The Experts below are selected from a list of 99 Experts worldwide ranked by ideXlab platform
Andrew J Mclachlan - One of the best experts on this subject based on the ideXlab platform.
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muscle distribution of the neuromuscular blocker Gallamine using microdialysis
Journal of Pharmaceutical Sciences, 2002Co-Authors: Lucy Sasongko, Iqbal Ramzan, Kenneth M Williams, Andrew J MclachlanAbstract:Abstract Measurement of drug concentrations in target tissue has the potential to provide insight into the pharmacokinetics and pharmacodynamics of a drug. In this study, the distribution of the neuromuscular blocker, Gallamine, into muscle tissue was investigated in urethane‐anesthetized rats after an intravenous bolus dose (6 mg/kg). Microdialysis sampling was used to continuously determine Gallamine concentrations in muscle interstitial fluid (MIF). In vivo microdialysis recovery of Gallamine was determined as the relative loss of Gallamine from the perfusate into muscle tissue after perfusion with Gallamine (2 μg/mL). Recovery was determined in each rat before the pharmacokinetic studies. Terminal muscle sampling followed by homogenization was also performed to examine Gallamine distribution within muscle tissue. All samples were assayed for Gallamine using a validated high‐performance liquid chromatography assay. Gallamine was rapidly distributed into MIF with a MIF–plasma partition coefficient of 0.9 ± 0.1 ( n = 6). By contrast, the estimated Gallamine concentration in muscle tissue homogenate was only 23 ± 5% ( n = 5) of the concentration in MIF as estimated by microdialysis sampling at the terminal sampling time. These findings suggest that Gallamine is not distributed uniformly within muscle but selectively distributes into MIF. Simulations using a hybrid physiologically based pharmacokinetic model which describes uptake of drug only into the interstitial space showed good agreement between predicted and observed concentration data obtained from microdialysis sampling, supporting the findings that Gallamine selectively distributes into MIF. These studies demonstrate microdialysis combined with conventional terminal tissue sampling provides valuable information on intra‐tissue drug distribution. © 2002 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 91: 769–775, 2002
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assessment of in vitro and in vivo recovery of Gallamine using microdialysis
Journal of Pharmacological and Toxicological Methods, 2000Co-Authors: Lucy Sasongko, Iqbal Ramzan, Kenneth M Williams, Andrew J MclachlanAbstract:Abstract The application of microdialysis technique for the investigation of pharmacokinetics and pharmacodynamics of drugs requires careful assessment of probe performance to ensure validity of the data obtained using this technique. The aim of this study was to establish and validate the microdialysis technique for investigation of the pharmacokinetics and pharmacodynamics of the neuromuscular blocker, Gallamine. In vitro recovery of Gallamine from the microdialysis probe when different perfusion flow rates were employed was evaluated leading to selection of a flow rate of 2 μl/min with 15-min sampling intervals for the subsequent studies. In vitro recovery of Gallamine from the microdialysis probe was independent of concentration, stable over an 8-h period and reproducible. Comparable in vitro recoveries were obtained by different established approaches including recovery estimation by gain, loss and the zero-net flux (ZNF) method. Recovery by loss was used to study the in vivo recovery of Gallamine from rat muscle tissue. The in vivo recovery was stable over a 5.5-h sampling period. In vitro performance of the probe subsequent to the in vivo study remained stable supporting reusage of the probe. These data highlight the importance of a systematic examination of microdialysis probe validation.
Klaus Mohr - One of the best experts on this subject based on the ideXlab platform.
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first Gallamine tacrine hybrid design and characterization at cholinesterases and the m2 muscarinic receptor
Journal of Medicinal Chemistry, 2007Co-Authors: Paul W Elsinghorst, Julia S Cieslik, Klaus Mohr, Christian Trankle, Michael GutschowAbstract:Gallamine and tacrine are allosteric antagonists at muscarinic M2 acetylcholine receptors and inhibitors of acetylcholinesterase. At both acetylcholine-binding proteins, Gallamine and tacrine are known to occupy two different binding sites: in M2 receptors within the allosteric binding area and in acetylcholinesterase at its catalytic and its peripheral site. To find new ligands of both targets, we designed a Gallamine−tacrine dimer and several derived hybrid compounds to address the two binding sites. Their M2 receptor allosteric and acetylcholinesterase inhibitory potential was determined. The hybrid compounds revealed an allosteric potency in the low nanomolar range exceeding the allosteric potency of Gallamine and tacrine by factors of 100 and 4800, respectively. Cholinesterase inhibition was augmented by hybrid formation, and all compounds exhibited IC50 values in the lower nanomolar range. Thus, Gallamine−tacrine hybrid formation is a valuable approach toward high affinity ligands concurrently targ...
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critical amino acid residues of the common allosteric site on the m2 muscarinic acetylcholine receptor more similarities than differences between the structurally divergent agents Gallamine and bis ammonio alkane type hexamethylene bis dimethyl 3 pht
Molecular Pharmacology, 2005Co-Authors: Xi Ping Huang, Klaus Mohr, Stefanie Prilla, John EllisAbstract:The structurally divergent agents Gallamine and hexamethylene- bis -[dimethyl-(3-phthalimidopropyl)ammonium]dibromide (W84) are known to interact competitively at a common allosteric site on muscarinic receptors. Previous studies reported that the M2 selectivity of Gallamine depended largely on the EDGE (172-175) sequence in the second outer loop (o2) and on 419Asn near the junction of o3 and the seventh transmembrane domain (TM7), whereas the selectivity of W84 depended on nearby residues 177Tyr and 423Thr. However, it has so far proven difficult to confer the high sensitivity for allosteric modulation of the M2 subtype onto the weakly sensitive M5 subtype by substituting these key residues. We now have found that M2 423Thr, not 419Asn, is the dominant residue in the o3/TM7 region for Gallamine's high potency, although 419Asn can substitute for 423Thr in some contexts; in contrast, the presence of 419Asn reduces the potency of W84 in every context we have studied. In addition, the orientation of 177Tyr is crucial to high sensitivity toward W84, and it seems that the proline residue at position 179 in M5 (corresponding to M2 172Glu) may interfere with that orientation. Consistent with these observations, a mutant M5 receptor with these three key mutations, M5P179E, Q184Y, and H478T, showed dramatically increased sensitivity for W84 (>100-fold), compared with the wild-type M5 receptor. This same mutant receptor approached M2 sensitivity toward Gallamine. Thus, Gallamine and W84 derive high potency from the same receptor domains (epitopes in o2 and near the junction between o3 and TM7), even though these allosteric agents have quite different structures.
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muscarinic allosteric modulation m2 m3 subtype selectivity of Gallamine is independent of g protein coupling specificity
Naunyn-schmiedebergs Archives of Pharmacology, 2001Co-Authors: Christian Trankle, Evi Kostenis, Klaus MohrAbstract:Among the five subtypes of muscarinic acetylcholine receptors, the sensitivity towards allosteric modulation is generally higher in M2 and M4 receptors that preferentially couple to inhibitory G-proteins of the Gi/o type than in M1, M3, and M5 that preferentially couple to stimulatory G-proteins such as Gq/11. We aimed to check whether the high allosteric sensitivity of the M2 receptor compared to M3 is related to the differential G-protein coupling preference. As the third intracellular loop (i3) is known to be the major determinant in receptor G-protein coupling specificity, we used wild-type M2 and M3 receptors and the related chimeric constructs with exchanged i3-loops, i.e., M2 containing M3-i3 (M2/M3-i3) and M3 containing M2-i3 (M3/M2-i3). The allosteric effect of the archetypal modulator Gallamine on the dissociation and the equilibrium binding of [3H]N-methylscopolamine ([3H]NMS) was measured in membranes of mouse A9L cells stably expressing the wild-type and the chimeric receptors (4 mM Na2HPO4, 1 mM KH2PO4, pH 7.4, 23°C). The dissociation of [3H]NMS was monophasic under all conditions studied. Control values of t 1/2 were (means ± SEM, n=4–7): M2: 3.8±0.2 min, M2/M3-i3: 4.8±0.3 min, M3: 43.3±4.2 min, M3/M2-i3: 41.1±3.6 min. At M2 receptors, 0.2 µM Gallamine allosterically reduced the apparent rate constant of dissociation k –1 to 51±5% of the control value (n=5). At M2/M3-i3 the allosteric potency of Gallamine was not significantly changed (0.2 µM Gallamine → k –1=61±4%, n=7). At M3, a 20-fold higher concentration was required for an equieffective allosteric action (10 µM Gallamine → k –1=51±5%, n=5). The potency of Gallamine at M3/M2-i3 was not increased compared with M3 receptors (10 µM Gallamine → k –1=73±2%, n=4) but even significantly diminished. [3H]NMS equilibrium binding experiments revealed that neither the binding constants of Gallamine at free receptor subtypes (pK A,M2: 7.57±0.04, n=4; pK A,M3: 5.56±0.13, n=3) nor the factors of negative cooperativity with [3H]NMS (αM2=31±1, αM3=3±0.4) were affected by the exchanged i3-loops (pK A,M2/M3-i3: 7.65±0.03, pK A,M3/M2-i2: 5.35±0.24, αM2/M3-i3=30±2, αM3/M2-i2=3±0.7).
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using a radioalloster to test predictions of the cooperativity model for Gallamine binding to the allosteric site of muscarinic acetylcholine m2 receptors
Molecular Pharmacology, 1999Co-Authors: Christian Trankle, Oliver Weyand, Alexandra Schroter, Klaus MohrAbstract:The muscarinic M 2 receptor contains an orthosteric and an allosteric site. Binding of an allosteric agent may induce a shift α of the equilibrium dissociation constant K D of a radioligand for the orthosteric site. According to the cooperativity model, the K A of alloster binding is expected to be shifted to an identical extent depending on whether the orthosteric site is occupied by the orthoster or not. Here, the novel radioalloster [ 3 H]dimethyl-W84 ( N , N ′-bis[3-(1,3-dihydro-1,3-dioxo-4-methyl-2 H -isoindol-2-yl)propyl]- N , N , N ′, N ′-tetramethyl-1,6-hexanediaminium diiodide) was applied to directly measure the K A shift induced for the prototype allosteric modulator Gallamine by binding of N- methylscopolamine (NMS) to the orthosteric site of porcine heart M 2 receptors (4 mM Na 2 HPO 4 , 1 mM KH 2 PO 4 , pH 7.4; 23°C; data are means ± S.E.). First, in the common way, the concentration-dependent inhibition by Gallamine of [ 3 H]NMS equilibrium binding was measured and analyzed using the cooperativity model, which yielded for the affinity of Gallamine binding at free receptors a p K A = 8.35 ± 0.09 and a cooperativity factor α = 46 ( n = 5). The dissociation constant for Gallamine binding at NMS-occupied receptors was predicted as p(α · K A ) = 6.69. Labeling of the allosteric site by [ 3 H]dimethyl-W84 allowed the measure of competitive displacement curves for Gallamine. The K i for Gallamine at free receptors amounted to p K i,−NMS = 8.27 ± 0.39 ( n = 5), which is in line with the prediction of the cooperativtiy model. In the presence of 1 μM NMS, to occupy the orthosteric site, Gallamine displaced [ 3 H]dimethyl-W84 with p K i,+NMS = 6.60 ± 0.19 ( n = 3). Thus, the NMS-induced p K i shift amounted to 47, which matches the predicted value of α = 46. These results validate the cooperativity model.
Lucy Sasongko - One of the best experts on this subject based on the ideXlab platform.
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muscle distribution of the neuromuscular blocker Gallamine using microdialysis
Journal of Pharmaceutical Sciences, 2002Co-Authors: Lucy Sasongko, Iqbal Ramzan, Kenneth M Williams, Andrew J MclachlanAbstract:Abstract Measurement of drug concentrations in target tissue has the potential to provide insight into the pharmacokinetics and pharmacodynamics of a drug. In this study, the distribution of the neuromuscular blocker, Gallamine, into muscle tissue was investigated in urethane‐anesthetized rats after an intravenous bolus dose (6 mg/kg). Microdialysis sampling was used to continuously determine Gallamine concentrations in muscle interstitial fluid (MIF). In vivo microdialysis recovery of Gallamine was determined as the relative loss of Gallamine from the perfusate into muscle tissue after perfusion with Gallamine (2 μg/mL). Recovery was determined in each rat before the pharmacokinetic studies. Terminal muscle sampling followed by homogenization was also performed to examine Gallamine distribution within muscle tissue. All samples were assayed for Gallamine using a validated high‐performance liquid chromatography assay. Gallamine was rapidly distributed into MIF with a MIF–plasma partition coefficient of 0.9 ± 0.1 ( n = 6). By contrast, the estimated Gallamine concentration in muscle tissue homogenate was only 23 ± 5% ( n = 5) of the concentration in MIF as estimated by microdialysis sampling at the terminal sampling time. These findings suggest that Gallamine is not distributed uniformly within muscle but selectively distributes into MIF. Simulations using a hybrid physiologically based pharmacokinetic model which describes uptake of drug only into the interstitial space showed good agreement between predicted and observed concentration data obtained from microdialysis sampling, supporting the findings that Gallamine selectively distributes into MIF. These studies demonstrate microdialysis combined with conventional terminal tissue sampling provides valuable information on intra‐tissue drug distribution. © 2002 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 91: 769–775, 2002
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assessment of in vitro and in vivo recovery of Gallamine using microdialysis
Journal of Pharmacological and Toxicological Methods, 2000Co-Authors: Lucy Sasongko, Iqbal Ramzan, Kenneth M Williams, Andrew J MclachlanAbstract:Abstract The application of microdialysis technique for the investigation of pharmacokinetics and pharmacodynamics of drugs requires careful assessment of probe performance to ensure validity of the data obtained using this technique. The aim of this study was to establish and validate the microdialysis technique for investigation of the pharmacokinetics and pharmacodynamics of the neuromuscular blocker, Gallamine. In vitro recovery of Gallamine from the microdialysis probe when different perfusion flow rates were employed was evaluated leading to selection of a flow rate of 2 μl/min with 15-min sampling intervals for the subsequent studies. In vitro recovery of Gallamine from the microdialysis probe was independent of concentration, stable over an 8-h period and reproducible. Comparable in vitro recoveries were obtained by different established approaches including recovery estimation by gain, loss and the zero-net flux (ZNF) method. Recovery by loss was used to study the in vivo recovery of Gallamine from rat muscle tissue. The in vivo recovery was stable over a 5.5-h sampling period. In vitro performance of the probe subsequent to the in vivo study remained stable supporting reusage of the probe. These data highlight the importance of a systematic examination of microdialysis probe validation.
A C Olito - One of the best experts on this subject based on the ideXlab platform.
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differential effects of the muscarinic m2 antagonists af dx 116 and Gallamine on single neurons of rabbit sympathetic ganglia
Neuropharmacology, 1990Co-Authors: C A Yarosh, J H Ashe, A C OlitoAbstract:Abstract Intracellular recording techniques were used to compare the effects of the M2 muscarinic antagonists, AF-DX 116 and Gallamine, on membrane potential (Vm), input resistance (Ri), responses induced by methacholine, muscarinic slow postsynaptic potentials and action potentials in the superior cervical ganglion of the rabbit. Gallamine or AF-DX 116 antagonized methacholine-induced or synaptically-evoked muscarinic hyperpolarization, without having significant effect on depolarization induced by methacholine or synaptically. The drug AF-DX 116 reduced evoked muscarinic hyperpolarizing potentials, without significant change in Vm or Ri, recorded in the absence of muscarinic stimulation. In contrast to AF-DX 116, Gallamine elicited a concentration-dependent depolarization of the membrane, with a corresponding increase in Ri, when tested in the absence of muscarinic stimulation. These effects of Gallamine were accompanied by an increase in duration and decrease in the slope of the descending phase of the action potential. Blockade by Gallamine of evoked hyperpolarization was independent of membrane depolarization and readily occurred when Gallamine-induced depolarization was prevented by clamping Vm at its pre-Gallamine level. The effects of Gallamine were maintained during its presence and reversed upon washing with Gallamine-free physiological solution. These results indicate that AF-DX 116 and Gallamine have a specificity for antagonism of muscarinic responses, mediated by receptors of the M2 type in the superior cervical ganglion. However, Gallamine, while an effective antagonist of M2 responses, also has the ability to modify the electrical characteristics of ganglion cells and thus may modify ganglionic transmission by mechanisms other than antagonism of receptors.
Christian Trankle - One of the best experts on this subject based on the ideXlab platform.
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first Gallamine tacrine hybrid design and characterization at cholinesterases and the m2 muscarinic receptor
Journal of Medicinal Chemistry, 2007Co-Authors: Paul W Elsinghorst, Julia S Cieslik, Klaus Mohr, Christian Trankle, Michael GutschowAbstract:Gallamine and tacrine are allosteric antagonists at muscarinic M2 acetylcholine receptors and inhibitors of acetylcholinesterase. At both acetylcholine-binding proteins, Gallamine and tacrine are known to occupy two different binding sites: in M2 receptors within the allosteric binding area and in acetylcholinesterase at its catalytic and its peripheral site. To find new ligands of both targets, we designed a Gallamine−tacrine dimer and several derived hybrid compounds to address the two binding sites. Their M2 receptor allosteric and acetylcholinesterase inhibitory potential was determined. The hybrid compounds revealed an allosteric potency in the low nanomolar range exceeding the allosteric potency of Gallamine and tacrine by factors of 100 and 4800, respectively. Cholinesterase inhibition was augmented by hybrid formation, and all compounds exhibited IC50 values in the lower nanomolar range. Thus, Gallamine−tacrine hybrid formation is a valuable approach toward high affinity ligands concurrently targ...
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muscarinic allosteric modulation m2 m3 subtype selectivity of Gallamine is independent of g protein coupling specificity
Naunyn-schmiedebergs Archives of Pharmacology, 2001Co-Authors: Christian Trankle, Evi Kostenis, Klaus MohrAbstract:Among the five subtypes of muscarinic acetylcholine receptors, the sensitivity towards allosteric modulation is generally higher in M2 and M4 receptors that preferentially couple to inhibitory G-proteins of the Gi/o type than in M1, M3, and M5 that preferentially couple to stimulatory G-proteins such as Gq/11. We aimed to check whether the high allosteric sensitivity of the M2 receptor compared to M3 is related to the differential G-protein coupling preference. As the third intracellular loop (i3) is known to be the major determinant in receptor G-protein coupling specificity, we used wild-type M2 and M3 receptors and the related chimeric constructs with exchanged i3-loops, i.e., M2 containing M3-i3 (M2/M3-i3) and M3 containing M2-i3 (M3/M2-i3). The allosteric effect of the archetypal modulator Gallamine on the dissociation and the equilibrium binding of [3H]N-methylscopolamine ([3H]NMS) was measured in membranes of mouse A9L cells stably expressing the wild-type and the chimeric receptors (4 mM Na2HPO4, 1 mM KH2PO4, pH 7.4, 23°C). The dissociation of [3H]NMS was monophasic under all conditions studied. Control values of t 1/2 were (means ± SEM, n=4–7): M2: 3.8±0.2 min, M2/M3-i3: 4.8±0.3 min, M3: 43.3±4.2 min, M3/M2-i3: 41.1±3.6 min. At M2 receptors, 0.2 µM Gallamine allosterically reduced the apparent rate constant of dissociation k –1 to 51±5% of the control value (n=5). At M2/M3-i3 the allosteric potency of Gallamine was not significantly changed (0.2 µM Gallamine → k –1=61±4%, n=7). At M3, a 20-fold higher concentration was required for an equieffective allosteric action (10 µM Gallamine → k –1=51±5%, n=5). The potency of Gallamine at M3/M2-i3 was not increased compared with M3 receptors (10 µM Gallamine → k –1=73±2%, n=4) but even significantly diminished. [3H]NMS equilibrium binding experiments revealed that neither the binding constants of Gallamine at free receptor subtypes (pK A,M2: 7.57±0.04, n=4; pK A,M3: 5.56±0.13, n=3) nor the factors of negative cooperativity with [3H]NMS (αM2=31±1, αM3=3±0.4) were affected by the exchanged i3-loops (pK A,M2/M3-i3: 7.65±0.03, pK A,M3/M2-i2: 5.35±0.24, αM2/M3-i3=30±2, αM3/M2-i2=3±0.7).
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using a radioalloster to test predictions of the cooperativity model for Gallamine binding to the allosteric site of muscarinic acetylcholine m2 receptors
Molecular Pharmacology, 1999Co-Authors: Christian Trankle, Oliver Weyand, Alexandra Schroter, Klaus MohrAbstract:The muscarinic M 2 receptor contains an orthosteric and an allosteric site. Binding of an allosteric agent may induce a shift α of the equilibrium dissociation constant K D of a radioligand for the orthosteric site. According to the cooperativity model, the K A of alloster binding is expected to be shifted to an identical extent depending on whether the orthosteric site is occupied by the orthoster or not. Here, the novel radioalloster [ 3 H]dimethyl-W84 ( N , N ′-bis[3-(1,3-dihydro-1,3-dioxo-4-methyl-2 H -isoindol-2-yl)propyl]- N , N , N ′, N ′-tetramethyl-1,6-hexanediaminium diiodide) was applied to directly measure the K A shift induced for the prototype allosteric modulator Gallamine by binding of N- methylscopolamine (NMS) to the orthosteric site of porcine heart M 2 receptors (4 mM Na 2 HPO 4 , 1 mM KH 2 PO 4 , pH 7.4; 23°C; data are means ± S.E.). First, in the common way, the concentration-dependent inhibition by Gallamine of [ 3 H]NMS equilibrium binding was measured and analyzed using the cooperativity model, which yielded for the affinity of Gallamine binding at free receptors a p K A = 8.35 ± 0.09 and a cooperativity factor α = 46 ( n = 5). The dissociation constant for Gallamine binding at NMS-occupied receptors was predicted as p(α · K A ) = 6.69. Labeling of the allosteric site by [ 3 H]dimethyl-W84 allowed the measure of competitive displacement curves for Gallamine. The K i for Gallamine at free receptors amounted to p K i,−NMS = 8.27 ± 0.39 ( n = 5), which is in line with the prediction of the cooperativtiy model. In the presence of 1 μM NMS, to occupy the orthosteric site, Gallamine displaced [ 3 H]dimethyl-W84 with p K i,+NMS = 6.60 ± 0.19 ( n = 3). Thus, the NMS-induced p K i shift amounted to 47, which matches the predicted value of α = 46. These results validate the cooperativity model.
