Chick Biventer Cervicis Nerve-muscle Preparation Research Articles

Neuromuscular effects of fractions isolated from the venom of the Thailand cobra, Naja naja siamensis

Four fractions (α, β, γ and δ) were isolated from the venom of the Thailand cobra, Naja naja siamensis by ion-exchange chromatography. Each fraction was tested for activity on the isolated chick biventer cervicis nerve-muscle preparation. Fractions α, β and γ inhibited neuromuscular transmission by blocking acetylcholine receptors, but had no direct action on muscle contractility. Fraction α produced neuromuscular blockade in about 30 min when added at a concentration of 1 μg/ml, and was about 100 times more potent than fractions β and γ. The blockade of indirect stimulation was apparently irreversible, although the responses to acetylcholine and carbachol recovered to about 50% after 4 hr. Fraction δ produced contracture of the muscle followed by blockade of responses to both direct and indirect stimulation. The effects of fraction δ could be inhibited by raising the calcium concentration of the bathing solution; on return to normal calcium concentration the effects of fraction δ also returned. It is concluded that fractions α, β and γ are postjunctional “neurotoxins” and that fraction δ contains a “cardiotoxin” which may act by displacing calcium from membranes.

Reversibility of neuromuscular blockade produced by toxins isolated from the venom of the seasnake Laticauda semifasciata

The neuromuscular blocking potencies and reversibilities of purified toxins from the venom of the seasnake Laticauda semifasciata were assessed on the chick biventer cervicis nerve-muscle preparation. All toxins inhibited the responses to indirect stimulation and to carbachol but not the responses to direct muscle stimulation or to raised KCl concentrations. Fraction 0·06 M and 0·09 M were about 100 times less potent than the most potent compound, erabutoxin b but they were similarly partially and slowly reversible. In contrast LsIII toxin was less potent than erabutoxin b but its effect could be readily reversed by washing. Since the toxins differ in chemical structure, it is suggested that separate parts of the toxin molecule may determine affinity for the acetylcholine receptor and reversibility.

Neuromuscular blocking activity of two fractions isolated from the venom of the seasnake, Laticauda semifasciata

Two components (0·06 M and 0·09 M fractions) have been isolated from the venom of the seasnake, Laticauda semifasciata. Both are similar polypeptides, each with about 100 amino acids and a mol. wt of approximately 10,000. When tested on the isolated chick biventer cervicis nerve-muscle preparation, both fractions were post-junctional cholinoceptor antagonists, although about 100 times less active than erabutoxin b from the same venom. The fractions chemically resemble venom phospholipase A, but they differed from the enzyme in some effects at the neuromuscular junction. The fractions were very slowly and only partially reversible, whereas the effects of phospholipase A were reversed rapidly on washing.

The actions of aminopyridines on avian muscle

The effects of 2-, 3-, and 4-aminopyridines were investigated on the isolated chick biventer cervicis nerve-muscle preparation and on nerve-free cell cultures of embryonic chick skeletal muscle. All 3 compounds reversed tubocurarine blockade and augmented twitch height in indirectly stimulated biventer cervicis preparations. 4-Aminopyridine was approximately 10 times more potent than 2-, or 3-aminopyridine. Twitch augmentation was also seen in directly stimulated preparations but to a much lesser extent. The compounds did not have significant anticholinesterase activity, nor did they have any depolarizing activity when tested on nerve-free cultured muscle fibres. At high concentrations the aminopyridines produced a maintained contracture in the biventer preparations which was enhanced by neostigmine and inhibited by tubocurarine. It is suggested that the aminopyridines facilitate neuromuscular transmission by increasing acetylcholine release in response to nerve stimulation, and that the compounds can also increase spontaneous transmitter release.

An analysis of the mode of action of carbachol on the chick biventer cervicis nerve-muscle preparation

The mechanism of contracture evoked by carbachol in the isolated chick biventer cervicis nerve-muscle preparation was studied. At concentrations lower than 11 μM, carbachol progressively induced contracture at a rate much slower than did acetylcholine. A spontaneous increase of the response to carbachol, but not to acetylcholine, was observed 2–4 hr after isolation of the muscle. By contrast, no change of the response occured in the denervated muscle. Anticholinesterase treatment shifted the dose-response curve for carbachol markedly to the left as far as the contracture attained after 4–6 min incubation was concerned. The shift was much less marked for the dose-response curve plotted against the initial rate of response defined as the contracture obtained after 1 min incubation. No enhancement of response was detected by inhibition of acetylcholinesterase in the denervated muscle. β-Bungarotoxin which blocked neuromuscular transmission within 30 min, caused a transient enhancement of the response to carbachol. After 2–3 hr treatment, however, the sponteneous increase of response to carbachol was counteracted by the toxin. No potentiation of the response by anticholinesterase agents was observed after toxin treatment. When added in the presence of physostigmine or echothiophate, β-bungarotoxin reduced the response to carbachol to the control level in 2 hr. The response to carbachol in the muscle treated with α-bungarotoxin and washed subsequently was also not potentiated by anticholinesterase treatment. Repetitive nerve stimulation in the presence of hemicholinium-3 caused a neuromuscular blockade but did not appreciably antagonize the response to carbachol more than that to acetylcholine either in the absence or presence of physostigmine. When calcium ion in the medium was decreased from 2.7 to 0.54 mM, the response to nerve stimulation was nearly completely inhibited, but the response to carbachol was not affected and could still be potentiated by anticholinesterase. It is concluded that carbachol has both direct and indirect effects on the chick biventer cervicis muscle. At a low concentration, particularly in the presence of an anticholinesterase and if sufficient time of incubation is allowed, the indirect effect caused by release of acetylcholine may become more prominent than the direct action on the post-synaptic receptor. Both the mechanism and the store of acetylcholine for this indirect action appear to be different from those for the nerve impulse.

Inhibition of nicotinic responses at neuromuscular junctions by 3,3-dimethyl-1-butanol carbamate (DBC)

The action of 3,3-dimethyl-1-butanol carbamate (DBC), a synthetic carbachol analogue in which the quaternary nitrogen atom has been replaced by a carbon atom, has been studied in relation to its effects on nicotinic responses. In the thick biventer cervicis nerve-muscle preparation and frog rectus abdominis muscle preparation, DBC inhibited the responses elicited by nicotine and dimethylphenylpiperazinium at concentrations that did not affect responses induced by acetylcholine. DBC ( −4–10 −3M) depressed markedly the uptake of 3H-nicotine (2 × 10 −3M) into the frog rectus abdominis muscle preparation. In the innervated chick biventer cervicis nerve-muscle preparation, DBC (10 −3M) also decreased the uptake of 3H-nicotine (2 × 10 −3M), but in denervated preparations, DBC had no effect. It is concluded that the peripheral antinicotinic actions of DBC require an intact nerve-muscle junction and are due to a blockade in the uptake of nicotine into nerve fibers.

The effects of 2-(4-phenylpiperidino)-cyclohexanol (AH5183) on the acetylcholine content of, and output from, the chick biventer cervicis muscle preparation.

The effects of the tertiary base 2-(4-phenylpiperidino)-cyclohexanol (AH5183) were studied on the acetylcholine content of, and output from, the chick biventer cervicis nerve-muscle preparation. Neither treatment with AH5183 (4 μg/ml) alone, nor continued rapid nerve stimulation alone, produced significant changes in the acetylcholine content of the preparations. However, after ninety minutes rapid nerve stimulation in the presence of AH5183 the acetylcholine content of the preparations was significantly reduced. The output of acetylcholine from the nerve-muscle preparations was markedly reduced after two and three hours nerve stimulation in the presence of AH5183. The results provide further evidence that AH5183 possesses a potent prejunctional action at the neuromuscular junction. The potential usefulness of AH5183 in producing acute chemical denervation is indicated.

Cholinolytic effects of 3,3-dimethylbutanol carbamate (DBC)

3,3-Dimethylbutanol carbamate (DBC) was shown to block cholinergic responses of acetylcholine (ACh) on frog rectus abdominis muscle, guinea pig vas deferens and guinea pig ileum preparations. The blockade was reversed by washing and was non-competitive in nature. However, DBC did not block ACh responses on chick biventer cervicis preparation with concentrations up to 5.5 × 10 −3 M. In lower concentrations, DBC blocked neuromuscular transmission in the chick biventer cervicis nerve-muscle preparation, but in contrast to d-tubocurarine, it preserved completely the muscular responses to exogenously applied ACh. The neuroeffector transmission in the guinea pig nerve ileum preparation was also blocked by DBC. However, in contrast to atropine, which totally abolished responses to exogenous ACh during neuroeffector blockade, DBC preserved somewhat diminished responses to exogenous ACh. DBC (7 × 10 −6 M − 7 × 10 −5 M) incubated with minced guinea pig cerebral cortex caused a marked release of ACh. Since hemicholinium, hexamecholinium and tetrodotoxin inhibit ACh release from nerve endings, DBC does not appear to work like these compounds either.

The interaction of diazepam with myoneural blocking agents.

The interaction of diazepam with neuromuscular blocking agents was studied in patients and in several animal nerve-muscle preparations. This drug did not alter the recovery slope of blockade produced by d-tubocurarine, decamethonium, or gallamine. In higher doses diazepam produced a decrease in muscle contractions and blocked the actions of exogenously applied nicotine and acetylcholine in the superfused chick biventer cervicis nerve—muscle preparation. In low doses the commercially available drug given intra-arterially reversed the myoneural blockades produced by both depolarizing and nondepolarizing blockers. However, this property was due to the solvent system of the drug.

Neuromuscular pharmacology of oxotremorine

Oxotremorine caused contracture of rat phrenic nerve-diaphragm followed at high doses by blockade. Similar results were obtained on the sciatic nerve-tibialis anterior muscle preparation of the cat. The slowly developing spastic paralysis obtained with high doses was long lasting. Qualitatively similar results but of lower intensity were observed on chick biventer-cervicis nerve-muscle preparation. There was no effect on frog rectus abdominis preparation. On rat diaphragm contracture and paralysis were potentiated by halving the calcium chloride concentration and by physostigmine respectively. Depolarising effect was further proven by potentiation of reduction in the force and duration of tetanic response by physostigmine on rat diaphragm. An indirect mechanism involving release of acetylcholine is postulated.