Evidence for nonacetylcholinesterase targets of organophosphorus nerve agent: supersensitivity of acetylcholinesterase knockout mouse to VX lethality.
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The possibility that organophosphate toxicity is due to inhibition of targets other than acetylcholinesterase (AChE, EC 3.1.1.7) was examined in AChE knockout mice. Mice (34-55 days old) were grouped for this study, after it was determined that AChE, butyrylcholinesterase (BChE), and carboxylesterase activities had reached stable values by this age. Mice with 0, 50, or 100% AChE activity were treated subcutaneously with the nerve agent VX. The LD50 for VX was 10 to 12 microg/kg in AChE-/-, 17 microg/kg in AChE+/-, and 24 microg/kg in AChE+/+ mice. The same cholinergic signs of toxicity were present in AChE-/- mice as in wild-type mice, even though AChE-/- mice have no AChE whose inhibition could lead to cholinergic signs. Wild-type mice, but not AChE-/- mice, were protected by pretreatment with atropine. Tissues were extracted from VX-treated and untreated animals and tested for AChE, BChE, and acylpeptide hydrolase activity. VX treatment inhibited 50% of the AChE activity in brain and muscle of AChE+/+ and +/- mice, 50% of the BChE activity in all three AChE genotypes, but did not significantly inhibit acylpeptide hydrolase activity. It was concluded that the toxicity of VX must be attributed to inhibition of nonacetylcholinesterase targets in the AChE-/- mouse. Organophosphorus ester toxicity in wild-type mice is probably due to inhibition or binding to several proteins, only one of which is AChE. (+info)
New thiocholine ester substrates for the assay of human serum cholinesterase.
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BACKGROUND: Several thiocholine alkanoyl esters were newly synthesized and explored as substrates for the assay of human serum cholinesterase after being subjected to the Ellman reaction (Arch Biochem Biophys 1958;74:443-50 and Arch Biochem Biophys 1959;82:70-7). METHODS: We synthesized thiocholine ester iodides by the method of Renshow et al. (J Am Chem Soc 1938;60:1765-70). We examined solubility in H(2)O, substrate specificity serum for cholinesterase, (spontaneous) self-hydrolysis, storage stability, and reaction conditions for measurement of the activity of the enzyme. RESULTS: Isobutyryl and cyclohexane-carboxyl esters showed the best efficiency for the specific and stable assay of human serum cholinesterase. Aqueous solubility of each was >10 mmol/L, and the reactivity with acetylcholinesterase was negligible. For isobutyryl and cyclohexane-carboxyl esters, respectively, spontaneous hydrolysis in the aqueous phase was approximately 1/25 and approximately 1/175 slower than the enzymatic hydrolysis, and assays with these substrates were linear to 1800 and 3000 U/L, respectively. The K(m) values of these acylthiocholines with human cholinesterase were almost equivalent (6.9 x 10(-3) mmol/L). The substrates were stable in aqueous solution and in the solid state as the iodides for at least 5 years at 5 degrees C. CONCLUSIONS: The isobutyrate and cyclohexane-carboxylate of thiocholine are suitable for the specific assay of human serum cholinesterase. (+info)
Effects of salinity on aldicarb toxicity in juvenile rainbow trout (Oncorhynchus mykiss) and striped bass (Morone saxatilis x chrysops).
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Fluctuations in several environmental variables, such as salinity, can influence the interactions between organisms and pollutants in aquatic organisms, and, therefore, affect the toxicity of xenobiotics. In this study, after 2 species of fish, rainbow trout (Oncorhynchus mykiss) and hybrid striped bass (Morone saxatilis x chrysops) were acclimated to 4 salinity regimens of 1.5, 7, 14, and 21 ppt for 1 week and then exposed to 0.5 mg/l aldicarb. Mortality, brain, and muscle cholinesterase levels were measured after 96 h. Rates of (14)C-aldicarb sulfoxide formation were determined in kidney (trout only), liver, and gill microsomes from each species acclimated to the 4 salinity regimens. Salinity significantly enhanced aldicarb toxicity, cholinesterase inhibition, and (14)C-aldicarb sulfoxide formation in rainbow trout but not in striped bass. In vitro incubations with (14)C-aldicarb and the cytochrome P450 (CYP) inhibitor, N-benzylimidazole, did not significantly alter aldicarb sulfoxide formation in tissue microsomes from either species of fish, indicating CYP did not contribute to aldicarb sulfoxidation. Salinity increased flavin-containing monooxygenase (FMO) mRNA expression and catalytic activities in microsomes of liver, gill, and kidney of rainbow trout, which was consistent with the salinity-induced enhancement of aldicarb toxicity. Salinity did not alter FMO mRNA expression and catalytic activities in striped bass, which was also consistent with the lack of an effect of salinity on aldicarb toxicity in this species. These results suggest that salinity-mediated enhancement of aldicarb toxicity is species-dependent, and at least partially due to the salinity-related upregulation of FMOs, which, in turn, increases the bioactivation of aldicarb to aldicarb sulfoxide, which is a more potent inhibitor of cholinesterase than aldicarb. (+info)
Cholinesterase inhibition by aluminium phosphide poisoning in rats and effects of atropine and pralidoxime chloride.
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AIM: To investigate the cholinesterase inhibition and effect of atropine and pralidoxime (PAM) treatment on the survival time in the rat model of aluminium phosphide (AlP) poisoning. METHODS: The rats were treated with AlP (10 mg/kg; 5.55 x LD50; ig) and the survival time was noted. The effect of atropine (1 mg/kg, ip) and PAM (5 mg/kg, ip) was noted on the above. Atropine and PAM were administered 5 min after AlP. Plasma cholinesterase levels were measured spectrophotometrically in the control and AlP treated rats 30 min after administration. RESULTS: Treatment with atropine and PAM increased the survival time by 2.5 fold (1.4 h+/-0.3 h vs 3.4 h+/-2.5 h, P < 0.01) in 9 out of 15 animals and resulted in total survival of the 6 remaining animals. Plasma cholinesterase levels were inhibited by 47 %, (438+/-74) U/L in AlP treated rats as compared to control (840+/-90) U/L (P < 0.01). CONCLUSION: This preliminary study concludes that AlP poisoning causes cholinesterase inhibition and responds to treatment with atropine and PAM. (+info)
Neurotoxic effect of dermally-applied chlorpyrifos and cypermethrin in Wistar rats.
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The aim of the study was to evaluate the neurotoxic effect of a dermally-applied mixture of chlorpyrifos and cypermethrin in rats based on cognitive function, activity of the blood cholinesterase and brain acetylcholinesterase, as well as histologic brain examination. Nurelle D 550 EC (500 g of chlorpyrifos and 50 g of cypermethrin) was used in the study. The application liquid was in the form of a water solution. The investigation covered eight groups of animals: six experimental groups and two control groups, of 15 rats each. Experimental groups received 5.6 mg/cm2 chlorpyrifos and 0.5 mg/cm2 cypermethrin, or 27.8 mg/cm2 chlorpyrifos and 2.7 mg/cm2 cypermethrin dermally, for one day, one week and four weeks, except for Saturdays and Sundays. The preparations examined were applied to the tail skin of rats. The animals were anaesthetized at the end of exposure period. Plasma cholinesterase and brain acetylcholinesterase activities were determined. The brain for histological examination was perfused with a solution of methanol, formalin and glacial acetic acid, and the sections stained by the Nissel method. The behaviour of the animals was evaluated in the open field test four times: before exposure, and after one, two and four weeks of the experiment. The results of the study showed that chlorpyrifos and cypermethrin applied in a mixture caused an inhibition of cholinesterase and acetylcholinesterase activity and elicited the pycnosis of brain neurocytes. (+info)
Serum cholinesterase is an early and sensitive marker of graft-versus host-disease (GVHD) and transplant-related mortality (TRM).
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Serum cholinesterase (CHE) has been reported to be a significant indicator of liver function and prognosis in patients with cirrhosis. On the other hand, liver complications are frequent following allogeneic stem cell transplantation (HSCT). We therefore tested whether CHE was predictive of graft-versus-host disease and outcome in HSCT recipients. We studied 689 patients receiving a HSCT from an HLA-identical sibling (SIB) (n = 511), an alternative donor (n = 173) or a syngeneic twin (n = 5). Acute graft-versus-host disease (GVHD) was scored as 0-I, II, III-IV in 325 (47%), 279 (41%), and 85 patients (12%) respectively; 190 (28%) patients died of transplant-related complications (TRM). On day -7 the median CHE serum level was comparable in patients who either survived or died of TRM (5900 IU/l). On day 0, serum CHE levels were respectively 2310 and 2120 IU/l (P = NS) indicating the impact of the conditioning regimen. On day +7 after HSCT, the median level for surviving patients was 2598 IU/l vs 2309 IU/l for patients who subsequently died (P = 0.0002), on day +21 CHE levels were respectively 3348 vs 2528 IU/l (P < 0.00001), on day +50, 3575 vs 2358 IU/l (P < 0.00001) and on day +100 4193 vs 2729 IU/l (P < 0.00001). CHE levels on day +50 strongly correlated with aGVHD (3803 vs 3070 vs 1933 IU/l for patients with GVHD grade 0-I, II, and III-IV, respectively (P < 0.00001) and relapse (3569 for patients relapsing vs 3115 IU/l for patients not relapsing, P = 0.0006). In conclusion, (1) serum cholinesterase is a simple and reliable marker of acute GVHD and transplant-related complications; and (2) high CHE levels on day +50 predict relapse. If confirmed, the latter patients may be eligible for early reduction of immunosuppressive therapy. (+info)
Peripheral cholinergic pathway modulates hyperthermia induced by stress in rats exposed to open-field stress.
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Exposure to an open field is psychologically stressful and leads to an elevation in core temperature (T(c)). Methyl scopolamine (MS), a muscarinic antagonist, and pyridostigmine (PYR), a carbamate that inhibits acetylcholinesterase, do not cross the blood-brain barrier and have little effect on T(c) in resting, nonstressed animals. However, we have found that MS has an antipyretic effect on T(c) that is caused by handling and cage-switch stress. PYR should act pharmacologically to reverse the effects of MS. To this end, we assessed the effects of MS and PYR on stress-induced hyperthermia. Male Sprague-Dawley rats at 90 days of age were housed individually at an ambient temperature of 22 degrees C. T(c) and motor activity were monitored by radiotelemetry in an open-field chamber. Rats were dosed intraperitoneally at 1200 with 1.0 mg/kg MS, 0.1 mg/kg PYR, a combination of MS and PYR, or saline and placed immediately inside the open-field chamber for 60 min. Stress-induced hyperthermia was suppressed immediately by MS and enhanced by PYR. T(c) only increased by 0.3 degrees C in the MS-treated animals. The hyperthermic response in the PYR group was nearly 0.6 degrees C above that of rats dosed with saline. Coadministration of PYR and MS led to a stress-induced hyperthermia response nearly identical to that of rats injected with saline. Overall, open-field stress exacerbated the effects of MS and PYR on body T(c) and provides support for a peripheral cholinergic mechanism that mediates stress-induced hyperthermia. (+info)
Structure and innervation of extraocular muscles of Carassius.
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The extraocular muscles of the carp Carassius contain two types of muscle fibre. Large white fibres have ribbon-shaped peripheral myofibrils and triads located at the Z line. Small red fibres, rich in mitochondria, have polygonal-shaped myofibrils and triads at the A-I junction. Silver- and cholinesterase-stained preparations show that the large fibres are innervated by axons which spiral around them and exhibit intense cholinesterase activity over long distances. Axons supplying small muscle fibres run across bundles of fibres, making one contact with each fibre. By electron microscopy the nerve endings on each fibre type appear identical, both having a smooth post-junctional muscle membrane. The differences in structure and innervation pattern of the two fibre types are discussed in relation to their possible functional roles. (+info)