Muscarinic receptor modulation of GABA-mediated giant depolarizing potentials in the neonatal rat hippocampus.
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1. The whole-cell patch clamp technique was used to study the role of muscarinic receptors in regulating the frequency of giant depolarizing potentials (GDPs) in CA3 hippocampal neurones in slices from postnatal (P) P1-P8 rats. 2. Atropine (1 microM) reduced the frequency of GDPs by 64.2 +/- 2.9 %. The acetylcholinesterase inhibitor edrophonium (20 microM) increased the frequency of GDPs in a developmentally regulated way. This effect was antagonized by the M1 muscarinic receptor antagonist pirenzepine. 3. In the presence of edrophonium, tetanic stimulation of cholinergic fibres induced either an enhancement of GDP frequency (179 +/- 79 %) or a membrane depolarization (27 +/- 16 mV) associated with an increase in synaptic noise. These effects were prevented by atropine. 4. Application of carbachol (3 microM) produced an increase in GDP frequency that at P5-P6 was associated with a membrane depolarization and an increase in synaptic noise. These effects were prevented by atropine, pirenzepine (3 microM) and bicuculline (10 microM). 5. In the presence of pirenzepine, carbachol reduced GDP frequency by 50 +/- 4 %. Conversely, in the presence of methoctramine (3 microM), carbachol enhanced GDP frequency by 117 +/- 4 %. 6. It is concluded that endogenous acetylcholine, through the activation of M1 receptors, enhances the release of gamma-aminobutyric acid (GABA), in a developmentally regulated way. On the other hand, carbachol exerts both an up- and downregulation of GABA release through the activation of M1 and M2 receptors, respectively. (+info)
Antagonism of rapacuronium using edrophonium or neostigmine: pharmacodynamics and pharmacokinetics.
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We have studied the pharmacodynamics and pharmacokinetics of rapacuronium (Org 9487) in 70 healthy patients. Neuromuscular transmission was monitored using TOF stimulation of the ulnar nerve and mechanomyography of the adductor pollicis muscle. Half of the patients were given a single dose of rapacuronium 1.5 mg kg-1 and the remainder rapacuronium 1.5 mg kg-1 with three incremental doses of 0.5 mg kg-1, each given when T1/T0 had recovered to 25%. In all patients, neuromuscular block was antagonized using neostigmine 0.05 mg kg-1 or edrophonium 1.0 mg kg-1 (allocated randomly), 2 min after the final dose of rapacuronium. All patients developed complete block after rapacuronium 1.5 mg kg-1. Mean onset time was 66 (SD 24) s. In patients who received an antagonist 2 min after the first dose of rapacuronium, time to recovery of T1/T0 to 25% was similar after neostigmine (9.8 (3.8) min) and edrophonium (10.3 (4.3) min): in patients who received incremental doses of rapacuronium, spontaneous recovery of T1/T0 to 25% after the first dose was 18.9 (4.7) min. In those who received an antagonist 2 min after the first dose of rapacuronium, times to recovery of T4/T1 to 0.7 were also similar after neostigmine (23.7 (7.7) min) and edrophonium (29.1 (10.7) min). After three incremental doses of rapacuronium, there was a longer time to recovery of T1/T0 = 25% after neostigmine compared with edrophonium (5.1 (1.0) vs 3.3 (1.3) min; P < 0.05) but more rapid recovery to T1/T0 = 75% (10.1 (2.9) vs 16.8 (10.1) min; P < 0.05) and T4/T1 = 0.7 (19.8 (6.3) vs 35.1 (10.4) min; P < 0.05). A three-compartment pharmacokinetic model was justified. Typical values for clearance and initial volume of distribution (V1) were 4.4 ml kg-1 min-1 and 94.8 ml kg-1, respectively. In females, clearance was decreased by 38.5% compared with males and V1 was decreased by 25% in patients aged more than 65 yr. (+info)
Effect of edrophonium and neostigmine on the pharmacokinetics and neuromuscular effects of mivacurium.
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BACKGROUND: Previous studies demonstrated that both edrophonium and neostigmine affect mivacurium's pharmacokinetics, thereby potentially affecting its recovery profile. However, those studies were not clinically relevant because mivacurium was still infused after the antagonists were given. In the present study, the authors gave antagonists (or placebo) after discontinuing a mivacurium infusion, thereby obtaining data that are more clinically relevant. METHODS: In 18 patients, mivacurium was infused at 10 microg kg(-1) x min(-1) for 40 min, the infusion was discontinued for 15 min and then restarted at the same rate for another 40 min. Patients were randomized to receive 500 microg/kg edrophonium, 50 microg/kg neostigmine, or saline at discontinuation of the second infusion; all subjects received 1 mg atropine. Plasma was sampled during the final 10 min of each infusion to determine steady state mivacurium concentrations and for 15 min after each infusion. Twitch tension was recorded. Mivacurium concentrations after each of the two infusions were compared. RESULTS: After discontinuation of the second infusion, mivacurium concentrations were larger than those after the first infusion at 2 min with edrophonium and at 2, 4, and 7 min with neostigmine. With both neostigmine and edrophonium, twitch tension recovered after infusion #2 more rapidly than after infusion #1; however, the magnitude of this effect was small CONCLUSION: Edrophonium transiently slows the rate at which mivacurium concentrations decrease; this is consistent with our previous findings. Neostigmine has a similar, although longer, effect. Despite altering mivacurium's elimination characteristics, both drugs facilitate neuromuscular recovery, although their benefit is small. (+info)
A regional technique for the study of sensitivity to curare in human muscle.
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A regional technique for the study of curare sensitivity of human muslce in vivo is described. 0-5 mg d-tubocurarine is given intravenously at the wrist while the circulation to the hand and forearm is occluded. Neuromuscular transsmission is then studied by delivering trains of stimuli to the ulnar nerve and recording changes in evoked muscle action potential (map) amplitude from the abductor digiti minimi. In most normal subjects, this dose causes a recognizable block in neuromuscular transmission. The amplitude of a single evoked MAP is depressed and declines further during trains of 3/s repetitive stimulation. Recovery usually takes place gradually during the 20-30 minutes after the release of the tourniquet. The technique will be of value in the study of latent disturbance of neuromuscular transmission in neurological and metabolic disorders. (+info)
Thioflavin T is a fluorescent probe of the acetylcholinesterase peripheral site that reveals conformational interactions between the peripheral and acylation sites.
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Three-dimensional structures of acetylcholinesterase (AChE) reveal a narrow and deep active site gorge with two sites of ligand binding, an acylation site at the base of the gorge, and a peripheral site near the gorge entrance. Recent studies have shown that the peripheral site contributes to catalytic efficiency by transiently binding substrates on their way to the acylation site, but the question of whether the peripheral site makes other contributions to the catalytic process remains open. A possible role for ligand binding to the peripheral site that has long been considered is the initiation of a conformational change that is transmitted allosterically to the acylation site to alter catalysis. However, evidence for conformational interactions between these sites has been difficult to obtain. Here we report that thioflavin T, a fluorophore widely used to detect amyloid structure in proteins, binds selectively to the AChE peripheral site with an equilibrium dissociation constant of 1.0 microm. The fluorescence of the bound thioflavin T is increased more than 1000-fold over that of unbound thioflavin T, the greatest enhancement of fluorescence for the binding of a fluorophore to AChE yet observed. Furthermore, when the acylation site ligands edrophonium or m-(N, N,N-trimethylammonio)trifluoroacetophenone form ternary complexes with AChE and thioflavin T, the fluorescence is quenched by factors of 2.7-4.2. The observation of this partial quenching of thioflavin T fluorescence is a major advance in the study of AChE for two reasons. First, it allows thioflavin T to be used as a reporter for ligand reactions at the acylation site. Second, it indicates that ligand binding to the acylation site initiates a change in the local AChE conformation at the peripheral site that quenches the fluorescence of bound thioflavin T. The data provide strong evidence in support of a conformational interaction between the two AChE sites. (+info)
Interaction of edrophonium with muscarinic acetylcholine M2 and M3 receptors.
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BACKGROUND: It has been reported that edrophonium can antagonize the negative chronotropic effect of carbachol. This study was undertaken to evaluate in detail the interaction of edrophonium with muscarinic Mz and M3 receptors. METHODS: A functional study was conducted to evaluate the effects of edrophonium on the concentration-response curves for the negative chronotropic effect and the bronchoconstricting effect of carbachol in spontaneously beating right atria and tracheas of guinea pigs. An electrophysiologic study was conducted to compare the effects of edrophonium on carbachol-, guanosine triphosphate (GTP)gama S-, and adenosine-induced outward K+ currents in guinea pig atrial cells by whole cell voltage clamp technique. A radioligand binding study was conducted to examine the effects of edrophonium on specific [3HIN-methylscopolamine (NMS) binding to guinea pig atrial (M2) and submandibular gland (M3) membrane preparations, and on atropine-induced dissociation of [3H]NMS. RESULTS: Edrophonium shifted rightward the concentration-response curves for the negative chronotropic and bronchoconstricting effects of carbachol in a competitive manner. The pA2 values for cardiac and tracheal muscarinic receptors were 4.61 and 4.03, respectively. Edrophonium abolished the carbachol-induced outward current without affecting the GTPgamma S- and adenosine-induced currents in the atrial cells. Edrophonium inhibited [3H]NMS binding to M2 and M3 receptors in a concentration-dependent manner. The pseudo-Hill coefficient values and apparent dissociation constants of edrophonium for M2 and M3 receptors were 1.02 and 1.07 and 21 and 34 microM, respectively. Edrophonium also changed dissociation constant values of [3H]NMS without affecting its maximum binding capacities. CONCLUSION: Edrophonium binds to muscarinic M2 and M3 receptors nonselectively, and acts as a competitive antagonist. (+info)
Passive transfer of experimental autoimmune myasthenia by lymph node cells in inbred guinea pigs.
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Passive transfer of experimental autoimmune myasthenia (EAM) was performed with lymph node cells from donor guinea pigs immunized with purified acetylcholine receptor (AChR) from Torpedo californica. Recipient animals revealed the same clinical signs and electromyographic patterns as observed in actively challenged animals. These phenomena are parallel to the clinical manifestations of the human disease myasthenia gravis, in which cellular response to AChR was recently demonstrated. (+info)
Effects of the anticholinesterase edrophonium on spectral analysis of heart rate and blood pressure variability in humans.
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Edrophonium, an anticholinesterase, exerts a biphasic effect on cardiovascular autonomic drive in humans (lower doses enhance; higher doses reduce). Twenty-five anesthetized, mechanically respired (10 breaths. min(-1), constant tidal volume) patients were given either saline (n = 10) or edrophonium (0.01-1.0 mg. kg(-1), n = 15) following surgery. ECG, radial arterial pressure, and respiratory rate were sampled at 250 Hz to obtain time series for consecutive R-R intervals (RRIs), and systolic (SBP) and diastolic blood pressure (DBP). A Wigner distribution was used for time frequency mapping of spectral powers at high (HFP, 0.15-0.5 Hz) and low (LFP, 0.0-0.05 Hz) frequency. Edrophonium produced a dose-dependent decrease in heart rate [baseline 66.8 +/- 1.9 (S.E.M.) beats per minute; maximum decrease to 55.8 +/- 1.4 beats per minute with 1.0 mg. kg(-1), P < 0.01]. HFP of the RRI increased at low doses (0.2-0.4 mg. kg(-1); maximum increase to 111.0 +/- 58.2% baseline; P < 0.01) but decreased (-49.5 +/- 35.5% baseline; P < 0.01) at higher (1.0 mg. kg(-1)) doses. Edrophonium had no effect on SBP and DBP. HFP of SBP decreased with increasing doses (maximal decrease to -26.2 +/- 7.5% baseline, P < 0.01, 1.0 mg. kg(-1)). LFP of SBP was also decreased (-46.3 +/- 10.9% baseline, P < 0.01, 1.0 mg. kg(-1)). Edrophonium may enhance (lower dose) or reduce (higher dose) cardiovascular autonomic drive in humans, as evidenced by the significant changes it evokes in HFP of the RRI (parasympathetic drive), and in the HFP and LFP of SBP (sympathetic drive). These observations may account for the modest autonomic side effects of edrophonium when this drug is used clinically. (+info)