Cholinesterase activity in Gammarus pulex (Crustacea Amphipoda): characterization and effects of chlorpyrifos

The aim of this study was to characterize cholinesterase (ChE) activity in Gammarus pulex, an abundant and ecologically relevant species of the European stream environment. Biochemical and pharmacological properties were tested using different substrates (acetylthiocholine iodide, propionylthiocholine iodide and butyrylthiocholine iodide) and selective inhibitors (eserine sulfate, BW284c51 and iso-OMPA). In a second part, the in vitro and in vivo effects of a widely used organophosphorous pesticide, chlorpyrifos, on ChE activity were investigated. The results suggest that G. pulex possess only one ChE which displays the typical properties of an acetylcholinesterase, since: (1) it hydrolyses to the substrate acetylthiocholine at a higher rate than all other tested substrates and (2) it is highly sensitive to eserine sulphate and BW284c51, but not to iso-OMPA. In vitro and in vivo inhibitions were observed for highly different contamination levels, which suggests that bioaccumulation and biotransformation mechanisms are involved. In vivo AChE inhibition was observed at realistic environmental concentrations, with lethal effects appearing at inhibitions higher than 50%. The results of this study show the value of G. pulex as a sentinel organism for environmental assessment.     

Behavioral Changes and Acute Toxicity to the Freshwater Shrimp Atyaephyra desmaresti Millet (Decapoda: Natantia) from Exposure to Acid Mine Drainage

Short-term 48 h laboratory bioassays with water from an acid mine drainage (AMD: pH 3.3, 4.4, 5.0, 5.5, 6.4, control) and water from an arsenic containing reservoir were performed with the freshwater shrimp Atyaephyra desmaresti Millet, validated in situ and compared to acidified control water (ACID). Behaviour, mortality and time to death were monitored with the Multispecies Freshwater Biomonitor (MFB). The shrimps had equal 24 h-LC₅₀s at pH 4 in AMD and ACID. However, after 48 h AMD proved more toxic (48 h-LC₅₀ at pH 5.2) than ACID (48 h-LC₅₀ at pH 4.5). Stress behaviour in AMD consisted at pH ≤ 6.4 of a pH-dependent decrease in activity, with disappearance of circadian rhythmicity, and at pH 4.4 a clear increase of ventilation. At pH 5.5 bioaccumulation of metals was higher and locomotion lower than at pH 5.0. In ACID, only at pH ≤ 4.4 locomotion became abated and arythmic. Locomotion in the field was equal or higher compared to the laboratory, whereas the ventilation was higher in the laboratory. A. desmaresti is a valuable species to be used in short term behavioural bioassays of AMD in Europe.     

Studies on zinc sulphate microcapsules: (III) in vivo evaluation

Zinc sulphate capsules and syrup were prepared as conventional dosage forms and in vivo experiments were performed for both on the conventional dosage forms as well as on microcapsules. Blood samples were taken from healthy volunteers at 1, 2, 3, 4 and 5 hours and AUC3 and AUC5 were calculated by trapezoidal rule. Relative bioavailability of zinc was calculated and the 5th hour relative bioavailability difference was found to be significant. The results show that zinc sulphate in microcapsule form was able to prolong the action.     

The effects of copper, zinc, mercury, and cadmium on rabbit erythrocytic porphobilinogen synthase in vivo

The interactions of Zn2+, Hg2+, Cd2+, and Cu2+ with normal and lead-intoxicated rabbit erythrocytic porphobilinogen synthase (PBG-S) were studied in vivo. The results obtained show that each divalent ion has a characteristic effect on the pH-activity relationship of PBG-S. The effects for a given ion were both pH and concentration dependent and were similar to those obtained in vitro. Lead was found to be the most potent effector in vivo.     

The effects of trinitrotoluene toxicity on zinc and copper metabolism

Wistar rats were exposed to trinitrotoluene (TNT, 200 mg/kg/d x 6 d/wk) for 6 weeks, and a 2-week recovery period was continued after treatment. After initiation of TNT exposure, zinc and copper concentrations in rat testes, liver and serum, and serum caeruloplasmin activity were assayed every 2 weeks. Testicular size and zinc concentrations were decreased throughout the exposure and recovery periods. After 6 weeks of TNT exposure, rat testicular copper and serum zinc concentrations were significantly decreased and increased, respectively. The activities of caeruloplasmin were decreased after 4 and 6 weeks of exposure. Liver zinc and copper concentrations and serum copper concentrations had not changed significantly in rats throughout the exposure and recovery periods. Copper concentration in hair samples from TNT-exposed workers was lower than that in control workers, but zinc content was significantly increased compared with that in control workers.     

The in vivo effects of manganese and zinc on δ-aminolevulinic acid dehydratase activity inhibited by lead

The in vivo effects of zinc (Zn) and manganese (Mn) on the activity of δ-aminolevulinic acid dehydratase (5-aminolevulinate hydrolyase, ALAD, EC 4.2.1.24) in blood with and without inhibition by lead and on the functions of the liver and kidney were studied in rabbits. Administration of Zn or Mn alone caused little change in any of the 29 parameters tested at concentrations of 5, 10, 25 or 50 μmol element/kg body weight. Administration of 5 μmol lead/kg inhibited ALAD activity in blood, but it did not induce any marked effects on the indices of hepatic and renal functions. In simultaneous i.v. injection with lead and manganese, both at a dose of 5 μmol/kg, however, the recovery process of ALAD activity was delayed. Three out of 29 parameters tested, β-lipoprotein, triglyceride and aldolase, increased significantly one day after simultaneous injection with lead and manganese. Equimolar zinc did not show either an antagonistic or cooperative effect on these three parameters and the ALAD activity in blood.     

锌、铜对大鼠红细胞超氧化物歧化酶活性的影响

为探讨锌、铜在抗衰老中的作用及观察老龄大鼠的抗氧化能力的变化,我们测定了给予锌、铜的成年与老年大鼠红细胞超氧化物歧化酶(SOD)活性。结果表明,给锌、铜组红细胞SOD活性比对照组明显增高(P<0.01),但给铜组较给锌组高(P<0.01)。老龄大鼠抗氧化能力是降低的。这说明锌、铜可以提高机体的抗氧化能力。     

Synthesis and evaluation of antibacterial activity of water-soluble copper, nickel and zinc tetra (n-carbonylacrylic) aminephthalocyanines

Water-soluble copper, nickel, and zinc tetra (n-carbonylacrylic) aminephthalocyanines were synthesized with an aim to evaluate their role as therapeutic materials. They were characterized by UV–Vis spectrophotometer, infra-red spectra, elemental analysis, nuclear magnetic resonance, and thermogravimetry. Efforts were made to optimize the synthesis of intermediates and final products. Photoluminescence studies were carried out on these compounds. The investigation of antibacterial activity of these aqueous soluble phthalocyanines was carried out on Staphylococcus citreus, Serratia marcescens, Proteus vulgaris, Bacillus subtilis, and Pseudomonas fluorescence, showed good inhibitory activity.     

Cholinesterase activity in gilthead seabream (Sparus aurata) larvae: Characterization and sensitivity to the organophosphate azinphosmethyl

Assessment of cholinesterase (ChE) inhibition is widely used as a specific biomarker for evaluating the exposure and effects of non-target organisms to anticholinesterase agents. Cholinesterase and carboxylesterase activities have been measured in larvae of gilthead seabream, Sparus aurata, during the endogenous feeding stage, and ChE was characterized with the aid of diagnostic substrates and inhibitors. The results of the present study showed that whole-body ChE of yolk-sac seabream larvae possesses typical properties of acetylcholinesterase (AChE) with a apparent affinity constant (K(m)) of 0.163+/-0.008 mM and a maximum velocity (V(max)) of 332.7+/-2.8 nmol/min/mg protein. Moreover, sensibility of this enzyme was investigated using the organophosphorus insecticide azinphosmethyl. Static-renewal toxicity tests were conducted over 72 h and larval survival and AChE inhibition were recorded. Mean mortality of seabream larvae increased with increasing concentrations of azinphosmethyl and exposure duration. The estimated 72-h LC50 value to azinphosmethyl was 4.59 microg/l (95% CI=0.46-8.71 microg/l) and inhibition of ChE activity gave an IC50 of 3.04 microg/l (95% CI=2.73-3.31 microg/l). Larvae exposed to azinphosmethyl for 72h showed a 70% inhibition of the whole-body acetylcholinesterase activity at approximately the LC50. In conclusion, the results of the present study suggested that monitoring ChE activity is a valuable tool indicating OP exposure in S. aurata larvae and that acetylthiocholine is the most appropriate substrate for assessing ChE inhibition in this early-life stage of the fish.     

Crystallochemistry of copper (II) and zinc (II) chelates by nonsteroidal antiinflammatory drugs

Copper(II) and zinc(II) chelates by some non-steroidal antiinflammatory drugs NSAIDs (niflumic acid, indomethacin) and 3,5-diisopropylsalicylic acid (DIPS) were characterized by single X-ray diffraction methods. Copper(II) complexes by these two types of chelates are binuclear compounds, with Cu(2)(DIPS)(4)L(2) or Cu(2)(AINS)(4)L(2) formula (L=axial non-NSAID ligand such as diethylether, dimethylsulfoxide DMSO). In zinc(II) complex by DIPS, the metal ion is tetrahedrally coordinated and the corresponding compound is mononuclear with Zn(DIPS)(2)(DMSO)(2) formula. These copper(II) and zinc(II) complexes were found to be more active than their parent drugs from the antiinflammatory and anticonvulsant properties. It was pointed out that the Cu(2)(DIPS)(4)L(2) complexes (L=diethylether, N,N-dimethylformamide) exhibited no rotorod toxicity when examined for anticonvulsant activity using the seizure produced by maximal electroshock, following oral administration to rats.     

Effects of copper,zinc and paraquat on acetylcholinesterase activity in carp (Cyprinus carpio L.)

The effects of pH and substrate concentration on acetylcholinesterase (AChE) activity were studied in serum, brain, heart and muscle of common carp (Cyprinus carpio L.). Effects of zinc chloride, paraquat (1,1′-dimethyl-4,4′-dipyridinium dichloride) and copper sulphate from in vivo and in vitro exposure were studied on these AChE activities. Zinc chloride did not decrease AChE activity in any of the organs studied in vivo or in vitro. In contrast, paraquat, competitively, and copper sulphate, in a mixed way, inhibited acetylcholinesterase activity. Inhibition of AChE in fish exposed to these two pesticides may serve as an indicator of hazard due to application of these chemicals in the natural environment.     

Rufloxacin (MF-934): in vitro and in vivo antibacterial activity

Rufloxacin (MF-934) is a new quinolone which shows in vitro antibacterial activity against E. coli, Salmonella, Klebsiella, Proteus and Staphylococcus spp. Lower in vitro activity was observed with Pseudomonas, Serratia, Enterobacter and the streptococci group D. The antimicrobial activity of MF-934 in vitro is higher than that of nalidixic acid but lower than that of ciprofloxacin, ofloxacin, pefloxacin or norfloxacin. The protective effects of MF-934 in systemic infections in mice are lower than those of ciprofloxacin and ofloxacin. In respiratory infections in rats and in subcutaneous infections in guinea-pigs, the protective effects of MF-934 are of the same order as ciprofloxacin and ofloxacin. This is probably due to the pharmacokinetic properties of MF-934, i.e. the long half-life and tissue concentrations higher than plasma levels.     

Effects of lead,copper, and zinc on the rat's lactate dehydrogenase in vivo and in vitro

Following subacute intoxication of rats with Pb-, Cu-, and Zn-salts (separately or in mixture) for 5 weeks, the chelating agent D-penicillamine was administered for 3 weeks. In the course of the 3-month experiment, lactate dehydrogenase (LDH) was estimated in serum and in cytoplasmic fraction of the kidney. Pb²⁺ treatment resulted in an increase of LDH activity, Cu²⁺ in a slight decrease, whereas Zn²⁺ had no effect, respectively. Mixture of these metals caused a significant rise in the enzymatic activity. Seven weeks after the stoppage of the administration of toxic substances, altered LDH activity, both in serum and in kidney returned to normal. D-penicillamine treatment was found to accelerate a restoration of the enzyme activity.In the experiments in vitro, Cu²⁺ inhibited significantly the kidney LDH activity, Pb²⁺ and Zn²⁺ being 100- and 400-times less efficient, respectively. Cu²⁺ inhibition was reversed by D-penicillamine, whereas inhibition of LDH by Zn²⁺ or Pb²⁺ was irreversible.     

Functional effects of corticosterone on 5-HT(1A) and 5-HT(1B) receptor activity in rat brain: in vivo microdialysis studies

Glucocorticoid hormones are known to be elevated in depression, and to interact with serotonin 5-HT(1A) receptors at both the presynaptic and postsynaptic levels. Since one of the presumed mechanisms of action of antidepressant drugs is induction of changes in sensitivity of 5-HT(1A) and also 5-HT(1B) receptors, the effects of repeated administration of corticosterone (50 mg/kg s.c. b.i.d. for 10 days) on activities of these receptors were determined using in vivo microdialysis in freely moving rats. Presynaptic 5-HT(1A) receptor activity, as measured by the effect of a challenge dose (0.2 mg/kg s.c.) of the 5-HT(1A) agonist 8-hydroxy-2 (di-n-propylamino) tetralin (8-OH-DPAT) to reduce 5-HT levels in the hypothalamus, was not affected by corticosterone administration. Presynaptic 5-HT(1B) receptor activity, as measured by the effect of the 5-HT(1B) receptor antagonist (N-[4-methoxy-3-(4-methyl-1-piperizinyl)phenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazole-3-yl)[1,1'-biphenyl]-carboxamide (GR 127935) (5 mg/kg s.c.) to increase 5-HT levels, was increased in hypothalamus but not hippocampus of corticosterone-treated rats. Postsynaptic 5-HT(1A) receptor activity, as measured by the effect of 8-OH-DPAT to increase cyclic AMP levels in the hippocampus, was not affected by corticosterone administration. The decrease in presynaptic 5-HT(1B) receptor activity after chronic administration of antidepressant drugs complements the increases in 5-HT(1B) receptor number observed in animal models of depression.     

Comparison of the radiorecovery activity of copper(II)2(3,5-diisopropylsalicylate)4 and copper(II) (chloride)2

Copper(II)₂(3,5-diisopropylsalicylate)₄ [Cu(II)₂(3,5-DIPS)₄] and copper(II)(chloride)₂ [Cu(II)Cl₂ were used to treat γ-irradiated female C57BL/6 mice after irradiation at levels LD50/30 to compare their efficacy in facilitating recovery from radiation-induced systemic inflammatory disease accompanied by loss of body mass and in increasing survival of irradiated mice. Doses of 5, 10 or 20 μmol Cu(II)Cl₂ or 5, 10 or 20 μmol [Cu(II)₂(3,5-DIPS)₄]/kg were administered subcutaneously 3 h after LD_50/30 irradiation and body mass and survival determined throughout the 30-day post-irradiation period compared with controls. Treatment with Cu(II)₂(3,5-DIPS)₄ or Cu(II)Cl₂ facilitated recovery of radiation-induced systemic inflammatory disease, recovery of body mass, and increased survival. Treatment with 5, 10 or 20 (μmol [Cu(II)₂(3,5-DIPS)₄]/kg produced a 44%, 67% or 44% increase in survival, respectively, compared with the vehicle-treated control group. Treatment with 5,10 or 20 μmol Cu(II)Cl₂/kg produced a 7%, 21% or 29% increase in survival, respectively, compared with the vehicle-treated control group. The recovery of radiation-induced loss in body mass and an increase in survival document that both Cu(II)₂(3,5-DIPS)₄ and Cu(II)Cl₂ are effective radiorecovery agents. In addition, Cu(II)₂(3,5-DIPS)₄ is a more effective radiorecovery agent than Cu(II)Cl₂.     

Cholinesterase activity:muscarinic receptor ratios in canine and feline brains

In response to findings that brain cholinesterase (ChE): muscarinic cholinergic receptor density (mChR) ratio is highly conserved between bird species (I), this study investigated the conservation of brain ChE:mChR between 2 mammalian species. Whole brains of 7 normal greyhounds, 14 normal non-greyhound dogs, and 8 normal cats, were assayed for ChE, mChR and protein. The same assays were made on cerebrum, cerebellum and brainstem from the greyhounds and 6 of the non-greyhound dogs. The Ellman assay was used to measure serum ChE activity, mChR were labelled with tritiated quinuclidinyl benzilate, and protein was measured by Lowry assay. Brain ChE:mChR was not highly conserved between the species. There was a statistically significant difference in brain mChR between greyhounds and non-greyhounds but this did not translate to a statistically significant difference in mean ChE:mChR. Measurement of ChE:mChR is unlikely to be useful in endangered carnivores for which normal ChE levels are unknown, because it cannot be assumed that endangered species would have the same ChE:mChR as their domestic relatives.     

In vivo effects of black cohosh and genistein on estrogenic activity and lipid peroxidation in Japanese Medaka (Oryzias latipes)

This study was designed to assay the estrogenic activities and the antioxidant potential of ethanol extracts from the herbal dietary supplement black cohosh (Cimicifuga racemosa) relative to the natural phytoestrogen genistein. The in vivo mechanisms of action of these two natural products have not been completely elucidated, and Japanese medaka (Oryzias latipes) provides a useful organism for initial in vivo screening of natural products. While both genistein and estradiol altered ovarian and testicular steroid release and decreased circulating testosterone levels in males, neither black cohosh total extract (75-30,000 ng/fish), cimiracemoside A, 25-O-methyl-cimigenoside, actein, nor 26-deoxy-actein caused any differences in estrogenic activity compared to control fish. To assess antioxidant potential, animals were treated with natural products then challenged with 2-acetylaminofluorene (2-AAF) to induce lipid peroxidation (LPO) in the liver. Neither the total ethanol extracts from black cohosh nor its individual components showed an inhibitory effect in 2-AAF induced LPO. However, genistein manifested potent antioxidative activity in the LPO assay, with similar potency to a high dose of á-tocopherol. In contrast to genistein, black cohosh did not exhibit traditional estrogenic effects nor significant in vivo anti-oxidant potential in this fish model system.     

Anticonvulsive effects of carbenoxolone on penicillin-induced epileptiform activity: an in vivo study

Epilepsy is an important problem in neurological disorders. Recent studies claimed that gap junctions have a critical role in epileptic neuronal events. The aim of present study is to investigate the effects of gap junction blocker carbenoxolone on penicillin-induced experimental epilepsy. For this purpose, 4-month-old male Wistar rats were used in the present study. Permanent screw electrodes allowing EEG monitoring from conscious animals and permanent cannula providing the administration of the substances to the brain ventricle were placed into the cranium of rats under general anesthesia. At the end of the postoperative recovery period, epileptiform activity was generated by injecting 300 IU crystallized penicillin through the ventricular cannula. Epileptiform activity monitored from a digital recording system, when it reached its maximum intensity, carbenoxolone (100, 200, 500 nmol) was applied in the same way with penicillin. Effects of carbenoxolone on epileptiform activity were assessed by both electrophysiological and behavioral analysis. Carbenoxolone suppressed epileptiform activity by decreasing the amplitude and frequency of epileptiform spikes and by attenuating the epileptiform behavior. The results of this study suggest that the blockade of electrical synapses may contribute to the prevention and amelioration of epileptic activity.