Mary Broadfoot Walker: 83 years since a historical discovery / Mary Broadfoot Walker: 83 anos desde uma descoberta histórica

ABSTRACT Mary Broadfoot Walker was a Scottish physician who, in 1935, described in great detail the effect of an anticholinesterase drug (physostigmine) on the signs and symptoms of myasthenia gravis. An original five-minutes movie is available online and the skepticism of her contemporary British medical doctors is understandable when the drastic effect of the treatment is shown in this movie. What Mary Walker taught us, more than eight decades ago, about myasthenia gravis continues to be the basis of a pharmacological diagnostic test and treatment of this disease.

RESUMO Mary Broadfoot Walker foi uma médica escocesa que em 1935 descreveu em grande detalhe o efeito de uma droga anticolinesterásica (fisostigmina) nos sinais e sintomas da myasthenia gravis. Um filme original com cinco minutos de duração está disponível online e a reação cética dos colegas médicos contemporâneos de Mary é compreensível dado o drástico efeito terapêutico mostrado neste filme. O que Mary Walker nos ensinou mais de oito décadas atrás continua a ser a base de um teste diagnóstico farmacológico e do tratamento da myasthenia gravis.

Insecticidal Activity of Ethyl Acetate Extracts from Culture Filtrates of Mangrove Fungal Endophytes

In the search for novel potent fungi-derived bioactive compounds for bioinsecticide applications, crude ethyl acetate culture filtrate extracts from 110 mangrove fungal endophytes were screened for their toxicity. Toxicity tests of all extracts against brine shrimp (Artemia salina) larvae were performed. The extracts with the highest toxicity were further examined for insecticidal activity against Spodoptera litura larvae and acetylcholinesterase (AChE) inhibition activity. The results showed that the extracts of five isolates exhibited the highest toxicity to brine shrimp at 50% lethal concentration (LC50) values of 7.45 to 10.24 ppm. These five fungal isolates that obtained from Rhizophora mucronata were identified based on sequence data analysis of the internal transcribed spacer region of rDNA as Aspergillus oryzae (strain BPPTCC 6036), Emericella nidulans (strains BPPTCC 6035 and BPPTCC 6038), A. tamarii (strain BPPTCC 6037), and A. versicolor (strain BPPTCC 6039). The mean percentage of S. litura larval mortality following topical application of the five extracts ranged from 16.7% to 43.3%. In the AChE inhibition assay, the inhibition rates of the five extracts ranged from 40.7% to 48.9%, while eserine (positive control) had an inhibition rate of 96.8%, at a concentration of 100 ppm. The extracts used were crude extracts, so their potential as sources of AChE inhibition compounds makes them likely candidates as neurotoxins. The high-performance liquid chromatography profiles of the five extracts differed, indicating variations in their chemical constituents. This study highlights the potential of culture filtrate ethyl acetate extracts of mangrove fungal endophytes as a source of new potential bioactive compounds for bioinsecticide applications.

Pattern of Maxillofacial Fractures in Severe Multiple Trauma Patients: A 7-year Prospective Study

The incidence of facial trauma is high. This study has the primary objective of documenting and cataloging maxillofacial fractures in polytrauma patients. From a total of 1229 multiple trauma cases treated at the Emergency Room of the Santo Antonio Hospital - Oporto Hospital Center, Portugal, between August 2001 and December 2007, 251 patients had facial wounds and 209 had maxillofacial fractures. Aged ranged form 13 to 86 years. The applied selective method was based on the presence of facial wound with Abbreviated Injury Scale ≥1. Men had a higher incidence of maxillofacial fractures among multiple trauma patients (86.6%) and road traffic accidents were the primary cause of injuries (69.38%). Nasoorbitoethmoid complex was the most affected region (67.46%) followed by the maxilla (57.42%). The pattern and presentation of maxillofacial fractures had been studied in many parts of the world with varying results. Severe multiple trauma patients had different patterns of maxillofacial injuries. The number of maxillofacial trauma is on the rise worldwide as well as the incidence of associated sequelae. Maxillofacial fractures on multiple trauma patients were more frequent among males and in road traffic crashes. Knowing such data is elementary. The society should have a key role in the awareness of individuals and in prevention of road traffic accidents.

É alta a incidência de traumas na face. Este estudo teve por objetivo documentar e catalogar as fraturas maxilofaciais em pacientes com politraumatismos. De um total de 1229 casos de politraumatizados tratados na Sala de Emergência do Hospital de Santo António - Centro Hospitalar do Porto, Portugal, entre Agosto de 2001 e Dezembro de 2007, 251 pacientes tiveram ferimentos na face e 209 apresentaram fraturas maxilofaciais. As idades variaram de 13 a 86 anos. O método de seleção baseou-se na presença de ferimentos na face com Abreviated Injury Scale ≥1. Os homens apresentaram maior incidência de fraturas maxilofaciais (86,6%) entre os pacientes com múltiplos traumatismos na face e os acidentes de trânsito foram a causa principal dos traumatismos (69,38%). A região mais afetada foi o complexo naso-órbito-etmoidal (67,46%), seguido pela maxila (57,42%). O padrão e a apresentação das fraturas maxilofaciais tem sido estudado em muitas regiões do mundo com resultados variados. Pacientes com politraumatizados graves apresentaram padrões diferentes de traumatismos maxilofaciais. O número de traumatismos maxilofaciais tem aumentado à escala mundial, assim como a incidência das sequelas associadas. Entre os pacientes com traumatismos múltiplos, a maioria pertencia ao sexo masculino, assim como a causa mais frequente foram os acidentes automobilísticos. É elementar o conhecimento destes dados. A sociedade tem um papel primordial nos cuidados individuais e na prevenção dos acidentes de trânsito.

Alzheimer's Disease: Clinical Trials and Future Perspectives / 대한정신약물학회지

Alzheimer's disease is increasingly common in elderly population with a large socioeconomic burden. Current available drugs for Alzheimer's disease are acetylcholinesterase inhibitors and N-methyl-D-aspartate receptor antagonist. Much effort is directed towards not just symptomatic treatments but disease-modifying treatments. Several drugs with differing targets and mechanisms of action are under development for the treatment of Alzheimer's disease. Phase III trials of dimebon, Ginkgo biloba, non-steroidal anti-inflammatory drugs, phenserine, statins, semagacestat, tarenflurbil, tramiprosate, valproate, xaliproden have been completed without demonstrating adequate efficacy. Encouraging results would be expected from ongoing phase III trials of bapineuzumab and solanezumab. The clinical trials for the disease-modifying treatment of Alzheimer's disease have resulted in both promise and disappointment.

Differences between Physostigmine- and Yohimbine-induced States Are Visualized in Canonical Space Constructed from EEG during Natural Sleep-wake Cycle in Rats

Although quantitative EEG parameters, such as spectral band powers, are sensitive to centrally acting drugs in dose- and time-related manners, changes of the EEG parameters are redundant. It is desirable to reduce multiple EEG parameters to a few components that can be manageable in a real space as well as be considered as parameters representing drug effects. We calculated factor loadings from normalized values of eight relative band powers (powers of 0.5, 1.0~2.0, 2.5~4.0, 4.5~5.5, 6.0~8.0, 8.5~12.0, 12.5~24.5, and 25~49.5 Hz bands expressed as ratios of the power of 0.5-49.5 Hz band) of EEG during pre-drug periods (11:00~12:00) by factor analysis and constructed a two-dimensional canonical space (reference canonical space) by canonical correlation analysis. Eight relative band powers of EEG produced by either physostigmine or yohimbine were reduced to two canonical scores in the reference canonical space. While changes of the band powers produced by physostigmine and yohimbine were too redundant to describe the difference between two drugs, locations of two drugs in the reference canonical space represented the difference between two drug's effects on EEG. Because the distance between two locations in the canonical space (Mahalanobis distance) indicates the magnitude of difference between two different sets of EEG parameters statistically, the canonical scores and the distance may be used to quantitatively and qualitatively describe the dose-dependent and time-dependent effects and also tell similarity and dissimilarity among effects. Then, the combination of power spectral analysis and statistical analysis may help to classify actions of centrally acting drugs.

A Case of Seizure in Diphenhydramine Overdose

A 21-year-old woman ingested 1,250 mg of diphenhydramine in a single overdose. Diphenhydramine, a rare ingredient in over-the-counter medication, is used to treat insomnia in Korea. Toxicity is usually limited to anticholinergic symptoms. The standard approach to therapy for the treatment of diphenhydramine overdose is supportive care, including physostigmines and sodium bicarbonates. Here, we review the literature and for the first time report a case of acute diphenhydramine overdosage in Korea, complicated with seizures.

The History of Myasthenia Gravis

Since Willis described 'fatigable weakness' in 1672, most physicians consider it as a kind of hysteria due to the inconsistent fluctuation of symptoms. Erb presented three cases of 'bulbal palsy' in the 1870s, and Oppenheim and Hopper considered myasthenia gravis as a disease similar to curare poisoning and as a disease induced by attack of the motor centers by intrinsic toxins, respectively. In 1903, Elliot suggested that a 'chemical substance' mediates the nerve impulses at synapse. However, it was not until 1921 that this was demonstrated by Loewi, who provided evidence from the famous two-frog-hearts experiment. Dale later revealed the substance to be acetylcholine, and he also suggested that myasthenia gravis is due to a problem with the motor end plate. In 1934, Walker was prompted by the resemblance between myasthenia gravis and curare poisoning to apply physostigmine, a curare-poisoning antidote, to a patient, which produced a dramatic result. Since then the use of anticholinesterase inhibitors has been adopted for standard therapeutic modality. Some prominent surgeons have also applied thymectomy as a surgical modality. The most recent focus of myasthenia gravis has been immunological. In 1960, Simpson proposed the autoimmune hypothesis, and Chang et al. showed that snake venom contained a selective antagonist of the nicotinic acetylcholine receptor, alpha-bungarotoxin. The immunization of rabbits with acetylcholine receptor purified from the electrical organs of electric eels by Patrick et al. induced myasthenic symptoms and signs, and these were reversed by acetylcholinesterase inhibitors. The role of the autoimmune system has led to the introduction of an immunosuppressive modality and plasma exchange to the field of clinical neurology.

Correlative Changes of Endothelial Nitric Oxide Synthase and Choline Acetyltransferase in the Hippocampus after Exercise / 대한해부학회지

We found that the expression and activity of endothelial nitric oxide synthase (eNOS) is increased in the hippocampus during exercise (Moon et al., 2006). However, the upstream regulatory factor on the eNOS expression in the hippocampus during exercise has not been clear. In this study, we investigate the role of acetylcholine (ACh) as a regulatory factor for the eNOS expression and activity in the hippocampus during exercise. The results of the present study demonstrate that voluntary wheel running exercise for two weeks increases the expression and activity eNOS. In addition, choline acetyltransferase (ChAT) immnunoreacitvity within the hippocampus was increased after 2 weeks exercise. We further found that the upregulation of ACh with treatment of physostigmine, a booster of ACh releasing, increase the expression and activity of eNOS in the hippocampus. This present study provides the evidence that the upregulation of eNOS during exercise may be mediated by ACh in the hippocampus.

Effect of the carbamate [physostigmine] on the activity of nitric oxide synthase in different parts of rat brain [invivo studies]

Physostigmine [eserine] is the methyl carbamic ester of phenolic trimethyl ammonium compound. It is a powerful inhibitor of acetylcholinesterase enzyme activity and used in some therapeutic preparation. In this work, experiments were carried out in-vivo to study the intraperitoneal [i.p.] infusion of eserine on the activity of nitric oxide synthase [NOS] enzyme, obtained from whole and five different parts of rat brain, namely: basal ganglia, frontal cortex medulla oblongata, pons, and cerebellum. In this work ,two experiments were carried Experiment A: to study the dose dependence of i.p. infusion of eserine on NOS activity. Experiment B: to study the time dependence post-infusion of constant dose of eserine [the dose which caused 50% inhibition of the enzyme activity: 150]. The results showed that the "inhibition of the enzyme occurred in each part studied, and the inhibition increased with increasing the infused dose of eserine, and the time post i.p. infusion, i.e., the inhibition is dose and time dependent. The highest inhibition occurred in the pons and medulla oblongata extracts; these parts are responsible for the reflex centers of cough and vital centers

[Effects of intramedial septum infusion of physostigmine on hippocamal EEG and spatial memory in animal model of Alzheimer's disease]

The basalis magnocellularis nucleus [NBM] cholinergic projections to amygdala and forntal cortex have a crucial role in spatial learning and memory. There are relations between septum, hippocampus, amygdala and cerebral cortex. The role of NBM cholinergic projections to medial septum and then to hippocampus on spatial learning and memory, hippocamal EEG in animal model of Alzheimer's disease was assessed after unilateral lesion of NBM with phtalic acid [300 ng/kg]. Physostigmine was infused into the medial septum. Forty wistar male rats were divided in 4 groups: control, lesioned, lesioned received saline and lesioned treated with physostigmine [5microg/microl]. Animals were operated stereotaxicaly for NBM lesioning, intramedial septum cannulation and hippocamal electrode implantation. Rats were trained one session daily into T-maze and alterations of hippocampal EEG amplitude were evaluated. The results showed intramedial septum infusion of physostigmine improves spatial learning and memory in lesioned animals significantly [p<0.01]. NBM cholinergic projections to medial septum and then the hippocampus as well as its projections to amygdala and cortex have a role in spatial learning and memory. Administration of physostigmine improves decrease of hippocampal EEG amplitiude, spatial learning and memory impairment that was induced by NMB lesioning in male rats

Vias dopaminérgicas e somatostatinérgicas diminuem o TSH sérico em ratos portadores de carcinoma mamário walker-256 / Dopaminergic and somatostatinergic pathways decrease serum thyrotropin in rats bearing the 256-walker mammary carcinoma

A funcão do eixo hipotálamo-hipófise-tireóide em animais portadores da "síndrome do T3 baixo", foi estudada em ratos implantados com o tumor de Walker-256. Ratos machos adultos foram injetados com 1 x 106 células tumorais viáveis, por via SC, e sacrificados após 10 dias. A intensidade da síndrome guardou relacão positiva com o tamanho do tumor desenvolvido. Houve diminuicão da atividade tireoideana documentada pela diminuicão da área nuclear das células foliculares, das concentracões plasmáticas do T4, da rTg e da captacão do 131I. Mesmo o implante SC de um pellet de TSH de liberacão lenta causou menor estimulacão tireoideana, avaliada após 2 e 24h nos ratos com tumor. A secrecão do rTSH avaliada através da administracão IV de TRH mostrou-se significativamente diminuída nestas condicões, indicando aumento do tônus inibidor hipotalâmico sobre a secrecão deste hormônio. A participacão de outros neuro-mediadores hipotalâmicos foi verificada através da administracão prévia de metoclopramida e/ou fisostigmina, com ou sem estímulo subseqüente pelo TRH. Nos animais tratados com metoclopramida, os valores do rTSH aumentaram significativamente, assim como a resposta ao estímulo de secrecão pelo TRH. A fisostigmina mostrou-se mais eficiente na mediacão da resposta de secrecão do rTSH, bem como na resposta ao estímulo de secrecão pelo TRH. A administracão concomitante dos dois fármacos, seguida do estímulo pelo TRH, normalizou a secrecão do rTSH. Conclui-se que, além das alteracões conhecidas do metabolismo das iodotironinas, a secrecão de TSH encontra-se diminuída nos animais portadores de tumor de Walker-256, sugerindo diminuicão global do tônus tireoideano.

Sildenafil improves acquisition and retention of memory in mice.

Sildenafil (Viagra) has been introduced recently in market to correct male impotency and has gained immense popularity for its dramatic effects all over the world. The present study was designed to investigate the effect of sildenafil on learning and memory in mice using elevated plus maze. A total of XV groups of animals were employed in the present study. Central cholinergic pathways play a crucial role in learning and memory processes. Physostigmine, an anticholinesterase agent (0.5 mg, 1.0 mg kg(-1), i.p) was employed for its memory enhancing property and alprazolam a benzodiazepine receptor agonist served as a memory-impairing agent. In the present study, alprazolam produced anterograde amnesia (at 0.5 mg kg(-1), i.p) and retrograde amnesia (at 0.25 mg, 0.5 mg, 0.75 mg kg(-1), i.p.) in separate groups of animals. Caffeine at 5 mg, 10 mg and 20 mg kg(-1), i.p. (an established psychostimulant) did not show any significant change in learning and memory of mice. Sildenafil (at 8 mg kg(-1), i.p.) administered 30 minutes prior to training on first day produced a marginal decrease in transfer latency time on first day; whereas, sildenafil (at 2 mg, 4 mg, 8 mg kg(-1), i.p.) administered immediately after training on first day produced a dose-dependent improvement of memory in mice. However, further studies need to be carried out to elucidate the underlying mechanism of sildenafil as a memory enhancer.

new, simple and economical approach to analyze the inhibition kinetics of acetylcholinesterase using tolserine

Tolserine is a novel and highly selective synthetic inhibitor of the enzyme acetylcholinesterase [AchE] that is being developed towards clinical trials for the treatment of Alzheimer's disease. We recently characterized its enzyme kinetics for inhibiting purified human erythrocyte AChE by utilizing classical methodology. In the current study, we describe a new approach to analyze its mechanism of inhibition of AChE. This was undertaken with purified human erythrocyte AChE using low to higher substrate concentrations in both the absence and presence of dual tolserine concentrations. The optical density of the generated reaction-product then was monitored during the initial reaction time after addition of each concentration of substrate. Thereafter, the new kinetic parameters [Kneidc, Kesic, Kslxx, Krss, Kslm and Kslx] were calculated from the resulting experimental data. These kinetic constants will hopefully aid our understanding of the mechanism of inhibittion and kinetic analysis of a variety of critical physiological enzymes by a wide assortment of inhibitors in vitro, during health, aging and disease. A low Kneidc value of 2.31 nM for tolserine indicates that it is a highly potent inhibitor of human erythrocyte AChE, which supports previous reports of its unusually high potency for inhibiting AChE both in vitro and in vivo, as compared to its structural analogues, physostigmine and phenserine

Influence of Tacrine on Catecholamine Secretion in the Perfused Rat Adrenal Gland

The present study was designed to clarify whether tacrine affects the release of catecholamines (CA) from the isolated perfused model of rat adrenal gland or not and to elucidate the mechanism of its action. Tacrine (3 X 10(-5)~3 X 10(-4) M) perfused into an adrenal vein for 60 min inhibited CA secretory responses evoked by ACh (5.32 X 10(-3) M), DMPP (a selective neuronal nicotinic agonist, 10(-4) M for 2 min) and McN-A-343 (a selective muscarinic M1-agonist, 10(-4) M for 2 min) in relatively dose- and time- dependent manners. However, tacrine failed to affect CA secretion by high K+ (5.6 X 10(-2) M). Tacrine itself at concentrations used in the present experiments did not also affect spontaneous CA output. Furthermore, in the presence of tacrine (10(-4) M), CA secretory responses evoked by Bay-K-8644 (an activator of L-type Ca2+ channels, 10(-4) M), but not by cyclopiazonic acid (an inhibitor of cytoplasmic Ca2+-ATPase, 10(-4) M), was relatively time-dependently attenuated. Also, physostigmine (10(-4) M), given into the adrenal gland for 60 min, depressed CA secretory responses evoked by ACh, McN-A-343 and DMPP while did not affect that evoked by high K+. Collectively, these results obtained from the present study demonstrate that tacrine greatly inhibits CA secretion from the perfused rat adrenal gland evoked by stimulation of cholinergic (both nicotinic and muscarinic) receptors, but does fail to affect that by direct membrane-depolarization. It is suggested that this inhibitory effect of tacrine may be exerted by blocking both the calcium influx into the rat adrenal medullary chromaffin cells without Ca2+ release from the cytoplasmic calcium store, that is relevant to the cholinergic blockade. Also, the mode of action between tacrine and physostigmine in rat adrenomedullary CA secretion seems to be similar.

Postcardiotomy Central Anticholinergic Syndrome: Report of A Case / 대한흉부외과학회지

Central anticholinergic syndrome is defined as an absolute or relative reduction in cholinergic activity in the central nervous system and has a wide variety of manifestations. It is associated with almost any drug given during anesthesia, except neuromuscular relaxants, and treated with the cholinesterase inhibitor physostigmine. The diagnosis of central anticholinergic syndrome is often made when symptoms resolve promptly after the administration of physostigmine. We present a case of a central anticholinergic syndrome diagnosed by treatment with physostigmine, in a patient who received closure of patent foramen ovale associated with stroke.

Central Anticholinergic Syndrome Confirmed with Physostigmine after Open Heart Surgery / 대한마취과학회지

Central anticholinergic syndrome (CAS) can be caused by many anesthetic drugs. Early diagnosis and treatment are very important because untreated CAS may result in a life-threatening condition. Physostigmine, though not available in Korea, is the only drug which can confirm and treat CAS. A forty five year old patient underwent open heart surgery due to patent foramen ovale. Anesthetic agents which were used for anesthetic induction and maintenance were midazolam, fentanyl and isoflurane. Following anesthesia, he showed irritated and excited behavior and delayed recovery from anesthesia more than 3 h after operation in the ICU, even though flumazenil and naloxone were given to rule out the residual anesthetic effect. After physostigmine 4 mg was administered intravenously, he calmed down and became more coherent. There was no evidence of neurologic deficit in the following brain MRI and neurologic examination. We report the first case of CAS confirmed with physostigmine in Korea.

Organophosphate Cholinesterase Inhibitor (Diisopropylfluorophosphate) Induces Acetylcholinesterase-mediated Nicotinic Receptor Facilitation

BACKGROUND: Cholinesterase inhibitors (ChEIs) which have been widely used clinically are known to have diverse actions on the neuromuscular synaptic transmissions, suggesting that inhibiting cholinesterase (ChE) might not be their only mode of action. ChEIs interact with the nicotinic acetylcholine receptor (nAChR) macromolecule as a weak agonist, and as a modulator inducing desensitization and blockade at high concentrations. In a previous study, we reported that carbamate ChEIs, Pyridostigmine and Physostigmine could facilitate the ionic influx through nAChRs, when precluding the Ach-hydrolyzing effect of acetylChE (AChE) by applying carbachol as an agonist. The facilitation of the nAChR function was supposed to be achieved by AChE-mediated nAChR modulation and possibly by the up-regulation of nAChRs. METHODS: In this study, we analyzed the effect of irreversible organophosphate ChEI, diisopropylfluorophosphate (DFP) on the function of muscular nAChRs in TE671 cells, quantifying carbachol-induced intracellular 22 Na+ influx through nAChRs, using radioassay. RESULTS: Preincubation of cells with 1 mM DFP at 37 degrees C for 10 min as well as the simultaneous application of carbachol and DFP, decreased the carbachol-induced influx dose-dependently.However, preincubation of cells with 10 micrometer DFP potentiated the influx to 132.5+/-7.4% CPM. Moreover, Najar Tx completely inhibited the potentiated 22 Na + influx. CONCLUSIONS: Organophosphate ChEI can facilitate nAChR functions at low concentrations with a yet discovered mechanism, which is supposed to necessitate cellular metabolism, and be possibly mediated by AChE. The inhibition of DFP on nAChR functions at high concentration is attributable to its remained curare-like actions and direct cellular toxicity.

The Antinociceptive Effect of Intrathecal Anticholinesterase on the Formalin Test in Rats / 대한마취과학회지

BACKGROUND: Spinal anticholinesterase has been shown to have an antinociceptive action to acute noxious stimuli. The purpose of this study was to determine the effect of intrathecal anticholinesterase on the facilitated state developed after tissue injury evoked by formalin injection. METHODS: Rats were implanted with lumbar intrathecal catheters. For nociceptive test, 50 microliter of 5% formalin solution was injected into the hindpaw. The effect of pretreatment with intrathecal neostigmine, physostigmine and edrophonium, administered 10 min before formalin injection, was observed for 60 min. For the evaluation of the effect of posttreatment with intrathecal anticholinesterase, administered 9 min after formalin injection, the response was observed for 50 min. RESULTS: Formalin injection into the paw resulted in a biphasic incidence of flinching of the injected paw. Intrathecal pretreatment with neostigmine, physostigmine and edrophonium produced a dose- dependent suppression of the flinching during phase 1 and phase 2 on the formalin test. Posttreatment with three intrathecal anticholinesterases reduced the phase 2 flinching response. CONCLUSIONS: Both pretreatment and posttreatment with intrathecal anticholinesterase produced an antinociception on the formalin test. These results point out the usefulness of anticholinesterase to acute nociception and facilitated state.