Aracava Y

References (46)

Title : (R,S)-trihexyphenidyl, acting via a muscarinic receptor-independent mechanism, inhibits hippocampal glutamatergic and GABAergic synaptic transmissions: Potential relevance for treatment of organophosphorus intoxication - Aracava_2023_Neuropharmacol__109684
Author(s) : Aracava Y , Albuquerque EX , Pereira EFR
Ref : Neuropharmacology , :109684 , 2023
Abstract : Preclinical studies have reported that, compared to the muscarinic receptor (mAChR) antagonist atropine, (R,S)-trihexyphenidyl (THP) more effectively counters the cholinergic crisis, seizures, and neuropathology triggered by organophosphorus (OP)-induced acetylcholinesterase (AChE) inhibition. The greater effectiveness of THP was attributed to its ability to block mAChRs and N-methyl-d-aspartate-type glutamatergic receptors (NMDARs) in the brain. However, THP also inhibits alpha7 nicotinic receptors (nAChRs). The present study examined whether THP-induced inhibition of mAChRs, alpha7 nAChRs, and NMDARs is required to suppress glutamatergic synaptic transmission, whose overstimulation sustains OP-induced seizures. In primary hippocampal cultures, THP (1-30 microM) suppressed the frequency of excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs, respectively) recorded from neurons in nominally Mg(2+)-free solution. A single sigmoidal function adequately fit the overlapping concentration-response relationships for THP-induced suppression of IPSC and EPSC frequencies yielding an IC50 of 6.3 +/- 1.3 microM. Atropine (1 microM), the NMDAR antagonist d,l-2-amino-5-phosphonopentanoic acid (D,L-AP5, 50 microM), and the alpha7 nAChR antagonist methyllycaconitine (MLA, 10 nM) did not prevent THP-induced inhibition of synaptic transmission. THP (10 microM) did not affect the probability of transmitter release because it had no effect on the frequency of miniature IPSCs and EPSCs recorded in the presence of tetrodotoxin. Additionally, THP had no effect on the amplitudes and decay-time constants of miniature IPSCs and EPSCs; therefore, it did not affect the activity of postsynaptic GABA(A) and glutamate receptors. This study provides the first demonstration that THP can suppress action potential-dependent synaptic transmission via a mechanism independent of NMDAR, mAChR, and alpha7 nAChR inhibition.
ESTHER : Aracava_2023_Neuropharmacol__109684
PubMedSearch : Aracava_2023_Neuropharmacol__109684
PubMedID: 37549771

Title : Oral Pretreatment with Galantamine Effectively Mitigates the Acute Toxicity of a Supralethal Dose of Soman in Cynomolgus Monkeys Posttreated with Conventional Antidotes - Lane_2020_J.Pharmacol.Exp.Ther_375_115
Author(s) : Lane M , Carter D , Pescrille JD , Aracava Y , Fawcett WP , Basinger GW , Pereira EFR , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 375 :115 , 2020
Abstract : Earlier reports suggested that galantamine, a drug approved to treat mild-to-moderate Alzheimer's disease (AD), and other centrally acting reversible acetylcholinesterase (AChE) inhibitors can serve as adjunct pretreatments against poisoning by organophosphorus compounds, including the nerve agent soman. The present study was designed to determine whether pretreatment with a clinically relevant oral dose of galantamine HBr mitigates the acute toxicity of 4.0xLD(50) soman (15.08 microg/kg) in Macaca fascicularis posttreated intramuscularly with the conventional antidotes atropine (0.4 mg/kg), 2-pyridine aldoxime methyl chloride (30 mg/kg), and midazolam (0.32 mg/kg). The pharmacokinetic profile and maximal degree of blood AChE inhibition (-25%-40%) revealed that the oral doses of 1.5 and 3.0 mg/kg galantamine HBr in these nonhuman primates (NHPs) translate to human-equivalent doses that are within the range used for AD treatment. Subsequent experiments demonstrated that 100% of NHPs pretreated with either dose of galantamine, challenged with soman, and posttreated with conventional antidotes survived 24 hours. By contrast, given the same posttreatments, 0% and 40% of the NHPs pretreated, respectively, with vehicle and pyridostigmine bromide (1.2 mg/kg, oral), a peripherally acting reversible AChE inhibitor approved as pretreatment for military personnel at risk of exposure to soman, survived 24 hours after the challenge. In addition, soman caused extensive neurodegeneration in the hippocampi of saline- or pyridostigmine-pretreated NHPs, but not in the hippocampi of galantamine-pretreated animals. To our knowledge, this is the first study to demonstrate the effectiveness of clinically relevant oral doses of galantamine to prevent the acute toxicity of supralethal doses of soman in NHPs. SIGNIFICANCE STATEMENT: This is the first study to demonstrate that a clinically relevant oral dose of galantamine effectively prevents lethality and neuropathology induced by a supralethal dose of the nerve agent soman in Cynomolgus monkeys posttreated with conventional antidotes. These findings are of major significance for the continued development of galantamine as an adjunct pretreatment against nerve agent poisoning.
ESTHER : Lane_2020_J.Pharmacol.Exp.Ther_375_115
PubMedSearch : Lane_2020_J.Pharmacol.Exp.Ther_375_115
PubMedID: 32759369

Title : Gestational exposures to organophosphorus insecticides: From acute poisoning to developmental neurotoxicity - Todd_2020_Neuropharmacol__108271
Author(s) : Todd SW , Lumsden EW , Aracava Y , Mamczarz J , Albuquerque EX , Pereira EFR
Ref : Neuropharmacology , :108271 , 2020
Abstract : For over three-quarters of a century, organophosphorus (OP) insecticides have been ubiquitously used in agricultural, residential, and commercial settings and in public health programs to mitigate insect borne diseases. Their broad-spectrum insecticidal effectiveness is accounted for by the irreversible inhibition of acetylcholinesterase (AChE), the enzyme that catalyzes acetylcholine (ACh) hydrolysis, in the nervous system of insects. However, because AChE is evolutionarily conserved, OP insecticides are also toxic to mammals, including humans, and acute OP intoxication remains a major public health concern in countries where OP insecticide usage is poorly regulated. Environmental exposures to OP levels that are generally too low to cause marked inhibition of AChE and to trigger acute signs of intoxication, on the other hand, represent an insidious public health issue worldwide. Gestational exposures to OP insecticides are particularly concerning because of the exquisite sensitivity of the developing brain to these insecticides. The present article overviews and discusses: (i) the health effects and therapeutic management of acute OP poisoning during pregnancy, (ii) epidemiological studies examining associations between environmental OP exposures during gestation and health outcomes of offspring, (iii) preclinical evidence that OP insecticides are developmental neurotoxicants, and (iv) potential mechanisms underlying the developmental neurotoxicity of OP insecticides. Understanding how gestational exposures to different levels of OP insecticides affect pregnancy and childhood development is critical to guiding implementation of preventive measures and direct research aimed at identifying effective therapeutic interventions that can limit the negative impact of these exposures on public health.
ESTHER : Todd_2020_Neuropharmacol__108271
PubMedSearch : Todd_2020_Neuropharmacol__108271
PubMedID: 32814088

Title : Galantamine prevents long-lasting suppression of excitatory synaptic transmission in CA1 pyramidal neurons of soman-challenged guinea pigs - Alexandrova_2014_Neurotoxicol_44_270
Author(s) : Alexandrova EA , Alkondon M , Aracava Y , Pereira EF , Albuquerque EX
Ref : Neurotoxicology , 44 :270 , 2014
Abstract : Galantamine, a drug currently approved for the treatment of Alzheimer's disease, has recently emerged as an effective pretreatment against the acute toxicity and delayed cognitive deficits induced by organophosphorus (OP) nerve agents, including soman. Since cognitive deficits can result from impaired glutamatergic transmission in the hippocampus, the present study was designed to test the hypothesis that hippocampal glutamatergic transmission declines following an acute exposure to soman and that this effect can be prevented by galantamine. To test this hypothesis, spontaneous excitatory postsynaptic currents (EPSCs) were recorded from CA1 pyramidal neurons in hippocampal slices obtained at 1h, 24h, or 6-9 days after guinea pigs were injected with: (i) 1xLD50 soman (26.3mug/kg, s.c.); (ii) galantamine (8mg/kg, i.m.) followed 30min later by 1xLD50 soman, (iii) galantamine (8mg/kg, i.m.), or (iv) saline (0.5ml/kg, i.m.). In soman-injected guinea pigs that were not pretreated with galantamine, the frequency of EPSCs was significantly lower than that recorded from saline-injected animals. There was no correlation between the severity of soman-induced acute toxicity and the magnitude of soman-induced reduction of EPSC frequency. Pretreatment with galantamine prevented the reduction of EPSC frequency observed at 6-9 days after the soman challenge. Prevention of soman-induced long-lasting reduction of hippocampal glutamatergic synaptic transmission may be an important determinant of the ability of galantamine to counter cognitive deficits that develop long after an acute exposure to the nerve agent.
ESTHER : Alexandrova_2014_Neurotoxicol_44_270
PubMedSearch : Alexandrova_2014_Neurotoxicol_44_270
PubMedID: 25064080

Title : Animal models that best reproduce the clinical manifestations of human intoxication with organophosphorus compounds - Pereira_2014_J.Pharmacol.Exp.Ther_350_313
Author(s) : Pereira EF , Aracava Y , DeTolla LJ, Jr. , Beecham EJ , Basinger GW, Jr. , Wakayama EJ , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 350 :313 , 2014
Abstract : The translational capacity of data generated in preclinical toxicological studies is contingent upon several factors, including the appropriateness of the animal model. The primary objectives of this article are: 1) to analyze the natural history of acute and delayed signs and symptoms that develop following an acute exposure of humans to organophosphorus (OP) compounds, with an emphasis on nerve agents; 2) to identify animal models of the clinical manifestations of human exposure to OPs; and 3) to review the mechanisms that contribute to the immediate and delayed OP neurotoxicity. As discussed in this study, clinical manifestations of an acute exposure of humans to OP compounds can be faithfully reproduced in rodents and nonhuman primates. These manifestations include an acute cholinergic crisis in addition to signs of neurotoxicity that develop long after the OP exposure, particularly chronic neurologic deficits consisting of anxiety-related behavior and cognitive deficits, structural brain damage, and increased slow electroencephalographic frequencies. Because guinea pigs and nonhuman primates, like humans, have low levels of circulating carboxylesterases-the enzymes that metabolize and inactivate OP compounds-they stand out as appropriate animal models for studies of OP intoxication. These are critical points for the development of safe and effective therapeutic interventions against OP poisoning because approval of such therapies by the Food and Drug Administration is likely to rely on the Animal Efficacy Rule, which allows exclusive use of animal data as evidence of the effectiveness of a drug against pathologic conditions that cannot be ethically or feasibly tested in humans.
ESTHER : Pereira_2014_J.Pharmacol.Exp.Ther_350_313
PubMedSearch : Pereira_2014_J.Pharmacol.Exp.Ther_350_313
PubMedID: 24907067

Title : Magnetic resonance imaging reveals that galantamine prevents structural brain damage induced by an acute exposure of guinea pigs to soman - Gullapalli_2010_Neurotoxicol_31_67
Author(s) : Gullapalli RP , Aracava Y , Zhuo J , Helal Neto E , Wang J , Makris G , Merchenthaler I , Pereira EF , Albuquerque EX
Ref : Neurotoxicology , 31 :67 , 2010
Abstract : Galantamine, a drug used to treat Alzheimer's disease, has recently emerged as a potential medical countermeasure against the toxicity of organophosphorus (OP) compounds, including the nerve agent soman. Here, magnetic resonance imaging (MRI) was used to characterize the neurotoxic effects of soman and the ability of galantamine to prevent these effects in guinea pigs, the best non-primate model to predict the effectiveness of antidotes against OP toxicity in humans. The brains of treated and untreated guinea pigs were imaged using a clinical 3.0 Tesla MRI scanner at 48 h before and 6-7 h, 48 h and 7 days after their challenge with 1.0xLD50 soman (26.6 microg/kg, sc). Significant brain atrophy was observed among all untreated animals at 7 days after their challenge with soman. In mildly intoxicated animals, significant shortening of spin-spin relaxation times (T2) was observed in the thalamus and amygdala at 7h after the challenge. In severely intoxicated animals, T2 values and T2-weighted signal intensities increased significantly in the piriform cortex, hippocampus, thalamus and amygdala; in most regions, changes were long-lasting. Voxel-based morphometric analysis of the images revealed that other brain regions were also damaged in these animals. Neuronal loss was confirmed histopathologically. In animals that were treated with galantamine (8 mg/kg, im) 30 min prior to the exposure to soman, T2, T2-weighted signal intensities, and CSF volumes were largely unaffected. It is, therefore, concluded that galantamine can effectively prevent the structural brain damage induced by an acute exposure to soman.
ESTHER : Gullapalli_2010_Neurotoxicol_31_67
PubMedSearch : Gullapalli_2010_Neurotoxicol_31_67
PubMedID: 19782102

Title : Pretreatment of Guinea pigs with galantamine prevents immediate and delayed effects of soman on inhibitory synaptic transmission in the hippocampus - Alexandrova_2010_J.Pharmacol.Exp.Ther_334_1051
Author(s) : Alexandrova EA , Aracava Y , Pereira EF , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 334 :1051 , 2010
Abstract : Galantamine has emerged as a potential antidote to prevent the acute toxicity of organophosphorus (OP) compounds. Changes in inhibitory GABAergic activity in different brain regions can contribute to both induction and maintenance of seizures in subjects exposed to the OP nerve agent soman. Here, we tested the hypothesis that galantamine can prevent immediate and delayed effects of soman on hippocampal inhibitory synaptic transmission. Spontaneous inhibitory postsynaptic currents (IPSCs) were recorded from CA1 pyramidal neurons in hippocampal slices obtained at 1 h, 24 h, or 6 to 9 days after the injection of guinea pigs with saline (0.5 ml/kg i.m.), 1xLD(50) soman (26.3 microg/kg s.c.), galantamine (8 mg/kg i.m.), or galantamine at 30 min before soman. Soman-challenged animals that were not pretreated showed mild, moderate, or severe signs of acute intoxication. At 1 h after the soman injection, the mean IPSC amplitude recorded from slices of mildly intoxicated animals and the mean IPSC frequency recorded from slices of severely intoxicated animals were larger and lower, respectively, than those recorded from slices of control animals. Regardless of the severity of the acute toxicity, at 24 h after the soman challenge the mean IPSC frequency was lower than that recorded from slices of control animals. At 6 to 9 days after the challenge, the IPSC frequency had returned to control levels, whereas the mean IPSC amplitude became larger than control. Pretreatment with galantamine prevented soman-induced changes in IPSCs. Counteracting the effects of soman on inhibitory transmission can be an important determinant of the antidotal effectiveness of galantamine.
ESTHER : Alexandrova_2010_J.Pharmacol.Exp.Ther_334_1051
PubMedSearch : Alexandrova_2010_J.Pharmacol.Exp.Ther_334_1051
PubMedID: 20554906

Title : An acute exposure to a sub-lethal dose of soman triggers anxiety-related behavior in guinea pigs: interactions with acute restraint - Mamczarz_2010_Neurotoxicol_31_77
Author(s) : Mamczarz J , Pereira EF , Aracava Y , Adler M , Albuquerque EX
Ref : Neurotoxicology , 31 :77 , 2010
Abstract : In this study, we tested the hypothesis that a single exposure of guinea pigs to sub-lethal doses of soman triggers anxiety-related behavior that is modifiable by acute stress. Prepubertal male guinea pigs were subjected to one of the following treatments: (i) saline (0.5 ml/kg, sc), (ii) soman (0.6x or 0.8xLD50, sc), (iii) saline followed 30 min later by 2-h restraint, or (iv) soman followed 30 min later by 2-h restraint. Behavior of the animals was examined 2 and 3 months later in a large open field and in the elevated plus maze. Animals that had been exposed to restraint stress alone or soman alone showed decreased exploratory activity when tested in the open field with bare floor at light intensity of 20-30 lx. Total distance traveled and distance traveled in the center of the field were shorter for animals that were exposed to either restraint stress or soman than for saline-injected animals. In addition, animals challenged with soman or restraint stress remained immobile for a longer time in the open field than did saline-injected guinea pigs. Performance in the elevated plus maze test revealed that exposure of guinea pigs to soman or restraint stress decreased their number of entries and the time spent in the open arms of the maze (measures of anxiety) and reduced their overall locomotor activity. Soman exposure and restraint stress cancelled out each other's effect on locomotion, while only attenuating one another's effect on anxiety-related behavior. It is concluded that a single exposure to sub-lethal doses of soman triggers long-lasting anxiogenesis and decreased locomotor activity and that acute restraint stress modifies the magnitude of these effects.
ESTHER : Mamczarz_2010_Neurotoxicol_31_77
PubMedSearch : Mamczarz_2010_Neurotoxicol_31_77
PubMedID: 19883683

Title : Molecular and cellular actions of galantamine: clinical implications for treatment of organophosphorus poisoning - Pereira_2010_J.Mol.Neurosci_40_196
Author(s) : Pereira EF , Aracava Y , Alkondon M , Akkerman M , Merchenthaler I , Albuquerque EX
Ref : Journal of Molecular Neuroscience , 40 :196 , 2010
Abstract : There have been continued efforts to develop effective antidotal therapies against poisoning with organophosphorus (OP) compounds, including nerve agents and pesticides. We reported recently that galantamine, a drug used to treat Alzheimer's disease, administered before (up to 3 h) or soon after (up to 5 min) an exposure of guinea pigs to 1.5-2 x LD50 soman or sarin effectively counteracted the acute toxicity and lethality of the nerve agents provided that the animals were also post-treated with atropine. Here, we demonstrate that administered to guinea pigs at 30 min before or up to 15 min after an acute challenge with 1 x LD50 soman, galantamine (8 mg/kg, intramuscular) alone is sufficient to counteract the lethality and acute toxicity of the nerve agent. Evidence is also provided that 100% survival can be attained when the association of appropriate doses of galantamine and atropine is administered 30-45 min after the challenge of the guinea pigs with 1 x LD50 soman. Galantamine counteracts the neurodegeneration and the changes in the nicotinic cholinergic system that result from an acute exposure of guinea pigs to 1 x LD50 soman. The results presented herein corroborate that galantamine is an effective antidote against OP poisoning.
ESTHER : Pereira_2010_J.Mol.Neurosci_40_196
PubMedSearch : Pereira_2010_J.Mol.Neurosci_40_196
PubMedID: 19690988

Title : A single in vivo application of cholinesterase inhibitors has neuron type-specific effects on nicotinic receptor activity in guinea pig hippocampus - Alkondon_2009_J.Pharmacol.Exp.Ther_328_69
Author(s) : Alkondon M , Aracava Y , Pereira EF , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 328 :69 , 2009
Abstract : The present study was designed to test the hypothesis that an acute in vivo treatment with reversible or irreversible acetylcholinesterase (AChE) inhibitors modifies the activities of nicotinic receptors (nAChRs) in hippocampal neurons. Here, whole-cell nicotinic responses were recorded from CA1 interneurons in hippocampal slices obtained from male guinea pigs at 1, 7, or 14 days after treatment with the irreversible AChE inhibitor, soman (1x LD(50) s.c.), and/or the reversible AChE inhibitor, galantamine (8 mg/kg i.m.). Naive animals were used as controls. Three types of nAChR responses, namely types IA, II, and III, which were mediated by alpha 7, alpha 4 beta 2, and alpha 3 beta 2 beta 4 nAChRs, respectively, could be recorded from the interneurons. The magnitude of alpha 7 nAChR currents was neuron-type dependent. Stratum radiatum interneurons (SRIs) with thick initial dendrites had the largest alpha 7 nAChR currents. Acute challenge with soman caused sustained reduction of type IA current amplitudes recorded from stratum oriens interneurons and increased the ratio of acetylcholine- to choline-evoked current amplitudes recorded from SRIs. In guinea pigs that developed long-lasting convulsions after the soman challenge, there was a sustained reduction of alpha 3 beta 2 beta 4 nAChR responses. Acute treatment with galantamine had no effect on type IA or III responses, whereas it decreased the incidence of type II currents. Pretreatment of the guinea pigs with galantamine prevented the suppressive effect of soman on type III responses. The neuron type-specific changes in nAChR activity induced by soman, some of which could be prevented by galantamine, may contribute to the maintenance of pathological rhythms in the hippocampal neuronal network.
ESTHER : Alkondon_2009_J.Pharmacol.Exp.Ther_328_69
PubMedSearch : Alkondon_2009_J.Pharmacol.Exp.Ther_328_69
PubMedID: 18842705

Title : Effectiveness of donepezil, rivastigmine, and (+\/-)huperzine A in counteracting the acute toxicity of organophosphorus nerve agents: comparison with galantamine - Aracava_2009_J.Pharmacol.Exp.Ther_331_1014
Author(s) : Aracava Y , Pereira EF , Akkerman M , Adler M , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 331 :1014 , 2009
Abstract : Galantamine, a centrally acting cholinesterase (ChE) inhibitor and a nicotinic allosteric potentiating ligand used to treat Alzheimer's disease, is an effective and safe antidote against poisoning with nerve agents, including soman. Here, the effectiveness of galantamine was compared with that of the centrally active ChE inhibitors donepezil, rivastigmine, and (+/-)huperzine A as a pre- and/or post-treatment to counteract the acute toxicity of soman. In the first set of experiments, male prepubertal guinea pigs were treated intramuscularly with one of the test drugs and 30 min later challenged with 1.5 x LD(50) soman (42 microg/kg s.c.). All animals that were pretreated with galantamine (6-8 mg/kg), 3 mg/kg donepezil, 6 mg/kg rivastigmine, or 0.3 mg/kg (+/-)huperzine A survived the soman challenge, provided that they were also post-treated with atropine (10 mg/kg i.m.). However, only galantamine was well tolerated. In subsequent experiments, the effectiveness of specific treatment regimens using 8 mg/kg galantamine, 3 mg/kg donepezil, 6 mg/kg rivastigmine, or 0.3 mg/kg (+/-)huperzine A was compared in guinea pigs challenged with soman. In the absence of atropine, only galantamine worked as an effective and safe pretreatment in animals challenged with 1.0 x LD(50) soman. Galantamine was also the only drug to afford significant protection when given to guinea pigs after 1.0 x LD(50) soman. Finally, all test drugs except galantamine reduced the survival of the animals when administered 1 or 3 h after the challenge with 0.6 or 0.7 x LD(50) soman. Thus, galantamine emerges as a superior antidotal therapy against the toxicity of soman.
ESTHER : Aracava_2009_J.Pharmacol.Exp.Ther_331_1014
PubMedSearch : Aracava_2009_J.Pharmacol.Exp.Ther_331_1014
PubMedID: 19741148

Title : Acute toxicity of organophosphorus compounds in guinea pigs is sex- and age-dependent and cannot be solely accounted for by acetylcholinesterase inhibition - Fawcett_2009_J.Pharmacol.Exp.Ther_328_516
Author(s) : Fawcett WP , Aracava Y , Adler M , Pereira EF , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 328 :516 , 2009
Abstract : This study was designed to test the hypothesis that the acute toxicity of the nerve agents S-[2-(diisopropylamino)ethyl]-O-ethyl methylphosphonothioate (VX), O-pinacolyl methylphosphonofluoridate (soman), and O-isopropyl methylphosphonofluoridate (sarin) in guinea pigs is age- and sex-dependent and cannot be fully accounted for by the irreversible inhibition of acetylcholinesterase (AChE). The subcutaneous doses of nerve agents needed to decrease 24-h survival of guinea pigs by 50% (LD(50) values) were estimated by probit analysis. In all animal groups, the rank order of LD(50) values was sarin > soman > VX. The LD(50) value of soman was not influenced by sex or age of the animals. In contrast, the LD(50) values of VX and sarin were lower in adult male than in age-matched female or younger guinea pigs. A colorimetric assay was used to determine the concentrations of nerve agents that inhibit in vitro 50% of AChE activity (IC(50) values) in guinea pig brain extracts, plasma, red blood cells, and whole blood. A positive correlation between LD(50) values and IC(50) values for AChE inhibition would support the hypothesis that AChE inhibition is a major determinant of the acute toxicity of the nerve agents. However, such a positive correlation was found only between LD(50) values and IC(50) values for AChE inhibition in brain extracts from neonatal and prepubertal guinea pigs. These results demonstrate for the first time that the lethal potencies of some nerve agents in guinea pigs are age- and sex-dependent. They also support the contention that mechanisms other than AChE inhibition contribute to the lethality of nerve agents.
ESTHER : Fawcett_2009_J.Pharmacol.Exp.Ther_328_516
PubMedSearch : Fawcett_2009_J.Pharmacol.Exp.Ther_328_516
PubMedID: 18984651

Title : Effective countermeasure against poisoning by organophosphorus insecticides and nerve agents - Albuquerque_2006_Proc.Natl.Acad.Sci.U.S.A_103_13220
Author(s) : Albuquerque EX , Pereira EF , Aracava Y , Fawcett WP , Oliveira M , Randall WR , Hamilton TA , Kan RK , Romano JA, Jr. , Adler M
Ref : Proc Natl Acad Sci U S A , 103 :13220 , 2006
Abstract : The nerve agents soman, sarin, VX, and tabun are deadly organophosphorus (OP) compounds chemically related to OP insecticides. Most of their acute toxicity results from the irreversible inhibition of acetylcholinesterase (AChE), the enzyme that inactivates the neurotransmitter acetylcholine. The limitations of available therapies against OP poisoning are well recognized, and more effective antidotes are needed. Here, we demonstrate that galantamine, a reversible and centrally acting AChE inhibitor approved for treatment of mild to moderate Alzheimer's disease, protects guinea pigs from the acute toxicity of lethal doses of the nerve agents soman and sarin, and of paraoxon, the active metabolite of the insecticide parathion. In combination with atropine, a single dose of galantamine administered before or soon after acute exposure to lethal doses of soman, sarin, or paraoxon effectively and safely counteracted their toxicity. Doses of galantamine needed to protect guinea pigs fully against the lethality of OPs were well tolerated. In preventing the lethality of nerve agents, galantamine was far more effective than pyridostigmine, a peripherally acting AChE inhibitor, and it was less toxic than huperzine, a centrally acting AChE inhibitor. Thus, a galantamine-based therapy emerges as an effective and safe countermeasure against OP poisoning.
ESTHER : Albuquerque_2006_Proc.Natl.Acad.Sci.U.S.A_103_13220
PubMedSearch : Albuquerque_2006_Proc.Natl.Acad.Sci.U.S.A_103_13220
PubMedID: 16914529

Title : Memantine blocks alpha7* nicotinic acetylcholine receptors more potently than n-methyl-D-aspartate receptors in rat hippocampal neurons - Aracava_2005_J.Pharmacol.Exp.Ther_312_1195
Author(s) : Aracava Y , Pereira EF , Maelicke A , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 312 :1195 , 2005
Abstract : The N-methyl-d-aspartate (NMDA) receptor antagonist memantine is an approved drug for treatment of Alzheimer's disease (AD). Other such treatments are cholinesterase inhibitors and nicotinic acetylcholine receptor (nAChR)-sensitizing agents such as galantamine. The present study was designed to test whether memantine exerts any effect on the cholinergic system, in particular the Ca(2+)-conducting alpha7(*) nAChR, in cultured hippocampal neurons. Memantine caused a concentration-dependent reduction of the amplitudes of whole-cell currents evoked by the alpha7(*) nAChR-selective agonist choline (10 mM) or by N-methyl-d-aspartate (NMDA) (50 muM) plus glycine (10 muM). It also inhibited tonically activated NMDA receptors. Memantine was more potent in inhibiting alpha7(*) nAChRs than NMDA receptors; at -60 mV, the IC(50) values for memantine were 0.34 and 5.1 muM, respectively. Consistent with an open-channel blocking mechanism, memantine-induced NMDA receptor inhibition was voltage and use-dependent; the Hill coefficient (n(H)) was approximately 1. Memantine-induced alpha7(*) nAChR inhibition had an n(H) < 1 and showed a variable voltage dependence; the effect was voltage-independent at 0.1 muM, becoming voltage-dependent at >/=1 muM. Thus, memantine interacts with more than one class of sites on the alpha7(*) nAChRs. One is voltage-sensitive and therefore likely to be within the receptor channel. The other is voltage-insensitive and therefore likely to be in the extracellular domain of the receptor. It is suggested that blockade of alpha7(*) nAChRs by memantine could decrease its effectiveness for treatment of AD, particularly at early stages when the degrees of nAChR dysfunction and of cognitive decline correlate well.
ESTHER : Aracava_2005_J.Pharmacol.Exp.Ther_312_1195
PubMedSearch : Aracava_2005_J.Pharmacol.Exp.Ther_312_1195
PubMedID: 15522999

Title : Spine density and dendritic branching pattern of hippocampal CA1 pyramidal neurons in neonatal rats chronically exposed to the organophosphate paraoxon - Santos_2004_Neurotoxicol_25_481
Author(s) : Santos HR , Cintra WM , Aracava Y , Maciel CM , Castro NG , Albuquerque EX
Ref : Neurotoxicology , 25 :481 , 2004
Abstract : The organophosphate cholinesterase (ChE) inhibitor paraoxon is the oxidized active metabolite of parathion, a pesticide whose use in agriculture has been matter of increasing concern. The present work was aimed at reproducing a prolonged exposure to low concentrations of paraoxon and assessing possible damage to the hippocampus during the period of most significant cholinergic development. Male Wistar rats were given, from P8 to P20, subcutaneous daily injections of paraoxon (0.1, 0.15 and 0.2mg/kg). The rate of body weight gain was reduced by all doses of paraoxon and brain ChE activity progressively decreased up to 60% by P21. Some deaths occurred in the beginning of the treatment, but the surviving animals showed neither convulsions nor overt signs of cholinergic hyperstimulation. Morphometric analysis of Lucifer Yellow-stained CA1 pyramidal neurons in coronal sections of the hippocampus showed that by P21 paraoxon caused a decrease in spine density on basal but not on secondary apical dendrites. The dendritic arborization and the pyramidal and granular cell body layers were not altered by paraoxon. ChE staining decreased in all hippocampal and dentate gyrus regions studied, whereas choline acetyltransferase (ChAT) and zinc-positive fibers remained as in control. In summary, chronic exposure to low paraoxon concentrations during the period of rapid brain development caused significant and selective decrease in basal dendritic spine density of the CA1 pyramidal neurons. Distinct modulation of the basal tree at the stratum oriens by the interplay of cholinergic afferent and GABAergic interneurons, as well as the remodeling process in response to a repetitive and rather mild paraoxon insult, may account for this selective susceptibility of basal dendritic spines. The hippocampal alterations described here occurred in the absence of toxic cholinergic signs and may affect brain development and cause functional deficits that could continue into adulthood.
ESTHER : Santos_2004_Neurotoxicol_25_481
PubMedSearch : Santos_2004_Neurotoxicol_25_481
PubMedID: 15019311

Title : Low concentrations of pyridostigmine prevent soman-induced inhibition of GABAergic transmission in the central nervous system: involvement of muscarinic receptors - Santos_2003_J.Pharmacol.Exp.Ther_304_254
Author(s) : Santos MD , Pereira EF , Aracava Y , Castro NG , Fawcett WP , Randall WR , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 304 :254 , 2003
Abstract : This study was designed to investigate the effects of the cholinesterase inhibitors soman and pyridostigmine bromide (PB) on synaptic transmission in the CA1 field of rat hippocampal slices. Soman (1-100 nM, 10-15 min) decreased the amplitude of GABAergic postsynaptic currents (IPSCs) evoked by stimulation of Schaffer collaterals and recorded from CA1 pyramidal neurons. It also decreased the amplitude and frequency of spontaneous IPSCs recorded from pyramidal neurons. Whereas the maximal effect of soman on evoked GABAergic transmission was observed at 10 nM, full cholinesterase inhibition was induced by 1 nM soman. After 10-15-min exposure of hippocampal slices to 100 nM PB, GABAergic transmission was facilitated and cholinesterase activity was not significantly affected. At nanomolar concentrations, soman and PB have no direct effect on GABA(A) receptors. The effects of soman and PB on GABAergic transmission were inhibited by the m2 receptor antagonist 11-[[[2-diethylamino-O-methyl]-1-piperidinyl] acetyl]-5,11-dihydrol-6H-pyridol[2,3-b][1,4]benzodiazepine-6- one (1 nM) and the m3 receptor antagonist 4-diphenylacetoxy-N-methyl-piperidine (100 nM), respectively, and by the nonselective muscarinic receptor antagonist atropine (1 microM). Thus, changes in GABAergic transmission are likely to result from direct interactions of soman and PB with m2 and m3 receptors, respectively, located on GABAergic fibers/neurons synapsing onto the neurons under study. Although the effects of 1 nM soman and 100 nM PB were diametrically opposed, they only canceled one another when PB was applied to the neurons before soman. Therefore, PB, acting via m3 receptors, can effectively counteract effects arising from the interactions of soman with m2 receptors in the brain.
ESTHER : Santos_2003_J.Pharmacol.Exp.Ther_304_254
PubMedSearch : Santos_2003_J.Pharmacol.Exp.Ther_304_254
PubMedID: 12490599

Title : Design, synthesis, and pharmacological profile of novel fused pyrazolo[4,3-d]pyridine and pyrazolo[3,4-b][1,8]naphthyridine isosteres: a new class of potent and selective acetylcholinesterase inhibitors - Barreiro_2003_J.Med.Chem_46_1144
Author(s) : Barreiro EJ , Camara CA , Verli H , Brazil-Mas L , Castro NG , Cintra WM , Aracava Y , Rodrigues CR , Fraga CA
Ref : Journal of Medicinal Chemistry , 46 :1144 , 2003
Abstract : A new family of tacrine (THA) analogues (7-9, 12), containing the azaheterocyclic pyrazolo[4,3-d]pyridine or pyrazolo[3,4-b][1,8]naphthyridine systems as isosteres of the quinoline ring of THA, has been synthesized. The compounds were tested in rat brain cholinesterases using Ellman's method, and all were fully efficacious in inhibiting the enzymes. Compounds 9 and 12b were the most potent against acetylcholinesterase (AChE), showing IC(50) of 6.0 and 6.4 microM, and were less active against rat brain butyrylcholinesterase, showing selectivity indexes of 5.3 and 20.9, respectively. Compounds 7-9 and 12 were also tested for their acute neurotoxicity in vitro, using cultured rat cortical cells. Compounds 7 and 8 were not significantly toxic; 9 was toxic at 500 microM, but not at 100 microM. The naphthyridine derivatives 12a and 12b showed a significant concentration-dependent neurotoxicity, being able to kill most cells at 500 microM. Molecular dynamic simulation using the X-ray crystal structure of AChE from Torpedo californica was used to explain the possible binding mode of these new THA isosteres.
ESTHER : Barreiro_2003_J.Med.Chem_46_1144
PubMedSearch : Barreiro_2003_J.Med.Chem_46_1144
PubMedID: 12646025

Title : The nicotinic allosteric potentiating ligand galantamine facilitates synaptic transmission in the mammalian central nervous system - Santos_2002_Mol.Pharmacol_61_1222
Author(s) : Santos MD , Alkondon M , Pereira EF , Aracava Y , Eisenberg HM , Maelicke A , Albuquerque EX
Ref : Molecular Pharmacology , 61 :1222 , 2002
Abstract : In this study, the patch-clamp technique was used to determine the effects of galantamine, a cholinesterase inhibitor and a nicotinic allosteric potentiating ligand (APL) used for treatment of Alzheimer's disease, on synaptic transmission in brain slices. In rat hippocampal and human cerebral cortical slices, 1 microM galantamine, acting as a nicotinic APL, increased gamma-aminobutyric acid (GABA) release triggered by 10 microM acetylcholine (ACh). Likewise, 1 microM galantamine, acting as an APL on presynaptically located nicotinic receptors (nAChRs) that are tonically active, potentiated glutamatergic or GABA-ergic transmission between Schaffer collaterals and CA1 neurons in rat hippocampal slices. The cholinesterase inhibitors rivastigmine, donepezil, and metrifonate, which are devoid of nicotinic APL action, did not affect synaptic transmission. Exogenous application of ACh indicated that high and low levels of nAChR activation in the Schaffer collaterals inhibit and facilitate, respectively, glutamate release onto CA1 neurons. The finding then that the nAChR antagonists methyllycaconitine and dihydro-beta-erythroidine facilitated glutamatergic transmission between Schaffer collaterals and CA1 neurons indicated that in a single hippocampal slice, the inhibitory action of strongly, tonically activated nAChRs in some glutamatergic fibers prevails over the facilitatory action of weakly, tonically activated nAChRs in other glutamatergic fibers synapsing onto a given neuron. Galantamine is known to sensitize nAChRs to activation by low, but not high agonist concentrations. Therefore, at 1 microM, galantamine is likely to increase facilitation of synaptic transmission by weakly, tonically activated nAChRs just enough to override inhibition by strongly, tonically activated nAChRs. In conclusion, the nicotinic APL action can be an important determinant of the therapeutic effectiveness of galantamine.
ESTHER : Santos_2002_Mol.Pharmacol_61_1222
PubMedSearch : Santos_2002_Mol.Pharmacol_61_1222
PubMedID: 11961141

Title : A novel thienylhydrazone, (2-thienylidene)3,4-methylenedioxybenzoylhydrazine, increases inotropism and decreases fatigue of skeletal muscle - Gonzalez-Serratos_2001_J.Pharmacol.Exp.Ther_299_558
Author(s) : Gonzalez-Serratos H , Chang R , Pereira EF , Castro NG , Aracava Y , Melo PA , Lima PC , Fraga CA , Barreiro EJ , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 299 :558 , 2001
Abstract : This study was designed to investigate the effects on single skeletal muscle fibers of a novel thienylhydrazone, referred to as LASSBio-294, which is a bioisoster of pyridazinone compounds that inhibit the cyclic AMP-specific phosphodiesterase (PDE) 4. Twitch and fatigue were analyzed in single skeletal muscle fibers isolated from either the semitendinous or the tibialis anterior muscles dissected from the frog Rana pipiens. LASSBio-294 (12.5-100 microM) increased twitch tension, accelerated the maximal rate of tension decay during relaxation, and had very little effect in the maximal rate of tension development of muscle fibers directly stimulated at < or =30 Hz. The positive inotropic effect of LASSBio-294 developed slowly, reaching its maximum at 40 min and was inversely proportional to the frequency of stimulation, becoming negligible at 60 and 90 Hz. The concentration-response relationship for LASSBio-294-induced potentiation of twitch tension was bell-shaped, with maximal effect occurring at 25 microM. In addition, LASSBio-294 reduced development of fatigue induced by tetanic stimulation of the muscle fibers and reduced the time needed for 80% prefatigue tension recovery after fatigue had developed to 50% of the maximal pretetanic force. These effects of LASSBio-294 can be fully explained by stimulation of the sarcoplasmic reticulum Ca2+ pump and could be ascribed to an increase in cellular levels of cyclic AMP due to PDE inhibition. The novel thienylhydrazone LASSBio-294 may be useful for treatment of patients suffering from conditions in which muscle fatigue is a debilitating symptom (e.g., chronic heart failure).
ESTHER : Gonzalez-Serratos_2001_J.Pharmacol.Exp.Ther_299_558
PubMedSearch : Gonzalez-Serratos_2001_J.Pharmacol.Exp.Ther_299_558
PubMedID: 11602667

Title : Low concentrations of the organophosphate VX affect spontaneous and evoked transmitter release from hippocampal neurons: toxicological relevance of cholinesterase-independent actions - Rocha_1999_Toxicol.Appl.Pharmacol_159_31
Author(s) : Rocha ES , Santos MD , Chebabo SR , Aracava Y , Albuquerque EX
Ref : Toxicol Appl Pharmacol , 159 :31 , 1999
Abstract : In the present study, the patch-clamp technique was applied to cultured hippocampal neurons to evaluate the effects of the nerve agent VX on evoked and spontaneous postsynaptic currents mediated by gamma-aminobutyric acid (GABA) and glutamate. At 0.01 nM, VX reduced the amplitude of evoked GABAergic currents, and only at concentrations >1 nM did it decrease the amplitude of evoked glutamatergic currents. The effect of VX on GABAergic currents, which was partially reversible upon washing of the neurons with VX-free external solution, could be prevented by the muscarinic antagonist atropine. In contrast, the effect of VX on glutamatergic currents, which was not reversible upon washing, appears to be related to the VX-induced reduction of the amplitude and frequency of repetitively firing by action potentials. In the presence of the Na(+)-channel blocker tetrodotoxin (TTX), VX (>/=10 nM) increased the frequency of GABA- and glutamate-mediated miniature postsynaptic currents (MPSCs). This effect of VX was unrelated to cholinesterase inhibition and was Ca(2+) dependent. The lack of effect of VX on MPSC kinetics indicates that VX-induced alterations of evoked and spontaneous currents are exclusively due to alterations of the transmitter release processes. The ability of VX to affect transmitter release in the brain may underlie some of its neurotoxic effects and may provide the basis for the development of therapeutic countermeasures to treat and/or prevent VX-induced neurotoxicity.
ESTHER : Rocha_1999_Toxicol.Appl.Pharmacol_159_31
PubMedSearch : Rocha_1999_Toxicol.Appl.Pharmacol_159_31
PubMedID: 10448123

Title : An analysis of low level doses of cholinesterase inhibitors in cultured neurons and hippocampal slices of rats - Rocha_1998_Drug.Chem.Toxicol_21 Suppl 1_191
Author(s) : Rocha ES , Chebabo SR , Santos MD , Aracava Y , Albuquerque EX
Ref : Drug & Chemical Toxicology , 21 Suppl 1 :191 , 1998
Abstract : Recent data indicate that the neurotoxic effects of organophosphate compounds, including those of the nerve agents VX and sarin, are not solely due to irreversible cholinesterase inhibition. In this study we applied the patch clamp technique to hippocampal neurons in culture and slices to investigate the effects of VX, sarin and huperzine A on transmitter release and the mechanisms related with such effects. The nerve agents VX and sarin at very low concentrations significantly reduced the evoked release of GABA and glutamate. This effect was dependent of the activation of muscarinic receptors. In the presence or absence of the Na(+)-channel blocker tetrodotoxin (TTX), VX increased the frequency of spontaneous glutamate and GABA-induced postsynaptic currents. The effect of VX on TTX-insensitive spontaneous currents appears to be unrelated to cholinesterase inhibition, because it could be detected even after cholinesterase was blocked by high concentrations of the nerve agent soman. The ability of the nerve gases to decrease evoked release of GABA and increase spontaneous transmitter release may underlie some of the neurotoxic effects of the compounds. Huperzine A did not affect spontaneous or evoked release of GABA and glutamate, suggesting that this compound may be a pure cholinesterase inhibitor and had no effect on postsynaptic GABAA or AMPA receptors.
ESTHER : Rocha_1998_Drug.Chem.Toxicol_21 Suppl 1_191
PubMedSearch : Rocha_1998_Drug.Chem.Toxicol_21 Suppl 1_191
PubMedID: 10028411

Title : Properties of neuronal nicotinic acetylcholine receptors: pharmacological characterization and modulation of synaptic function -
Author(s) : Albuquerque EX , Alkondon M , Pereira EF , Castro NG , Schrattenholz A , Barbosa CT , Bonfante-Cabarcas R , Aracava Y , Eisenberg HM , Maelicke A
Ref : Journal of Pharmacology & Experimental Therapeutics , 280 :1117 , 1997
PubMedID: 9067295

Title : Methamidophos: an anticholinesterase without significant effects on postsynaptic receptors or transmitter release - Camara_1997_Neurotoxicol_18_589
Author(s) : Camara AL , Braga MF , Rocha ES , Santos MD , Cortes WS , Cintra WM , Aracava Y , Maelicke A , Albuquerque EX
Ref : Neurotoxicology , 18 :589 , 1997
Abstract : Methamidophos (O,S-dimethyl phosphoroamidothiolate, Tamaron), an organophosphate (OP) anticholinesterase of limited toxicity, is widely used as an insecticide and acaricide. To provide additional insight into the molecular basis of its action, we have used electrophysiological and biochemical techniques to study the effects of methamidophos on the neuromuscular junction of rat and frog and on the central nervous system of rat. Methamidophos has a relatively weak inhibitory action on cholinesterases in rat diaphragm muscle, brain and hippocampal homogenates, with IC50 values on the order of 20-20 microM. An even weaker anticholinesterase activity was found in frog muscle homogenates, with the IC50 being above 300 microM. As further evidence of anticholinesterase activity, methamidophos (1-100 microM) was able to reverse the blockade by d-tubocurarine (0.5-0.7 microM) of neuromuscular transmission in rat phrenic nerve-hemidiaphragm preparations. Inhibition of cholinesterase activity by methamidophos was long lasting, which is consistent with the formation by the agent of a covalent bond with the enzyme's active serine residue. The action was also slowly reversible, which suggests spontaneous reactivation of the enzyme. electrophysiological studies at the rat neuromuscular junction showed that, due to its anticholinesterase activity, methamidophos increased the amplitude and prolonged the decay phase of nerve-evoked and spontaneous miniature end-plate potentials. In contrast to other OP compounds, e.g., paraoxon (Rocha et al., 1996a), methamidophos did not affect neurotransmitter release, nor did it interact directly with the muscle nicotinic acetylcholine receptor. Moreover, it contrast to paraoxon, methamidophos did not affect the whole-cell currents induced by application of acetylcholine, glutamate or gamma-aminobutyric acid recorded to cultured hippocampal neurons. Based on these data, methamidophos appears to have a selective effect on cholinesterase.
ESTHER : Camara_1997_Neurotoxicol_18_589
PubMedSearch : Camara_1997_Neurotoxicol_18_589
PubMedID: 9291508

Title : Glycine and calcium-dependent effects of lead on N-methyl-D-aspartate receptor function in rat hippocampal neurons - Marchioro_1996_J.Pharmacol.Exp.Ther_279_143
Author(s) : Marchioro M , Swanson KL , Aracava Y , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 279 :143 , 1996
Abstract : The effects of lead (Pb++) on N-methyl-D-aspartate (NMDA) receptor function of rat hippocampal neurons in culture were studied by use of the whole-cell patch-clamp technique. Currents activated by NMDA (100 microM) in the presence of nonsaturating concentrations of glycine (0.01-0.05 microM) were potentiated in a voltage-independent manner by Pb++ (1-10 microM), and the potentiation was antagonized by 50 microM kynurenic acid. Increasing extracellular Ca++ from 1 to 10 mM similarly potentiated the NMDA-activated currents in the presence of a nonsaturating concentration of glycine (0.2 microM). The potentiation of NMDA-activated currents by low micromolar concentrations of Pb++ may be mediated by this cation's ability to increase the affinity of the NMDA receptor for glycine. In the presence of 10 microM glycine and 2 mM Ca++, Pb++ reduced the peak amplitudes of currents activated by NMDA (100 microM) in a voltage-independent manner (IC50 = 5.9 microM Pb++, Hill coefficient (nH) = 1.2). Also, steady-state currents activated by NMDA (50 microM) were inhibited by rapid application of Pb++ (IC50 = 3.2 microM, nH = 0.7). Increasing extracellular Ca++, in the presence of 10 microM glycine, reduced the NMDA-activated currents and shifted the Pb++ concentration-response curves to the right: at 0.2, 2 and 20 mM Ca++, the IC50 values of Pb++ were 3.0, 5.9 and 12.5 microM and the nH values were 0.9, 1.2 and 1.1, respectively. The finding that external Ca++ antagonized the inhibitory effect of Pb++ suggests that the noncompetitive inhibitory action of Pb++ with respect to glycine and NMDA may be mediated by Pb++ competition with Ca++ for a site on the NMDA receptor.
ESTHER : Marchioro_1996_J.Pharmacol.Exp.Ther_279_143
PubMedSearch : Marchioro_1996_J.Pharmacol.Exp.Ther_279_143
PubMedID: 8858987

Title : Paraoxon: cholinesterase-independent stimulation of transmitter release and selective block of ligand-gated ion channels in cultured hippocampal neurons - Rocha_1996_J.Pharmacol.Exp.Ther_278_1175
Author(s) : Rocha ES , Swanson KL , Aracava Y , Goolsby JE , Maelicke A , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 278 :1175 , 1996
Abstract : Paraoxon (O,O-diethyl O-p-nitrophenyl phosphate) is the neurotoxic metabolite of the insecticide parathion (O,O-diethyl O-p-nitrophenyl phosphorothioate). The effects of organophosphorus compounds on peripheral synapses have been attributed to inhibition of cholinesterase and to direct actions on muscarinic and nicotinic receptors, but less is known about the actions of organophosphorus compounds, including paraoxon, in the central nervous system. We investigated initially the effects of paraoxon on spontaneous transmitter release by recording miniature postsynaptic currents (MPSCs) from cultured rat hippocampal neurons using the whole-cell mode of the patch-clamp technique. Paraoxon (0.3 microM) in the presence of tetrodotoxin (0.3 microM) and atropine (1 microM) caused a significant increase in the frequency of gamma-aminobutyric acid- and glutamate-mediated MPSCs, but did not change the peak amplitudes or decay-time constants of these MPSCs. In contrast, application of nicotinic agonists or antagonists did not change the MPSC frequency. The presynaptic effect of paraoxon shown here was not mediated by actions on muscarinic or nicotinic receptors, or by elevated acetylcholine levels secondary to inhibition of cholinesterase. In addition, agonists were applied to assess the postsynaptic effects of paraoxon on excitatory and inhibitory amino acid receptors. Paraoxon (30 microM-1 mM) blocked the ion channels of glycine, gamma-aminobutyric acidA, N-methyl-D-aspartic acid and nicotinic acetylcholine receptors, but not the ion channels of kalnate- and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptors. The combined effects of paraoxon on spontaneous transmitter release and on the functions of several ligand-gated receptors may constitute mechanisms relevant to the neurotoxicity of paraoxon.
ESTHER : Rocha_1996_J.Pharmacol.Exp.Ther_278_1175
PubMedSearch : Rocha_1996_J.Pharmacol.Exp.Ther_278_1175
PubMedID: 8819500

Title : Ligand-gated ion channels in acutely dissociated rat hippocampal neurons with long dendrites - Barbosa_1996_Neurosci.Lett_210_177
Author(s) : Barbosa CT , Alkondon M , Aracava Y , Maelicke A , Albuquerque EX
Ref : Neuroscience Letters , 210 :177 , 1996
Abstract : A technique for dissociation of hippocampi of 3-25-day-old rats is described by which pyramidal and bipolar neurons with many long (up to 200 microns) dendrites can be obtained. Dissociation of CA1 neurons was achieved by mechanical means, in the absence of Ca2+, and without the use of proteolytic enzymes. The functional properties of the dissociated neurons were assessed using the whole-cell patch-clamp technique. Whole-cell currents were elicited by U-tube application of the agonists N-methyl-D-aspartate (NMDA), gamma-aminobutyric acid (GABA), and acetylcholine (ACh), and spontaneous miniature currents were also observed in these neurons. ACh-elicited currents were blocked by methyllycaconitine (MLA, 1 nM) and Pb2+ (0.1-10 microM). These results establish acutely dissociated neurons as a simple and reliable preparation for the study of the pharmacology, kinetics and subcellular distribution of ligand-gated ion channels.
ESTHER : Barbosa_1996_Neurosci.Lett_210_177
PubMedSearch : Barbosa_1996_Neurosci.Lett_210_177
PubMedID: 8805124

Title : Pyrazole, an alcohol dehydrogenase inhibitor, has dual effects on N-methyl-D-aspartate receptors of hippocampal pyramidal cells: agonist and noncompetitive antagonist - Pereira_1992_J.Pharmacol.Exp.Ther_261_331
Author(s) : Pereira EF , Aracava Y , Aronstam RS , Barreiro EJ , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 261 :331 , 1992
Abstract : Electrophysiological and biochemical studies demonstrated that pyrazole, an inhibitor of alcohol dehydrogenase and a proposed therapeutic agent for treatment of alcoholic intoxication, activated and blocked the N-methyl-D-aspartate (NMDA) receptor and did not interact significantly with the end-plate nicotinic acetylcholine receptor (AChR). Pyrazole, at concentrations as low as 0.5 microM, applied to outside-out patches excised from the membrane of cultured rat hippocampal neurons, elicited single-channel currents of 48 pS which were blocked by DL-2-amino-5-phosphorovaleric acid, a competitive antagonist of NMDA. In addition, binding studies showed that pyrazole displaced 1-(cis-2-carboxypiperidine-4-yl)methyl-1-phosphoric acid from the agonist recognition site of the NMDA receptor in a concentration-dependent manner and enhanced the binding of (+)-5-methyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-imine to this complex. These data indicate that pyrazole is an agonist at NMDA receptors. However, at higher concentrations, open and burst times as well as the frequency of single-channel currents activated by pyrazole were reduced significantly, a finding which suggests that this compound is also an open channel blocker. In agreement with these results, it was shown biochemically that pyrazole was able to stimulate influx of Ca++ into rat brain microsomes via NMDA receptors and on the other hand to block the influx of Ca++ induced by NMDA. Pyrazole was unable to affect the neuromuscular transmission of frog sartorius muscle-sciatic nerve preparations. Additionally, pyrazole did not interact either with the agonist recognition site or with noncompetitive sites of the AChR. However, this drug had a very weak agonist-like action on the AChR of the Torpedo electric organ, most likely via binding sites different from those described previously for acetylcholine. Therefore, the therapeutic efficacy of pyrazole may be related at least in part to its effects on the NMDA receptor. Furthermore, this compound, because of the small size and rigidity of its molecular structure, becomes a promising drug for the study of the NMDA receptor. Indeed its use may allow a better understanding of the physiological and pathological processes involving this receptor.
ESTHER : Pereira_1992_J.Pharmacol.Exp.Ther_261_331
PubMedSearch : Pereira_1992_J.Pharmacol.Exp.Ther_261_331
PubMedID: 1313873

Title : Sensitivity of N-methyl-D-aspartate (NMDA) and nicotinic acetylcholine receptors to ethanol and pyrazole -
Author(s) : Aracava Y , Froes-Ferrao MM , Pereira EF , Albuquerque EX
Ref : Annals of the New York Academy of Sciences , 625 :451 , 1991
PubMedID: 1711814

Title : Functional properties of the nicotinic and glutamatergic receptors - Albuquerque_1991_J.Recept.Res_11_603
Author(s) : Albuquerque EX , Costa AC , Alkondon M , Shaw KP , Ramoa AS , Aracava Y
Ref : J Recept Res , 11 :603 , 1991
Abstract : Several important physiological processes such as plasticity, memory, cell death, and rhythmic firing involve the N-methyl-D-aspartate (NMDA)-type of glutamatergic receptor. Nicotinic acetylcholine receptors (AChR), recently demonstrated in the central nervous system (CNS), are also of great interest. We have used several ligands to study the physiology and pharmacology of the agonist recognition sites of these receptors and kinetic properties of associated ion channels using whole-cell, cell-attached or outside-out variants of the patch-clamp technique. Enzymatically dissociated frog interosseal muscles were used to study peripheral AChRs, and tissue cultured or acutely dissociated hippocampal neurons and retinal ganglion cells (RGCs) for CNS receptors. For reproducible and fast solution changes when recording in the whole-cell configuration, we modified the "U"-shaped tube system to obtain different outputs from the same outflow port. We used fluorescent rhodamine-labeled latex microspheres to identify RGCs. Our studies provide important information regarding the molecular mechanisms of several clinically used agents. Additionally, similar actions of noncompetitive agents on the ion channels of the nicotinic ACh and NMDA receptors support the concept of a receptor ion channel superfamily.
ESTHER : Albuquerque_1991_J.Recept.Res_11_603
PubMedSearch : Albuquerque_1991_J.Recept.Res_11_603
PubMedID: 1715922

Title : Activation and blockade of the acetylcholine receptor-ion channel by the agonists (+)-anatoxin-a, the N-methyl derivative and the enantiomer - Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
Author(s) : Kofuji P , Aracava Y , Swanson KL , Aronstam RS , Rapoport H , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 252 :517 , 1990
Abstract : The effects of (+)- and (-)-anatoxin-a (AnTX) and N-methylanatoxin (M-AnTX) on peripheral nicotinic ion channel activity were studied using high micromolar concentrations. Whereas (+)-AnTX is an effective agonist at nanomolar concentrations, (-)-AnTX and M-AnTX were effective at low micromolar concentrations. The binding of [3H]perhydrohistrionicotoxin to the nicotinic acetylcholine receptor-ion channel was stimulated by the above agonist concentrations, but [3H]perhydrohistrionicotoxin binding was inhibited at high micromolar concentrations of each of the toxins. In single channel recordings, these toxins exhibited ion channel blocking properties; the concentration- and voltage-dependent kinetics of each were essentially the same. In the case of (+)-AnTX, desensitization was also present at micromolar concentrations. These data show that ion channel blockade may be a property of many anatoxin-a analogs, and that in the particular case of analogs with low agonist potency, ion channel blockade may be a concomitant primary effect of the toxins. Stereospecificity and number of amine moieties did not influence the ion channel blocking characteristics in this series of molecules, although these factors strongly modified agonist potency.
ESTHER : Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
PubMedSearch : Kofuji_1990_J.Pharmacol.Exp.Ther_252_517
PubMedID: 1690292

Title : Molecular effects of dimethylanatoxin on the peripheral nicotinic acetylcholine receptor - Costa_1990_J.Pharmacol.Exp.Ther_252_507
Author(s) : Costa AC , Swanson KL , Aracava Y , Aronstam RS , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 252 :507 , 1990
Abstract : N,N-dimethylanatoxin (DMAnTX), the quaternary derivative of the potent nicotinic agonist (+)-anatoxin-a (AnTX), has been evaluated for potency and efficacy at nicotinic acetylcholine receptors of frog motor endplates and Torpedo electric organs. DMAnTX was only weakly effective in eliciting contracture of the frog rectus abdominis and was orders of magnitude less potent than AnTX. Biochemical assay showed that DMAnTX was a weak inhibitor of [125I]alpha-bungarotoxin binding to the receptors in frog muscle and Torpedo electroplaque membranes: the IC50 values were 60 and 14 microM, respectively. A low frequency of single channel currents recorded from isolated interosseal fibers at concentrations from 20 to 100 microM of DMAnTX and the stimulation of [3H]perhydrohistrionicotoxin [( 3H]H12-HTX) binding (half-maximal at 0.3 microM) confirmed the weak activation of the receptor. DMAnTX also exhibited antagonist effects. In muscle twitch assays, 100 microM of DMAnTX effectively decreased the tension induced by nerve stimulation, although DMAnTX did not affect muscle membrane action potentials. The binding of [3H] perhydrohistrionicotoxin was also inhibited at high micromolar concentrations of DMAnTX. Combination of DMAnTX with acetylcholine in single channel current experiments demonstrated that DMAnTX possesses ion channel blocking properties, which become apparent at low micromolar concentrations, and DMAnTX enhances the desensitization induced by acetylcholine above 10 microM AnTX. The difference in agonist potency between AnTX and DMAnTX may be attributed to a change in conformation of the molecular skeleton induced by the N-methyl groups.
ESTHER : Costa_1990_J.Pharmacol.Exp.Ther_252_507
PubMedSearch : Costa_1990_J.Pharmacol.Exp.Ther_252_507
PubMedID: 1690291

Title : The anticonvulsant MK-801 interacts with peripheral and central nicotinic acetylcholine receptor ion channels - Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
Author(s) : Ramoa AS , Alkondon M , Aracava Y , Irons J , Lunt GG , Deshpande SS , Wonnacott S , Aronstam RS , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 254 :71 , 1990
Abstract : The effects of MK-801 [( +]-5-methyl-10,11-dihydro-5H-di-benzo[a, d]cyclohepten-5,10-imine) on peripheral and central nicotinic receptors were studied using electrophysiological and biochemical techniques. MK-801 depressed the peak amplitude and accelerated the decay of end-plate currents. The drug (1-10 microM) decreased the frequency of activation of acetylcholine (ACh)-induced single-channel currents in addition to shortening the mean open and burst times of channels activated by either ACh or (+)anatoxin-a (AnTX). MK-801 (10-40 microM) depressed the single potentials and trains of ACh and AnTX-induced potentials in chronically denervated rat soleus muscles. MK-801 blocked the twitch responses (20-100 microM) of both frog sartorius and rat diaphragm muscles evoked by stimulation of their respective nerves. Also this drug (less than 1 microM) decreased the frequency of channels activated by AnTX or ACh in outside-out patch membranes of rat retinal ganglion cells with minimal changes in the channel open time. MK-801 (10-25 microM) depressed (-)nicotine-evoked gamma-amino[2,3-3H]butyric acid release from rat hippocampal synaptosomes; however, it failed to affect the binding of [3H](-)nicotine to brain membranes and also failed to interfere with the binding of [125I]alpha-bungarotoxin to either frog muscle or Torpedo membranes. On the other hand, MK-801 inhibited the binding of [3H]perhydrohistrionicotoxin to Torpedo membranes and such an effect was more pronounced in the presence of carbamylcholine. Neither AnTX nor any other nicotinic agonist increased the binding of [3H]MK-801 to the N-methyl-D-aspartate receptor ion channel complex. The actions of MK-801 were evident at concentrations comparable with those needed to block N-methyl-D-aspartate receptors. These results demonstrate the existence of at least three different types of nicotinic AChR, all of which were blocked noncompetitively by MK-801.
ESTHER : Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
PubMedSearch : Ramoa_1990_J.Pharmacol.Exp.Ther_254_71
PubMedID: 1694895

Title : Agonist recognition site of the peripheral acetylcholine receptor ion channel complex differentiates the enantiomers of nicotine - Rozental_1989_J.Pharmacol.Exp.Ther_251_395
Author(s) : Rozental R , Aracava Y , Scoble GT , Swanson KL , Wonnacott S , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 251 :395 , 1989
Abstract : The multiple actions of nicotine enantiomers at the peripheral nicotinic acetylcholine receptor were evaluated using electrophysiological and biochemical techniques. The alpha-bungarotoxin binding site showed a 6-fold greater affinity for (-)-nicotine than for the (+)-isomer, and this stereoselectivity was reflected in differences in the ability of the alkaloids to activate physiological responses in the forms of single ion channel currents, endplate depolarizations and muscle contractures. (-)-Nicotine was also more potent to induce slow desensitization. In contrast, both (-)- and (+)-nicotine were equipotent as ion channel blockers. Ion channel blockade occurred at effective agonist concentrations for (+)-nicotine but above the effective concentration for (-)-nicotine. The rapid and reversible interaction of nicotine enantiomers with the ion channel occurred at concentrations which implicate a significant contribution of channel blockade to the inhibition of indirect muscle twitch. The agonistic and ion channel blocking effects of the nicotine enantiomers provide important clues regarding the mechanisms by which nicotine may affect central nervous system nicotinic receptors.
ESTHER : Rozental_1989_J.Pharmacol.Exp.Ther_251_395
PubMedSearch : Rozental_1989_J.Pharmacol.Exp.Ther_251_395
PubMedID: 2478693

Title : N-methylanatoxinol isomers: derivatives of the agonist (+)-anatoxin-a block the nicotinic acetylcholine receptor ion channel - Swanson_1989_Mol.Pharmacol_35_223
Author(s) : Swanson KL , Aracava Y , Sardina FJ , Rapoport H , Aronstam RS , Albuquerque EX
Ref : Molecular Pharmacology , 35 :223 , 1989
Abstract : Using biochemical and patch-clamp techniques, we investigated the pharmacology of S- and R-epimers of N-methylanatoxinol, which are analogs of the semi-rigid, stereoselective, nicotinic agonist (+)-anatoxin-a. In contrast to (+)-anatoxin-a, both isomers had poor ability to inhibit the binding of 125I-alpha-bungarotoxin or to open acetylcholine channels, and they were unable to elicit contracture of frog rectus abdominis muscles. However, both isomers were able to demonstrate significant concentration-dependent blockade of the nicotinic acetylcholine receptor ion channel. The R-isomer was approximately 4-fold more potent in causing inhibition of [3H]H12HTX binding than was the S-isomer, in the absence of carbamylcholine. In the presence of carbamylcholine, the affinity of the R-isomer of N-methylanatoxinol for the ion channel sites was further enhanced, so that its affinity became much greater than that of the S-isomer. Refinement of voltage- and concentration-dependent terms for the ion channel blocking and unblocking rates yielded functions that were able to predict the channel open times and short closed times well. The S-isomer bound and dissociated from the ion channel site of the nicotinic acetylcholine receptor more rapidly and with greater voltage sensitivity than the R-isomer. The present characterization of the antagonistic properties of these new analogs of (+)-anatoxin-a introduces a new aspect to the molecular pharmacology of (+)-anatoxin-a analogs; the semi-rigid compounds could be useful in describing the allosteric binding sites of the acetylcholine receptor, as well as in delimiting the agonist binding site.
ESTHER : Swanson_1989_Mol.Pharmacol_35_223
PubMedSearch : Swanson_1989_Mol.Pharmacol_35_223
PubMedID: 2465486

Title : Structure-activity relationship of reversible cholinesterase inhibitors: activation, channel blockade and stereospecificity of the nicotinic acetylcholine receptor-ion channel complex - Albuquerque_1988_Braz.J.Med.Biol.Res_21_1173
Author(s) : Albuquerque EX , Aracava Y , Cintra WM , Brossi A , Schonenberger B , Deshpande SS
Ref : Brazilian Journal of Medical & Biological Research , 21 :1173 , 1988
Abstract : 1. We have shown that all cholinesterase (ChE) inhibitors, in addition to their well-known anti-ChE activity, have multiple effects on the nicotinic acetylcholine receptor-ion channel (AChR) macromolecule resulting from interactions with the agonist recognition site and with sites located at the ion channel component. Activation, competitive antagonism and different types of noncompetitive blockade occurring at similar concentration ranges and contributing in different proportions result in complex and somewhat unpredictable alterations in AChR function. The question is now raised as to how each effect of these compounds contributes to their antidotal property against organophosphorus (OP) poisoning, and what set of actions makes one reversible ChE inhibitor a better antidote. Many lines of evidence support the importance of direct interactions with various sites on the AChR: 1) morphological and toxicological studies with (+) physostigmine showed that anti-ChE activity is not essential to protect animals against toxicity by irreversible ChE inhibitors; 2) (-)physostigmine is far more effective against OP poisoning; 3) open channel blockers such as mecamylamine with no significant anti-ChE activity enhance the protective action of (-)physostigmine; 4) neostigmine, pyridostigmine, (-)physostigmine and (+)physostigmine showed qualitatively and quantitatively distinct toxicity and damage to endplate morphology and function. 2. In prophylaxis and during the very early phase of OP poisoning, carbamates, especially (-)physostigmine combined with mecamylamine and atropine, could protect almost 100% of the animals exposed to multiple lethal doses of OPs. Electrophysiological data showed that (-)physostigmine, among several reversible ChE inhibitors, showed greater potency in depressing both endplate current (EPC) peak amplitude and tau EPC. Therefore, concerning neuromuscular transmission, it seems that the higher the potency of a drug in reducing endplate permeability, the better is its protection against OP toxicity. A reversible open channel blockade combined with some agonist property helps to decrease the effect of ACh at its agonist site and to reduce the ion permeability of open channels. It should be pointed out that, during the later phase of OP poisoning, AChR desensitization should be most prevalent. Thus, a drug that can remove the AChR from this rather irreversible state to a more reversible blocked state should be a better protector. Indeed, oximes such as 2-PAM and a more potent analog, HI-6, produce multiple alterations in AChR function that comprise increased channel activation and open-channel blockade.(ABSTRACT TRUNCATED AT 400 WORDS)
ESTHER : Albuquerque_1988_Braz.J.Med.Biol.Res_21_1173
PubMedSearch : Albuquerque_1988_Braz.J.Med.Biol.Res_21_1173
PubMedID: 3074841

Title : Nicotinic acetylcholine receptors in cultured neurons from the hippocampus and brain stem of the rat characterized by single channel recording - Aracava_1987_FEBS.Lett_222_63
Author(s) : Aracava Y , Deshpande SS , Swanson KL , Rapoport H , Wonnacott S , Lunt G , Albuquerque EX
Ref : FEBS Letters , 222 :63 , 1987
Abstract : Single channel recording techniques have been applied to neurons cultured from the hippocampus and the respiratory area of the brain stem of fetal rats in order to search for nicotinic acetylcholine receptors (nAChR) in the central nervous system. In addition to acetylcholine (ACh), the potent and specific agonist (+)-anatoxin-a was also used to characterize nicotinic channels. nAChRs were concentrated on the somal surface near the base of the apical dendrite, and in some patches their density was sufficient to record 2 or more channel openings simultaneously. Although a multiplicity of conductance states was also evident, the predominant population showed a single channel conductance of 20 pS at 10 degrees C. Thus, these neuronal nAChRs resembled the embryonic or denervated-type nAChRs in muscle. However, channel opening and closing kinetics were faster than reported for similar conductance channels in muscle. Therefore the nicotinic channels described here are similar but not identical to those of the well-characterized muscle nAChR, in agreement with biochemical, pharmacological, and molecular genetic studies on brain AChR.
ESTHER : Aracava_1987_FEBS.Lett_222_63
PubMedSearch : Aracava_1987_FEBS.Lett_222_63
PubMedID: 2443390

Title : The molecular basis of anticholinesterase actions on nicotinic and glutamatergic synapses -
Author(s) : Aracava Y , Deshpande SS , Rickett DL , Brossi A , Schonenberger B , Albuquerque EX
Ref : Annals of the New York Academy of Sciences , 505 :226 , 1987
PubMedID: 2446549

Title : Noncompetitive blockade of the nicotinic acetylcholine receptor-ion channel complex by an irreversible cholinesterase inhibitor - Rao_1987_J.Pharmacol.Exp.Ther_240_337
Author(s) : Rao KS , Aracava Y , Rickett DL , Albuquerque EX
Ref : Journal of Pharmacology & Experimental Therapeutics , 240 :337 , 1987
Abstract : Interactions of O-ethyl S-[2-(diisopropylamino)ethyl]methylphosphonothioate (VX), an irreversible, organophosphorus, anticholinesterase (anti-ChE) agent, with the nicotinic acetylcholine receptor-ion channel complex (AChR) of the frog Rana pipiens were investigated using electrophysiological techniques. At low concentrations (0.1-0.5 microM) of VX, typical effects due to cholinesterase (ChE) inhibition, such as potentiation of indirect muscle twitches as well as increases in the peak amplitude and decay time constant (tau EPC) of end-plate currents (EPC), were observed. At concentrations greater than or equal to 1.0 microM, VX produced opposite effects. The indirectly elicited muscle twitches and EPC peak amplitude were depressed with an IC50 of about 33 microM. tau EPC was reduced, and at a higher concentration of 100 microM, VX split the decays into faster and slower components. Similar results were also obtained with the amplitude and decays of miniature end-plate currents (MEPC). However, although the MEPC peak amplitude and tau MEPC were not decreased to levels below control values, EPC peak amplitude (but not its tau EPC) was markedly depressed beyond the control values, in a concentration-dependent manner. The fact that nerve action potential-evoked events were more affected than the spontaneous MEPCs suggested an interference with the depolarization-evoked release process. Indeed, VX caused a reduction of the quantal content and, in addition, induced a significant increase in the frequency of MEPCs. All these effects, except the anti-ChE activity, were reversible upon wash. The results from both EPC fluctuation analysis and single channel recordings disclosed a concentration-dependent shortening of the channel open times without change in single channel conductance.(ABSTRACT TRUNCATED AT 250 WORDS)
ESTHER : Rao_1987_J.Pharmacol.Exp.Ther_240_337
PubMedSearch : Rao_1987_J.Pharmacol.Exp.Ther_240_337
PubMedID: 2433427

Title : Activation and Inhibition of the Nicotinic Receptor: Actions of Physostigmine, Pyridostigmine and Meproadifen -
Author(s) : Albuquerque EX , Allen CN , Aracava Y , Akaike A , Shaw KP , Rickett DL
Ref : Advances in Behavioral Biology , 30 :677 , 1986
PubMedID:

Title : A possible involvement of cyclic AMP in the expression of desensitization of the nicotinic acetylcholine receptor. A study with forskolin and its analogs - Albuquerque_1986_FEBS.Lett_199_113
Author(s) : Albuquerque EX , Deshpande SS , Aracava Y , Alkondon M , Daly JW
Ref : FEBS Letters , 199 :113 , 1986
Abstract : Forskolin, an activator of adenylate cyclase, and its analogs were studied on the nicotinic acetylcholine receptor-ion channel complex (AChR) of rat and frog skeletal muscles. At nanomolar concentrations, forskolin caused desensitization of the AChR located at the junctional region of innervated and the extrajunctional region of chronically denervated rat soleus muscles. The desensitization of the AChR occurred without alteration of the conducting state (channel lifetime, conductance or bursting) as shown by single channel currents. Accordingly, forskolin decreased the peak amplitude of the repetitive evoked endplate currents in frog sartorius muscles. These findings taken together with the good correlation found between the effects of forskolin and its analogs on the desensitization of the nicotinic AChR and their ability to activate adenylate cyclase suggested a possible involvement of phosphorylation of AChR via cyclic AMP on the desensitization process.
ESTHER : Albuquerque_1986_FEBS.Lett_199_113
PubMedSearch : Albuquerque_1986_FEBS.Lett_199_113
PubMedID: 2420646

Title : The acetylcholine receptor of the neuromuscular junction recognizes mecamylamine as a noncompetitive antagonist - Varanda_1985_Mol.Pharmacol_28_128
Author(s) : Varanda WA , Aracava Y , Sherby SM , VanMeter WG , Eldefrawi ME , Albuquerque EX
Ref : Molecular Pharmacology , 28 :128 , 1985
Abstract : The secondary amine, mecamylamine, interacts with the nicotinic receptor ionic channel complex as a noncompetitive antagonist. Mecamylamine (1-10 microM) blocked indirect muscle twitches with no discernible effect on the membrane potential, overshoot, or amplitude of the action potential. It also produced a voltage- and concentration-dependent depression of the peak amplitude of the endplate currents (EPC) and induced nonlinearity in the current-voltage relationship. The decay time constant of the EPC (TEPC) was significantly shortened. The linear relationship between the reciprocal of TEPC and the drug concentration suggested an open channel blockade. Patch-clamp studies, in agreement with the noise analysis results, revealed that mecamylamine (1-8 microM) shortened the lifetime of the open channels. Further, the single channel studies showed that at high concentrations mecamylamine reduced the double exponential nature of the distribution of open times characteristic of channels recorded from myoballs. Closed times had a complex distribution that could not be fitted to a single exponential function because of the presence of short closures or "flickers" during the open state. Although the frequency of channel openings progressively decreased with increasing drug concentration, the single channel conductance remained unchanged at all the concentrations tested. Biochemical studies showed that mecamylamine (up to 100 microM) did not block [3H]acetylcholine binding to the nicotinic receptor of the Torpedo electroplax, but inhibited the binding of [3H]perhydrohistionicotoxin to its channel site, both in the resting and the activated state. These results suggested that, at the nicotinic receptors of the neuromuscular junction, mecamylamine acted as a noncompetitive blocker, binding primarily to the receptor's open channel conformation. Most of the alterations of EPCs were consistent with the predictions of a sequential model for open channel blockade. Biochemical and patch-clamp results, however, could not be fully explained by this model and provided some evidence of the existence of additional blocked states most likely through pathways into desensitized species. In contrast to a competitive antagonism of acetylcholine receptors reported at autonomic ganglia, there was no such action of the drug at the neuromuscular junction; thus, mecamylamine is a useful tool to characterize the nicotinic receptors from different synapses.
ESTHER : Varanda_1985_Mol.Pharmacol_28_128
PubMedSearch : Varanda_1985_Mol.Pharmacol_28_128
PubMedID: 2410768

Title : Multiple actions of anticholinesterase agents on chemosensitive synapses: molecular basis for prophylaxis and treatment of organophosphate poisoning - Albuquerque_1985_Fundam.Appl.Toxicol_5_S182
Author(s) : Albuquerque EX , Deshpande SS , Kawabuchi M , Aracava Y , Idriss M , Rickett DL , Boyne AF
Ref : Fundamental & Applied Toxicology , 5 :S182 , 1985
Abstract : The present study demonstrates that the reversible and irreversible anti-ChE agents have direct actions on the nicotinic acetylcholine receptor-ionic channel (AChR) and on the locust glutamatergic neuromuscular junction. In addition, the prophylaxis of lethality of organophosphorus anti-ChE compounds was studied. The lethality of VX and sarin was diminished when the rats were pretreated with physostigmine and atropine. The effectiveness of this protection, however, was markedly increased when a ganglionic blocker, either mecamylamine or chlorisondamine, was added, such that all the animals survived after receiving four times a lethal dose of VX. Pretreated animals receiving sarin showed significant recovery of morphological and functional properties of the neuromuscular junction as compared to the damage of structures from animals without pretreatment. Blood ChE inhibition was slightly decreased while brain and muscle AChE levels were significantly recovered (from 98 and 70% to 56 and 32%, respectively) by the pretreatment. This effect may partially explain the protection given by physostigmine but not that afforded by addition of a non-anti-ChE agent. Physostigmine, at concentrations greater than 20 microM, showed both a marked depression of the peak amplitudes of the endplate current (EPC) and a shortening of the decay time constants tau EPC. These effects were mostly due to a direct drug interaction with the nicotinic AChR blocking the ionic channel in its open conformation. Single-channel recordings showed that physostigmine decreases conductance and open times of the channels activated in the presence of ACh and in addition has an agonistic property on the nicotinic AChR. VX, on the other hand, only shortened the open times of ACh-activated channels without affecting the conductance. No agonist property was detected with VX. On glutamatergic synapses, the ChE inhibitors generated spontaneous firing of end-plate potentials (EPPs) and action potentials (APs). This effect was blocked in the presence of low external Ca2+ concentration or tetrodotoxin. It seems that the spontaneous EPP and AP firing resulted from an increased transmitter release induced by an increase in Na+ influx at the presynpatic nerve terminal. Physostigmine and some irreversible ChE inhibitors (VX and DFP) also blocked the postjunctional glutamate receptors. Similar to the nicotinic AChR, this effect was mostly related to a blockade of the open channels. In conclusion, the present studies showed significant protection of rats by physostigmine in combination with some ganglionic antagonists against lethality by organophosphate agents.(ABSTRACT TRUNCATED AT 400 WORDS)
ESTHER : Albuquerque_1985_Fundam.Appl.Toxicol_5_S182
PubMedSearch : Albuquerque_1985_Fundam.Appl.Toxicol_5_S182
PubMedID: 2868960

Title : The reversible cholinesterase inhibitor physostigmine has channel-blocking and agonist effects on the acetylcholine receptor-ion channel complex - Shaw_1985_Mol.Pharmacol_28_527
Author(s) : Shaw KP , Aracava Y , Akaike A , Daly JW , Rickett DL , Albuquerque EX
Ref : Molecular Pharmacology , 28 :527 , 1985
Abstract : The actions of the carbamate cholinesterase inhibitors, physostigmine (Phy) and physostigmine methiodide (MetPhy), were studied on the acetylcholine receptor-ion channel complex (AChR) of skeletal muscles. Low concentrations of these agents produced cholinesterase inhibition which resulted in potentiation of nerve-elicited muscle twitches and an increased peak amplitude and prolongation of the decay time constant (tau EPC) of endplate currents (EPCs) elicited in frog (Rana pipiens) sartorius muscles. However, increasing concentrations of Phy depressed the peak amplitude and shortened the decay phase of the EPC with an apparent loss in the voltage dependence of tau EPC. At higher concentrations and depolarized potentials, EPC decays were double exponential. The effects of both Phy and MetPhy on the postsynaptic AChR complex were also evident in preparations pretreated with diisopropylfluorophosphate. Under these conditions, a linear relationship between the reciprocal of tau EPC and the concentration of these agents was observed. Single channel studies revealed that Phy (20-600 microM) shortened channel lifetime and decreased channel conductance at very high concentrations. In addition, Phy (0.5 microM) induced the appearance of channel openings with conductance similar to that of acetylcholine. High concentrations (greater than 50 microM) of this agent activated channel openings with decreased conductance. Similar results were obtained with MetPhy. Thus, the reversible cholinesterase inhibitors Phy and MetPhy altered the properties of the AChR by interacting as agonists capable of inducing desensitization and blockade.
ESTHER : Shaw_1985_Mol.Pharmacol_28_527
PubMedSearch : Shaw_1985_Mol.Pharmacol_28_527
PubMedID: 2417099

Title : Interactions of bupivacaine with ionic channels of the nicotinic receptor. Electrophysiological and biochemical studies - Ikeda_1984_Mol.Pharmacol_26_293
Author(s) : Ikeda SR , Aronstam RS , Daly JW , Aracava Y , Albuquerque EX
Ref : Molecular Pharmacology , 26 :293 , 1984
Abstract : The actions of the tertiary local anesthetic bupivacaine were studied on the nicotinic receptor-ionic channel complex (AChR) using electrophysiological and biochemical methods. Voltage clamp studies of the frog sartorius and cutaneous pectoris neuromuscular junction revealed a concentration-dependent depression of the decay time constant of the end-plate (tau EPC) and spontaneous miniature end-plate (tau MEPC) currents. The relationship of the reciprocal of either tau EPC or tau MEPC and bupivacaine concentration up to 100 microM was linear. Voltage dependence of EPC over the range +60 to -150 mV was reduced, whereas both EPC and MEPC decays were adequately described by a single exponential function at all concentrations tested. Peak MEPC and EPC amplitudes were also depressed in a concentration-dependent manner such that 100 microM bupivacaine reduced peak amplitude by about 50%. The current-voltage relationship remained linear under all conditions tested. Nerve-evoked responses were difficult to study at concentrations greater than 100 microM because of apparent blockade of nerve conduction. Extracellular recording of the MEPC afforded results similar to those obtained with EPCs. The tau MEPC could be reduced to less than 300 mu sec at a bupivacaine concentration of 400 microM. Fluctuation analysis showed that bupivacaine at concentrations of 10 and 25 microM did not change channel conductance but decreased single-channel lifetime to 76% and 39% of control values, respectively. Biochemical studies were performed on Torpedo californica membrane fragments using [3H]phencyclidine ([3H]PCP) and [3H]perhydrohistrionicotoxin ([3H]H12-HTX) as channel probes. Bupivacaine inhibited the binding of [3H]PCP and [3H]H12-HTX with inhibition constants (Ki) of 32 and 25 microM, respectively. The corresponding inhibition constants for bupivacaine methiodide were 1.8 and 3.2 microM. The preincubation of the membranes with carbamylcholine increased the affinity of bupivacaine for the ionic channel sites 5- to 8-fold and the affinity of bupivacaine methiodide 3- to 4-fold. Bupivacaine, however, had no affinity for the agonist recognition site as determined by [3H]ACh and [125I]alpha-bungarotoxin bindings. The electrophysiological and biochemical studies indicate that bupivacaine reacts primarily with the ionic channel of the nicotinic AChR. The results are consistent with a sequential model in which the drug interacts with the sites at the ionic channel of AChR in its open conformation, producing species with little or no conductance. From the present studies there is no evidence for an interaction of bupivacaine with the agonist binding site or closed states of AChR.
ESTHER : Ikeda_1984_Mol.Pharmacol_26_293
PubMedSearch : Ikeda_1984_Mol.Pharmacol_26_293
PubMedID: 6090884

Title : Meproadifen enhances activation and desensitization of the acetylcholine receptor-ionic channel complex (AChR): single channel studies - Aracava_1984_FEBS.Lett_174_267
Author(s) : Aracava Y , Albuquerque EX
Ref : FEBS Letters , 174 :267 , 1984
Abstract : The effects of the quaternary agent meproadifen on ACh-activated channel currents were studied on myoballs cultured from hind limb muscles of neonatal rats. Meproadifen (0.02-0.1 microM) combined with ACh (0.1-0.3 microM) in the patch pipette caused an increase, followed by a decrease, in the frequency of channel openings. At concentrations greater than 0.2 microM the initial phase was not detected and a rapid and marked reduction in the opening frequency was observed. Meproadifen (up to 2.5 microM) produced no change in the duration or conductance of the open state of ACh-activated channels. In addition, this agent induced the appearance of events with a marked increase in the 'noise' during the opening phase. The lack of effect under inside-out patch conditions suggested that meproadifen binds to a site located at the external portion of the nicotinic macromolecule and has no access to it through the cell membrane. This study indicated that non-competitive antagonists such as meproadifen can facilitate receptor activation and desensitization.
ESTHER : Aracava_1984_FEBS.Lett_174_267
PubMedSearch : Aracava_1984_FEBS.Lett_174_267
PubMedID: 6088290

Title : Interactions of bupivacaine with ionic channels of the nicotinic receptor. Analysis of single-channel currents - Aracava_1984_Mol.Pharmacol_26_304
Author(s) : Aracava Y , Ikeda SR , Daly JW , Brookes N , Albuquerque EX
Ref : Molecular Pharmacology , 26 :304 , 1984
Abstract : Bupivacaine and its quaternary derivative, bupivacaine methiodide, were studied on acetylcholine (ACh)-activated single-channel currents recorded in myoballs from neonatal rat muscles using the patch-clamp technique. Under control conditions, the ACh-induced channels had three conductance states, 10, 20, and 33 pS, at a temperature of 10 degrees. The intermediate conductance state (20 pS) was the most prevalent. Moreover, an excessive number of very short events was observed which contributed to a deviation of the channel open-time distribution from a single-exponential function. At 20 degrees, the amplitude of these currents was increased (Q10 = 1.4), and the mean channel open time was decreased (Q10 = 3). Bupivacaine and its quaternary derivative (5-50 microM), when inside the patch micropipette with ACh, caused shortening of the channel open time, but the single-channel conductance remained unchanged at all concentrations studied. In the presence of bupivacaine, there was a loss of voltage dependence of the mean channel open time seen under control conditions; i.e., the shortening of the channel open time was more pronounced at more negative potentials. The plot of the reciprocal of mean channel open time versus bupivacaine concentration was linear. Similar effects were observed when bupivacaine was added to the bathing medium in both cell-attached and inside-out patch conditions, but in this case the onset of the drug action occurred at a later time and its potency was lower. Application of bupivacaine methiodide via the bathing medium after the establishment of the gigaohm seals, however, had no effect on the kinetics of ACh-activated single channels under both patch conditions (cell-attached and inside-out). The patch-clamp results indicated that the charged form of bupivacaine blocks the open state of ACh-activated ionic channels interacting with sites at the extracellular segment of the ACh receptor-ionic channel complex and creating a species with little or no conductance. A sequential model can be used to explain the interactions of these noncompetitive antagonists of the ACh receptor-ionic channel complex with the open channel. This interpretation of the action of bupivacaine and its quaternary analogue as open channel blockers also was reached based on an analysis of macroscopic events in nicotinic synapses of frog muscle.
ESTHER : Aracava_1984_Mol.Pharmacol_26_304
PubMedSearch : Aracava_1984_Mol.Pharmacol_26_304
PubMedID: 6090885