ethanol oxidation which were also dependent upon the concentration of ethanol. The rates were higher in the hepatocytes from the ethanol-fed rats than in the controls. The addition of inhibitors of ADH and catalase lowered the rates, but abolished neither the differences nor the concentration dependency. In the in vivo studies, ethanol elimination rates were measured in alcohol-fed and control baboons by using a constant ethanol infusion to maintain blood ethanol at three different levels: 5, 10 or 50 mM. Ethanol elimination rate was accelerated with increasing concentration, particularly in alcohol-fed baboons. These observations indicate that a pathway other than the low Km alcohol dehydrogenase participates in alcohol oxidation and is responsible in part for the adaptive increase in ethanol metabolism associated with chronic ethanol consumption.

Alcoholism↗

Fipronil: environmental fate, ecotoxicology, and human health concerns.

Fipronil is a highly effective, broad-spectrum insecticide with potential value for the control of a wide range of crop, public hygiene, amenity, and veterinary pests. It can generally be applied at low to very low dose rates to achieve effective pest control. Application rates vary between 0.6 and 200 g a.i./ha, depending on the target pest and formulation. It belongs to the phenyl pyrazole or fiprole group of chemicals and is a potent disrupter of the insect central nervous system via interference with the gamma-aminobutyric acid (GABA-) regulated chloride channel. Fipronil degrades slowly on vegetation and relatively slowly in soil and in water, with a half-life ranging between 36 hr and 7.3 mon depending on substrate and conditions. It is relatively immobile in soil and has low potential to leach into groundwater. One of its main degradation products, fipronil desulfinyl, is generally more toxic than the parent compound and is very persistent. There is evidence that fipronil and some of its degradates may bioaccumulate, particularly in fish. Further investigation on bioaccumulation is warranted, especially for the desulfinyl degradate. The suitability of fipronil for use in IPM must be evaluated on a case-by-case basis. In certain situations, fipronil may disrupt natural enemy populations, depending on the groups and species involved and the timing of application. The indications are that fipronil may be incompatible with locust IPM; hence, this possibility requires further urgent investigation. It is very highly toxic to termites and has severe and long-lasting negative impacts on termite populations. It thus presents a long-term risk to nutrient cycling and soil fertility where termites are "beneficial" key species in these ecological processes. Its toxicity to termites also increases the risk to the ecology of habitats in which termites are a dominant group, due to their importance as a food source to many higher animals. This risk has been demonstrated in Madagascar, where two endemic species of lizard and an endemic mammal decline in abundance because of their food chain link to termites. Fipronil is highly toxic to bees (LD50 = 0.004 microgram/bee), lizards [LD50 for Acanthodactylus dumerili (Lacertidae) is 30 micrograms a.i./g bw], and gallinaceous birds (LD50 = 11.3 mg/kg for Northern bobwhite quail), but shows low toxicity to waterfowl (LD50 > 2150 mg/kg for mallard duck). It is moderately toxic to laboratory mammals by oral exposure (LD50 = 97 mg/kg for rats; LD50 = 91 mg/kg for mice). Technical fipronil is in toxicity categories II and III, depending on route of administration, and is classed as a nonsensitizer. There are indications of carcinogenic action in rats at 300 ppm, but it is not carcinogenic to female mice at doses of 30 ppm. The acute toxicity of fipronil varies widely even in animals within the same taxonomic groups. Thus, toxicological findings from results on standard test animals are not necessarily applicable to animals in the wild. Testing on local species seems particularly important in determining the suitability of fipronil-based products for registration in different countries or habitats and the potential associated risk to nontarget wildlife. Risk assessment predictions have shown that some fipronil formulations present a risk to endangered bird, fish, and aquatic and marine invertebrates. Great care should thus be taken in using these formulations where they may impact any of these endangered wildlife groups. Work in Madagascar has highlighted field evidence of this risk. The dose levels at which fipronil produces thyroid cancer in rats are very high and are unlikely to occur under normal conditions of use. There is also dispute as to whether this is relevant to human health risk. However, as fipronil is a relatively new insecticide that has not been in use for long enough to evaluate the risk it may pose to human health, from data on human exposure to the product, a precautionary approach may be warranted. The use of some fipronil-based products on domestic animals is not recommended where handlers spend significant amounts of time grooming or handling treated animals. In general, it would appear unwise to use fipronil-based insecticides without accompanying environmental and human health monitoring, in situations, regions, or countries where it has not been used before, and where its use may lead to its introduction into the wider environment or bring it into contact with people. Further work is needed on the impacts of fipronil on nontarget vertebrate fauna (amphibians, reptiles, birds, and mammals) in the field before the risk to wildlife from this insecticide can be adequately validated. Further field study of the effects of fipronil on the nutrient cycling and soil water-infiltration activities of beneficial termites is required to assess the ecological impacts of the known toxicity of fipronil to these insects.

Animals↗

Investigations on the mechanism of liver tumour induction by peroxisome proliferators.

Further understanding of the mechanism by which peroxisome proliferators induce liver tumours is essential to assessing the risks of such compounds to exposed humans. To this end the effects of nafenopin upon the liver have been investigated. Nafenopin was shown to induce certain drug metabolising enzymes, but sub-cellular fractions from induced animals did not form reactive metabolites which could be detected as mutagens. Nafenopin treatment slightly increased the rate of alkaline elution of hepatic nuclear DNA from polycarbonate filters. However, simultaneous administration of sodium glycolate to stimulate H2O2 production or pyrazole to inhibit catalase activity had no further effects. These findings demonstrate that nafenopin is not activated to a mutagen and argue against the hypothesis that indirect DNA damage as a result of excess H2O2 production is responsible for tumour induction.

7-Alkoxycoumarin O-Dealkylase↗

Inhibition of long-term potentiation in rat hippocampal slices by anandamide and WIN55212-2: reversal by SR141716 A, a selective antagonist of CB1 cannabinoid receptors.

It has been reported previously that delta 9-tetrahydrocannabinol and the synthetic cannabinoid agonist HU-210 [(--)-11-OH-delta 8-dimethylheptyl tetrahydrocannabinol] prevent long-term potentiation (LTP) induction in rat hippocampal slices. In this study we confirm that both WIN55212-2 ¿R-(+)-(2,3-dihydro-5-methyl-3-[¿4-morpholinyl¿ methyl] pyrol [1,2,3-de]-1,4-benzoxazin-6-yl) (1-naphtalenyl) methanone monomethanesulphonate¿ (3 and 10 microM), another synthetic cannabinoid agonist, and anandamide (10 microM), considered to be the endogenous ligand of cannabinoid receptors, inhibit LTP formation in the Schaffer collateral-CA1 field complex. In addition, we show that SR141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4- methyl-1H-pyrazole-3-carboxamide hydrochloride] at 0.1-10 microM, a potent and selective antagonist of CB1 cannabinoid receptors, concentration-dependently reversed the inhibition of LTP induced by both WIN55212-2 and anandamide. These data indicate that cannabinoid receptor agonists inhibit hippocampal LTP formation through CB1 receptor activation and that anandamide could be a candidate for an endogenous neuromessenger involved in memory processes.

Animals↗