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James P Dilger - One of the best experts on this subject based on the ideXlab platform.
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site selectivity of competitive antagonists for the mouse adult muscle nicotinic acetylcholine receptor
Molecular Pharmacology, 2009Co-Authors: James P DilgerAbstract:The nicotinic acetylcholine receptor (nAChR) is a prototypical member of a superfamily of pentameric ligand-gated ion channels, which includes muscle and neuronal nAChRs, 5-HT3 receptors, GABA receptors, and glycine receptors (Unwin, 2005; Sine and Engel, 2006). The adult muscle type has the subunit stoichiometry (α1)2β1eδ. This type of nAChR mediates rapid synaptic transmission at the neuromuscular junction. The binding of two molecules of ACh to sites in the extracellular domain promotes transmembrane cation channel opening. The two binding sites for ACh, situated between the αe- and αδ-subunit interfaces of adult mouse nAChR, have similar affinities for ACh (Akk and Auerbach, 1996). Antagonists such as (+)-tubocurarine and pancuronium compete with ACh for binding to these sites; the binding of a single antagonist is sufficient to prevent channel opening (see Discussion). Several competitive antagonists are known to distinguish between the two binding sites. The αe-interface has been identified as the high-affinity binding site for (+)-tubocurarine and Metocurine based on evidence from chimeras, mutations (Bren and Sine, 1997) and computational docking (Wang et al., 2003) in mouse or human nAChR. The affinity of the αδ-interfacial site for these antagonists is 20-170-fold lower (Fletcher and Steinbach, 1996; Bren and Sine, 1997). There is less information about the site selectivity of other antagonists. Binding experiments have demonstrated that pancuronium and atracurium discriminate between the two sites in adult mouse nAChR by factors of 26 and 2.5, respectively (Fletcher and Steinbach, 1996). (Atracurium is a mixture of 10 stereoisomers; cisatracurium is the isomer that is currently used clinically). Vecuronium and cisatracurium are sensitive to mutations in the e-subunit, whereas vecuronium, cisatracurium, and pancuronium are all sensitive to mutations in the δ-subunit (Dilger et al., 2007). These results suggest that antagonist combinations such as (+)-tubocurarine + vecuronium should not be completely competitive with each other and have a synergistic effect on inhibition. However, it has been reported that the inhibition produced by this combination in adult mouse nAChR is additive (Paul et al., 2002). Here, we examine the site specificity of competitive antagonist binding to adult mouse nAChR measuring current inhibition in the presence of pairs of antagonists. In contrast to the previous report (Paul et al., 2002), our measurements were made under conditions of high receptor inhibition by one or both antagonists. This provides a more sensitive test for synergistic effects (Waud and Waud, 1985; Nigrovic and Amann, 2004). We have used this approach in a study of adult human nAChR (Liu and Dilger, 2008) and found evidence for synergy; in some cases, synergy at the receptor level correlated with the clinical phenomenon of muscle relaxant synergy in which pairs of antagonist are more effective at inducing paralysis than expected from their individual potencies (Lebowitz et al., 1980; Waud and Waud, 1984).
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roles of amino acids and subunits in determining the inhibition of nicotinic acetylcholine receptors by competitive antagonists
Anesthesiology, 2007Co-Authors: James P Dilger, Ana Maria Vidal, Claire Mettewie, Takahiro Suzuki, Anh Pham, Deeptankar DemazumderAbstract:THE muscle-type nicotinic acetylcholine receptor channel (nAChR) is the prototypical member of the Cys-loop ligand-gated ion channel superfamily.1 These proteins are composed of α1, β1, δ, and γ or e subunits arranged as transmembrane pentamers: (α1)(β1)δ(α1)γ in the embryonic nAChR subtype and (α1)(β1)δ(α1)e in the adult nAChR subtype (clockwise, as viewed from the synapse). There are several reasons why so much is known about this protein. Biochemical and structural studies take advantage of the abundance of nAChR in tissue from Torpedo electric organ.2 Electrophysiologic measurements are aided by its stability in patch clamp experiments. And the discovery3 and structure determination4 of a water-soluble analog to the extracellular domain of the receptor, acetylcholine binding protein (AChBP), has led to inferences about structure–function relations of the nAChR.5 Mutagenesis, functional, labeling, and structural studies have provided detailed information about the ligand binding site in the nAChR (fig. 1A). Binding sites for agonists and competitive antagonists are located at the α–δ and α–e (or α–γ) subunit interfaces.** Studies have identified seven noncontinuous “loops” of amino acids that participate in agonist and competitive antagonist binding to the nAChR: loops A–C are on the α subunit (primary component of the binding site), and loops D–G are on the non-α subunit (complementary component of the binding site). The three dimensional arrangement of these loops became apparent by making analogies to the structure of AChBP.4,6 AChBP is composed of 10 β strands arranged in an immunoglobulin-like topology. Several of the loops are located between the β strands, and the loops from the primary and complementary components come together at the interface between subunits. The binding site is centered around a conserved tryptophan in loop B that provides stability for the quaternary nitrogen moiety of agonists and antagonists via a strong π–cation interaction.7 Four additional aromatic amino acids are distributed around the primary and complementary components. The C loop may act as a cap that closes upon binding of agonist and greatly decreases the agonist dissociation rate.8 The amino acids on the complementary component are not conserved between the γ, δ, and e subunits; this leads to subunit specificity for ligand binding. Fig. 1 (A) A model of the ligand binding site at the α–e interface of the human muscle nicotinic acetylcholine receptor channel. The α subunit is on the left, and the e subunit is on the right. The backbone and molecular ... Agonists such as acetylcholine have a higher affinity for the α–δ interface compared with the α–e interface; the extent of the difference depends on both species (Torpedo receptors exhibit a 100-fold preference for acetylcholine binding to the α–δ interface9) and subtype (fetal mouse receptors exhibit a 32-fold preference,10 and adult mouse receptors exhibit very little preference11). Both sites must be occupied for efficient opening of the channel.11 Some competitive antagonists are known to bind differently to the two sites. Metocurine, for example, has a 70- to 170-fold higher affinity for the α–e interface compared with the α–δ interface.12,13 Occupation of just one of the sites by a competitive antagonist is sufficient to prevent channel opening. Most of the information about the residues that participate in antagonist binding comes from studies of either (+)-tubocurarine or Metocurine.14 Even for these similar ligands (Metocurine has three additional methyl groups; fig. 1), there are differences in sensitivities to mutations. Using computational chemistry to dock ligands to a homology model of the nAChR, Wang et al. 15 showed that (+)-tubocurarine and Metocurine bind with different orientations within the α–e interface and have different contacts with the amino acid residues there. Although the clinical action of competitive nAChR antagonists, muscle paralysis, is a straightforward consequence of their molecular action, there are outstanding questions about the effects of nondepolarizing muscle relaxants. Specific issues include the mechanisms of tetanic fade16 and of muscle relaxant synergy.17 Although these phenomena are often ascribed to actions on presynaptic nAChRs, postsynaptic explanations may still be viable. As a first step to address these questions, we use a functional determination of current inhibition to examine competitive antagonism by (+)-tubocurarine, Metocurine, pancuronium, vecuronium, and cisatracurium (fig. 1B) on mouse adult wild-type and five mutant nAChRs. We chose these mutations because they have been shown to affect Metocurine and/or (+)-tubocurarine binding.13,15,18,19 The αY198F mutation is in loop C, and the eD59 mutations and eD173 mutations are in loops E and G, respectively. We used the δD180K mutation to test the strength of binding to the α–δ interface. We postulated that the antagonists would show different sensitivities to mutations but that all of them would have a higher affinity for the α–e interface compared with the α–δ interface.
Gerald A. Gronert - One of the best experts on this subject based on the ideXlab platform.
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Allometry of pharmacokinetics and pharmacodynamics of the muscle relaxant Metocurine in mammals
2016Co-Authors: Gerald A. Gronert, Steven L. Shafer, Dennis L Fung, James H Jones, Sue V. Hildebrand, A. Disbrow, Gerald AAbstract:namics of the muscle relaxant Metocurine in mammals. Am. J. Physiol. 268 (Regulatory Integrative Comp. PhysioZ. 37): R85-R91, 1995.We investigated the effects of body size on the pharmacokinetics and pharmacodynamics of the renally cleared muscle relaxant Metocurine. We hypothesized that pharmaco-kinetics of the drug would change allometrically in proportion to physiological time [m&2:.25, where Mb is body mass] and that pharmacodynamics would be independent of size because of the highly conserved structure of the acetylcholine receptor. Metocurine effects during general anesthesia were examined in 17 rats, 8 cats, 6 dogs, 5 pigs, 7 sheep, and 12 horses. Allometric analysis demonstrated size dependence for pharma-cokinetics, which were affected by physiological time (ME.25). Pharmacodynamics were size independent, except for the value for effect compartment concentration associated wit
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deep sedation and mechanical ventilation without paralysis for 3 weeks in normal beagles exaggerated resistance to Metocurine in gastrocnemius muscle
Anesthesiology, 1999Co-Authors: Gerald A. Gronert, Dennis L Fung, Steve C Haskins, Eugene SteffeyAbstract:Background: Patients in the intensive care unit may have muscle weakness in the recovery phase, and disuse atrophy may play a role in this weakness. To assess this problem, the authors measured changes in the potency of the nondepolarizing neuromuscular blocking agent Metocurine in a a canine model that involved 3 weeks of intensive care, nonparalyzing anesthesia with pentobarbital, and positive-pressure ventilation. Methods: Six dogs were anesthetized with pentobarbital to a sufficient depth that spontaneous and reflex muscle movements were absent. Their tracheas were intubated, their lungs were mechanically ventilated, and they received round-the-clock intensive medical and nursing care for 3 weeks. Transduced gastrocnemius muscle responses to Metocurine were determined weekly. A 4- to 15-min infusion of 148-4,300 μg/min (longer durations and greater concentrations on progressive weeks) yielded more than 80% paralysis. Serial Metocurine plasma concentrations during the onset of the block and recovery provided data to determine pharmacokinetics using NON-MEM. Metocurine plasma concentrations and the degree of paralysis were used to model the effect compartment equilibration constant, and the Hill equation was used to yield the slope factor and potency within the effect compartment. Results: The Metocurine effect compartment concentration associated with a 50% diminution of twitch height after 3 weeks was 1,716 ± 1,208 ng/ml (mean ± SD), which was significantly different from 257 ± 34 ng/ml, the value on day 0. There were no pharmacokinetic differences. Conclusion: The absence of muscle tone and reflex responsiveness for 3 weeks was associated with exaggerated resistance to the neuromuscular blocker Metocurine.
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Metocurine pharmacokinetics and pharmacodynamics in goats
Journal of Veterinary Pharmacology and Therapeutics, 1995Co-Authors: J F Antognini, R Wood, Gerald A. GronertAbstract:Non-depolarizing muscle relaxants can facilitate surgery and anaesthesia in numerous species. and volatile inhalational anaesthetics such as isoflurane potentiate their action. We studied the effect of isoflurane on the pharmacodynamics and pharmacokinetics of Metocurine in six goats. Each was studied twice: once during barbiturate-opiate anaesthesia and once during isoflurane anaesthesia. The evoked response to sciatic nerve stimulation was measured using a force transducer attached to the hoof. Metocurine was infused until approximately 80-90% blockade. Plasma Metocurine concentration was determined by high-performance liquid chromatography. Isoflurane increased the potency of Metocurine significantly; IC50 (the concentration in the effect compartment at 50% paralysis) was 70 +/- 15 ng/mL during isoflurane anaesthesia and 129 +/- 42 ng/mL during barbiturate-opiate anaesthesia (P < 0.03). Volume of distribution (63 +/- 18 mL/kg), clearance (1.6 +/- 0.4 mL/min.kg) and elimination half-life (99 +/- 9 min) during barbiturate-opiate anaesthesia were not significantly different during isoflurane anaesthesia: 64 +/- 25 mL/kg, 1.5 +/- 0.7 mL/kg.min, 116 +/- 16 min respectively. We conclude that, relative to barbiturate-opiate anaesthesia, isoflurane potentiates Metocurine in goats.
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the changing pharmacodynamics of Metocurine identify the onset and offset of canine gastrocnemius disuse atrophy
Anesthesiology, 1995Co-Authors: Dennis L Fung, Gerald A. Gronert, D A White, E DisbrowAbstract:BackgroundImmobilization of skeletal muscle results in disuse atrophy and resistance to nondepolarizing muscle relaxants. We studied the pharmacodynamics of Metocurine (MTC) to identify the development and recovery of disuse-related resistance to MTC.
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allometry of pharmacokinetics and pharmacodynamics of the muscle relaxant Metocurine in mammals
American Journal of Physiology-regulatory Integrative and Comparative Physiology, 1995Co-Authors: Gerald A. Gronert, Steven L. Shafer, Dennis L Fung, Susan Hildebrand, James H Jones, E A DisbrowAbstract:We investigated the effects of body size on the pharmacokinetics and pharmacodynamics of the renally cleared muscle relaxant Metocurine. We hypothesized that pharmacokinetics of the drug would change allometrically in proportion to physiological time [infinity Mb0.25, where Mb is body mass] and that pharmacodynamics would be independent of size because of the highly conserved structure of the acetylcholine receptor. Metocurine effects during general anesthesia were examined in 17 rats, 8 cats, 6 dogs, 5 pigs, 7 sheep, and 12 horses. Allometric analysis demonstrated size dependence for pharmacokinetics, which were affected by physiological time (Mb0.25). Pharmacodynamics were size independent, except for the value for effect compartment concentration associated with 50% twitch paralysis (IC50). Data from individual species had a bimodal distribution that was significant: pigs and sheep were more sensitive than other large species, and their IC50 appeared size independent. IC50 was size dependent in more active species (horse, dog, cat, rat). Although the mechanism is unknown, we speculate that this trend might relate to receptor density within the end plate. Thus pharmacokinetics changed in proportion to physiological time, and pharmacodynamics were in part size independent.
Dennis L Fung - One of the best experts on this subject based on the ideXlab platform.
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Allometry of pharmacokinetics and pharmacodynamics of the muscle relaxant Metocurine in mammals
2016Co-Authors: Gerald A. Gronert, Steven L. Shafer, Dennis L Fung, James H Jones, Sue V. Hildebrand, A. Disbrow, Gerald AAbstract:namics of the muscle relaxant Metocurine in mammals. Am. J. Physiol. 268 (Regulatory Integrative Comp. PhysioZ. 37): R85-R91, 1995.We investigated the effects of body size on the pharmacokinetics and pharmacodynamics of the renally cleared muscle relaxant Metocurine. We hypothesized that pharmaco-kinetics of the drug would change allometrically in proportion to physiological time [m&2:.25, where Mb is body mass] and that pharmacodynamics would be independent of size because of the highly conserved structure of the acetylcholine receptor. Metocurine effects during general anesthesia were examined in 17 rats, 8 cats, 6 dogs, 5 pigs, 7 sheep, and 12 horses. Allometric analysis demonstrated size dependence for pharma-cokinetics, which were affected by physiological time (ME.25). Pharmacodynamics were size independent, except for the value for effect compartment concentration associated wit
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deep sedation and mechanical ventilation without paralysis for 3 weeks in normal beagles exaggerated resistance to Metocurine in gastrocnemius muscle
Anesthesiology, 1999Co-Authors: Gerald A. Gronert, Dennis L Fung, Steve C Haskins, Eugene SteffeyAbstract:Background: Patients in the intensive care unit may have muscle weakness in the recovery phase, and disuse atrophy may play a role in this weakness. To assess this problem, the authors measured changes in the potency of the nondepolarizing neuromuscular blocking agent Metocurine in a a canine model that involved 3 weeks of intensive care, nonparalyzing anesthesia with pentobarbital, and positive-pressure ventilation. Methods: Six dogs were anesthetized with pentobarbital to a sufficient depth that spontaneous and reflex muscle movements were absent. Their tracheas were intubated, their lungs were mechanically ventilated, and they received round-the-clock intensive medical and nursing care for 3 weeks. Transduced gastrocnemius muscle responses to Metocurine were determined weekly. A 4- to 15-min infusion of 148-4,300 μg/min (longer durations and greater concentrations on progressive weeks) yielded more than 80% paralysis. Serial Metocurine plasma concentrations during the onset of the block and recovery provided data to determine pharmacokinetics using NON-MEM. Metocurine plasma concentrations and the degree of paralysis were used to model the effect compartment equilibration constant, and the Hill equation was used to yield the slope factor and potency within the effect compartment. Results: The Metocurine effect compartment concentration associated with a 50% diminution of twitch height after 3 weeks was 1,716 ± 1,208 ng/ml (mean ± SD), which was significantly different from 257 ± 34 ng/ml, the value on day 0. There were no pharmacokinetic differences. Conclusion: The absence of muscle tone and reflex responsiveness for 3 weeks was associated with exaggerated resistance to the neuromuscular blocker Metocurine.
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the changing pharmacodynamics of Metocurine identify the onset and offset of canine gastrocnemius disuse atrophy
Anesthesiology, 1995Co-Authors: Dennis L Fung, Gerald A. Gronert, D A White, E DisbrowAbstract:BackgroundImmobilization of skeletal muscle results in disuse atrophy and resistance to nondepolarizing muscle relaxants. We studied the pharmacodynamics of Metocurine (MTC) to identify the development and recovery of disuse-related resistance to MTC.
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allometry of pharmacokinetics and pharmacodynamics of the muscle relaxant Metocurine in mammals
American Journal of Physiology-regulatory Integrative and Comparative Physiology, 1995Co-Authors: Gerald A. Gronert, Steven L. Shafer, Dennis L Fung, Susan Hildebrand, James H Jones, E A DisbrowAbstract:We investigated the effects of body size on the pharmacokinetics and pharmacodynamics of the renally cleared muscle relaxant Metocurine. We hypothesized that pharmacokinetics of the drug would change allometrically in proportion to physiological time [infinity Mb0.25, where Mb is body mass] and that pharmacodynamics would be independent of size because of the highly conserved structure of the acetylcholine receptor. Metocurine effects during general anesthesia were examined in 17 rats, 8 cats, 6 dogs, 5 pigs, 7 sheep, and 12 horses. Allometric analysis demonstrated size dependence for pharmacokinetics, which were affected by physiological time (Mb0.25). Pharmacodynamics were size independent, except for the value for effect compartment concentration associated with 50% twitch paralysis (IC50). Data from individual species had a bimodal distribution that was significant: pigs and sheep were more sensitive than other large species, and their IC50 appeared size independent. IC50 was size dependent in more active species (horse, dog, cat, rat). Although the mechanism is unknown, we speculate that this trend might relate to receptor density within the end plate. Thus pharmacokinetics changed in proportion to physiological time, and pharmacodynamics were in part size independent.
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determination of sensitivity to Metocurine in exercised horses
American Journal of Veterinary Research, 1992Co-Authors: D A White, Dennis L Fung, Susan Hildebrand, James H Jones, G A GronertAbstract:On the basis of results in dogs, conditioning exercise may increase sensitivity to nondepolarizing muscle relaxants. Five Thoroughbreds were exercised/conditioned 3 times weekly on a treadmill for 8 months. Increasing maximal rate of O2 consumption verified that the horses were responding to exercise conditioning. Six nonexercised Thoroughbreds served as the control group. Studies were done with horses under general anesthesia by use of halothane during partial paralysis by a brief constant-rate infusion with the muscle relaxant, Metocurine iodide. Quantification of degree of paralysis of the hoof twitch (eg, digital extensor) occurred with simultaneous quantification of blood values of Metocurine. Pharmacokinetic and pharmacodynamic analyses of the data were done by a nonlinear regression program, using the Hill equation. There were no differences in findings between exercised and nonexercised horses. The mean blood concentration for the 50% paralyzing dose of Metocurine was 0.44 +/- 0.11 (SD) microgram/ml in exercised horses, and 0.58 +/- 0.22 microgram/ml in nonexercised horses. Despite evidence for a response to conditioning, a significant change in the sensitivity of the neuromuscular junction to Metocurine was not found.
J. M. Hunter - One of the best experts on this subject based on the ideXlab platform.
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PHARMACOKINETICS OF ATRACURIUM AND LAUDANOSINE IN THE ELDERLY
2016Co-Authors: A.p. Kent, C. J. R. Parker, J. M. HunterAbstract:Atracurium breaks down spontaneously in plasma by Hofmann elimination and ester hydrolysis, and is thought not to be dependent on hepatic or renal function for its elimination [1]. Such properties suggest that this drug would be suitable for use in the elderly patient, in whom liver and kidney function are often impaired [2]. Advancing age has been shown to affect the pharmacokinetics of other neuromuscular blocking drugs, including vecuronium, pancuronium [3-5], Metocurine and tubocurarine [6]. In previous pharmacokinetic studies of atracurium reported in the elderly, deBros and colleagues [7] suggested results in this age group similar to those reported previously in young adults [8, 9]. In contrast, however, Kitts and colleagues [10] found a difference in the elimination half-life, organ clearance and volume of distribution of atracurium at steady state between a young and an elderly group of patients. Laudanosine, a major metabolite of atracurium, was not measured in these studies. All other pharmacokinetic studies of atracurium have been restricted to young populations [8,9]. In a pharmacodynamic study, d'Hollander and his colleagues [11] found no significant difference in the infusion rate of atracurium required to maintain 10 % control twitch height in the elderly compared with a young, or middle-aged popu-lation. Laudanosine in pharmacological doses has been shown in animals to produce excitation of the central nervous system [12]. Its excretion is mediated in part by the liver [13] and in part by the kidney [14] and, in contrast with those o
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ADVERSE EFFECTS OF NEUROMUSCULAR BLOCKING DRUGS
2016Co-Authors: J. M. HunterAbstract:Neuromuscular blocking drugs are not as notori-ous for producing adverse reactions as the i.v. induction agents; nevertheless, to a varying degree they all produce unfavourable or harmful effects. The newer non-depolarizing neuromuscular blockers, atracurium and vecuronium, have been developed in an attempt to overcome the disad-vantages of the earlier drugs, but although much more specific agents, they are not completely free from side-effects. The adverse effects of the neuromuscular blocking drugs available in Great Britain today will be discussed in this paper. Those which are not available in this country (such as pipecuronium and Metocurine), and obsolete agents (such as decamethonium) will not be discussed. Although what follows is concerned mainly with the non-depolarizing agents, suxa-methonium is included where appropriate, whilst what might be regarded as the unique features of the drug are discussed later in the article. DIRECT CARDIOVASCULAR EFFECTS With the possible exception of vecuronium, all the neuromuscular blocking drugs have some effect on the cardiovascular system. Such effects ar
Rao V L Papineni - One of the best experts on this subject based on the ideXlab platform.
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interaction of d tubocurarine analogs with the torpedo nicotinic acetylcholine receptor methylation and stereoisomerization affect site selective competitive binding and binding to the noncompetitive site
Journal of Biological Chemistry, 1995Co-Authors: Steen E Pedersen, Rao V L PapineniAbstract:Analogs of d-tubocurarine were used to determine the individual effects of methylation, stereoisomerization, and halogenation of d-tubocurarine on the affinity for each of the two acetylcholine (ACh) binding sites of the Torpedo nicotinic acetylcholine receptor (AChR) and for the noncompetitive antagonist site. Eight analogs were synthesized, including three new compounds: 7'-O-methyl-chondocurarine, 12'-O-methyl-chondocurarine, and 13'-bromo-d-tubocurarine. The two ACh sites differ in their affinities for d-tubocurarine by 400-fold, as shown by inhibition of [3H]ACh binding, whereas the affinity ratio for Metocurine, the trimethylated derivative of d-tubocurarine, is reduced to 30 due to a decreased affinity for the high affinity site. Binding analysis of five d-tubocurarine analogs demonstrates that methylation of the phenols alone is responsible for the observed changes in affinity. Substitution with bromine or iodine at the 13'-position affected affinity at both sites with a net increase in site selectivity. Stereoisomers of d-tubocurare had decreased affinity for only the high affinity ACh site. Thus, the ring systems, including the 12'- and 13'-positions and the 1-position stereocenter, appear to be important in discriminating between the two ACh binding sites. Desensitization of the AChR was measured by increased affinity for [3H]phencyclidine. Binding to only the single, high affinity acetylcholine binding site, comprised by the alpha gamma-subunits, was required for partial desensitization of the AChR by d-tubocurarine and its analogs. Stronger desensitization, to the same extent observed in the presence of the agonist carbamylcholine, occurred upon binding by iodonated or brominated d-tubocurarine. Interaction of the analogs at the noncompetitive antagonist site of the AChR was also measured by [3H]phencyclidine binding. The bis-tertiary ammonium analogs of either the d- or l-stereoisomers bound to the noncompetitive antagonist binding site of the AChR with 100-fold higher affinity than the corresponding quaternary ammonium analogs.