Myasthenia Gravis: Brief Guide to Diagnosis and Management

Physicians should consider the possibility of myasthenia gravis in patients complaining of fluctuating muscle weakness and easy fatigability. The disorder may occur in either sex at any age. Repeated measurements of multiple muscle groups and functions to include baseline strength and response to a control as well as anticholinesterase drugs are desirable in reaching a diagnosis. Appropriate doses of anticholinesterase medication reduce weakness and fatigability in most patients. Selected patients may benefit from thymectomy. Patients in crisis failing to respond to anticholinesterase drugs, having difficulty maintaining a patent airway or adequate respiratory exchange are best managed by prompt tracheostomy using a cuffed tube, with adequate tracheobronchial toilet and mechanical respiratory assistance.     

Brain cholinesterase activity of apparently normal wild birds

Organophosphorus and carbamate pesticides are potent anticholinesterase substances that have killed large numbers of wild birds of various species. Cause of death is diagnosed by demonstration of depressed brain cholinesterase (ChE) activity in combination with chemical detection of anticholinesterase residue in the affected specimen. ChE depression is determined by comparison of the affected specimen to normal ChE activity for a sample of control specimens of the same species, but timely procurement of controls is not always possible. Therefore, a reference file of normal whole brain ChE activity is provided for 48 species of wild birds from North America representing 11 orders and 23 families for use as emergency substitutes in diagnosis of anticholinesterase poisoning. The ChE values, based on 83 sets of wild control specimens from across the United States, are reproducible provided the described procedures are duplicated. Overall, whole brain ChE activity varied nearly three-fold among the 48 species represented, but it was usually similar for closely related species. However, some species were statistically separable in most families and some species of the same genus differed as much as 50%.     

Ketamine antagonizes toxic action of anticholinesterase agents

Ketamine is an anaesthetic interacting with several neurotransmitters. Among others, ketamine exerts some cholinergic actions (ACh). This paper presents the results of studying the interaction of ketamine with ACh in two animal species. Atropine slightly increased the time of immobility produced by ketamine injections in rats. Meanwhile, neostigmine slightly decreased such immobility. Ketamine resulted similar in behavioral actions and shared some electroencephalographic (EEG) actions of scopolamine in cats. The most striking interaction consisted on an antagonism of ketamine on the action of anticholinesterase agents. In both species, ketamine blocked the EEG and the behavioral toxic effects of neostigmine and physostigmine. Notwithstanding, the anticholinesterase agents were unable in reducing the actions of ketamine. This partial cholinergic agonist action of ketamine support certain but limited use of the anesthetic against insecticidal anticholinesterase poisoning.     

Anticholinesterase effect of organophosphorus and pyrethroid insecticides on some phytophages, their predators, and parasites

The normal catalytic activity of insects cholinesterases was determined: Apanteles glomeratus L., Coccinella septempunctata L., Rhopalosiphum padi L. and Pieris brassicae L. Constants of bimolecular interaction of three insecticides for insects were calculated: cholinesterase of C. septempunctata were the most sensitive to Bi-58 new [Kii = (4.54 +/- 0.23). 10(4)], cholinesterase of aphids was revealed to be the most sensitive to acetec [Kii = (2.9 +/- 0.14). 10(5)]. The studies have been carried aimed to determination of anticholinesterase actioncoefficients: acetec demonstrated the selective anticholinesterase activity in the system Aphididae--Coccinellidae (Kache = 118.9). Its anticholinesterase activity underlies in the toxicity mechanism for the given species.     

Cholinergic mechanisms related to REM sleep. III. Tonic and phasic inhibition of monosynaptic reflexes induced by an anticholinesterase in the decerebrate cat

The depression of the postural activity induced by intravenous injection of eserine sulphate (0.1 mg/kg), an anticholinesterase, has been studied in precollicular decerebrate cats. The extensor and flexor monosynaptic reflexes elicited by single shock stimulation of the GS, P1-FDHL and DP nerves are tonically depressed during the episodes of postural atonia induced by the anticholinesterase. A further phasic depression of the monosynaptic reflexes occurs during the bursts of rapid eye movements (REM) typical of these episodes. These changes in spinal reflex activity closely resemble the tonic depression of the spinal reflexes described in the unrestrained cats during the desynchronized sleep as well as the phasic depression of the spinal reflexes characteristic of the hypnic bursts of REM. Results obtained after spinal cord section indicate that both the tonic and the phasic depression of the spinal reflexes induced by eserine are due to active inhibitory influences originating from supraspinal structures. A complete bilateral destruction of the vestibular nuclei or limited to the medial and descending vestibular nuclei abolishes not only the cholinergically induced bursts of REM, as reported in a previous paper, but also the related phasic depression of the monosynaptic reflexes. These findings can be related with previous observations showing that a bilateral lesion of the vestibular nuclei abolishes the REM bursts of desynchronized sleep, as well as the related phasic inhibition of the spinal reflexes. The tonic depression of the monosynaptic reflexes induced by the anticholinesterase, on the other hand, remains unmodified by this vestibular lesion. This depression, therefore, can be attributed to supraspinal descending inhibitory volleys originating from extravestibular structures.     

Tetramethonium derivatives as reversible inhibitors of various cholinesterases

To study the effect of the onium atom nature on anticholinesterase efficiency, we tested elementorganic derivatives of tetramethylenbisonium compounds as reversible inhibitors of the following cholinesterases (ChE): acetyl-ChE from human erythrocytes, butyryl-ChE from horse serum, ChE from the brain of the grass frog Rana temporaria, ChEs from visual ganglia of the Pacific squid Todarodes pacificus, and ChE from visual ganglia of the commander squid Berryteuthis magister from different habitats in the Northwestern Pacific Ocean. Bisphosphonium inhibitors were found to be much stronger effectors than bisammonum compounds, although this may be due to a significantly increased size and hydrophobicity of their onium groups. Bisammonium organosilicon compound and its monoammonium analog were equally active as reversible ChE inhibitors in mammals. The first studied bis(phenyliodonium) derivative, which is characterized by a significantly increased hydrophobicity due to the introduction of fluorine atoms to the interonium tetramethylene chain, also exhibited a pronounced anticholinesterase effect on mammalian ChE.     

Determination of trace quantities of anticholinesterase pesticides

A procedure has been described which is applicable to the screening of large numbers of samples for anticholinesterase activity. The method is sufficiently sensitive to detect cholinesterase depression due to as little as 30 ppt paraoxon; 3 ppb malaoxon or 2 ppm of carbaryl in either ethylene glycol or water. While the method does not allow the absolute differentiation between compounds which might be present, some classification is possible, i.e., for compounds which must be activated before they possess anticholinesterase activity.     

Combined effect of short-term dehydration and sublethal acute oral dicrotophos exposure confounds the diagnosis of anticholinesterase exposure in common quail (Coturnix coturnix) using plasma cholinesterase activity

Common Quail (Coturnix coturnix) were subjected to controlled and replicated experiments in the summer of 2008 to investigate the effects of short-term dehydration on cholinesterase activity in brain and plasma and the interaction between dehydration and exposure to the organophosphorus pesticide dicrotophos in these same tissues. Our objective was to determine if dehydration could confound the diagnosis of anticholinesterase exposure using inhibition of cholinesterase activity in quail tissues. The effect of dehydration was quantified using measures of plasma osmolality and hematocrit. Dicrotophos exposure caused significant inhibition of cholinesterase activity in brain, while the effects of dehydration and interaction were not significant. Dehydration caused significant duration-dependent increases in plasma osmolality and hematocrit. Dehydration also caused a significant increase in plasma cholinesterase activity. Variation in the change in plasma cholinesterase activity in response to dehydration was significantly and positively correlated with dehydration-induced variation in both the change in plasma osmolality and the change in hematocrit. These correlations suggest that plasma cholinesterase activity in quail is not limited to plasma but occupies some larger pool of the extracellular fluid volume, and we suggest lymph is part of that pool. The effects of dehydration on plasma cholinesterase activity masked the inhibitory effects of dicrotophos. Here, the combination of dehydration and dicrotophos exposure produced plasma cholinesterase activity that was not significantly different from reference and pre-exposure values, confounding the diagnosis of anticholinesterase exposure in dehydrated, dicrotophos-exposed quail. A method to adjust plasma cholinesterase activities for the confounding effects of dehydration and enable the diagnosis of anticholinesterase exposure in dehydrated, dicrotophos-exposed quail was developed. Clinicians and practitioners responsible for the diagnosis of anticholinesterase exposure in birds are cautioned that dehydration, commonly observed in sick wildlife, may mask the effect of anticholinesterases on plasma cholinesterase activity.     

Tissue distribution of cholinesterases and anticholinesterases in native and transgenic tomato plants

Acetylcholinesterase, a major component of the central and peripheral nervous systems, is ubiquitous among multicellular animals, where its main function is to terminate synaptic transmission by hydrolyzing the neurotransmitter, acetylcholine. However, previous reports describe cholinesterase activities in several plant species and we present data for its presence in tomato plants. Ectopic expression of a recombinant form of the human enzyme and the expression pattern of the transgene and the accumulation of its product in transgenic tomato plants are described. Levels of acetylcholinesterase activity in different tissues are closely effected by and can be separated from -tomatine, an anticholinesterase steroidal glycoalkaloid. The recombinant enzyme can also be separated from the endogenous cholinesterase activity by its subcellular localization and distinct biochemical properties. Our results provide evidence for the co-existence in tomato plants of both acetylcholinesterase activity and a steroidal glycoalkaloid with anticholinesterase activity and suggest spatial mutual exclusivity of these antagonistic activities. Potential functions, including roles in plant-pathogen interactions are discussed.     

Non-quantal acetylcholine release is increased after nitric oxide synthase inhibition

After anticholinesterase treatment, depolarization of the postsynaptic muscle membrane by about 5 mV develops due to non-quantally released acetylcholine from the motor nerve terminal and can be revealed as hyperpolarization by the addition of curare (H-effect). The H-effect increases significantly to 8.7 mV after inhibition of NO-synthase by L-nitroarginine methylester (L-NAME) whilst no changes in the amplitude and frequency of quantal miniature endplate potentials are observed.     

Differences in conformational stability between native and phosphorylated acetylcholinesterase as evidenced by a monoclonal antibody.

Monoclonal antibody 25B1 generated against diisopropyl phosphorofluoridate inhibited fetal bovine serum acetylcholinesterase has been extensively characterized with respect to its anticholinesterase properties. This antibody demonstrated considerably different properties from previously reported inhibitory antibodies raised against acetylcholinesterase in terms of the degree of inhibition (greater than 98%), the high degree of specificity, and the stability of the antigen-antibody complex. Monoclonal antibody 25B1 appears to be directed against a conformational epitope located in close proximity to the catalytic center of the enzyme and was found to be most suitable for studying the stabilization of the active site of acetylcholinesterase against denaturation by heat or guanidine following phosphorylation by organophosphorus anticholinesterase compounds. This approach allowed the determination of stability rank order of various phosphorylated acetylcholinesterases. Among all the organophosphates tested, the combination of a methyl group and a negatively charged oxygen attached to the P atom, CH3P(O)(O-)-AChE, conferred the greatest protection to the active site of aged or nonaged organophosphoryl conjugates of acetylcholinesterase.     

An integrated model for the interaction of muscle relaxants with their antagonists

An integrated model describing the interaction of nondepolarizing neuromuscular blocking agents with reversible anticholinesterase agents is derived and compared with a naive model using experimental data obtained from four anesthetized dogs. Three consecutive but separate steady-state d-tubocurarine blocks (approximately 50, 70, and 90%) were induced in each of the four dogs and reversed by short edrophonium infusions. Edrophonium arterial concentrations and twitch tension of the anterior tibialis muscle were measured. Both the integrated and the naive model were fit to the twitch tension data using a model with a hypothetical "effect" compartment. The integrated model consistently fit the twitch tension data better than the naive model; the sum of squared deviations was lower by 46, 45, 87, and 69%, respectively, with the integrated model than with the naive model. Also, in contrast to the naive model, the integrated model is capable of describing the interaction of the anticholinesterase agent and the neuromuscular blocking agent when the concentration of either varies with time.     

Anabasine derivatives as reversible and irreversible inhibitors of cholinesterases from different animals

A comparative study was carried out of action of 18 specifically synthesized anabasine derivatives and their analogs, including diiodomethylates of anabasine acylates and bis-anabasine derivatives of dicarboxylic glutaric, adipic, azelaic, and sebacic acids, on activity of cholinesterase of brain of frog Rana temporaria and visual ganglia of the Pacific squid Todarodes pacificus and Commander squid Berryteuthis magister from different zones of habitations in the northwest part of the Pacific Ocean as well as of erythrocyte acetylcholinesterase and serum butyrylcholinesterase. These compounds were not submitted to cholinesterase hydrolysis and turned out to be efficient reversible inhibitors that have to certain degree specificity toward the studied enzymes both by their potency and by the type of their inhibitory action. Specificity of effects of the key structural fragments of the anabasine grouping on anticholinesterase efficacy was checked. Also performed was analysis of action of 24 reversible and irreversible organophosphorus inhibitors, anabasine derivatives, toward erythrocyte acetylcholinesterase and serum butyrylcholinesterase. The anticholinesterase effects of anabasine-containing inhibitors differing in structure and action mechanism was compared.     

石蒜属植物生物碱成分研究进展

石蒜属为东亚特有属。该属植物富含生物碱,且此类次生代谢产物具有细胞毒、抗疟疾、抗病毒以及对乙酰胆碱酯酶的抑制作用等活性。本文概述了该属中生物碱类成分的化学结构、药理活性及其生源途径。     

Synthesis and biological evaluation of some dibenzofuran-piperazine derivatives

In the present paper, a novel series of dibenzofuran-piperazine derivatives were synthesized via the treatment of N-(2-methoxy-3-dibenzofuranyl)-2-chloroacetamide with substituted piperazine derivatives. The chemical structures of the compounds were elucidated by ¹H NMR, ¹³C NMR, mass spectral data; elemental analysis and HPLC analysis. Each derivative was evaluated for antiplatelet activity and anticholinesterase activity. Compound 2?m with 2-furoyl moiety exhibited high percentage inhibition as much as standard drug aspirin on arachidonic acid (AA)-induced platelet aggregation. None of the compounds presented significant inhibitor effect on collagen-induced platelet aggregation. Furthermore, the anticholinesterase activity of the compounds was determined and they did not show promising inhibitor activity compared with standard drug donepezil.     

Congenital Myasthenic Syndrome in the Dog Breed Gammel Dansk Hønsehund: Clinical,Electrophysiological, Pharmacological and Immunological Comparison with Acquired Myasthenia gravis

The effect of anticholinesterase drugs on the clinical and electrophysiological features in a canine congenital myasthenic syndrome is compared with findings in acquired myasthenia gravis in dogs. Anticholinesterase treatment had no effect on muscle weakness or electrophysiological parameters in the congenital myasthenic syndrome in contrast to its effect on clinical signs and electrophysiological parameters in acquired myasthenia gravis. The lack of effect of anticholinesterase in congenital myasthenia suggests a presynaptic defect as the aetiological factor. No antibodies to acetylcholine receptors were found in the Danish dog breed Gammel Dansk Hønsehund with the myasthenic syndrome. This classifies the disease in the group of canine and human congenital myasthenic diseases.     

Comparative analysis of sensitivity of proteases (chymotrypsin and trypsin) and cholinesterases of different origin to some organophosphorus inhibitors

The antichymotrypsin, antitrypsin, and anticholinesterase potencies of four homologous series of organophosphorus inhibitors are compared: O-(n-alkyl) methylthiophosphonates, O-(n-alkyl)-S-(n-butyl) methylthiophosphonates, O-(n-alkyl)-S-(β-ethylmercaptoethylene) methylth-iophosphonates, and their methylsulfomethylates. As sources of α-chymotrypsin and trypsin, commercial preparations from Worthington Biochemical Corporation and Leningrad Myasokombinat were tested. Bimolecular constant of the reaction rate was used as the measure of antienzyme potency. In all cases, the antichymotrypsin efficiency was lower, while the antitrypsin-essentially higher than the anticholinesterase activity of the studied inhibitors. These differences were found to much depend both on the inhibitor structure and on the nature of the cholinesterase preparations.     

Increased organophosphate scavenging in a butyrylcholinesterase mutant

Nicotiana benthamiana plant lines expressing a reengineered human butyrylcholinesterase (BChE) with enhanced cocaine hydrolase activity were created. Subsequent purification and biochemical analysis revealed that compared to wild-type butyrylcholinesterase, the cocaine hydrolase displayed increased affinity to the organophosphate (OP) pesticides paraoxon (6.84x10(-10)M vs. 1.11x10(-8)M) and malaoxon (9.81x10(-8)M vs. 5.99x10(-7)M). Furthermore, the cocaine hydrolase retained identical anticholinesterase binding profiles for all other compounds tested. Thus we have demonstrated a potential large-scale production platform for a multivalent antidote for cocaine and anticholinesterase poisoning.     

Rapid induction of supersensitivity to muscarinic antagonists-induced motor excitation by continuous stimulation of cholinergic receptors

Following single or repeated treatment with the irreversible anticholinesterase, DFP or, during infusion of the muscarinic receptor agonist, oxotremorine, and the reversible anticholinesterase physostigmine, effects of challenges with muscarinic antagonists were studied in rats. The antagonists, atropine, scopolamine, benztropine, orphenadrine and trihexyphenidyl induced, to a low degree, limb-shakes (myoclonus) and stereotyped behaviors in normal rats. However, within 24-72 hr after the above pretreatments, this myoclonus was significantly enhanced. The anticholinergic-stereotypies were also increased but only by severe cholinergic pretreatment and at a time later than that for the myoclonus. Myoclonus and stereotypies are known to be produced by treatments which directly enhance serotonergic and dopaminergic activities, respectively. It is suggested that during prolonged cholinergic stimulation, the cholinergic-monoaminergic balance in the brain can be altered depending upon the degree of stimulation. This could be responsible for the observed differential onset of changes in the anticholinergic-behavioral responses, which could, in turn, be mediated by different monoaminergic (serotonin and dopamine) systems.