Eldefrawi AT


Full name : Eldefrawi Amira T

First name : Amira T

Mail : University of Maryland School of Medicine, 655 W. Baltimore Street, Baltimore, Maryland 21201

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References (47)

Title : Design, synthesis and pharmacological profile of novel dopamine D2 receptor ligands - Menegatti_2003_Bioorg.Med.Chem_11_4807
Author(s) : Menegatti R , Cunha AC , Ferreira VF , Pereira EF , El-Nabawi A , Eldefrawi AT , Albuquerque EX , Neves G , Rates SM , Fraga CA , Barreiro EJ
Ref : Bioorganic & Medicinal Chemistry , 11 :4807 , 2003
Abstract : The present study describes the synthesis and pharmacological profile of three novel heterocyclic compounds originally designed, on the basis of bioisosterism, as dopamine D2 receptor ligands: 1-[1-(4-chlorophenyl)-1H-pyrazol-4-ylmethyl]-4-phenyl-piperazine (LASSBio-579), 1-phenyl-4-(1-phenyl-1H-[1,2,3]triazol-4-ylmethyl)-piperazine (LASSBio-580) and 1-[1-(4-chlorophenyl)-1H-[1,2,3]triazol-4-ylmethyl]-4-phenyl-piperazine (LASSBio-581). Binding studies performed on brain homogenate indicated that all three compounds bind selectively to D2 receptors. In addition, electrophysiological studies carried out in cultured hippocampal neurons suggested that LASSBio-579 and 581 act as D2 agonists, whereas LASSBio-580 acts as a D2 antagonist.
ESTHER : Menegatti_2003_Bioorg.Med.Chem_11_4807
PubMedSearch : Menegatti_2003_Bioorg.Med.Chem_11_4807
PubMedID: 14556797

Title : Cytotoxicity of organophosphate anticholinesterases - Cao_1999_In.Vitro.Cell.Dev.Biol.Anim_35_493
Author(s) : Cao CJ , Mioduszewski RJ , Menking DE , Valdes JJ , Katz EJ , Eldefrawi ME , Eldefrawi AT
Ref : In Vitro Cell Developmental Biology Anim , 35 :493 , 1999
Abstract : Organophosphate (OP) anticholinesterases were found to modulate metabolic activities of human neuroblastoma cells and hepatocytes, which was detectable by the Cytosensor microphysiometer. The nerve gas ethyl-S-2-diisopropylaminoethyl methylphosphorothiolate (VX), at 10 microM, produced significant reduction in cell metabolism within 2 min, as measured by changes in the acidification rate of the medium. The reduction was dose- and time-dependent and irreversible after 4 h of exposure. Two alkaline degradation products of VX produced no cytotoxicity. Exposure for 24 h to 3 microM VX caused 36% and 94% irreversible loss of metabolism in hepatocytes and neuroblastoma cells, respectively. The insecticides parathion and chlorpyrifos stimulated hepatocyte metabolism but inhibited neuroblastoma cells. Their oxons were more active. Exposure of neuroblastoma cells for 4 h to VX, parathion, paraoxon, diisopropylfluorophosphate or chlorpyrifos gave an LC50 of 65, 775, 640, 340, or 672 microM, respectively, whereas 24 h gave an LC50 of 0.7, 3.7, 2.5, 29, and 31 microM, respectively. Preincubation of hepatocytes with phenobarbital enhanced their response to parathion and VX due to metabolic bioactivation. Atropine partially blocked the effects of VX and paraoxon on both cell types, which suggests the involvement of a muscarinic receptor as the target for cytotoxicity. There was no correlation between OP in vivo neurotoxicity and in vitro cytotoxicity. It is suggested that the former results from their cholinesterase inhibition, while the latter results from action on different targets and requires much higher concentrations.
ESTHER : Cao_1999_In.Vitro.Cell.Dev.Biol.Anim_35_493
PubMedSearch : Cao_1999_In.Vitro.Cell.Dev.Biol.Anim_35_493
PubMedID: 10548430

Title : Common mechanism of toxicity: a case study of organophosphorus pesticides - Mileson_1998_Toxicol.Sci_41_8
Author(s) : Mileson BE , Chambers JE , Chen WL , Dettbarn W , Ehrich M , Eldefrawi AT , Gaylor DW , Hamernik K , Hodgson E , Karczmar AG , Padilla S , Pope CN , Richardson RJ , Saunders DR , Sheets LP , Sultatos LG , Wallace KB
Ref : Toxicol Sci , 41 :8 , 1998
Abstract : The Food Quality Protection Act of 1996 (FQPA) requires the EPA to consider "available information concerning the cumulative effects of such residues and other substances that have a common mechanism of toxicity ... in establishing, modifying, leaving in effect, or revoking a tolerance for a pesticide chemical residue." This directive raises a number of scientific questions to be answered before the FQPA can be implemented. Among these questions is: What constitutes a common mechanism of toxicity? The ILSI Risk Science Institute (RSI) convened a group of experts to examine this and other scientific questions using the organophosphorus (OP) pesticides as the case study. OP pesticides share some characteristics attributed to compounds that act by a common mechanism, but produce a variety of clinical signs of toxicity not identical for all OP pesticides. The Working Group generated a testable hypothesis, anticholinesterase OP pesticides act by a common mechanism of toxicity, and generated alternative hypotheses that, if true, would cause rejection of the initial hypothesis and provide criteria for subgrouping OP compounds. Some of the alternative hypotheses were rejected outright and the rest were not supported by adequate data. The Working Group concluded that OP pesticides act by a common mechanism of toxicity if they inhibit acetylcholinesterase by phosphorylation and elicit any spectrum of cholinergic effects. An approach similar to that developed for OP pesticides could be used to determine if other classes or groups of pesticides that share structural and toxicological characteristics act by a common mechanism of toxicity or by distinct mechanisms.
ESTHER : Mileson_1998_Toxicol.Sci_41_8
PubMedSearch : Mileson_1998_Toxicol.Sci_41_8
PubMedID: 9520337

Title : Toxicity of sea nettle toxin to human hepatocytes and the protective effects of phosphorylating and alkylating agents - Cao_1998_Toxicon_36_269
Author(s) : Cao CJ , Eldefrawi ME , Eldefrawi AT , Burnett JW , Mioduszewski RJ , Menking DE , Valdes JJ
Ref : Toxicon , 36 :269 , 1998
Abstract : The sea nettle jellyfish toxin (SNTX), which contains several polypeptides, was highly toxic to human hepatocytes. The Cytosensor microphysiometer was used continuously to monitor cell media acidification rate as an index of cellular metabolic activity. Cells exposed to > 1 microg SNTX protein/ml media exhibited a transient increase in metabolic activity, followed by a sharp decrease and cell death within minutes. The kinetics of the transient increase and subsequent decline increased with higher concentrations of SNTX. The biphasic and time-dependent response of hepatocytes to SNTX suggests that more than one mechanism may be involved in the toxicity of its different polypeptides. SNTX-induced cytotoxicity of hepatocytes was reduced by the presence of high titer antibodies against a heterologous jellyfish. Phenobarbital-induced cells became more vulnerable to SNTX, suggesting that some toxin component(s) require(s) bioactivation. Short-term exposure (1-2 h) to 10 microg/ml of the calcium ionophore calcimycin, or the non-selective monovalent cation ionophore gramicidin, had no effect on metabolic activity. However, 165 microg/ml gramicidin or 53 microg/ml calcimycin produced slight transient activation followed by steady decline in metabolic activity, while 20 h exposure to either ionophore produced total cell death. Exposure to even a 10-fold lower concentration of either ionophore killed 88% and 75%, respectively. This contrasts with the toxicity of SNTX which is detectable in minutes with as little as 3 microg/ml. Since pre-exposure to the organophosphate anticholinesterases VX and paraoxon, or the chemotherapeutic alkylating agents cyclophosphamide and mechlorethamine reduced the cytotoxic effects of SNTX, it suggests that phosphorylation or alkylation of cell protein(s) interferes with SNTX toxicity.
ESTHER : Cao_1998_Toxicon_36_269
PubMedSearch : Cao_1998_Toxicon_36_269
PubMedID: 9620575

Title : Chlorpyrifos, parathion, and their oxons bind to and desensitize a nicotinic acetylcholine receptor: relevance to their toxicities - Katz_1997_Toxicol.Appl.Pharmacol_146_227
Author(s) : Katz EJ , Cortes VI , Eldefrawi ME , Eldefrawi AT
Ref : Toxicol Appl Pharmacol , 146 :227 , 1997
Abstract : The nicotinic acetylcholine receptor (nAChR) of the electric organ of the electric ray. Torpedo sp., the richest source of nAChR, with similar structure and pharmacology to the mammalian skeletal muscle nAChR, carries several binding sites for different ligands. Incubation of Torpedo membrane-bound nAChRs with the agonist carbamylcholine (Carb) stimulated the binding of [3H]thienyl-cyclohexylpiperidine ([3H]TCP), which binds to the receptor's noncompetitive antagonist binding site in its ionic channel, with high affinity (Kd of 196 nM). The agonist-stimulated binding of [3H]TCP (i.e., binding to activated nAChRs) was inhibited in a concentration-dependent manner by four organophosphate (OP) anticholinesterases, chlorpyrifos oxon (CPO), chlorpyrifos (CPS), parathion (PS), and paraoxon (PO) with IC50 (concentration that inhibits 50% of the effect) values of 5, 150, 200, and 300 microM, respectively. The binding of CPO was totally reversible. The OPs had no effect on equilibrium binding of [alpha-125I]bungarotoxin ([alpha-125I]BGT) to the receptor's acetylcholine (ACh)-binding site, but preincubation of the membranes with the OPs increased this site's affinity for Carb. In absence of agonist, 100 microM of the OPs increased the binding of [3H]TCP by two- to fivefold with the following order of decreasing potency: PS > CPO > CPS > PO. The data suggest that in addition to inhibition of acetylcholinesterase, these OPs bind to a site on the nAChR that is different from the sites that bind ACh or TCP and that this binding induces nAChR desensitization. The relevance of this direct action of OPs on nAChRs on their acute toxicities is discussed.
ESTHER : Katz_1997_Toxicol.Appl.Pharmacol_146_227
PubMedSearch : Katz_1997_Toxicol.Appl.Pharmacol_146_227
PubMedID: 9344890

Title : Differential regulation of muscarinic receptor subtypes in rat brain regions by repeated injections of parathion - Jett_1994_Toxicol.Lett_73_33
Author(s) : Jett DA , Fernando JC , Eldefrawi ME , Eldefrawi AT
Ref : Toxicol Lett , 73 :33 , 1994
Abstract : Repeated injections with increasing moderate doses of parathion into adult male rats for 21 days resulted in 84-90% inhibition of acetylcholinesterase in the brain without overt signs of toxicity. Muscarinic acetylcholine receptor (mAChR) affinities for ligands were unaffected, but there was significant down-regulation of the m4 receptor subtype gene product, m1 mRNA and m3 mRNA in the frontal cortex as well as the m4 subtype and m4 mRNA in the striatum. However, in the hippocampus, there were no significant reductions in either the m1 receptor subtype nor its mRNA. The data suggest that the receptor subtype down-regulations in the cortex and striatum are due to reductions in mRNA expression. Since the degrees of inhibition of acetylcholinesterase were similar in the 3 brain regions, it is suggested that the in situ concentrations of paraoxon were also similar. Accordingly, the absence of down-regulation of the m1 receptor in the hippocampus is not due to a lower concentration of paraoxon than in the cortex or striatum. It is possible that injections of higher parathion doses would produce down-regulation of mAChRs in the hippocampus, and that the hippocampus may have differences in the feed-back mechanisms for receptor regulation from those in the frontal cortex and the striatum.
ESTHER : Jett_1994_Toxicol.Lett_73_33
PubMedSearch : Jett_1994_Toxicol.Lett_73_33
PubMedID: 8042201

Title : Choline derivatives and sodium fluoride protect acetylcholinesterase against irreversible inhibition and aging by DFP and paraoxon - Dehlawi_1994_J.Biochem.Toxicol_9_261
Author(s) : Dehlawi MS , Eldefrawi AT , Eldefrawi ME , Anis NA , Valdes JJ
Ref : Journal of Biochemical Toxicology , 9 :261 , 1994
Abstract : A light addressable potentiometric sensor was used to measure acetylcholinesterase (AChE) activity in order to evaluate the protective effects of quaternary compounds and NaF against enzyme phosphorylation and aging by two organophosphates. The use of the immobilized AChE made possible the quick removal of reagents (i.e., organophosphate, 2-pralidoxime, and protectant), thereby permitting accurate determination of AChE activity before and after phosphorylation and aging. Paraoxon was 15-fold more potent in inhibiting AChE than DFP, while the percent aging following phosphorylation by diisopropylfluorophosphate (DFP) was much higher. Sodium fluoride (NaF), the most effective protectant against phosphorylation and aging, and the quaternary ammonium compounds reduced significantly AChE inhibition by DFP and paraoxon, to similar degrees. Even though the percent AChE activity that was lost to aging was reduced by these agents, aging as a percent of phosphorylated AChE was not reduced. Thus, their major effect was in reducing the percent AChE phosphorylation, which consequently resulted in reduction of total aged AChE. The finding that quaternary ammonium compounds protect against phosphorylation is consonant with the proposed presence of the active site of AChE in an aromatic gorge.
ESTHER : Dehlawi_1994_J.Biochem.Toxicol_9_261
PubMedSearch : Dehlawi_1994_J.Biochem.Toxicol_9_261
PubMedID: 7853361

Title : Down-regulation of muscarinic receptors and the m3 subtype in white-footed mice by dietary exposure to parathion - Jett_1993_J.Toxicol.Environ.Health_39_395
Author(s) : Jett DA , Hill EF , Fernando JC , Eldefrawi ME , Eldefrawi AT
Ref : J Toxicol Environ Health , 39 :395 , 1993
Abstract : The effect of ad libitum dietary exposure (as occurs in the field) to parathion for 14 d was investigated on the muscarinic acetylcholine receptor (mAChR) in brains and submaxillary glands of adults of a field species, the white-footed mouse Peromyscus leucopus. Immunoprecipitation using subtype selective antibodies revealed that the relative ratios of the m1-m5 mAChR subtypes in Peromyscus brain were similar to those in rat brain. There was little variability in acetylcholinesterase (AChE) activity in control mice brains but large variability in 39 exposed mice, resulting from differences in food ingestion and parathion metabolism. Accordingly, data on radioligand binding to mAChRs in each mouse brain were correlated with brain AChE activity in the same mouse, and AChE inhibition served as a biomarker of exposure reflecting in situ paraoxon concentrations. Exposure to parathion for 14 d reduced maximal binding (Bmax) of [3H]quinuclidinyl benzilate ([3H]QNB), [3H]-N-methylscopolamine ([3H]NMS), and [3H]-4-diphenylacetoxy-N-methylpiperidine methiodide ([3H]-4-DAMP) by up to approximately 58% without affecting receptor affinities for these ligands. Maximal reduction in Bmax of [3H]QNB and [3H]-4-DAMP binding occurred in mice with highest AChE inhibition, while equivalent maximal reduction in Bmax of [3H]NMS occurred in mice with only approximately 10% AChE inhibition, without further change at higher parathion doses. This is believed to be due to the hydrophilicity of [3H]NMS, which limits its accessibility to internalized desensitized receptors. In submaxillary glands (mAChRs are predominantly m3 subtype), there were significant dose-dependent reductions in [3H]QNB binding and m3 mRNA levels in exposed mice, revealed by Northern blot analyses. The reduction in m3 receptors is suggested to result mostly from reduced synthesis at the transcription level, rather than from translational or posttranslational events. The data suggest that down-regulation of mAChRs occurs after dietary exposure for 14 d to sublethal concentrations of parathion in a field rodent species, and that significant though incomplete recovery in AChE and mAChRs occurs in 7 d following termination of exposure.
ESTHER : Jett_1993_J.Toxicol.Environ.Health_39_395
PubMedSearch : Jett_1993_J.Toxicol.Environ.Health_39_395
PubMedID: 8350385

Title : Differential effects of paraoxon on the M3 muscarinic receptor and its effector system in rat submaxillary gland cells - Abdallah_1992_J.Biochem.Toxicol_7_125
Author(s) : Abdallah EA , Jett DA , Eldefrawi ME , Eldefrawi AT
Ref : Journal of Biochemical Toxicology , 7 :125 , 1992
Abstract : The effects of the organophosphorus anticholinesterase paraoxon on the binding of radioactive ligands to the M3 subtype of the muscarinic receptor and receptor-coupled synthesis of second messengers in intact rat submaxillary gland (SMG) cells were investigated. The binding of [3H]quinuclidinyl benzilate ([3H]QNB) was most sensitive to atropine and the M3-specific antagonist 4-DAMP followed by pirenzepine and least sensitive to the cardioselective M2 antagonist AFDX116. This, and the binding characteristics of [3H]4-DAMP, confirmed that the muscarinic receptors in this preparation are of the M3 subtype. Activation of these muscarinic receptors by carbamylcholine (CBC) produced both stimulation of phosphoinositide (PI) hydrolysis and inhibition of cAMP synthesis, suggesting that this receptor subtype couples to both effector systems. Paraoxon (100 microM) reduced Bmax of [3H]4-DAMP binding from 27 +/- 4 to 13 +/- 3 fmol/mg protein with nonsignificant change in affinity, suggesting noncompetitive inhibition of binding by paraoxon. Like the agonist CBC, paraoxon inhibited the forskolin-induced cAMP formation in SMG cells with an EC50 of 200 nM, but paraoxon was greater than 500 fold more potent than CBC. However, while the inhibition by CBC was counteracted by 2 microM atropine, that by paraoxon was unaffected by up to 100 microM atropine. It suggested that this effect of paraoxon was not via binding to the muscarinic receptor. Paraoxon did not affect beta-adrenoreceptor function in the preparation, since it did not affect the 10 microM isoproterenol-induced cAMP synthesis, which was inhibited totally by 10 microM propranolol and partially by CBC. Paraoxon had a small but significant effect on CBC-stimulated PI metabolism in the SMG cells.
ESTHER : Abdallah_1992_J.Biochem.Toxicol_7_125
PubMedSearch : Abdallah_1992_J.Biochem.Toxicol_7_125
PubMedID: 1328639

Title : Acetylcholinesterase fiber-optic biosensor for detection of anticholinesterases - Rogers_1991_Fundam.Appl.Toxicol_16_810
Author(s) : Rogers KR , Cao CJ , Valdes JJ , Eldefrawi AT , Eldefrawi ME
Ref : Fundamental & Applied Toxicology , 16 :810 , 1991
Abstract : An optical sensor for anticholinesterases (AntiChEs) was constructed by immobilizing fluorescein isothiocyanate (FITC)-tagged eel electric organ acetylcholinesterase (AChE) on quartz fibers and monitoring enzyme activity. The pH-dependent fluorescent signal generated by FITC-AChE, present in the evanescent zone on the fiber surface, was quenched by the protons produced during acetylcholine (ACh) hydrolysis. Analysis of the fluorescence response showed Michaelis-Menten kinetics with a Kapp value of 420 microM for ACh hydrolysis. The reversible inhibitor edrophonium (0.1 mM) inhibited AChE and consequently reduced fluorescence quenching. The biosensor response immediately recovered upon its removal. The carbamate neostigmine (0.1 mM) also inhibited the biosensor response but recovery was much slower. In the presence of ACh, the organophosphate (OP) diisopropylfluorophosphate (DFP) at 0.1 mM did not interfere with the ACh-dependent fluorescent signal quenching, but preexposure of the biosensor to DFP in absence of ACh inhibited totally and irreversibly the biosensor response. However, the DFP-treated AChE biosensor recovered fully after a 10-min perfusion with pralidoxime (2-PAM). Echothiophate, a quaternary ammonium OP, inhibited the ACh-induced fluorescence quenching in the presence of ACh and the phosphorylated biosensor was reactivated with 2-PAM. These effects reflected the mechanism of action of the inhibitors with AChE and the inhibition constants obtained were comparable to those from colorimetric methods. The biosensor detected concentrations of the carbamate insecticides bendiocarb and methomyl and the OPs echothiophate and paraoxon in the nanomolar to micromolar range. Malathion, parathion, and dicrotophos were not detected even at millimolar concentrations; however, longer exposure or prior modification of these compounds (i.e., to malaoxon, paraoxon) may increase the biosensor detection limits. This AChE biosensor is fast, sensitive, reusable, and relatively easy to operate. Since the instrument is portable and can be self-contained, it shows potential adaptability to field use.
ESTHER : Rogers_1991_Fundam.Appl.Toxicol_16_810
PubMedSearch : Rogers_1991_Fundam.Appl.Toxicol_16_810
PubMedID: 1909249

Title : High-affinity activation by paraoxon of a muscarinic receptor subtype in rat brain striatum - Jett_1991_Pestic.Biochem.Physiol_39_149
Author(s) : Jett DA , Abdallah EAM , El-Fakahany EE , Eldefrawi ME , Eldefrawi AT
Ref : Pesticide Biochemistry and Physiology , 39 :149 , 1991
Abstract : The mechanism of action of the anticholinesterase paraoxon on the function of a muscarinic receptor subtype in rat brain striatum was investigated. Paraoxon inhibited binding of the muscarinic agonist cis-[3H]methyldioxolane, which binds to a high-affinity population of muscarinic receptors, with K0.5 of 80 nM, compared to a K0.5 of 7 uM for parathion. The inhibition was competitive, suggesting that paraoxon and CD bind to a common site. When this muscarinic receptor (possibly an M4 subtype) is activated it inhibits cAMP synthesis. Thus, function of the paraoxon-sensitive receptor was assayed by the inhibition of the forskolin-activated [3H]cAMP synthesis. Paraoxon inhibited cAMP synthesis in a dose-dependent manner as did the muscarinic agonist carbachol, and this inhibition was completely blocked by the muscarinic antagonist atropine. When low concentrations of carbachol and paraoxon were used together, there was additive inhibition of cAMP synthesis. However, there was no further increase when both paraoxon and carbachol were present in concentrations that individually produced maximal inhibition. The data suggest that paraoxon acts like carbachol, causing activation of the muscarinic receptor subtype in brain striatum and leading to inhibition of cAMP synthesis. Considering the high affinity that this muscarinic receptor has for paraoxon, it is suggested that this direct reversible action of paraoxon on the muscarinic receptor could affects its toxicity, expecially early on before most of the acetylcholinesterase is phosphorylated.
ESTHER : Jett_1991_Pestic.Biochem.Physiol_39_149
PubMedSearch : Jett_1991_Pestic.Biochem.Physiol_39_149

Title : Putative M2 muscarinic receptors of rat heart have high affinity for organophosphorus anticholinesterases - Silveira_1990_Toxicol.Appl.Pharmacol_103_474
Author(s) : Silveira CL , Eldefrawi AT , Eldefrawi ME
Ref : Toxicol Appl Pharmacol , 103 :474 , 1990
Abstract : The M2 subtype of muscarinic receptor is predominant in heart, and such receptors were reported to be located in muscles as well as in presynaptic cholinergic and adrenergic nerve terminals. Muscarinic receptors of rat heart were identified by the high affinity binding of the agonist (+)-[3H]cis-methyldioxolane ([3H]CD), which has been used to label a high affinity population of M2 receptors. A single population of sites (KD 2.74 nM; Bmax of 82 fmol/mg protein) was detected and [3H]CD binding was sensitive to the M2 antagonist himbacine but much less so to pirenzepine, the M1 antagonist. These cardiac receptors had different sensitivities to NiCl2 and N-ethylmaleimide from brain muscarinic receptors, that were also labeled with [3H]CD and considered to be of the M2 subtype. Up to 70% of the [3H]CD-labeled cardiac receptors had high affinities for several organophosphate (OP) anticholinesterases. [3H]CD binding was inhibited by the nerve agents soman, VX, sarin, and tabun, with K0.5 values of 0.8, 2, 20, and 50 nM, respectively. It was also inhibited by echothiophate and paraoxon with K0.5 values of 100 and 300 nM, respectively. The apparent competitive nature of inhibition of [3H]CD binding by both sarin and paraoxon suggests that the OPs bind to the acetylcholine binding site of the muscarinic receptor. Other OP insecticides had lower potencies, inhibiting less than 50% of 5 nM [3H]CD binding by 1 microM of EPN, coumaphos, dioxathion, dichlorvos, or chlorpyriphos. There was poor correlation between the potencies of the OPs in reversibly inhibiting [3H]CD binding, and their anticholinesterase activities and toxicities. Acetylcholinesterases are the primary targets for these OP compounds because of the irreversible nature of their inhibition, which results in building of acetylcholine concentrations that activate muscarinic and nicotinic receptors and desensitize them, thereby inhibiting respiration. Nevertheless, the high affinities that cardiac muscarinic receptors have for these toxicants point to their extra vulnerability. It is suggested that the success of iv administration of the muscarinic receptor inhibitor atropine in initial therapy of poisoning by OP anticholinesterases may be related in part to the extra sensitivity of M2 receptors to certain OPs.
ESTHER : Silveira_1990_Toxicol.Appl.Pharmacol_103_474
PubMedSearch : Silveira_1990_Toxicol.Appl.Pharmacol_103_474
PubMedID: 2339420

Title : Actions of the insecticide 2(nitromethylene)tetrahydro-1,3-thiazine on insect and vertebrate nicotinic acetylcholine receptors - Sattelle_1989_Proc.R.Soc.Lond.B.Biol.Sci_237_501
Author(s) : Sattelle DB , Buckingham SD , Wafford KA , Sherby SM , Bakry NM , Eldefrawi AT , Eldefrawi ME , May TE
Ref : Proc R Soc Lond B Biol Sci , 237 :501 , 1989
Abstract : The nitromethylene heterocyclic compound 2(nitromethylene)tetrahydro)1,3-thiazine (NMTHT) inhibits the binding of [125I]alpha-bungarotoxin to membranes prepared from cockroach (Periplaneta americana) nerve cord and fish (Torpedo californica) electric organ. Electrophysiological studies on the cockroach fast coxal depressor motorneuron (Df) reveal a dose-dependent depolarization in response to bath-applied NMTHT. Responses to ionophoretic application of NMTHT onto the cell-body membrane of motorneuron Df are suppressed by bath-applied mecamylamine (1.0 x 10(-4) M) and alpha-bungarotoxin (1.0 x 10(-7) M). These findings, together with the detection of a reversal potential close to that estimated for acetylcholine, provide evidence for an agonist action of this nitromethylene on an insect neuronal nicotinic acetylcholine receptor. The binding of [3H]H12-histrionicotoxin to Torpedo membranes was enhanced in the presence of NMTHT indicating an agonist action at this vertebrate peripheral nicotinic acetylcholine receptor. NMTHT is ineffective in radioligand binding assays for rat brain GABAA receptors, rat brain L-glutamate receptors and insect (Musca domestica) L-glutamate receptors. Partial block of rat brain muscarinic acetylcholine receptors is detected at millimolar concentrations of NMTHT. Thus nitromethylenes appear to exhibit selectivity for acetylcholine receptors and exhibit an agonist action at nicotinic acetylcholine receptors.
ESTHER : Sattelle_1989_Proc.R.Soc.Lond.B.Biol.Sci_237_501
PubMedSearch : Sattelle_1989_Proc.R.Soc.Lond.B.Biol.Sci_237_501
PubMedID: 2479949

Title : Allosteric inhibition of nicotinic acetylcholine receptors of vertebrates and insects by philanthotoxin - Rozental_1989_J.Pharmacol.Exp.Ther_249_123
Author(s) : Rozental R , Scoble GT , Albuquerque EX , Idriss M , Sherby S , Sattelle DB , Nakanishi K , Konno K , Eldefrawi AT , Eldefrawi ME
Ref : Journal of Pharmacology & Experimental Therapeutics , 249 :123 , 1989
Abstract : The effects of pure philanthotoxin (PhTX), a component of the venom of the wasp Philanthus triangulum, were studied on nicotinic acetylcholine receptors (nAChRs) of vertebrates and insects so as to compare their sensitivities and the mechanism of action of PhTX. Electrophysiological techniques were used on frog muscles and cockroach thoracic ganglia and biochemical techniques were applied to membranes from Torpedo electric organ and honeybee brain. PhTX (1-20 microM) inhibited reversibly the indirectly elicited muscle twitch and reduced the endplate current peak amplitude and its decay time constant in a concentration-dependent manner. In patch clamp studies, PhTX (1-5 microM) when combined with acetylcholine, induced a concentration-dependent decrease in frequency of channel openings and in channel open and burst times. The cockroach fast coxal depressor neuron was inhibited by PhTX in a time- and voltage-dependent manner. The initial rate of binding of [3H]perhydrohistrionicotoxin to Torpedo nAChR in the presence of carbamylcholine was inhibited competitively by PhTX. Binding of alpha-[125I] bungarotoxin to electric organ and honeybee brain membranes was inhibited by PhTX. Binding of [3H]acetylcholine to the electric organ receptor was potentiated by low concentrations of PhTX but inhibited by high concentrations. PhTX, therefore, inhibits both vertebrate and insect nAChRs, which may be important molecular targets for its toxicity. It is suggested that PhTX at high concentration may have some competitive action on nAChR, but it acts mainly as a blocker of the ion channel of the nAChR in its open conformation.
ESTHER : Rozental_1989_J.Pharmacol.Exp.Ther_249_123
PubMedSearch : Rozental_1989_J.Pharmacol.Exp.Ther_249_123
PubMedID: 2468760

Title : Direct actions of organophosphate anticholinesterases on nicotinic and muscarinic acetylcholine receptors - Bakry_1988_J.Biochem.Toxicol_3_235
Author(s) : Bakry NM , el-Rashidy AH , Eldefrawi AT , Eldefrawi ME
Ref : Journal of Biochemical Toxicology , 3 :235 , 1988
Abstract : Four nerve agents and one therapeutic organophosphate (OP) anticholinesterase (anti-ChE) bind to acetylcholine (ACh) receptors, inhibit or modulate binding of radioactive ligands to these receptors, and modify events regulated by them. The affinity of nicotinic (n) ACh receptors of Torpedo electric organs and most muscarinic (m) ACh receptors of rat brain and N1E-115 neuroblastoma cultures for the OP compounds was usually two to three orders of magnitude lower than concentrations required to inhibit 50% (IC-50) of ACh-esterase activity. However, a small population of m-ACh receptors had an affinity as high as that of ACh-esterase for the OP compound. This population is identified by its high-affinity [3H]-cis-methyldioxolane ([3H]-CD) binding. Although sarin, soman, and tabun had no effect, (O-ethyl S[2-(diisopropylamino)ethyl)] methyl phosphonothionate (VX) and echothiophate inhibited competitively the binding of [3H]-quinuclidinyl benzilate ([3H]-QNB) and [3H]-pirenzepine ([3H]-PZ) to m-ACh receptors. However, VX was more potent than echothiophate in inhibiting this binding and 50-fold more potent in inhibiting carbamylcholine (carb)-stimulated [3H]-cGMP synthesis in N1E-115 neuroblastoma cells--both acting as m receptor antagonist. All five OPs inhibited [3H]-CD binding, with IC-50s of 3, 10, 40, 100, and 800 nM for VX, soman, sarin, echothiophate, and tabun, respectively. The OP anticholinesterases also bound to allosteric sites on the n-ACh receptor (identified by inhibition of [3H]-phencyclidine binding), but some bound as well to the receptor's recognition site (identified by inhibition of [125I]-alpha-bungarotoxin binding). Soman and echothiophate in micromolar concentrations acted as partial agonists of the n-ACh receptor and induced receptor desensitization. On the other hand, VX acted as an open channel blocker of the activated receptor and also enhanced receptor desensitization. It is suggested that the toxicity of OP anticholinesterases may include their action on n-ACh as well as m-ACh receptors if their concentrations in circulation rise above micromolar levels. At nanomolar concentrations their toxicity is due mainly to their inhibition of ACh-esterase. However, at these low concentrations, many OP anticholinesterases (eg, VX and soman) may affect a small population of m-ACh receptors, which have a high affinity for CD. Such effects on m-ACh receptors may play an important role in the toxicity of certain OP compounds.
ESTHER : Bakry_1988_J.Biochem.Toxicol_3_235
PubMedSearch : Bakry_1988_J.Biochem.Toxicol_3_235
PubMedID: 3236334

Title : Action of organophosphates on GABAA receptor and voltage-dependent chloride channels - Gant_1987_Fundam.Appl.Toxicol_9_698
Author(s) : Gant DB , Eldefrawi ME , Eldefrawi AT
Ref : Fundamental & Applied Toxicology , 9 :698 , 1987
Abstract : The effects of several organophosphates were studied on the binding of t-[35S]butyl-bicyclophosphorothionate ([35S]TBPS) to rat brain GABAA receptor and receptor function as assayed by GABA-induced 36Cl-influx into membrane vesicles and on the binding of [35S]TBPS to a voltage-dependent Cl-channel in Torpedo californica electric organ. The organophosphate anticholinesterases diisopropylphosphorofluoridate, soman, sarin, tabun, and VX had little or no effect on GABA-regulated chloride channels. They also had no effect on [35S]TBPS binding to the voltage-dependent chloride channel, except for soman which inhibited it with an IC50 of 24 microM. Triphenyl phosphate was the only one of three organophosphate flame retardants tested that inhibited both GABA-regulated chloride channel and binding of [35S]TBPS to the voltage-dependent chloride channel with IC50s of 18 and 13 microM, respectively. The industrial organophosphate tri-o-cresyl phosphate and the anticholinesterase organophosphate insecticides leptophos, leptophos oxon, and O-ethyl O-4-nitrophenyl phenylphosphonothioate inhibited GABA-regulated chloride channels and bound with high affinity to the voltage-dependent chloride channels (IC50 = 0.3 to 8.7 microM). There was no apparent correlation between the affinities of the GABAA receptor chloride channel or the voltage-dependent chloride channel for the different organophosphates and their potencies in inhibiting acetylcholinesterase or in inducing delayed neurotoxicity. Nevertheless, although the voltage-dependent chloride channel and/or GABAA receptor are not primary targets for organophosphate anticholinesterases and flame retardants, it is suggested that the inhibition of these two proteins by certain organophosphates may contribute to their toxicities.
ESTHER : Gant_1987_Fundam.Appl.Toxicol_9_698
PubMedSearch : Gant_1987_Fundam.Appl.Toxicol_9_698
PubMedID: 2446940

Title : Oxadiazolidinones: irreversible inhibition of cholinesterases and effects on acetylcholine receptors - Bakry_1986_Neurotoxicol_7_1
Author(s) : Bakry NM , Sherby SM , Eldefrawi AT , Eldefrawi ME
Ref : Neurotoxicology , 7 :1 , 1986
Abstract : Inhibition of four acetylcholinesterases (AChE) and a butyrylcholinesterase (BCHE) by 3-(2,3-dihydro-2,2-dimethyl-benzofuran-'7-yl)-5-methoxy-1,3,4-oxadiaz ol-2(3H)-one (DBOX) and 3-(2-methoxyphenyl)-5-methoxy-1,3,4-oxadiazol-2(3H)-one (MPOX) was measured by the Ellman spectrophotometric method. Both oxadiazolidinones inhibited AChE and BCHE irreversibly and with quasi first order kinetics. DBOX was 2-3 orders of magnitude more potent than MPOX. Housefly brain AChE and horse serum BCHE were more sensitive than AChEs of red blood cells or eel and Torpedo electric organs. Aldicarb, a carbamate anticholinesterase, which protected Torpedo AChE against irreversible phosphorylation by DFP, also protected it against irreversible inhibition by DBOX and MPOX. It is suggested that the nonesteratic oxadiazolidinones are converted to carbanillates on the surface of the enzyme, then acylate the active site of ChEs, producing carbanillated enzymes. At higher concentrations, the two oxadiazolidinones also affected the specific binding of (125I) alpha-bungarotoxin (alpha-BGT) and [3H]perhydrohistrionicotoxin (H12-HTX) to Torpedo nicotinic ACh-receptors, but did not affect the specific binding of [3H]quinuclidinyl benzilate (QNB) to rat brain muscarinic ACh-receptors.
ESTHER : Bakry_1986_Neurotoxicol_7_1
PubMedSearch : Bakry_1986_Neurotoxicol_7_1
PubMedID: 3822252

Title : Comparison of the actions of carbamate anticholinesterases on the nicotinic acetylcholine receptor - Sherby_1985_Mol.Pharmacol_27_343
Author(s) : Sherby SM , Eldefrawi AT , Albuquerque EX , Eldefrawi ME
Ref : Molecular Pharmacology , 27 :343 , 1985
Abstract : Neostigmine (Neo), pyridostigmine (Pyr), and physostigmine (Phy) at low concentrations inhibited acetylcholine (ACh) esterase, thereby indirectly potentiating ACh enhancement of [3H]perhydrohistrionicotoxin (H12-HTX) binding to the channel sites of the nicotinic ACh receptor of Torpedo membranes. However, at higher concentrations, they inhibited ACh action due to their direct binding to the ACh receptor. They displaced binding of [3H]ACh and 125I-alpha-bungarotoxin (alpha-BGT) to the receptor sites with the following order of decreasing potency: Neo greater than Phy greater than Pyr. Furthermore, Neo and Pyr potentiated [3H] H12-HTX binding to the receptor's channel sites. Preincubation of ACh receptors with any of the three carbamates reduced the rate of binding of 125I-alpha-BGT and increased the potency of carbamylcholine in inhibiting 125I-alpha-BGT binding, suggesting that the three carbamates act as partial agonists and potentiate receptor desensitization. Although none of the three carbamates inhibited [3H]H12-HTX binding to the receptor's closed channel conformation, only Phy was a potent inhibitor of [3H]H12-HTX binding to the carbamylcholine-activated conformation. The potency of Phy was not due to the absence of positive charge since Phy methiodide acted similarly. The data suggest that the major action of the three carbamates at nicotinic cholinergic synapses is inhibition of ACh-esterase. Their interactions with the nicotinic ACh receptor are with its "receptor" as well as allosteric "channel" sites, but they differ in their effects. Neo and Pyr act mainly as partial agonists, while Phy is mostly an inhibitor of the channel in the activated receptor conformation.
ESTHER : Sherby_1985_Mol.Pharmacol_27_343
PubMedSearch : Sherby_1985_Mol.Pharmacol_27_343
PubMedID: 3974572

Title : Interactions of phencyclidine with crayfish muscle membranes. Sensitivity to calcium channel antagonists and other drugs - El-Fakahany_1984_Mol.Pharmacol_25_369
Author(s) : El-Fakahany EE , Eldefrawi AT , Murphy DL , Aguayo LG , Triggle DJ , Albuquerque EX , Eldefrawi ME
Ref : Molecular Pharmacology , 25 :369 , 1984
Abstract : [3H]Phencyclidine ( [3H]PCP) bound to crayfish abdominal muscle membranes at pH 7.4 with two affinities (Kd of 0.96 nM for 0.38 pmole/mg of protein, and 18.9 nM for 7.6 pmoles/mg of protein). Binding affinities increased at higher pH, suggesting that binding may be due mostly to the un-ionized form of [3H]PCP. This high-affinity [3H]PCP binding was sensitive to the actions of trypsin, protease, and dicyclohexylcarbodiimide, but insensitive to phospholipase A, concanavalin A,N-ethylmaleimide, and dithiothreitol. Calcium channel antagonists were most potent in inhibiting the high-affinity [3H]PCP binding with the following descending order of potencies: bepridil greater than nicardipine = diltiazem = verapamil greater than cinnarizine greater than (+)-D-600 greater than (-)-D-600 greater than 4-NO2-nifedipine greater than 2-NO2-nifedipine. The binding was also highly sensitive to several PCP analogues, antipsychotics, piperocaine , and tilorone, and moderately sensitive to d-tubocurarine, atropine, imipramine, nortryptyline , and tetracaine. Although verapamil and nifedipine inhibited the action potential of crayfish muscle, PCP did not and actually prolonged slightly the falling phase of the action potential. Although it is unlikely that the [3H]PCP binding protein in crayfish muscles is a Ca2+ channel, it is possible that it may be a K+ channel.
ESTHER : El-Fakahany_1984_Mol.Pharmacol_25_369
PubMedSearch : El-Fakahany_1984_Mol.Pharmacol_25_369
PubMedID: 6328263

Title : Neurotransmitter receptors as targets for pesticides - Eldefrawi_1983_J.Environ.Sci.Health.[B]_18_65
Author(s) : Eldefrawi ME , Eldefrawi AT
Ref : J Environ Sci Health [B] , 18 :65 , 1983
Abstract : Nicotinic and muscarinic acetylcholine (ACh) receptors have been identified biochemically by means of their specific binding of [3H] alpha-bungarotoxin ([3H]alpha-BGT) and [3H]quinuclidinyl benzilate, respectively. There are some differences in the drug specificities, and sensitivities to active group reagents, of these receptors in insects when compared to those in vertebrates. Also, insect brain contains more nicotinic than muscarinic receptors, while the reverse is found in mammalian brain. Insect brain contains a third kind of putative ACh-receptor that is relatively soluble and is both nicotinic and muscarinic in its pharmacology but does not bind alpha-BGT. Toxic nicotine and analogs bind to it with high affinities. Several organophosphorus and carbamate insecticides and nereistoxin bind with high affinities to the nicotinic ACh-receptor of the electric organ of Torpedo. A few chlorinated hydrocarbon insecticides and derivatives interact with Torpedo nicotinic ACh-receptors, not at their 'receptor' sites but at their allosteric or 'channel' sites (which are identified by their specific binding of [3H]perhydrohistrionicotoxin). A few also bind to mammalian brain muscarinic receptors. The most potent on both receptors is the acaricide chlorobenzilate. Pyrethrins and synthetic pyrethroids also bind with high affinities to the channel sites of the Torpedo nicotinic ACh-receptor, though not to its receptor sites. Another group that binds to ACh-receptors is the organic and inorganic mercury compounds, which interact with both the Torpedo nicotinic and rat brain muscarinic receptors. Thus, neurotransmitter receptors act as molecular targets, primary or secondary for different pesticides.
ESTHER : Eldefrawi_1983_J.Environ.Sci.Health.[B]_18_65
PubMedSearch : Eldefrawi_1983_J.Environ.Sci.Health.[B]_18_65
PubMedID: 6339601

Title : Interactions of d-tubocurarine with the nicotinic acetylcholine receptor\/channel molecule - Shaker_1982_J.Pharmacol.Exp.Ther_220_172
Author(s) : Shaker N , Eldefrawi AT , Aguayo LG , Warnick JE , Albuquerque EX , Eldefrawi ME
Ref : Journal of Pharmacology & Experimental Therapeutics , 220 :172 , 1982
Abstract : The interactions of d-tubocurarine (d-TC) with the ionic channel of the nicotinic acetylcholine receptor were studied by biochemical methods in Torpedo electric organ membranes and by electrophysiological methods on frog sciatic nerve-sartorius muscle preparation. Torpedo membranes were treated with alpha-bungarotoxin to inhibit the acetylcholine receptor sites, then binding of [3H]perhydrohistrionicotoxin to the ionic channel sites was studied and found to be inhibited by d-TC. At 37 degrees C, the Ki of d-TC was 10 microM, and at 22 degrees C it was 100 microM. The affinity of d-TC for the ionic channel sites relative to that of perhydrohistrionicotoxin was constant at temperatures from 2-20 degrees C, but increased at higher temperatures up to 37 degrees C. The peak endplate current amplitude was depressed with 1 to 2 microM d-TC in a voltage-dependent manner, with considerable departure from linearity at 10 and 30 degrees C. The effect of d-TC on spontaneous miniature endplate currents was similar and slightly more potent. The time constant of endplate current decay was decreased by d-TC (1 and 2 microM) at temperatures of 10, 15 and 30 degrees C. The channel lifetime was reduced by d-TC, but channel conductance was unaffected. It is suggested that d-TC interacts with both the acetylcholine receptor sites as well as its ionic channel sites in closed and open conformations.
ESTHER : Shaker_1982_J.Pharmacol.Exp.Ther_220_172
PubMedSearch : Shaker_1982_J.Pharmacol.Exp.Ther_220_172
PubMedID: 6273528

Title : Structure-activity relationships of amantadine. I. Interaction of the N-alkyl analogues with the ionic channels of the nicotinic acetylcholine receptor and electrically excitable membrane - Warnick_1982_Mol.Pharmacol_22_82
Author(s) : Warnick JE , Maleque MA , Bakry N , Eldefrawi AT , Albuquerque EX
Ref : Molecular Pharmacology , 22 :82 , 1982
Abstract : In this study the effects of amantadine (1-adamantanamine) and its N-alkyl-substituted analogues [N-methyl- (NMA), N-ethyl- (NEA), N-propyl- (NPA), N-butyl- (NBA), and N,N-diethyl-amantidine (NNDEA)] were investigated on ionic channels of the electrically excitable membrane and of the nicotinic acetylcholine (ACh) receptors in frog sartorius muscles and on the binding of perhydrohistrionicotoxin (H12-HTX) to isolated membranes of the electric organ of the electric ray Torpedo. Amantadine and each analogue blocked the indirectly elicited twitch, but NPA, NBA, and NNDEA also potentiated the directly elicited twitch. The order of potency in inhibiting the indirect twitch was: NEA = NPA = NNDEA (10 microM) greater than NMA (15 microM) greater than NBA (40 microM) much greater than amantadine (130 microM). Neither amantadine nor its N-alkyl analogues affected miniature end-plate potential frequency or resting membrane potential but decreased miniature end-plate potential amplitude. Each compound prolonged the directly elicited action potential but did not alter delayed rectification. All of the compounds induced a concentration-dependent depression of the peak end-plate current (EPC) amplitude at negative membrane potentials and induced nonlinearity in the response at membrane potentials more negative than -40 mV. The order of potency in inhibiting the EPC (at -90 mV) was NNDEA (less than 0.5 microM) greater than NPA (less than 1.0 microM) greater than NBA (less than 2.0 microM) greater than NEA (19 microM) greater than NMA (42 microM) greater than amantadine (64 microM). Only NPA, NBA, and NNDEA depressed the peak EPC amplitude at positive membrane potentials as well. The shortening of the time constant of EPC decay by all compounds was concentration-dependent. At the higher concentrations examined, the slope of the relationship between the time constant of decay and membrane potential was reversed for all compounds. Only NPA induced a double-exponential decay of the EPC at positive membrane potentials. Neither amantadine nor its N-alkyl analogues inhibited the binding of [3H]ACh to its receptor in Torpedo electroplax but they inhibited the binding of [3H]H12-HTX binding to the ionic channel sites of the ACh receptor. The Ki for inhibition of [3H]H12-HTX binding was NEA = NNDEA (15 microM) greater than NMA (30 microM) greater than NPA = NBA (40 microM) greater than amantadine (60 microM). A gross correlation exists between their ability to block the indirect muscle twitch, miniature end-plate potential amplitude, peak EPC amplitude and the binding of [3H]H12-HTX. But, no correlation was found between these potencies and their antiviral activity. It is suggested that these compounds may interact with the ionic channel of the ACh receptor in its open and closed conformation.
ESTHER : Warnick_1982_Mol.Pharmacol_22_82
PubMedSearch : Warnick_1982_Mol.Pharmacol_22_82
PubMedID: 6126806

Title : Regulation of [3H]perhydrohistrionicotoxin binding to Torpedo ocellata electroplax by effectors of the acetylcholine receptor - Aronstam_1981_J.Biol.Chem_256_2843
Author(s) : Aronstam RS , Eldefrawi AT , Pessah IN , Daly JW , Albuquerque EX , Eldefrawi ME
Ref : Journal of Biological Chemistry , 256 :2843 , 1981
Abstract : The specific binding of [3H]perhydrohistrionicotoxin ([3H]H12-HTX) to the ionic channel of the nicotinic receptor in membranes from the electric organ of the electric ray Torpedo ocellata was studied by use of a rapid filter assay. The time course of the binding was monitored and the initial rate of binding (i.e. within 30 s) was found to be increased up to several hundredfold by the presence of several receptor agonists in a dose-dependent manner. Receptor antagonists blocked this agonist-increased initial rate of binding. The presence of receptor antagonists, with the exception of alpha-bungarotoxin, also increased the initial rate of [3H]H12-HTX binding, though to a much lesser degree than agonists. Preincubation of the membranes with carbamylcholine reduced the initial rate of binding in a time-dependent manner. This was proposed to be due to receptor desensitization. Thus, it is suggested that the time course of [3H]H12-HTX binding to the ionic channel sites is influenced by the conformational state of the receptor-channel complex such that receptor activation, inactivation, and desensitization appear to be reflected in [3H]H12-HTX binding.
ESTHER : Aronstam_1981_J.Biol.Chem_256_2843
PubMedSearch : Aronstam_1981_J.Biol.Chem_256_2843
PubMedID: 7204378

Title : Interaction of imipramine with the ionic channel of the acetylcholine receptor of motor endplate and electric organ -
Author(s) : Eldefrawi ME , Warnick JE , Schofield GG , Albuquerque EX , Eldefrawi AT
Ref : Biochemical Pharmacology , 30 :1391 , 1981
PubMedID: 6268097

Title : Recent advances in the molecular mechanisms of human and animal models of myasthenia gravis - Albuquerque_1981_Ann.N.Y.Acad.Sci_377_496
Author(s) : Albuquerque EX , Warnick JE , Mayer RF , Eldefrawi AT , Eldefrawi ME
Ref : Annals of the New York Academy of Sciences , 377 :496 , 1981
Abstract : The receptor-channel molecule is a dynamic system which exists in multiple conformations and that is the way we should think of it when we study antibody interaction with the molecule. The results presented here suggest that some antibodies may affect receptor function by occupying sites other than the receptor site. Some of these sites may by exposed only in certain conformations, and occupation of some site by antibodies may effect conformational changes. These small but perhaps important differences in cholinergic channel properties of the myasthenic muscle from the normal one are revealed by studying the effect of myasthenic sera on drug interactions with the channel sites. The sera of myasthenics are able to react with certain channel conformations and are able to affect the interaction of channel antagonists such as H12HTX and QNB. The sera appear to act preferentially with the open conformation of the channel. As a consequence of such an effect, important conformational changes of the channel may fail to occur upon activation.
ESTHER : Albuquerque_1981_Ann.N.Y.Acad.Sci_377_496
PubMedSearch : Albuquerque_1981_Ann.N.Y.Acad.Sci_377_496
PubMedID: 6280564

Title : Activation, inactivation, and desensitization of acetylcholine receptor channel complex detected by binding of perhydrohistrionicotoxin - Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
Author(s) : Eldefrawi ME , Aronstam RS , Bakry NM , Eldefrawi AT , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 77 :2309 , 1980
Abstract : The effects of receptor activation were studied on the interaction of perhydrohistrionicotoxin (H(12)-HTX) with the ionic channel of the nicotinic acetylcholine (AcCho) receptor in membranes from the electric organ of Torpedo ocellata and with the endplate region of the soleus muscle of the rat. In Torpedo membranes, the initial rate (i.e., within 30 sec) of [(3)H]H(12)-HTX bindings to the ionic channel of the AcCho receptor was accelerated 10(2)- to 10(3)-fold in the presence of carbamoylcholine (Carb). H(12)-HTX also inhibited Carb-activated (22)Na(+) influx, over 95% inhibition at 10 muM H(12)-HTX. At this concentration H(12)-HTX did not inhibit [(3)H]AcCho binding to the AcCho-receptor sites. There was good correspondence between the degree of acceleration of [(3)H]H(12)-HTX binding and the stimulation of (22)Na(+) influx over a wide range of Carb concentrations (up to 100 muM). Preincubation of Torpedo membranes with Carb decreased the initial rate of [(3)H]H(12)-HTX binding, as well as the rate of (22)Na(+) influx, which may reflect desensitization of the AcCho-receptor. d-Tubocurarine inhibited the agonist-mediated acceleration of [(3)H]H(12)-HTX binding and (22)Na(+) influx. In the soleus muscle endplate, H(12)-HTX inhibited the transient depolarization induced by microiontophoretic application of AcCho; the more receptors activated and channels opened, the stronger was the inhibition by H(12)-HTX. These findings suggest that H(12)-HTX binds to closed and open ionic channels, with a preference for the latter conformation. It is also suggested that the conformational changes associated with activation or desensitization of the receptor can be monitored by studying binding of [(3)H]H(12)-HTX to the ionic channel sites as well as by the AcCho-receptor-regulated (22)Na(+) influx.
ESTHER : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
PubMedSearch : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_2309
PubMedID: 6246539

Title : Sites of action of phencyclidine. II. Interaction with the ionic channel of the nicotinic receptor -
Author(s) : Albuquerque EX , Tsai MC , Aronstam RS , Eldefrawi AT , Eldefrawi ME
Ref : Molecular Pharmacology , 18 :167 , 1980
PubMedID: 6252436

Title : Similarities in the binding sites of the muscarinic receptor and the ionic channel of the nicotinic receptor -
Author(s) : Aronstam RS , Eldefrawi AT , Eldefrawi ME
Ref : Biochemical Pharmacology , 29 :1311 , 1980
PubMedID: 6249329

Title : Sites of action of phencyclidine. III. Interactions with muscarinic receptors -
Author(s) : Aronstam RS , Eldefrawi ME , Eldefrawi AT , Albuquerque EX , Jim KF , Triggle DJ
Ref : Molecular Pharmacology , 18 :179 , 1980
PubMedID: 7421791

Title : Nereistoxin interaction with the acetylcholine receptor-ionic channel complex -
Author(s) : Eldefrawi AT , Bakry NM , Eldefrawi ME , Tsai MC , Albuquerque EX
Ref : Molecular Pharmacology , 17 :172 , 1980
PubMedID: 6248754

Title : Sites of action of phencyclidine. I. Effects on the electrical excitability and chemosensitive properties of the neuromuscular junction of skeletal muscle -
Author(s) : Tsai MC , Albuquerque EX , Aronstam RS , Eldefrawi AT , Eldefrawi ME , Triggle DJ
Ref : Molecular Pharmacology , 18 :159 , 1980
PubMedID: 6968397

Title : [3H]Phencyclidine: a probe for the ionic channel of the nicotinic receptor - Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
Author(s) : Eldefrawi ME , Eldefrawi AT , Aronstam RS , Maleque MA , Warnick JE , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 77 :7458 , 1980
Abstract : To evaluate [3H]phencyclidine ([3H]PCP)as a probe for the ionic channel of the nicotinic receptor, the characteristics of its binding to electric organ membranes od Torpedo ocellata and its effects on frog sartorius muscle were studied. Similar to PCP, [3H]PCP depressed the peak amplitude of endplate current, caused nonlinearity in the voltage-current relationship at negative potentials, accelerated the decay time of the end-plate current, and shortened the channel lifetime. Thus, [3H]PCP interacted with the ionic channel of the nicotinic receptor, although there were a few differences between its effect and that of PCP. Binding of [3H]PCP to Torpedo membranes was to sites on the ionic channel of acetylcholine (AcCho) receptor because it was saturable, dependent upon protein concentration, and inhibited by drugs that interact with the ionic channel, and the initial rate of binding was potentiated by receptor agonists. Equilibrium binding of [3H]PCP to Torpedo membranes was with two affinities, but in the presence of AcCho, [3H]PCP binding was with a single affinity. The affinities of channel drugs obtained by inhibition of binding of [3H]PCP and [3H[perhydrohistrionicotoxin to Torpedo membranes were different, with correlation coefficients of 0.52 and 0.82 in the absence and presence of a receptor agonist, respectively; this suggests differences in their binding sites on the ionic channel of the AcCho receptor.
ESTHER : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
PubMedSearch : Eldefrawi_1980_Proc.Natl.Acad.Sci.U.S.A_77_7458
PubMedID: 6261260

Title : Phencyclidine interactions with the ionic channel of the acetylcholine receptor and electrogenic membrane - Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
Author(s) : Albuquerque EX , Tsai MC , Aronstam RS , Witkop B , Eldefrawi AT , Eldefrawi ME
Ref : Proc Natl Acad Sci U S A , 77 :1224 , 1980
Abstract : The effects of phencyclidine (PCP) were studied on the electrogenic and chemosensitive properties of the neuromuscular junction of skeletal muscle as well as on the binding sites on the acetylcholine (AcCho) receptor and its ionic channel in the electric organ membranes of the electric ray. The directly elicited muscle twitch was markedly potentiated by prolonging the falling phase of the muscle action potential and blocking delayed rectification. The indirectly elicited muscle twitch was transiently potentiated and then blocked by PCP at concentrations below 60 muM. PCP blocked miniature endplate potentials and AcCho sensitivities at the junctional region of innervated muscle, blocked the extrajunctional sensitivity of the chronically denervated muscle, and significantly depressed the peak amplitude of the endplate current (EPC) in a voltage- and time-dependent manner. PCP also caused acceleration of the time course of EPC decay and shortening of the mean life-time of the open ionic channel. The effects of PCP were not due to inhibition of AcCho receptor sites because PCP did not protect against the quasi-irreversible inhibition of receptor sites by alpha-bungarotoxin, nor did it inhibit binding of [(3)H]AcCho or [(125)I-labeled alpha-bungarotoxin to the receptor sites. On the other hand, PCP blocked the binding of [(3)H]perhydrohistrionicotoxin to the sites of the ionic channel of the AcCho receptor. The data suggest that PCP reacts with the electrogenic K(+) channel and the ionic channel associated with the AcCho receptor in the open as well as the closed conformation.
ESTHER : Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
PubMedSearch : Albuquerque_1980_Proc.Natl.Acad.Sci.U.S.A_77_1224
PubMedID: 6928673

Title : Reaction of tetraethylammonium with the open and closed conformations of the acetylcholine receptor ionic channel complex - Adler_1979_J.Gen.Physiol_74_129
Author(s) : Adler M , Oliveira AC , Albuquerque EX , Mansour NA , Eldefrawi AT
Ref : Journal of General Physiology , 74 :129 , 1979
Abstract : The effect of tetraethylammonium (TEA) bromide on the neurally and iontophoretically evoked endplate current (EPC) of frog sartorius muscle was investigated using voltage-clamp and noise analysis techniques, and its binding to the acetylcholine (ACh) receptor ionic channel complex was determined on the electric organ of Torpedo ocellata. TEA (250-500 microM) produced an initial enhancement followed by a slow decline in the amplitude of the endplate potential and EPC, but caused only depression in the amplitude of the miniature endplate potential and current. In normal ringer's solution, the EPC current-voltage relationship was approximately linear, and the decay phase varied exponentially with membrane potential. Upon addition of 50-100 microM TEA, the current-voltage relationship became markedly nonlinear at hyperpolarized command potentials, and with 250-2000 microM TEA, there was an initial linear segment, an intermediate nonlinear segment, and a region of negative conductance. The onset of nonlinearity was dose-dependent, undergoing a 50 mV shift for a 10-fold increase in TEA concentration. The EPC decay phase was shortened by TEA at hyperpolarized but not depolarized potentials, and remained a single expotential function of time at all concentrations and membrane potentials examined. These actions of TEA were found to be independent of the sequence of polarizations, the length of the conditioning pulse, and the level of the initial holding potential. TEA shifted the power spectrum of ACh noise to higher frequencies and produced a significant depression of single channel conductance. The shortening in the mean channel lifetime agreed closely with the decrease in the EPC decay time constant. At the concentrations tested, TEA did not alter the EPC reversal potential, nor the resting membrane potential, and had little effect on the action potential duration. TEA inhibited the binding of both [3H] ACh (Ki = 200 microM) and [3H]perhydrohistrionicotoxin (Ki = 280 microM) to receptor-rich membranes from the electric organ of Torpedo ocellata, and inhibited the carbamylcholine-activated 22Na+ efflux from these microsacs. It is suggested that TEA reacts with the nicotinic ACh-receptor as well as its ion channel; the voltage-dependent actions are associated with blockade of the ion channel. The results are compatible with a kinetic model in which TEA first binds to the closed conformation of the receptor-ionicchannel complex to produce a voltage-depdndent depression of endplate conductance and sudsequently to its open conformation, giving rise to the shortening in the EPC decay and mean channel lifetime.
ESTHER : Adler_1979_J.Gen.Physiol_74_129
PubMedSearch : Adler_1979_J.Gen.Physiol_74_129
PubMedID: 486241

Title : Tetraethylammonium: voltage-dependent action on endplate conductance and inhibition of ligand binding to postsynaptic proteins - Adler_1979_Proc.Natl.Acad.Sci.U.S.A_76_531
Author(s) : Adler M , Oliveira AC , Eldefrawi ME , Eldefrawi AT , Albuquerque EX
Ref : Proc Natl Acad Sci U S A , 76 :531 , 1979
Abstract : Tetraethylammonium (Et(4)N(+)) ions depressed the amplitude and accelerated the decay rate of spontaneously occurring and nerve-evoked endplate currents (EPCs) in frog sartorius muscle. The relationship between peak EPC amplitude and membrane potential became nonlinear in the presence of 100 muM Et(4)N(+), and with drug concentrations of 250 muM or greater the current-voltage relationship exhibited negative conductance in the hyperpolarized region. Et(4)N(+) modified the exponential dependence of the EPC decay on membrane potential such that the decays between -150 and -50 mV were abbreviated and voltage independent but remained near control levels at more positive membrane potentials. The minimal effective concentration of Et(4)N(+) for altering the EPC time course was 10, and maximal effects were attained with 100 muM. Little additional shortening in the EPC decay phase was detected on raising the drug concentration to 1000 muM. Acetylcholine noise analysis revealed a voltage-dependent reduction in the mean channel open time, which was comparable in magnitude to the shortening in the EPC decay, and a depression of single-channel conductance. In concomitant biochemical studies, Et(4)N(+) was found to inhibit the binding of both [(3)H]acetylcholine and [(3)H]perhydrohistrionicotoxin to receptor-rich membranes from the electric organ of Torpedo ocellata with K(i) values of 200 muM and 280 muM, respectively. These results suggest that Et(4)N(+) interacts with both the acetylcholine receptor and its associated ionic channel. The voltage-dependent actions of Et(4)N(+) are attributed to blockade of the ionic channel in closed as well as open conformation.
ESTHER : Adler_1979_Proc.Natl.Acad.Sci.U.S.A_76_531
PubMedSearch : Adler_1979_Proc.Natl.Acad.Sci.U.S.A_76_531
PubMedID: 284372

Title : Myasthenia gravis animal model: quantitative analysis of the efficiency of pre- and postjunctional regions of intercostal muscles in rabbits immunized with Torpedo acetylcholine receptor -
Author(s) : Albuquerque EX , Eldefrawi AT , Oliveira AC , Copio DS , Eldefrawi ME
Ref : Experimental Neurology , 66 :109 , 1979
PubMedID: 225189

Title : Voltage- and time-dependent actions of piperocaine on the ion channel of the acetylcholine receptor -
Author(s) : Tiedt TN , Albuquerque EX , Bakry NM , Eldefrawi ME , Eldefrawi AT
Ref : Molecular Pharmacology , 16 :909 , 1979
PubMedID: 316855

Title : Mode of action of quinacrine on the acetylcholine receptor ionic channel complex -
Author(s) : Tsai MC , Oliveira AC , Albuquerque EX , Eldefrawi ME , Eldefrawi AT
Ref : Molecular Pharmacology , 16 :382 , 1979
PubMedID: 316101

Title : Amantadine: neuromuscular blockade by suppression of ionic conductance of the acetylcholine receptor - Albuquerque_1978_Science_199_788
Author(s) : Albuquerque EX , Eldefrawi AT , Eldefrawi ME , Mansour NA , Tsai MC
Ref : Science , 199 :788 , 1978
Abstract : Amantadine hydrochloride decreases the sensitivity of denervated mammalian muscle to iontophoretically applied acetylcholine. The drug depresses the amplitude of the end-plate current and reverses the slope of the relation between half-decay time and membrane potential suggesting that it alters the ionic conductance that is mediated by the acetylcholine receptor. Binding studies confirm that amantadine acts on the ion conductance modulator rather than the acetylcholine receptor.
ESTHER : Albuquerque_1978_Science_199_788
PubMedSearch : Albuquerque_1978_Science_199_788
PubMedID: 622570

Title : Acetylcholine receptor and ionic channel of Torpedo electroplax: binding of perhydrohistrionicotoxin to membrane and solubilized preparations - Eldefrawi_1978_Biochemistry_17_5474
Author(s) : Eldefrawi ME , Eldefrawi AT , Mansour NA , Daly JW , Witkop B , Albuquerque EX
Ref : Biochemistry , 17 :5474 , 1978
Abstract : The electric organ of the ray, Torpedo ocellata, can serve as a source for both the acetylcholine (ACh) receptor and its ionic channel. The two entities were identified by their specific binding of [3H]ACh and [3H]perhydrohistrionicotoxin ([3H]H12-HTX), respectively. Binding of [3H]H12-HTX was inhibited by certain drugs and toxins, e.g., histrionicotoxin (HTX), amantadine, and tetraethylammonium (TEA) ions at concentrations that did not inhibit [3H]ACh binding. However, the specific carbamoylcholine-induced 22Na efflux from microsacs from the electric organ membranes was blocked by inhibitors of either the receptor or its ionic channel. The ionic channel had the properties of a protein as judged by heat sensitivity and the inhibition of [3H]H12-HTX binding, after incubation of the electric organ membranes with protein reagents such as p-chloromercuribenzenesulfonic acid (PCMBS) or N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ). The "binding" of [3H]H12-HTX at 4 X 10(-8) M to lipids in the microsacs was 12% of the total binding to intact microsacs and was nonsaturable and insensitive to heat or specific drugs. After solubilization with cholate, the [3H]H12-HTX binding subunits retained the same affinities for toxins and drugs. The Kd for [3H]H12-HTX was 3 X 10(-7) M. The majority of the ionic channel could be separated from the ACh receptors in the cholate extract by incubation with ACh-receptor affinity gel and ACh-receptor antibodies. The ACh receptor purified by this affinity gel contained only a few active ionic channel units as judged by low levels of high affinity binding of [3H]H12-HTX. On the other hand, after solubilization with Triton X-100, all the ionic channel molecules were either separated or denatured so that the purified ACh receptor did not exhibit high affinity binding for [3H]H12-HTX.
ESTHER : Eldefrawi_1978_Biochemistry_17_5474
PubMedSearch : Eldefrawi_1978_Biochemistry_17_5474
PubMedID: 728410

Title : Effects of alpha-bungarotoxin and reversible cholinergic ligands on normal and denervated mammalian skeletal muscle - Sarvey_1978_Membr.Biochem_1_131
Author(s) : Sarvey JM , Albuquerque EX , Eldefrawi AT , Eldefrawi M
Ref : Membr Biochem , 1 :131 , 1978
Abstract : alpha-Bungarotoxin (BuTX; 5 micrograms/ml) completely blocked the endplate potential and extrajunctional acetylcholine (ACh) sensitivity of surface fibers in normal and chronically denervated mammalian muscles, respectively, in about 35 min. A 0.72 +/- 0.033 mV amplitude endplate potential returned in normal muscle fibers after 6.5 hr. of washout of alpha-BuTX, and an ACh sensitivity of 41.02 +/- 3.95 mV/nC was recorded in denervated muscle after 6.5 hr of wash (control being 1215 +/- 197 mV/nC). A two-step reaction of BuTX with binding sites which may allosterically interact is postulated. Several pharmacologic differences were noted between the ACh receptors at the normal endplate and those appearing extrajunctionally following denervation. In normal innervated muscles exposed to BuTX in the presence of 20 microM carbamylcholine or decamethonium, washout of both drugs restored twitch to control levels within 2 hr. Endplate potentials large enough to initiate action potentials were also recorded in most surface fibers. In contrast, these agents, in much higher concentrations (50 microM), were almost ineffective in preventing BuTX blockade of ACh sensitivity in denervated muscle. Hexamethonium (10 and 50 mM) depressed neuromuscular transmission and blocked the action of BuTX in normal muscle in a dose-dependent fashion. On the extrajunctional receptors, hexamethonium (50 mM) was ineffective in protecting against BuTX. We may conclude that at the normal endplate region there are two distinct populations of ACh receptors, both of which react with cholinergic ligands and BuTX, but that a small population (representing congruent to 1% of the total) reacts with BuTX reversibly. Our findings further suggest a clear distinction between ACh receptors located at the normal endplate region and those of the extrajunctional region of the chronically denervated mammalian muscle.
ESTHER : Sarvey_1978_Membr.Biochem_1_131
PubMedSearch : Sarvey_1978_Membr.Biochem_1_131
PubMedID: 756485

Title : Mechanism of action of amantadine on neuromuscular transmission -
Author(s) : Tsai MC , Mansour NA , Eldefrawi AT , Eldefrawi ME , Albuquerque EX
Ref : Molecular Pharmacology , 14 :787 , 1978
PubMedID: 213703

Title : Characterization of calcium-binding sites of the purified acetylcholine receptor and identification of the calcium-binding subunit -
Author(s) : Rubsamen H , Eldefrawi AT , Eldefrawi ME , Hess GP
Ref : Biochemistry , 17 :3818 , 1978
PubMedID: 698199

Title : Perhydrohistrionicotoxin: a potential ligand for the ion conductance modulator of the acetylcholine receptor - Eldefrawi_1977_Proc.Natl.Acad.Sci.U.S.A_74_2172
Author(s) : Eldefrawi AT , Eldefrawi ME , Albuquerque EX , Oliveira AC , Mansour N , Adler M , Daly JW , Brown GB , Burgermeister W , Witkop B
Ref : Proc Natl Acad Sci U S A , 74 :2172 , 1977
Abstract : Histrionicotoxin from the Colombian frog Dendrobates histrionicus and its perhydro derivative reversibly block the acetylcholine-sensitive ion conductance system in frog neuromuscular preparations. The perhydro derivative and [3H]perhydrohistrionicotoxin, like histrionicotoxin, caused a significant decrease in the peak amplitude of the end-plate current and shortened its rise time and half-decay time. In membrane preparations from Torpedo electroplax, [3H]perhydrohistrionicotoxin bound reversibly to a limited number of high-affinity sites [dissociation constant, (KD) = 0.4 micronM]. The ratio of perhydrohistrionicotoxin to acetylcholine binding sites in these membrane preparations approached 2. Histrionicotoxins, local anesthetics, and certain cholinergic agonists inhibited binding of perhydrohistrionicotoxin. Binding of perhydrohistrionicotoxin to membranes was decreased by heat or treatment with proteases. Treatment of membranes with Triton X-100 solubilized acetylcholine binding proteins and apparently also perhydrohistrionicotoxin-binding proteins. However, the detergent Triton X-100 also bound [3H]perhydrohistrionicotoxin. This nonspecific binding was not saturable and complicated studies on the antagonism by drugs of binding of [3H]perhydrohistrionicotoxin. In solubilized preparations the binding protein for acetylcholine could be removed by affinity chromatography or immunoprecipitation without affecting binding of perhydrohistrionicotoxin. Sephadex chromatography also separated acetylcholine- from perhydrohistrionicotoxin-binding proteins. Perhydrohistrionicotoxin did not bind significantly to purified acetylcholine-receptor protein but presumably bound to an ion conductance modulator protein that was associated with the acetylcholine-receptor in intact membrane and readily separable from the receptor protein after solubilization.
ESTHER : Eldefrawi_1977_Proc.Natl.Acad.Sci.U.S.A_74_2172
PubMedSearch : Eldefrawi_1977_Proc.Natl.Acad.Sci.U.S.A_74_2172
PubMedID: 301278

Title : Interaction between calcium and ligand-binding sites of the purified acetylcholine receptor studied by use of a fluorescent lanthanide -
Author(s) : Rubsamen H , Hess GP , Eldefrawi AT , Eldefrawi ME
Ref : Biochemical & Biophysical Research Communications , 68 :56 , 1976
PubMedID: 942851

Title : Structure and function of the acetylcholine receptor -
Author(s) : Eldefrawi ME , Eldefrawi AT
Ref : Croatica Chemica Acta , 47 :425 , 1975

Title : Isolation of acetylcholine receptors -
Author(s) : O'Brien RD , Eldefrawi ME , Eldefrawi AT
Ref : Annual Review of Pharmacology , 12 :19 , 1972