Neuromuscular Blocking Action Research Articles

Mode of neuromuscular blocking action of a toxic phospholipase A 2 from Pseudocerastes fieldi (Field's horned viper) snake venom

The effects of a toxic phospholipase A 2 (Fr.Cb) isolated from the venom of Pseudocerastes fieldi were studied on the chick biventer cervicis muscle and the mouse phrenic nerve — diaphragm preparations. In the chick muscle, Fr.Cb (10 μg/ml) caused complete neuromuscular blockade without producing contracture or affecting the response of the muscle to acetylcholine. In the mouse diaphragm, Fr.Cb blocked the indirectly elicited contraction without affecting that evoked directly. In a low calcium medium (0.5 mM), Fr.Cb produced a triphasic change of the indirectly elicited contractions. The frequency of miniature endplate potentials (m.e.p.p.s) in the mouse diaphragm was first increased 3 – 4 fold 40 min after toxin (10 μg/ml) application, then gradually decreased, while the amplitude of m.e.p.p.s. was not decreased, even after the evoked release of transmitter had failed. Giant m.e.p.p.s were frequently observed. The quantal content first increased and then decreased gradually. The resting membrane potential and the compound phrenic nerve action potential were not significantly affected by the toxin at 10 μg/ml after 2 hr of incubation. The motor nerve terminals in the Fr.Cb intoxicated mouse diaphragm showed swelling and vacuolization of both synaptic vesicles and mitochondria. It is concluded that the toxin produces a neuromuscular blockade by acting selectively on the presynaptic site.

Mode of neuromuscular blocking action of ceruleotoxin

Ceruleotoxin is an acidic toxin protein from a Bungarus venom which has recently been clarified as B. fasciatus venom. The toxin at a low concentration (10 −6g/ml) abolished the twitch response of the indirectly stimulated biventer cervicis muscle of the chick, without affecting the response to acetylcholine or carbamylcholine. At a higher concentration (10 -5g/ml), the toxin, in addition to inhibition of the twitch response, caused contracture in the chick biventer cervicis muscle and reduced the response to acetylcholine or carbamylcholine in both the innervated chick cervical and denervated rat diaphragm muscles. Electrophysiological studies on the rat phrenic nerve - diaphragm preparation showed that ceruleotoxin at a low concentration caused an initial inhibition followed by recovery of the quantal content of endplate potentials and then a second phase of inhibition leading to complete failure. At a higher concentration, the toxin gradually reduced the resting membrane potential of the diaphragm muscle to a level lower than 50 mV within 2 hr. In addition, enzyme assay showed that the toxin possessed phospholipase A activity comparable to that of other basic phospholipases A 2 of snake venom origin. It is concluded that ceruleotoxin is a phospholipase A 2 with presynaptic and myotropic actions on vertebrate nerve - muscle system similar to those of notexin from Notechis scutatus scutatus venom.

Amino acid sequences of three phospholipases A I, III and IV from the venom of the sea snake Laticauda semifasciata.

Amino acid sequences of three phospholipases A, I, III and IV, from the venom of the sea snake Laticauda semifasciata were elucidated. Each protein consisted of a single chain of 118 amino acid residues, including 14 half-cystine residues. They showed high homology among themselves, and with the other snake-venom phospholipases A and with the enzymes from mammalian pancreas. Phospholipases A III and IV were especially similar to each other, with only four differences out of their 118 amino acid residues. Phospholipase A I contained one tryptophan residue at position 64, which was important for enzymic activity, whereas III and IV did not contain tryptophan residues and their corresponding positions were occupied by leucine residues. The substitution by leucine resulted in a decreased, but definite, phospholipase A activity. The substituted enzymes have a more potent neuromuscular blocking activity. Full experimental details and evidence for the amino acid sequences of the proteins have been deposited as Supplementary Publication SUP 50118 (39 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem.J. (1981) 193, 5.

The Neuromuscular Blocking Actions of Coclaurine Derivatives and of Paeoniflorin Derivatives

The Neuromuscular Blocking Actions of Coclaurine Derivatives and of Paeoniflorin Derivatives

A new non-depolarizing myorelaxant with high activity and specificity

The curare-like activity and mode of action of tercuronium (p,p"-bistriethylammonium-p-terphenyl dibenzen-sulphonate) have been investigated in experiments with cats, rabbits, mice and pigeons as well as with rat phrenic diaphragm preparations. The curare-like activity of tercuronium was higher than that of tubocurarine and was close to that of pancuronium bromide. The curare-like activity of tercuronium was higher than that of tubocurarine and was close to that of pancuronium bromide. The neuromuscular blocking doses (ED95) of tercuronium, (+)-tubocurarine and pancuronium in cats, for example, were 0.08 microM/kg, 0.4 microM/kg and 0.04 microM/kg respectively. The time of development and duration of action were similar to those of (+)-tubocurarine and pancuronium. Tercuronium is a nondepolarizing myorelaxant. In experiments with cats the antagonism of neostigmine (0.1 mg/kg) to tercuronium was more pronounced than in the case of (+)-tubocurarine, pancuronium or gallamine. Tercuronium affected neither the arterial pressure nor the heart rate when given in neuromuscular blocking doses. Tercuronium did not block transmission through autonomic ganglia and had no atropine-like action. Only a 10-fold increase in the dose of tercuronium produced the ganglion blocking effect in cats. Under artificial respiration, cats and rabbits tolerated tercuronium in a dose 200 times exceeding its myoparalytic dose. It is concluded that tercuronium is distinguishable from (+)-tubocurarine by its high neuromuscular blocking activity as well as by its specificity and more pronounced neostigmine antagonism.

Observations on the neuromuscular blocking action of gallamine and pancuronium and their reversal by neostigmine

The neuromuscular blocking action of gallamine triethiodide and pancuronium bromide was investigated in the dog using train-of-four stimuli. The mean duration of gallamine was 29 minutes and that of pancuronium was 31 minutes. Reversal of the neuromuscular block was produced by atropine and neostigmine.

Site of action of caudoxin, a neurotoxic phospholipase A 2 from the horned puff adder ( Bitis caudalis) venom

Caudoxin, a single-chain phospholipase A 2 isolated from the venom of Bitis caudalis is a toxic polypeptide with a formula weight of 13,332 dalton. The ld 50 in mice (i.p.) was 0.18 (0.15–0.22) mg/kg. In the chick biventer cervicis muscle preparation the toxin (1–10 μg per ml) caused complete neuromuscular blockade without affecting the response of the muscle to acetylcholine. In the mouse phrenic nerve-diaphragm preparation, the toxin abolished the indirectly elicited contraction without inhibiting that evoked directly. When this preparation was bathed in a low calcium (0.6 mM) medium, the toxin induced a triphasic change in the indirectly evoked contractions: an immediate initial inhibition followed by augmentation and then a second phase of inhibition leading to irreversible neuromuscular blockade. Electrophysiological studies in the same preparation showed a similar triphasic change in the quantal content of endplate potentials. The frequency of miniature endplate potentials first increased and then decreased, while the resting membrane potential was not significantly decreased by the toxin. Histological study showed that the toxin caused local myonecrosis only at a higher dose (2 mg/kg mouse). It is concluded that caudoxin produced a neuromuscular block by acting selectively on a presynaptic site. However, the site of binding appears to be different from that of β-bungarotoxin since combination of the toxin with β-bungarotoxin caused potentiation of its neuromuscular blocking action rather than addition.

Presynaptic toxicity of the histidine-modified, phospholipase A 2-inactive, β-bungarotoxin, crotoxin and notexin

C. C. Chang and M. J. Su. Presynaptic toxicity of the histidine-modified, phospholipase A 2-inactive, β-bungarotoxin, crotoxin and notexin. Toxicon 20, 895–905, 1982.—β-Bungarotoxin, crotoxin and notexin were treated extensively with p-bromophenacyl bromide in order to modify the histidine group of the active site and to greatly decrease the phospholipase A 2(PLA) catalytic activity. They were studied for their residual presynaptic effects on the mouse isolated phrenic nerve-diaphragm and chick biventer cervicis muscle preparations. The modified toxins still had 1.7–5.2% PLA activity, which was inhibited by Sr 2+, as is the enzyme activity of the native toxins. The neuromuscular blocking activity of these modified toxins, which was reduced 30–60 fold in the mouse diaphragm, was due to a presynaptic effect, as judged from the unchanged amplitude of m.e.p.p.s in the blocked preparations. In contrast to the native toxins, however, the presynaptic effect of modified β-bungarotoxin and notexin was neither antagonized by Sr 2+ nor accelerated by increasing the rate of nerve stimulation, indicating that the presynaptic effects of the modified β-bungarotoxin and notexin are not likely to be due to their PLA enzyme activity. The modified toxins retained a much greater fraction of their early presynaptic effects in comparison to their enzymatic and neuromuscular blocking activities, although the time-course of the early effects was delayed. The results indicate that the modified neurotoxins per se have their own presynaptic effects, which are unrelated to the PLA enzyme activities.

Effects on muscle of a toxin from indian cobra ( Naja naja naja) venom

—The mode of action of a purified toxin from Naja naja naja (Indian cobra) venom was investigated in frog rectus abdominis muscle, chick biventer cervicis muscle, cat tibialis anterior muscle (close-arterial) and in both innervated and denervated rat diaphragm muscle preparations. The toxin inhibited the acetyl-choline responses of rectus abdominis muscle. The inhibition was antagonized by neostigmine and increasing concentrations of acetylcholine, suggesting a competitive binding of the toxin to cholinergic receptors. The toxin, even at high doses, did not produce depolarizing contractures in chronically denervated diaphragm, biventer cervicis muscle and rectus abdominis muscle preparations. In both cat tibialis anterior and denervated diaphragm muscles, the toxin abolished the acetylcholine sensitivity of the muscles at a faster rate than its effects on muscle contraction, suggesting a preferred action on the motor end-plate. A well-maintained tetanic contraction and very poor post-tetanic potentiation was observed in all preparations treated with toxin, indicating an atypical Wedensky inhibition. Anti-curare agents, such as K + and Ca 2+, were ineffective in antagonizing the curare-like neuromuscular block in phrenic nerve-diaphragm preparations. A frequency-independent neuromuscular block observed in these nerve-muscle preparations was suggestive of the absence of a possible presynaptic effect. These results demonstrate that although the neurotoxin in some cases can imitate d-tubocurarine, its neuromuscular blocking activity is different from that of curare in many respects.

Comparative evaluation of the neuromuscular blocking activity of three new aminoglycoside antibiotics in rats

The effects of three aminoglycoside antibiotics on the rat isolated phrenic nerve-diaphragm preparation and on the sciatic nerve-gastrocnemius muscle preparation were investigated. Tobramycin, amikacin and ribostamycin produced dose-dependent neuromuscular blockade of the diaphragm twitches. Comparison of results showed that the neuromuscular blocking potency was as follows: tobramycin > amikacin > ribostamycin. The neuromuscular blockade produced by these antibiotics was reversed by calcium chloride, whereas it was not influenced by neostigmine methylsulfate. Furthermore, the neuromuscular blocking potency in vitro of these three aminoglycosides was paralleled by their activity in vivo on the sciatic nerve-gastrocnemius muscle preparation.

INTERACTIONS BETWEEN THE NEUROMUSCULAR BLOCKING DRUG ORG NC45 AND SOME ANAESTHETIC, ANALGESIC AND ANTIMICROBIAL AGENTS

The effects of some anaesthetics and other agents used in anaesthetic practice on the neuromuscular blocking activity of cumulative doses of Org NC 45 have been studied in cats anaesthetized with chloralose. As expected from studies with other neuromuscular blocking drugs, both inhalation and i.v. anaesthetics reduced the doses of Org NC 45 required for neuromuscular blockade. An apparently specific potentiating interaction between Org NC 45 and metronidazole was observed.

Effects of corticosteroids on neuromuscular blocking actions of d-tubocurarine

Dexamethasone (50 μg/kg) significantly increased the LD 50 of d-tubocurarine (d-TC) when administered i.p. simultaneously with d-TC. Choline (50 and 100 mg/kg) gave some protection against the lethal effects of d-TC and the cholinesterase inhibitors neostigmine (250 μg/kg) and physostigmine (1000 μg/kg) provided full protection against doses of d-TC twice the LD 50. The blocking effect of d-TC (75 μg/kg) on the sciatic nerve-tibialis anterior muscle preparation was antagonized by dexamethasone. Prednisolone delayed the occurrence of a complete neuromuscular block caused by d-TC in the phrenic nerve-diaphragm preparation, and antagonized the effect of d-TC on short tetanic contractions. d-TC (5 μmol/l) inhibited the [ 14C]choline uptake in the endplate-rich region of the rat diaphragm during stimulation. This inhibition was antagonized by dexamethasone as well as by physostigmine. The incorporation of radioactive choline into acetylcholine was inhibited in the presence of d-TC (15 μmol/l), and both dexamethasone and physostigmine counteracted this inhibition. It is concluded from these experiments that d-TC very probably has an effect of the choline carrier system. These experimental results support the hypothesis that glucocoriticoids may improve reduced muscle performance by direct presynaptic effects at the neuromuscular junction.

Presynaptic actions of Mojave toxin isolated from Mojave rattlesnake ( Crotalus scutulatus) venom

As with other presynaptically acting toxins with phospholipase A 2 activity from snake venoms, Mojave toxin caused a triphasic change in both the indirectly elicited muscle contraction and the quantal content of endplate potentials. This led to irreversible transmission block in the mouse diaphragm bathed in low calcium medium. The frequency of miniature endplate potentials gradually decreased after an initial increase. The neuromuscular blocking action of this toxin was potentiated by β-bungarotoxin but not by crotoxin, suggesting that its binding site may be identical with that of crotoxin but different from that of β-bungarotoxin.

Cimetidine increases the neuromuscular blocking activity of aminoglycoside antibiotics: Antagonism by calcium

Cimetidine increases the neuromuscular blocking activity of aminoglycoside antibiotics: Antagonism by calcium

Synthesis, Stereochemical Analysis, and Neuromuscular Blocking Activity of Selected E,E- and Z,Z-Isomeric O-Ethers of 4-Androstene-3,17-Dione Dioximes

A number of selected geometric oxime ethers were synthesized as potential neuromuscular blocking agents. The study consisted of the synthesis, separation, and stereochemical analysis of selected compounds followed by a preliminary pharmacological evaluation for neuromuscular blockade. Synthesis of the compounds began with the double oximination of 4-androstene-3,17-dione followed by TLC, column chromatography, and fractional crystallization as the purification methods. These procedures yielded two of the possible four isomers. Configurational assignments on the isolated pair were based on experiments using IR, UV, and NMR spectroscopy. Isomerically pure diethers were prepared by two methods: (a) O-alkylation of the stereochemically pure dioximes with the appropriate aminoalkyl halide, and (b) O-alkylation of the dioxime mixture followed by quaternization and subsequent isolation of configurationally pure salts via fractional crystallization. A preliminary pharmacological evaluation was conducted by using mice on a treadmill apparatus. Some structure-activity relationships are discussed.

Structure—neuromuscular blocking activity relationships of ketophosphonium salts

Three ketophosphonium salts containing a morpholino group were prepared and evaluated for their neuromuscular blocking effect. Solution and crystal conformations were studied. An attempt was made to explain, in structural terms, different activity levels in connection with the interatomic distances.

Neuromuscular blocking activity of dibekacin, a new semisynthetic aminoglycoside antibiotic.

Dibekacin possesses the untoward effect of producing a neuromuscular blockade. Neostigmine is unable to reverse the neuromuscular blockade produced by dibekacin. Calcium has not only the ability to restore the neuromuscular transmission but also to exert protective action against the neuromuscular blocking activity of dibekacin.

Pharmacology of Org NC 45 Compared with Other Nondepolarizing Neuromuscular Blocking Drugs

From results of pharmacological tests on the neuromuscular and autonomic blocking actions of a series of pancuronium analogues, Org NC 45, the C16 monoquaternary analogue of pancuronium, was selected for detailed study. Org NC 45 has a non-depolarizing mechanism of action, is more rapid in onset and shorter in duration of action than pancuronium. It shows less cumulation than pancuronium or tubocurarine, and is easily antagonized by anticholinesterases and aminopyridines. Org NC 45 exhibits a low propensity to release histamine. Its ability to inhibit cholinesterases is not likely to be important at neuromuscular blocking doses. Org NC 45 possesses negligible ganglion-blocking activity and there is a wide margin between neuromuscular and vagal blocking doses. Thus cardiovascular side-effects are unlikely to occur with the use of Org NC 45. It will hydrolyse mainly to its 3-hydroxy analogue which, like Org NC 45, possesses a wide margin between neuromuscular and vagal blocking doses. Org NC 45 has a high selectivity for the neuromuscular junction and represents a potentially useful addition to the armamentarium of clinically useful muscle relaxants.

The Neuromuscular Blocking Action of ORG NC 45, A New Pancuronium Derivative, in Anaesthetized Patients: A pilot study

The neuromuscular blocking effects of the new monoquaternary analogue of pancuronium, Org NC 45, have been investigated in anaesthetized patients. In different doses administered as a single i.v. bolus or as an initial bolus followed by several small maintenance doses or by a continuous infusion. Org NC 45 appears to be approximately as potent as pancuronium, but has a more rapid onset of action, considerably shorter duration of action and faster recovery rate than pancuronium. It showed no cumulative effects even after 10 maintenance doses were injected in succession. Doses of 0.08 mg kg-1 provided ideal intubating conditions in 90--95 s. Infusions of Org NC 45 provided much smoother control of neuromuscular blockade than did pancuronium. No cardiovascular side-effects were noted even at the greatest dose (0.12 mg kg-1) used. Org NC 45 has clear advantages over pancuronium and represents a potentially valuable addition to the armamentarium of clinically useful muscle relaxants.

Use of the Human “Isolated Arm” Preparation to Indicate Qualitative Aspects of a New Neuromuscular Blocking Agent, ORG NC 45

In the human isolated arm preparation Org NC 45 had a significantly faster rate of recovery and a shorter duration of action (end of injection to 90% recovery of control twitch height) than pancuronium. The isolated arm preparation is suitable for the qualitative comparison of the neuromuscular blocking activities of new and established drugs.