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1.
Toxicol Lett ; 325: 67-76, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32017982

RESUMO

Racemic 3-quinuclidinyl-α-methoxydiphenylacetate (MB266) was synthesised. Its activity at muscarinic acetylcholine receptors (mAChRs), and muscle and neuronal nicotinic acetylcholine receptors (nAChRs), was compared to that of atropine and racemic 3-quinucidinyl benzilate (QNB) using a functional assay based on agonist-induced elevation of intracellular calcium ion concentration in CN21, Chinese Hamster Ovary (CHO) and SHSY5Y human cell lines. MB266 acted as an antagonist at acetylcholine receptors, displaying 18-fold selectivity for mAChR versus nAChR (compared to the 15,200-fold selectivity observed for QNB). Thus O-methylation of QNB reduced the affinity for mAChR antagonism and increased the relative potency at both muscle and neuronal nAChRs. Despite MB266 having a pharmacological profile potentially useful for the treatment of anticholinesterase poisoning, its administration did not improve the neuromuscular function in a soman-poisoned guinea-pig diaphragm preparation pretreated with the organophosphorus nerve agent soman. Consideration should be given to exploring the potential of MB266 for possible anticonvulsant action in vitro as part of a multi-targeted ligand approach.


Assuntos
Antídotos/farmacologia , Antídotos/uso terapêutico , Inibidores da Colinesterase/envenenamento , Antagonistas Muscarínicos/farmacologia , Antagonistas Muscarínicos/uso terapêutico , Agentes Neurotóxicos/envenenamento , Antagonistas Nicotínicos/farmacologia , Antagonistas Nicotínicos/uso terapêutico , Animais , Anticonvulsivantes/química , Anticonvulsivantes/uso terapêutico , Antídotos/síntese química , Células CHO , Linhagem Celular , Cricetinae , Cricetulus , Diafragma/efeitos dos fármacos , Cobaias , Humanos , Técnicas In Vitro , Masculino , Antagonistas Muscarínicos/síntese química , Músculo Esquelético/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Antagonistas Nicotínicos/síntese química , Convulsões/induzido quimicamente , Convulsões/prevenção & controle , Soman/envenenamento
2.
Toxicol Lett ; 322: 98-103, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-31954869

RESUMO

Patients intoxicated with organophosphorous compounds may need general anaesthesia to enable mechanical ventilation or for control of epileptiform seizures. It is well known that cholinergic overstimulation attenuates the efficacy of general anaesthetics to reduce spontaneous network activity in the cortex. However, it is not clear how propofol, the most frequently used intravenous anaesthetic today, is affected. Here, we investigated the effects of cholinergic overstimulation induced by soman and acetylcholine on the ability of propofol to depress spontaneous action potential activity in organotypic cortical slices measured by extracellular voltage recordings. Cholinergic overstimulation by co-application of soman and acetylcholine (10 µM each) did not reduce the relative inhibition of propofol (1.0 µM; mean normalized action potential firing rate 0.49 ± 0.06 of control condition, p < 0.001, Wilcoxon signed rank test) but clearly reduced its efficacy. Co-application of atropine (10 nM) did not improve the efficacy. Propofol preserved its relative inhibitory potential but did not produce a degree of neuronal depression which can be expected to assure hypnosis in humans. Since a combination with atropine did not improve its efficacy, an increase in dosage will probably be necessary when propofol is used in victims suffering from organophosphorous intoxication.


Assuntos
Acetilcolina/toxicidade , Potenciais de Ação/efeitos dos fármacos , Anestésicos Intravenosos/farmacologia , Rede Nervosa/efeitos dos fármacos , Propofol/farmacologia , Soman/toxicidade , Acetilcolina/administração & dosagem , Anestesia Geral , Anestésicos Intravenosos/administração & dosagem , Animais , Camundongos Endogâmicos C57BL , Neocórtex/efeitos dos fármacos , Neocórtex/fisiologia , Rede Nervosa/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Técnicas de Cultura de Órgãos , Intoxicação por Organofosfatos , Propofol/administração & dosagem , Soman/administração & dosagem
3.
Toxicol Lett ; 321: 138-145, 2020 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-31891759

RESUMO

Organophosphorus (OP)1 nerve agents pose a severe toxicological threat, both after dissemination in military conflicts and by terrorists. Hydrolytic enzymes, which may be administered into the blood stream of victims by injection and can decompose the circulating nerve agent into non-toxic metabolites in vivo, could offer a treatment. Indeed, for the phosphotriesterase found in the bacterium Brevundimonas diminuta (BdPTE),2 engineered versions with improved catalytic efficiencies have been described; yet, their biochemical stabilities are insufficient for therapeutic use. Here, we describe the application of rational protein design to develop novel mutants of BdPTE that are less susceptible to oxidative damage. In particular, the replacement of two unpaired cysteine residues by more inert amino acids led to higher stability while maintaining high catalytic activity towards a broad spectrum of substrates, including OP pesticides and V-type nerve agents. The mutant BdPTE enzymes were produced in Escherichia coli, purified to homogeneity, and their biochemical and enzymological properties were assessed. Several candidates both revealed enhanced thermal stability and were less susceptible to oxidative stress, as demonstrated by mass spectrometry. These mutants of BdPTE may show promise for the treatment of acute intoxications by nerve agents as well as OP pesticides.


Assuntos
Antídotos/farmacologia , Proteínas de Bactérias/farmacologia , Caulobacteraceae/enzimologia , Agentes Neurotóxicos/envenenamento , Intoxicação por Organofosfatos/tratamento farmacológico , Compostos Organofosforados/toxicidade , Hidrolases de Triester Fosfórico/farmacologia , Antídotos/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Caulobacteraceae/genética , Estabilidade de Medicamentos , Estabilidade Enzimática , Temperatura Alta , Mutação , Intoxicação por Organofosfatos/enzimologia , Compostos Organotiofosforados/envenenamento , Oxirredução , Hidrolases de Triester Fosfórico/genética , Hidrolases de Triester Fosfórico/metabolismo , Desnaturação Proteica , Proteínas Recombinantes/farmacologia , Sarina/envenenamento , Soman/envenenamento
4.
Toxicol Lett ; 321: 21-31, 2020 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-31830555

RESUMO

Nerve agents inhibit acetylcholinesterase (AChE), leading to a build-up of acetylcholine (ACh) and overstimulation at cholinergic synapses. Current post-exposure nerve agent treatment includes atropine to treat overstimulation at muscarinic synapses, a benzodiazepine anti-convulsant, and an oxime to restore the function of AChE. Aside from the oxime, the components do not act directly to reduce the overstimulation at nicotinic synapses. The false transmitters acetylmonoethylcholine (AMECh) and acetyldiethylcholine (ADECh) are analogs of ACh, synthesised similarly at synapses. AMECh and ADECh are partial agonists, with reduced activity compared to ACh, so it was hypothesised the false transmitters could reduce overstimulation. Synthetic routes to AMECh and ADECh, and their precursors, monoethylcholine (MECh) and diethylcholine (DECh), were devised, allowing them to be produced easily on a laboratory-scale. The mechanism of action of the false transmitters was investigated in vitro. AMECh acted as a partial agonist at human muscarinic (M1 and M3) and muscle-type nicotinic receptors, and ADECh was a partial agonist only at certain muscarinic subtypes. Their precursors acted as antagonists at muscle-type nicotinic, but not muscarinic receptors. Administration of MECh and DECh improved neuromuscular function in the soman-exposed guinea-pig hemi-diaphragm preparation. False transmitters may therefore help reduce nerve agent induced overstimulation at cholinergic synapses.


Assuntos
Acetilcolina/análogos & derivados , Antídotos/farmacologia , Colina/análogos & derivados , Inibidores da Colinesterase/envenenamento , Diafragma/inervação , Agentes Neurotóxicos/envenenamento , Neurotransmissores/farmacologia , Intoxicação por Organofosfatos/tratamento farmacológico , Soman/envenenamento , Sinapses/efeitos dos fármacos , Acetilcolina/síntese química , Acetilcolina/metabolismo , Acetilcolina/farmacologia , Acetilcolinesterase/metabolismo , Animais , Antídotos/síntese química , Células CHO , Linhagem Celular Tumoral , Colina/síntese química , Colina/farmacologia , Cricetulus , Agonismo Parcial de Drogas , Cobaias , Humanos , Masculino , Neurotransmissores/síntese química , Intoxicação por Organofosfatos/enzimologia , Intoxicação por Organofosfatos/fisiopatologia , Receptores Colinérgicos/efeitos dos fármacos , Receptores Colinérgicos/genética , Receptores Colinérgicos/metabolismo , Sinapses/enzimologia
5.
Chem Biol Interact ; 309: 108714, 2019 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-31228470

RESUMO

Acetylcholinesterase (AChE) is an enzyme which terminates the cholinergic neurotransmission, by hydrolyzing acetylcholine at the nerve and nerve-muscle junctions. The reversible inhibition of AChE was suggested as the pre-treatment option of the intoxications caused by nerve agents. Based on our derived 3D-QSAR model for the reversible AChE inhibitors, we designed and synthesized three novel compounds 8-10, joining the tacrine and aroylacrylic acid phenylamide moieties, with a longer methylene chain to target two distinct, toplogically separated anionic areas on the AChE. The targeted compounds exerted low nanomolar to subnanomolar potency toward the E. eel and human AChE's as well as the human BChE and showed mixed inhibition type in kinetic studies. All compounds were able to slow down the irreversible inhibition of the human AChE by several nerve agents including tabun, soman and VX, with the estimated protective indices around 5, indicating a valuable level of protection. Putative noncovalent interactions of the selected ligand 10 with AChE active site gorge were finally explored by molecular dynamics simulation suggesting a formation of the salt bridge between the protonated linker amino group and the negatively charged Asp74 carboxylate side chain as a significant player for the successful molecular recognition in line with the design strategy. The designed compounds may represent a new class of promising leads for the development of more effective pre-treatment options.


Assuntos
Substâncias para a Guerra Química/química , Inibidores da Colinesterase/química , Colinesterases/metabolismo , Substâncias Protetoras/química , Acetilcolinesterase/química , Acetilcolinesterase/metabolismo , Sítios de Ligação , Domínio Catalítico , Substâncias para a Guerra Química/metabolismo , Inibidores da Colinesterase/metabolismo , Colinesterases/química , Humanos , Cinética , Simulação de Dinâmica Molecular , Compostos Organofosforados/química , Compostos Organofosforados/metabolismo , Substâncias Protetoras/metabolismo , Relação Quantitativa Estrutura-Atividade , Soman/química , Soman/metabolismo
6.
Chem Biol Interact ; 308: 170-178, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31129133

RESUMO

Organophosphorus compounds (OP) pose a significant threat. Administration of human butyrylcholinesterase (HuBChE) may reduce or prevent OP toxicity. Thus, we evaluated the safety and efficacy of HuBChE in monkeys using sensitive neurobehavioral tests while concurrently characterizing absorption and elimination in the presence and absence of high-dose soman exposure to predict time course and degree of protection. Eight young adult male cynomolgus macaques were trained on two distinct automated tests of neurobehavioral functioning. HuBChE purified under current Good Manufacturing Practices (CGMP) was injected intramuscularly at 13.1 mg/kg, producing an average peak plasma value (Cmax) of over 27 Units/ml. The apparent time to maximum concentration (Tmax) approximated 7 h, the elimination half-life approximated 102 h, and plasma levels returned to pre-administration (baseline) levels by 14 days. No behavioral disruptions following HuBChE administration were observed on either neurobehavioral test, even in monkeys injected 24 h later with an otherwise lethal dose of soman. Thus, HuBChE provided complete neurobehavioral protection from soman challenge. The present data replicate and extend previous results from our laboratory that had used a different route of administration (intravenous), a different species (rhesus macaque), and a different BChE product (non-CGMP material). The addition of two sensitive neurobehavioral tests coupled with the PK/PD results convincingly demonstrates the neurobehavioral safety of plasma-derived HuBChE at therapeutic levels. Protection against an otherwise-lethal dose of soman by a pre-exposure treatment dose that is devoid of side effects establishes a foundation for additional testing using other exposure routes and treatment times, other challenge agents/routes, or other classes of organophosphate scavengers.


Assuntos
Comportamento Animal/efeitos dos fármacos , Butirilcolinesterase/administração & dosagem , Substâncias para a Guerra Química/metabolismo , Soman/metabolismo , Animais , Butirilcolinesterase/sangue , Butirilcolinesterase/farmacocinética , Substâncias para a Guerra Química/toxicidade , Meia-Vida , Humanos , Injeções Intramusculares , Macaca fascicularis , Masculino , Soman/toxicidade
7.
Neuropharmacology ; 155: 113-120, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31132436

RESUMO

The acute toxicity of organophosphorus-based compounds is primarily a result of acetylcholinesterase inhibition in the central and peripheral nervous systems. The resulting cholinergic crisis manifests as seizure, paralysis, respiratory failure and neurotoxicity. Though overstimulation of muscarinic receptors is the mechanistic basis of central organophosphorus (OP) toxicities, short-term changes in synapse physiology that precede OP-induced seizures have not been investigated in detail. To study acute effects of OP exposure on synaptic function, field excitatory postsynaptic potentials (fEPSPs) were recorded from Schaffer collateral synapses in the mouse hippocampus CA1 stratum radiatum during perfusion with various OP compounds. Administration of the OPs paraoxon, soman or VX rapidly and stably depressed fEPSPs via a presynaptic mechanism, while the non-OP proconvulsant tetramethylenedisulfotetramine had no effect on fEPSP amplitudes. OP-induced presynaptic long-term depression manifested prior to interictal spiking, occurred independent of recurrent firing, and did not require NMDA receptor currents, suggesting that it was not mediated by activity-dependent calcium uptake. Pharmacological dissection revealed that the presynaptic endocannabinoid type 1 receptor (CB1R) as well as postsynaptic M1 and M3 muscarinic acetylcholine receptors were necessary for OP-LTD. Administration of CB1R antagonists significantly reduced survival in mice after a soman challenge, revealing an acute protective role for endogenous CB1R signaling during OP exposure. Collectively these data demonstrate that the endocannabinoid system alters glutamatergic synaptic function during the acute response to OP acetylcholinesterase inhibitors.


Assuntos
Inibidores da Colinesterase/toxicidade , Organofosfatos/toxicidade , Receptor CB1 de Canabinoide/metabolismo , Receptores Muscarínicos/metabolismo , Animais , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Antagonistas Muscarínicos/farmacologia , Técnicas de Cultura de Órgãos , Distribuição Aleatória , Soman/toxicidade
8.
Sci Total Environ ; 683: 175-184, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31146057

RESUMO

A practical and efficient protocol for the derivatization and detection by GC-EI-MS of isopropyl-, pinacolyl- and cyclohexylmethylphosphonic acids, key diagnostic degradation products of the nerve agents sarin, soman and cyclosarin respectively, in six different types of soil matrices is presented. The method involves the in situ conversion of the phosphonic acids to their respective methyl esters using trimethyloxonium tetrafluoroborate when present in the soils at low levels (10 µg g-1) without any prior extractions or soil preparation. The soils employed in our study were Nebraska EPA soil, Georgia soil, silt, Virginia type A soil, regular sand and Ottawa sand and were chosen for their vast differences in composition and physical features. Appealing attributes of the protocol include its rapidity (t < 30 min), mildness (ambient temperature), and practicality that includes the production of the phosphonic methyl esters that can be easily detected by GC-EI-MS and corroborated with the instrument's internal NIST spectral library or the Organisation for the Prohibition of Chemical Weapons (OPCW) central analytical database (OCAD v.21_2019). The overall efficacy of the protocol was then tested on a soil sample featured in the 44th OPCW PT that our laboratory participated in. After preparing the soil so as to give pinacolyl methylphosphonic acid at a 5 µg g-1 concentration, the acid was successfully methylated and detected by GC-EI-MS. The protocol's performance mirrors that of the universally employed diazomethane protocol but accomplishes this without any of the explosive hazards and time consuming reagent preparation commonly associated with it.


Assuntos
Cromatografia Gasosa-Espectrometria de Massas/métodos , Compostos Organofosforados/análise , Poluentes do Solo/análise , Soman/análogos & derivados , Biomarcadores/análise , Metilação , Agentes Neurotóxicos/análise , Estudos Retrospectivos , Sarina/análise , Solo/química , Soman/análise
9.
Toxicology ; 423: 54-61, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31102696

RESUMO

The detrimental effects of organophosphate (OP) nerve agents have been reported but the mechanisms mediating these multiple effects are not well understood. Recent use of nerve agents in Syria and the UK illustrate their continuous threat to the modern world. Epigenetic and autophagy studies are useful to address the issues related to regulation of gene and protein expression by which nerve agents could impact on human health. These studies help to understand molecular mechanisms underlying the multiple neurotoxic effects of nerve agents. In the present study, changes in epigenetic (global DNA methylation and histone acetylation) and autophagic marker proteins were studied in the nerve agent sensitive rat brain areas (piriform cortex and hippocampus) after soman (1xLD50) exposure. Global DNA methylation analysis revealed that nerve agent induced hypomethylation in the brain regions at 1 and 7 days post exposure. In contrast, DNA hypermethylation was observed at 30 days post soman exposure, demonstrating a possible compensatory mechanism. Western blot analysis showed significant increase in the histone acetylation levels after soman exposure in the piriform cortex and hippocampus. The present study observed the changes in autophagic proteins of nerve agent poisoning for the first time to the best of our knowledge. Immunoreactivity levels of autophagic proteins (LC3-II, ATG-5 and p62) were transiently increased in the rat piriform cortex and hippocampus after soman exposure. In conclusion, this study provides insight into the epigenetic and autophagic changes in the brain following soman exposure and their possible role in the neuronal damage and development of multiple neurological effects.


Assuntos
Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Hipocampo/efeitos dos fármacos , Córtex Piriforme/efeitos dos fármacos , Soman/toxicidade , Acetilação/efeitos dos fármacos , Animais , Autofagia/efeitos dos fármacos , Colinesterases/sangue , Colinesterases/metabolismo , Metilação de DNA/efeitos dos fármacos , Epigênese Genética/efeitos dos fármacos , Hipocampo/metabolismo , Histonas/metabolismo , Masculino , Córtex Piriforme/metabolismo , Ratos Wistar
10.
Neurotox Res ; 36(2): 323-333, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31069755

RESUMO

Organophosphorus nerve agents (NAs) irreversibly inhibit acetylcholinesterase, which results in the accumulation of acetylcholine and widespread excitotoxic seizure activity. Because current medical countermeasures (anticholinergics, AChE reactivators, and benzodiazepines) lack sufficient anti-seizure efficacy when treatment is delayed, those intoxicated are at risk for severe brain damage or death if treatment is not immediately available. Toward developing a more effective anti-seizure treatment for NA intoxication, this study evaluated the efficacy of A1 adenosine (ADO) receptor (A1AR) agonists in a rat soman seizure model. One minute after exposure to soman (1.6 × LD50, subcutaneous), rats were treated intraperitoneally with one of the following agonists at increasing dose levels until anti-seizure efficacy was achieved: N6-cyclopentaladenosine (CPA), 2-chloro-N6-cyclopentyladenosine (CCPA), and (±)-5'-chloro-5'-deoxy-ENBA (ENBA). All A1AR agonists were efficacious in preventing seizure and promoting survival. The effective doses for the A1AR agonists were 60 mg/kg CPA, 36 mg/kg CCPA, and 62 mg/kg ENBA. Whereas vehicle-treated rats experienced 100% seizure and 21% survival (N = 28), ADO treatments reduced seizure occurrence and improved survival rates: 8% seizure and 83% survival with CPA (60 mg/kg, N = 12), 17% seizure and 75% survival with CCPA (36 mg/kg, N = 12), and 8% seizure, 83% survival with ENBA (62 mg/kg, N = 12). The brains of ADO-treated rats were also protected from damage as indicated by neurohistopathological analysis. While all ADO agonists provided neuroprotection, rats receiving CCPA and ENBA experienced less severe ADO-induced side effects (e.g., sedation, hypothermia, bradycardia) than with CPA. The data from this study suggest that the ADO signaling pathway is a promising mechanism for countering seizure activity induced by NAs.


Assuntos
Agonistas do Receptor A1 de Adenosina/uso terapêutico , Anticonvulsivantes/uso terapêutico , Contramedidas Médicas , Convulsões/induzido quimicamente , Convulsões/tratamento farmacológico , Soman/toxicidade , Animais , Relação Dose-Resposta a Droga , Masculino , Agentes Neurotóxicos/toxicidade , Ratos , Ratos Sprague-Dawley , Convulsões/fisiopatologia
11.
Molecules ; 24(5)2019 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-30813539

RESUMO

Here, we introduced a novel thiourea-based rhodamine compound as a chromo-fluorogenic indicator of nerve agent Soman and its simulant diethyl chlorophosphate (DCP). The synthesized probe N-(rhodamine B)-lactam-2-(4-cyanophenyl) thiourea (RB-CT), which has a rhodamine core linked by a cyanophenyl thiosemicarbazide group, enabled a rapidly and highly sensitive response to DCP with clear fluorescence and color changes. The detection limit was as low as 2 × 10-6 M. The sensing mechanism showed that opening of the spirolactam ring following the phosphorylation of thiosemicarbazides group formed a seven-membered heterocycle adduct, according to MS analysis and TD-DFT calculations. RB-CT exhibited high detecting selectivity for DCP, among other organophosphorus compounds. Moreover, two test kits were employed and successfully used to detect real nerve agent Soman in liquid and gas phase.


Assuntos
Corantes Fluorescentes/síntese química , Compostos Organofosforados/análise , Rodaminas/química , Soman/análise , Tioureia/química , Substâncias para a Guerra Química/análise , Substâncias para a Guerra Química/química , Corantes Fluorescentes/química , Limite de Detecção , Estrutura Molecular , Agentes Neurotóxicos/análise , Agentes Neurotóxicos/química , Compostos Organofosforados/química , Soman/química
12.
Artigo em Inglês | MEDLINE | ID: mdl-30790623

RESUMO

INTRODUCTION: Organophosphorus nerve agents (OPNAs) irreversibly block acetylcholinesterase activity, resulting in accumulation of excess acetylcholine at neural synapses, which can lead to a state of prolonged seizures known as status epilepticus (SE). Benzodiazepines, the current standard of care for SE, become less effective as latency to treatment increases. In a mass civilian OPNA exposure, concurrent trauma and limited resources would likely cause a delay in first response time. To address this issue, we have developed a rat model to test novel anticonvulsant/ neuroprotectant adjuncts at delayed time points. METHODS: For model development, adult male rats with cortical electroencephalographic (EEG) electrodes were exposed to soman and administered saline along with atropine, 2-PAM, and midazolam 5, 20, or 40 min after SE onset. We validated our model using three drugs: scopolamine, memantine, and phenobarbital. Using the same procedure outlined above, rats were given atropine, 2-PAM, midazolam and test treatment 20 min after SE onset. RESULTS: Using gamma power, delta power, and spike rate to quantify EEG activity, we found that scopolamine was effective, memantine was minimally effective, and phenobarbital had a delayed effect on terminating SE. Fluoro-Jade B staining was used to assess neuroprotection in five brain regions. Each treatment provided significant protection compared to saline + midazolam in at least two brain regions. DISCUSSION: Because our data agree with previously published studies on the efficacy of these compounds, we conclude that this model is a valid way to test novel anticonvulsants/ neuroprotectants for controlling benzodiazepine-resistant OPNA-induced SE and subsequent neuropathology.


Assuntos
Anticonvulsivantes/farmacologia , Benzodiazepinas/farmacologia , Memantina/farmacologia , Agentes Neurotóxicos/farmacologia , Fármacos Neuroprotetores/farmacologia , Fenobarbital/farmacologia , Escopolamina/farmacologia , Estado Epiléptico/tratamento farmacológico , Animais , Atropina/farmacologia , Encéfalo/efeitos dos fármacos , Eletroencefalografia/métodos , Masculino , Midazolam/farmacologia , Ratos , Ratos Sprague-Dawley , Convulsões/induzido quimicamente , Convulsões/tratamento farmacológico , Soman/farmacologia , Estado Epiléptico/induzido quimicamente
13.
Talanta ; 195: 728-731, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30625608

RESUMO

A new metal-affinity sorbent based on lanthanum stearate monolayers has been developed and characterized. The prospect of its application to specific extraction of organophosphorous compound (OP) adducts of blood proteins was demonstrated. For this, the patterns of soman adducts of human serum albumin (HSA) were comprehensively characterized by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS).


Assuntos
Lantânio/química , Albumina Sérica Humana/química , Soman/química , Estearatos/química , Adsorção , Adulto , Humanos , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
14.
Toxicology ; 416: 62-74, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30682440

RESUMO

BACKGROUND: Carbamates physostigmine and pyridostigmine have been used as a pretreatment against poisoning with nerve agents in order to reversibly inhibit and thus protect from irreversible inhibition a portion of acetylcholinesterase (AChE) in brain and respiratory muscles that is crucial for survival. Memantine, an adamantine derivative, has emerged as a promising alternative to carbamates, since it prevented the fasciculations and skeletal muscle necrosis induced by carbamates and organophosphates, including nerve agents. AIM: This experimental study was undertaken in order to investigate and compare the protective and behavioural effects of memantine and standard carbamates physostigmine and pyridostigmine in rats poisoned with soman and treated with atropine, oxime HI-6 and diazepam. Another goal was to elucidate the mechanisms of the antidotal effect of memantine and its potential synergism with standard antidotes against nerve agents. MATERIALS AND METHODS: Male Wistar rats were used throughout the experiments. In dose-finding experiments memantine was administered at dose interval 0-72 mg/kg sc 60 min before sc injection of soman. In time-finding experiments memantine was injected 18 mg/kg sc 0-1440 min before soman. Standard treatment antidotes - atropine 10 mg/kg, HI-6 50 mg/kg and diazepam 2.5 mg/kg - were administered im within 15 s post-exposure. Soman 0.75 LD50 was used to study its inhibitions of neuromuscular transmission on the phrenic nerve-diaphragm preparation in situ and of tissue AChE activity. Behavioural effects of the prophylactic antidotes were investigated by means of the rotarod test. Based on these data therapeutic index and therapeutic width was calculated for all three prophylactic agents. RESULTS: Memantine pretreatment (18 mg/kg sc) produced in rats poisoned with soman significantly better protective ratios (PRs) than the two carbamates - 1.25 when administered alone and 2.3 when combined with atropine pretreatment and 6.33 and 7.23 with atropine/HI-6 and atropine/HI-6/diazepam post-exposure therapy, respectively. The highest PR of 10.11 obtained in Atr/HI-6-treated rats was achieved after pretreatment with memantine 36 mg/kg. This additional protection lasted for 8 h. All three prophylactic regimens antagonised the soman-induced neuromuscular blockade, but the effect of memantine was fastest. Pretreatment with memantine assured higher AChE activity in brain and diaphragm than in unpretreated rats (46% vs 28% and 68% vs. 38%, respectively). All three prophylactic regimens affected the rotarod performance in rats, but the effect of memantine was relatively strongest. Memantine and pyridostigmine had lowest and highest therapeutic index and therapeutic width, respectively. CONCLUSIONS: Although memantine assures better and longer-lasting protection against soman poisoning in rats than the two carbamates, its small therapeutic index and narrow therapeutic width seriously limit its potential as a pretreatment agent. Despite its behavioural effects, memantine seems to be beneficial antidote when administered after soman, along with atropine/HI-6/diazepam therapy. Mechanism of the antidotal effect of memantine against soman poisoning appears to be a combination of AChE-protecting and NMDA receptor-blocking action.


Assuntos
Antídotos/farmacologia , Substâncias para a Guerra Química , Inibidores da Colinesterase , Memantina/farmacologia , Junção Neuromuscular/efeitos dos fármacos , Intoxicação por Organofosfatos/prevenção & controle , Soman , Acetilcolinesterase/metabolismo , Animais , Atropina/farmacologia , Comportamento Animal/efeitos dos fármacos , Diazepam/farmacologia , Modelos Animais de Doenças , Sinergismo Farmacológico , Quimioterapia Combinada , Proteínas Ligadas por GPI/metabolismo , Masculino , Junção Neuromuscular/enzimologia , Junção Neuromuscular/patologia , Junção Neuromuscular/fisiopatologia , Intoxicação por Organofosfatos/enzimologia , Intoxicação por Organofosfatos/patologia , Intoxicação por Organofosfatos/fisiopatologia , Oximas/farmacologia , Compostos de Piridínio/farmacologia , Ratos Wistar , Receptores de N-Metil-D-Aspartato/metabolismo , Transmissão Sináptica/efeitos dos fármacos
15.
Clin Toxicol (Phila) ; 57(5): 343-349, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30307341

RESUMO

BACKGROUND: In a military or terrorist scenario, combination of organophosphorus compounds (OP) poisoning with physical trauma requiring surgical treatment and thus general anaesthesia are possible. Previous in vitro studies showed an altered potency of relevant anaesthetics during cholinergic crisis. Hence, it is not clear, which anaesthetics are suitable to achieve the necessary stage of surgical anaesthesia in OP poisoning. METHODS: In the present study, different anaesthetic regimens (ketamine-midazolam, propofol-fentanyl, thiopental-fentanyl), relevant in military emergency medicine, were examined in soman-poisoned rats. Clinical signs and cardiovascular variables were recorded continuously. Blood samples for acetylcholinesterase (AChE) activity were drawn. After euthanasia or death of the animals, brain and diaphragm were collected for cholinesterase assays. RESULTS: Propofol-fentanyl and thiopental-fentanyl resulted in surgical anaesthesia throughout the experiments. With ketamine-midazolam, surgical anaesthesia without respiratory impairment could not be achieved in pilot experiments (no soman challenge) and was therefore not included in the study. Soman-poisoned and control animals required a comparable amount of propofol-fentanyl or thiopental-fentanyl. In combination with atropine, significantly less propofol was needed. Survival rate was higher with thiopental compared to propofol. Atropine improved survival in both groups. Blood and tissue AChE activities were strongly inhibited after soman administration with and without atropine treatment. DISCUSSION: The current in vivo study did not confirm concerns of altered potency of existing anaesthetic protocols for the application of propofol or thiopental with fentanyl due to soman poisoning. Despite severe cholinergic crisis, sufficient anaesthetic depth could be achieved in all animals. CONCLUSION: Further experiments in in vivo models closer to human pharmaco- and toxicokinetics (e.g., swine) are required for confirmation of the initial findings and for improving extrapolation to humans.


Assuntos
Analgésicos Opioides/farmacologia , Anestesia Intravenosa , Anestésicos Intravenosos/farmacologia , Inibidores da Colinesterase , Estado de Consciência/efeitos dos fármacos , Fentanila/farmacologia , Intoxicação por Organofosfatos/enzimologia , Propofol/farmacologia , Soman , Tiopental/farmacologia , Acetilcolinesterase/sangue , Animais , Biomarcadores/sangue , Modelos Animais de Doenças , Proteínas Ligadas por GPI/antagonistas & inibidores , Proteínas Ligadas por GPI/sangue , Masculino , Intoxicação por Organofosfatos/fisiopatologia , Ratos Wistar
16.
Toxicol In Vitro ; 54: 263-268, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30347232

RESUMO

BACKGROUND/AIMS: Following an incident involving toxic chemicals, deployment of countermeasures before the arrival of specialised services at the scene may provide a "therapeutic" window in which to mitigate skin absorption. METHODS: Five potential candidates (itaconic acid, N,N'-methylenebisacrylamide, 2-trifluoromethylacrylic acid, fuller's earth and Fast-Act®) previously found effective against a simulant (methyl salicylate) were evaluated against a 10 µL droplet of 14C-sulphur mustard (HD), soman (GD) or VX applied to the surface of porcine skin mounted on static skin diffusion cells. RESULTS: All the decontaminants applied to the skin 5 min post exposure achieved a marked reduction in the amount of 14C contaminant remaining within the skin at 24 h. Itaconic acid significantly (p < .05) reduced the amount of 14C-HD, GD and VX remaining in the skin at 24 h. Additionally, 2-trifluoromethylacrylic acid significantly reduced the amount of 14C-HD, whilst fuller's earth significantly reduced the amounts of 14C-HD and VX recovered within the skin at 24 h. CONCLUSION: All of the products evaluated in this study performed well in reducing the dermal absorption of all the chemical warfare agents tested.


Assuntos
Substâncias para a Guerra Química , Descontaminação/métodos , Pele , Animais , Feminino , Gás de Mostarda , Compostos Organotiofosforados , Absorção Cutânea , Soman , Suínos
17.
Redox Biol ; 20: 275-284, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30384261

RESUMO

Persistent inhibition of acetylcholinesterase resulting from exposure to nerve agents such as soman, is associated with prolonged seizure activity known as status epilepticus (SE). Without medical countermeasures, exposure to soman and resultant SE leads to high morbidity and mortality. Currently available therapeutics are effective in limiting mortality, however effects on morbidity are highly time-dependent and rely on the ability to suppress SE. We have previously demonstrated significant protection from secondary neuronal injury in surrogate nerve agent models by targeting oxidative stress. However, whether oxidative stress represents a relevant therapeutic target in genuine nerve agent toxicity is unknown. Here, we demonstrate that soman exposure results in robust region- and time-dependent oxidative stress. Targeting this oxidative stress in a post-exposure paradigm using a small molecular weight, broad spectrum catalytic antioxidant, was sufficient to attenuate brain and plasma oxidative stress, neuroinflammation and neurodegeneration. Thus, targeting of oxidative stress in a post-exposure paradigm can mitigate secondary neuronal injury following soman exposure.


Assuntos
Antioxidantes/farmacologia , Agentes Neurotóxicos/toxicidade , Fármacos Neuroprotetores/farmacologia , Animais , Biomarcadores , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Citocinas/sangue , Citocinas/metabolismo , Mediadores da Inflamação/sangue , Mediadores da Inflamação/metabolismo , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Espécies Reativas de Nitrogênio/sangue , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/sangue , Espécies Reativas de Oxigênio/metabolismo , Soman/farmacologia
18.
Environ Toxicol Pharmacol ; 64: 147-154, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30391876

RESUMO

Melatonin is a hormone with many different biological activities and therefore seems to be an important factor reducing the harmful effects caused by toxic organophosphorus compounds. In this study, we attempted to evaluate the protective effect of melatonin on liver cells of mice challenged with chemical warfare agent-soman. The study was conducted at the level of ultrastructural and biochemical changes (analysis of the activity of model lysosomal enzymes and assessment of the level of lipid peroxidation). Significant biochemical and ultrastructural changes were found in the studied mouse hepatocytes after administration of soman alone, and soman in combination with melatonin, and the scope of the disclosed changes was dependent on the time of action of the examined factors. Melatonin has shown protective action, shielding liver cells from toxic effects of soman, which may result from its antioxidant properties and stimulation of the lysosomal compartment, the system coordinating the isolation and removal of cell-threatening processes.


Assuntos
Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Hepatócitos/efeitos dos fármacos , Melatonina/farmacologia , Substâncias Protetoras/farmacologia , Soman/toxicidade , Animais , Autofagia/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/ultraestrutura , Peroxidação de Lipídeos/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Camundongos Endogâmicos BALB C
19.
Epilepsia ; 59(12): 2206-2218, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30368799

RESUMO

OBJECTIVE: Exposure to chemical warfare nerve agents (CWNAs), such as soman (GD), can induce status epilepticus (SE) that becomes refractory to benzodiazepines when treatment is delayed, leading to increased risk of epileptogenesis, severe neuropathology, and long-term behavioral and cognitive deficits. Rodent models, widely used to evaluate novel medical countermeasures (MCMs) against CWNA exposure, normally express plasma carboxylesterase, an enzyme involved in the metabolism of certain organophosphorus compounds. To better predict the efficacy of novel MCMs against CWNA exposure in human casualties, it is crucial to use appropriate animal models that mirror the human condition. We present a comprehensive characterization of the seizurogenic, epileptogenic, and neuropathologic effects of GD exposure with delayed anticonvulsant treatment in the plasma carboxylesterase knockout (ES1-/-) mouse. METHODS: Electroencephalography (EEG) electrode-implanted ES1-/- and wild-type (C57BL/6) mice were exposed to various seizure-inducing doses of GD, treated with atropine sulfate and the oxime HI-6 at 1 minute after exposure, and administered midazolam at 15-30 minutes following the onset of seizure activity. The latency of acute seizure onset and spontaneous recurrent seizures (SRS) was assessed, as were changes in EEG power spectra. At 2 weeks after GD exposure, neurodegeneration and neuroinflammation were assessed. RESULTS: GD-exposed ES1-/- mice displayed a dose-dependent response in seizure severity. Only ES1-/- mice exposed to the highest tested dose of GD developed SE, subchronic alterations in EEG power spectra, and SRS. Degree of neuronal cell loss and neuroinflammation were dose-dependent; no significant neuropathology was observed in C57BL/6 mice or ES1-/- mice exposed to lower GD doses. SIGNIFICANCE: The US Food and Drug Administration (FDA) animal rule requires the use of relevant animal models for the advancement of MCMs against CWNAs. We present evidence that argues for the use of the ES1-/- mouse model to screen anticonvulsant, antiepileptic, and/or neuroprotective drugs against GD-induced toxicity, as well as to identify mechanisms of GD-induced epileptogenesis.


Assuntos
Anticonvulsivantes/uso terapêutico , Carboxilesterase/genética , Substâncias para a Guerra Química , Midazolam/uso terapêutico , Soman , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/tratamento farmacológico , Animais , Contagem de Células , Reativadores da Colinesterase/uso terapêutico , Eletroencefalografia , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Degeneração Neural/patologia , Convulsões/fisiopatologia , Estado Epiléptico/genética
20.
J Chromatogr A ; 1577: 24-30, 2018 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-30297234

RESUMO

A highly sensitive method for the detection and identification of sarin (GB), soman (GD) and cyclosarin (GF) chemical warfare agents (CWAs) in environmental outdoor and indoor matrices such as soil, asphalt, linoleum, formica, concrete and cloth was developed. The method incorporates derivatization of the G-type nerve agent extracts with 2-[(dimethylamino)methyl]phenol (2-DMAMP), followed by LC-ESI(+)-MS/MS analysis. Four LC-amenable extraction solvents were explored in terms of their extraction efficiency and the reaction rate of the derivatizing agent. The reaction time, temperature and derivatization reagent amount were optimized. The optimal procedure was found to be extraction with water by agitation (2 min), followed by the addition of 2-DMAMP directly into the injection vial and stirring for 5 min prior to LC-ESI(+)-MS/MS analysis, without any other pretreatment. The method was applied to real-world samples and exhibited very low detection limits (LODs) of 0.8-20 pg/cm2 in asphalt, linoleum, cloth, formica and concrete and 4 pg/g in soil. The newly developed method demonstrated significantly superior sensitivity compared to conventional GC-MS- and LC-MS-based methods for the identification of G-nerve agents and allowed the determination of both G-nerve agents and their hydrolysis products within a single LC-MS/MS run. The proposed methodology may be practical for verifying contaminated matrices collected in the battlefield or terror scenes in forensic investigations where trace level analysis is required.


Assuntos
Cromatografia Líquida , Monitoramento Ambiental/métodos , Agentes Neurotóxicos/análise , Espectrometria de Massas em Tandem , Substâncias para a Guerra Química/análise , Cromatografia Gasosa-Espectrometria de Massas , Hidrólise , Limite de Detecção , Compostos Organofosforados/análise , Sarina/análise , Solo/química , Soman/análise , Água/química
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