Effects of calcium channel blocking agents on neostigmine-induced fasciculations

Male Sprague-Dawley rats were anesthetized with pentobarbital and prepared for monitoring contractions of the gastrocnemius muscle evoked by stimulation of the sciatic nerve. Animals received atropine prior to a dose of neostigmine of 0.02 mg/kg i.v. The effects on contractile strength and the number of fasciculations in a 2-min period were assessed. Pretreatment with phenytoin, 20 mg/kg, reduced the number of fasciculations to 32% of control without altering contractile strength. Both nifedipine and nitrendipine, 1 mg/kg each, virtually abolished fasciculations without altering twitch strength. Verapamil, 4 and 8 mg/kg, depressed fasciculation frequency to 50% of control without affecting pre-neostigmine twitch height. The dihydropyridine calcium blocking agents did however reduce the neostigmine-induced augmentation of contraction strength. These data suggest that a calcium-mediated current at presynaptic motor endings participates in the generation of repetitive nerve terminal discharges leading to muscle fasciculations.     

The neuromuscular blocking activity of isepamicin sulfate (HAPA-B) compared with other aminoglycoside antibiotics

Effects of isepamicin sulfate (HAPA-B), a new aminoglycoside antibiotic, on the neuromuscular transmission were studied in rats and compared with those of amikacin (AMK) or other aminoglycoside antibiotics. The HAPA-B, as well as other aminoglycoside antibiotics, depressed the twitch response of diaphragm to phrenic nerve stimulation in vitro. The depression effects of different drugs were compared and graded in the order of strengths of blocking action as: netilmicin (NTL) greater than gentamicin (GM) greater than streptomycin (SM) greater than kanamycin (KM) greater than AMK greater than HAPA-B. The IC50 (concentration which inhibited the response by 50%) of HAPA-B was 3.6 X 10(-3) g/ml. The neuromuscular blockade produced by HAPA-B was reversed by CaCl2, KCl or caffeine but not by neostigmine. D-Tubocurarine or MgCl2 augmented the neuromuscular effects of HAPA-B. Intramuscular (400 mg/kg) and intravenous (100 mg/kg) injections of HAPA-B did not affect the twitch response of gastrocnemius muscle to sciatic nerve stimulation in situ. Intravenous injection of 200 mg/kg caused death in some rats and depression of the twitch response in others. Intravenous AMK produced no significant effect on the twitch response at 50 mg/kg and caused death at 100 mg/kg. GM and SM caused death or significant degree of depression of the twitch response at intravenous doses of 50 mg/kg. In experiments of intravenous drug infusion for 60 minutes, the twitch response was depressed by HAPA-B at 400 mg/kg/hr and by AMK at 200 and 400 mg/kg/hr. In conclusion, HAPA-B has a neuromuscular blocking action presumably at the nerve terminal. However, its action was the weakest among the aminoglycoside antibiotics tested.     

Assessment of neuromuscular blockade produced by atracurium in frog sartorius muscle

The neuromuscular effects of atracurium in frog sartorius muscle were studied and the results were compared with those obtained by tubocurarine in the same preparation. Single twitch (at 1 per 5 s) and tetanic stimulation (at 1-100 per s) of the motor nerve, stimulated with 5-10 V (maximum) and 0.2 ms pulse duration, were used to assess the neuromuscular blockade produced by atracurium and tubocurarine in the frog sartorius muscle. The results showed that atracurium was twice as potent as tubocurarine in reducing the amplitude of the indirectly-elicited twitch contractions in the frog sartorius muscle. The mean IC50 values (concentration to produce 50% inhibition of twitch tension) of atracurium and tubocurarine-induced depression of the indirectly-elicited twitch tension were 0.64 +/- 0.1 microM and 1.2 +/- 0.2 microM respectively (means +/- s.e., n = 6, P less than 0.001). Furthermore, atracurium had a shorter time of onset, time to 50% block and time to 95% recovery than tubocurarine.     

Contribution of prostaglandins to the activity of guinea-pig phrenic and chicken esophageal parasympathetic nerves

In order to clarify the role of prostaglandins (PGs) in the activity of cholinergic neurones other than the ileal myenteric plexus, the effects of indomethacin (IND) and PGE2 on the contractile responses and the release of acetylcholine (ACh) induced by electrical stimulation were investigated in isolated guinea-pig phrenic nerve-diaphragm and chicken parasympathetically innervated oesophagus preparations. In the guinea-pig phrenic nerve-diaphragm preparations, IND at 56 microM did not affect the twitch responses induced by direct or indirect electrical stimulation. PGE2 at 1 microgram/ml augmented the twitch responses induced by direct or indirect stimulation of submaximal, but not of supramaximal intensity. The amounts of ACh released from the phrenic nerve by electrical stimulation were unaffected by IND (56 microM). PGE2 (0.01-1 microgram/ml) inhibited the ACh release by the nerve stimulation of high frequency (50 Hz) in a concentration-dependent manner. The inhibitory effect of PGE2 was less clear on the ACh release by lower frequency (1 Hz). Neither IND nor PGE2 affected the ACh release induced by 40 mM-K+. In the chicken parasympathetically innervated oesophagus preparation, IND (2.8-5.6 5.6 microM) augmented the twitch responses induced by the nerve stimulation at a frequency of 0.017 Hz, but not those produced by train pulse (5 sec at a frequency of 1 or 10 Hz). This augmentation was reversed by the application of PGE2 at 10 ng/ml. PGE2 at 10 ng/ml produced a transient inhibition of the twitch responses induced by 0.017 Hz or train pulses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Involvement of kappa-opioid receptors in peripheral response to nerve stimulation in kappa-opioid receptor knockout mice

1 The present study aimed to evaluate the role of kappa-opioid receptors at two peripheral sites, the vas deferens and the proximal colon, in kappa-opioid receptor knockout mice. We investigated the role of the kappa-opioid receptor in the vas deferens twitch response and in the colonic "off-contraction", a rebound contractile response which follows the inhibitory response to low frequencies stimulation (10, 20, 30 Hz) and which has been suggested to "locally" reproduce the contractile component of the peristaltic reflex. 2 Transmural stimulation of the vas deferens at lower frequencies (10 Hz, 10 V, 1 ms pulse trains lasting 0.5 s) evoked a contractile response that was significantly higher in the preparations from knockout mice because of lack of kappa-opioid receptors than in wild type mice. A selective kappa-opioid receptor agonist, U-50,488H, induced a dose-dependent inhibition of the electrically stimulated contraction in vas deferens. The percentages of reduction of the twitch response were significantly lower in knockout mice than in wild type mice after treatment with U-50,488H. The reduction of twitch response caused by U-50,488H was not reversed by administration of nor-binaltorphimine (nor-BNI) (5 x 10-6 m), a selective kappa-opioid receptor antagonist, in preparations from both knockout mice and wild type mice. U-50,488H has no effect on postsynaptic adrenergic receptors, as its administration did not affect the direct contractile response to noradrenaline. 3 Transmural stimulation (5 Hz, 20 V, 2 ms pulse trains lasting 30 s) induced inhibition of spontaneous activity of colonic strips during the period of stimulation, followed by an "off-contraction" after the cessation of stimulation. The statistical evaluation of the "off-contraction" responses between the two strains showed no significant difference. The off-contraction, measured in specimens from knockout mice, was inhibited concentration-dependently by U-50,488H (P < 0.01) and significantly less than from wild type mice. 4 The effect of U-50,488H was not reversed by administration of nor-BNI (5 x 10-6 m), either in preparations from knockout mice or from wild type mice. 5 Our data may suggest that kappa-opioid receptors are involved in some peripheral responses to the nerve stimulation, as indicated by the effect of U-50,488H, a selective kappa-opioid receptor agonist. However, the involvement of kappa-opioid receptor was also present, although less apparent, in kappa -opioid receptor knockout mice, suggesting either that this drug acts not only on kappa-opioid receptors but also on other receptor sites, such as kappa-like receptors. An alternative interpretation can be related to a sodium channel blocking action of U-50,488H, which could explain the inhibitory effects of twitch response still present but less evident in knockout strain and the lack of effect of the antagonist nor-BNI.     

吡喹酮对神经肌肉接头传递的作用

吡喹酮可使电刺激腓总神经所引起的胫前肌收缩反应明显加强。如给腓总神经施加超强刺激,静脉注射筒箭毒碱可减弱肌肉收缩反应,吡喹酮却能加强筒箭毒碱的抑制作用,使神经肌肉传递迅即完全阻断。采用微电极细胞内记录技术研究吡喹酮对离体大鼠膈神经隔肌的作用,发现吡喹酮不明显影响膈肌细胞的静息膜电位。低浓度吡喹酮(2×10^-4M)可使微终板电位(mepp)频率明显增加。高浓度吡喹酮8×10^-4M则使mepp振幅逐渐变小和消失。吡喹酮在使神经肌肉传递阻滞不断加深的同时,尚能使终板电位的振幅变小,终致完全消失。     

Effects of piperine on the motility of the isolated guinea-pig ileum: comparison with capsaicin.

Piperine (1 microM), a congener of capsaicin, produced an initial contraction blocked the capsaicin-sensitive contractile response to mesenteric nerve stimulation and inhibited the twitch response induced by field stimulation in the isolated guinea-pig ileum. These three effects of piperine (1 microM) were rapidly desensitized and significantly antagonized by ruthenium red (0.5-1 microM), an inorganic dye known to antagonize the effects of capsaicin. The contractile effect of piperine was abolished by application of tetrodotoxin plus desensitization with substance P or by extrinsic denervation. The inhibitory effect of piperine (1 microM) on the twitch response was antagonized by desensitization with calcitonin gene-related peptide (CGRP). Moreover, cross-tachyphylaxis between piperine and capsaicin was observed, suggesting that a similar mechanism may be involved in the effects of these agents. The contractile effects induced by piperine (10 microM) and the subsequent inhibitory effects on the twitch response were not desensitized and largely persisted after extrinsic denervation. The contractile effects of piperine (10 microM) were not strongly inhibited by tetrodotoxin plus desensitization with substance P. It was concluded that the lower concentration of piperine caused contraction and inhibited the twitch responses by releasing substance P and CGRP, respectively, from sensory nerves, and blocked the response to mesenteric nerve stimulation by a mechanism similar to that of capsaicin. At higher concentrations, piperine had non-specific direct actions on the smooth muscle.     

Neuromuscular blocking profile of the vecuronium analogue, Org-9487, in the rat isolated hemidiaphragm preparation.

1. The neuromuscular effects of the short-acting aminosteroid muscle relaxant Org-9487 have been studied in the in vitro rat phrenic nerve/hemidiaphragm preparation by use of twitch tension and electrophysiological recording techniques. 2. Org-9487 (5-100 microM) produced a concentration-dependent decrease in the amplitude of twitches (0.1 Hz) and tetanic contractions (50 Hz) evoked by motor nerve stimulation. The compound produced fade of force during both 50 Hz stimulation and train-of-four stimulation at 2 Hz, indicating a prejunctional component of action. 3. Anticholinesterases only partially reversed the effect of Org-9487 on twitch responses. This was possibly because, at the concentrations required to block twitches in the rat, Org-9487 itself was found to possess significant anticholinesterase activity. 4. Org-9487 (3 microM) increased the rundown of endplate current amplitudes during a 2 s train of 50 Hz nerve stimulation. This was because Org-9487 increased the quantal content of the first endplate current in the train without affecting acetylcholine release towards the latter part of the train. 5. Org-9487 (10 microM) produced a voltage-dependent decrease in the time constant of decay of endplate currents at 32 degrees C and 0.5 Hz, indicative of a block of endplate ion channels. The blocking rate constant increased with membrane hyperpolarization.     

The marine polyether gambierol enhances muscle contraction and blocks a transient K current in skeletal muscle cells

Gambierol is a complex marine toxin first isolated with ciguatoxins from cell cultures of the toxic dinoflagellate Gambierdiscus toxicus. Despite the chemical complexity of the polycyclic ether toxin, the total successful synthesis of gambierol has been achieved by different chemical strategies. In the present work the effects of synthetic gambierol on mouse and frog skeletal neuromuscular preparations and Xenopus skeletal myocytes have been studied. Gambierol (0.1-5 μM) significantly increased isometric twitch tension in neuromuscular preparations stimulated through the motor nerve. Less twitch augmentation was observed in directly stimulated muscles when comparing twitch tension-time integrals obtained by nerve stimulation. Also, gambierol induced small spontaneous muscle contraction originating from presynaptic activity that was completely inhibited by d-tubocurarine. Gambierol slowed the rate of muscle action potential repolarization, triggered spontaneous and/or repetitive action potentials, and neither affected action potential amplitude nor overshoot in skeletal muscle fibers. These results suggest that gambierol through an action on voltage-gated K⁺ channels prolongs the duration of action potentials, enhances the extent and time course of Ca²⁺ release from the sarcoplasmic reticulum, and increases twitch tension generation. Further evidence is provided that gambierol at sub-micromolar concentrations blocks a fast inactivating outward K⁺ current that is responsible for action potential prolongation in Xenopus skeletal myocytes.     

Inhibition of neuromuscular transmission in isolated mouse phrenic nerve-diaphragm by the enterotoxin of Clostridium perfringens type A.

The enterotoxin of Clostridium perfringens type A, a channel forming protein toxin, inhibited neuromuscular transmission under conditions of low calcium. Twitch tension of isolated phrenic nerve-diaphragm preparations elicited by electrical stimulations to the phrenic nerve was recorded isometrically, and the preparations were exposed to the purified enterotoxin. In Krebs solution containing 0.5 mM calcium, the enterotoxin (20 micrograms/ml) reduced within 10 min the amplitude of the twitch tension to 34 +/- 7% (mean +/- S.D., n = 11) of that recorded before the treatment. The effects of the enterotoxin on the twitch tension were irreversible and proceeded independently of stimulation. The reduction of the twitch tension by the enterotoxin was apparent in Krebs solution containing less than 0.6 mM calcium and the degree of reduction was inversely related to the concentration of calcium. The reduction of the twitch tension by the enterotoxin was also dependent on temperature and concentration of the toxin. At temperatures below 20 degrees C, no obvious reduction of twitch tension was observed with 20 micrograms/ml of the enterotoxin. Enterotoxin at a concentration of 0.4 micrograms/ml caused 16 +/- 2% (mean +/- S.D., n = 4) reduction of twitch tension, and the degree of the reduction in twitch tension increased with toxin concentration, reaching a plateau of 65 +/- 4% (mean +/- S.D., n = 7) at 6.5 micrograms/ml of the enterotoxin. The effects of the enterotoxin were antagonized by 2 microM physostigmine. Unlike curare, pretreatment of the preparation with enterotoxin did not antagonize the neuromuscular block by decamethonium. Neither the tension of muscular twitch elicited by direct electrical stimulation to the muscle nor the resting membrane potentials of muscle fibers recorded intracellularly were affected by the enterotoxin. The enterotoxin (2.2 micrograms/ml) reduced the frequency, but not mean amplitude or amplitude distribution, of miniature end-plate potentials, from 0.91 +/- 0.07/sec to 0.72 +/- 0.07 (mean +/- S.E., n = 5). The results suggest that the enterotoxin will provide a novel tool for the studies on the mechanism of the neuromuscular transmission because of the unique characteristics of the inhibition and of the known mechanism of its action on the cell membrane.     

The action of dantrolene on transmitter mobilization at the rat neuromuscular junction

Dantrolene sodium (20 microM) was found to decrease transmitter mobilization and the apparent available store of acetylcholine at frequencies of nerve stimulation of 50 and 100 Hz at the neuromuscular junction of the rat hemidiaphragm preparation treated with 2 microM (+)-tubocurarine. This effect of dantrolene sodium was not as marked at frequencies of nerve stimulation of 25 Hz or less, also no significant effect of the drug was observed on the amplitude of endplate potentials (EPPS) at any frequency of nerve stimulation. No effect of dantrolene sodium (20 microM) on mean miniature EPP amplitude or frequency was observed. It is suggested that the action of dantrolene sodium on mean EPP quantal content may be due to an action on stores of bound calcium within the motor nerve terminal. This effect of the drug is unlikely to be of physiological consequence in vivo, since it was only observed at high frequencies of nerve stimulation and did not cause a significant reduction of EPP amplitude during trains of stimuli.     

The action of dantrolene sodium on individual fast and slow motor units of the rat anaesthetized with urethane

Rats, anaesthetized with urethane, were injected intravenously with dantrolene sodium in a vehicle of 5% mannitol taken to pH 10 with NaOH. The muscle relaxant action of dantrolene sodium was measured from the contractions of individual motor units of the extensor digitorum longus (EDL), soleus (SOL) and segmental tail (ST) muscles. Data were also collected from their parent whole muscle preparations. The depressant action of dantrolene sodium on the percentage-normalized amplitude of contraction of the individual motor units was greater than its effect on the whole muscle twitch amplitude, in all three muscles. The twitch amplitude of fast contracting motor units was significantly more reduced (P less than 0.001) by dantrolene sodium than was that of slow contracting motor units. Dantrolene sodium reduced the contraction time of all motor units. The effect of the drug on half-relaxation time varied within and between groups of motor units studied. The drug was confirmed to have a greater depressant action on the twitch contraction than on the fused tetanus of whole muscle. This was true also for single motor units. With tetanic stimulation the effect of dantrolene sodium was also dependent upon the motor unit type, fast or slow. A maximum depression of contractile tension occurred at a stimulation frequency of 64 Hz for fast EDL motor units whereas the maximum depression for ST slow units, the slowest units tested, was at a stimulation frequency of 14 Hz.     

Pharmacological characterisation of the presynaptic alpha-adrenoceptor in the rat vas deferens.

The interactions between alpha-adrenoceptor agonists and antagonists on the twitch response of the rat isolated vas deferens to low frequency motor nerve stimulatioh have been examined. Oxymetazoline, clonidine, naphazoline and BAY-1470 caused a concentration-dependent inhibition of the twitch response at concentrations lower than those causing smooth muscle stimulation. The twitch-inhibitory effect of these compounds is thought to result from stimulation of presynaptically located alpha-adrenoceptors. In contrast, phenylephrine and methoxamine exerted little inhibitory effect at concentrations less than those which produced postsynaptic stimulation. The inhibitory effect of clonidine was antagonised by phentolamine, piperoxan, yohimbine and tolazoline, but not by thymoxamine. These results characterise the presynaptic alpha-adrenoceptors in the rat vas deferens as being of the same type as those present in the rabbit pulmonary artery and rat heart, but different from those located post-synaptically in sympathetically innervated tissues.     

Pharmacological studies on surugatoxin, the toxic principle from Japanese ivory mollusc (Babylonia japonica).

1 Some pharmacological properties of surugatoxin (SGTX), a purified toxic substance from the Japanese ivory mollusc (Babylonia japonica), have been investigated. SGTX (50 nmol/kg i.v.) produced a prolonged fall of blood pressure in anaesthetized cats. This hypotensive effect was neither blocked by atropine and propranolol nor by spinal cord transection. 2 SGTX (37-50 nmol/kg i.v.) inhibited the hypertensive and hypotensive response to 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP) and to electrical stimulation of the splanchnic and vagal nerve, whereas it usually augmented the hypertensive response to adrenaline and to 4-(m-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium chloride (McN-A-343) in anaesthetized cats. 3 Close intra-arterial injection of SGTX (6.2-12.3 nmol/kg) to the superior cervical ganglion blocked the contractile response of the nictitating membrane to preganglionic stimulation of cervical sympathetic nerve or injected DMPP, but not to postganglionic stimulation or to injected adrenaline and McN-A-343. 4 SGTX affected neither the indirectly nor the directly stimulated response of the rat isolated phrenic nerve-diaphragm at concentrations less than 12.3 mum. 5 The effect of SGTX on the contractile response to some agonists and on the twitch response to transmural stimulation in the guinea-pig isolated ileum was investigated. At less than 12.3 mumSGTX did not depress responses to acetylcholine or histamine. The curves for nicotine- and DMPP-induced contractions were shifted to the right and depressed gradually as the concentration of SGTX was increased (12.3 nm-1.23 mum). SGTX partially inhibited the contraction induced by 5-hyroxytryptamine and the transmurally-stimulated twitch response. 6 These results suggest that SGTX has a ganglion-blocking action. The mode of anti-nicotinic action of SGTX in the guinea-pig isolated ileum seems to differ from that of hexamethonium and tetraethylammonium and to resemble more closely that of mecamylamine.     

Studies on the repetitive discharges evoked in motor nerve and skeletal muscle after injection of anticholinesterase drugs

Repetitive discharges recorded from the ventral root and from the gastrocnemius muscle in response to single motor nerve shocks applied close to the muscle after injection of edrophonium, neostigmine or ambenonium were studied in cats anaesthetized with chloralose. Two closely spaced volleys with an interval of 1 to 5 msec between them produced more repetitive firing than did a single shock. With longer intervals, the repetitive firing was not potentiated by the second volley. All frequencies of tetanic stimulation depressed the repetitive firing and, for successive stimuli to produce a degree of repetitive firing equivalent to the first, it was necessary to stimulate at frequencies below 2 shocks/sec. With stimulation frequencies higher than 100 shocks/sec, repetitive firing did not occur unless the duration of the tetanus was shorter than about 30 msec when slight repetition followed the last stimulus of the train. With stimulation frequencies of 100 down to 20 shocks/sec, repetitive firing was produced only by the first volley of the tetanus. Subsequent nerve action potentials of the tetanus occurring during the repetitive firing in the nerve following the first volley were partially extinguished by collision with the back discharge. This effect contributed to the waning tetanus, which is characteristic of treatment with an anticholinesterase, but the main depression of tetanic contractions appeared to be a consequence of depolarization block through accumulating acetylcholine. Tubocurarine and benzoquinonium reversed the initial “extinction” phase of the depressed tetani by abolishing the repetitive discharge in the nerve and in larger doses reversed the secondary depressant phase presumably by reducing the excessive end-plate depolarization. The results are discussed in relation to the hypothesis that anticholinesterases may effect transmission by acting at three sites at the neuromuscular junction—on acetylcholinesterase, at the motor nerve ending and at the motor end-plate—and that reaction at any one site may be augmented by the production of reverberating activity across the junction.     

The role of prostaglandins in cholinergic neurotransmission in the guinea pig.

Prostaglandins have contrasting effects on neurotransmission at different cholinergic nerve endings. This is a report on the role of prostaglandins in a number of cholinergic preparations from the guinea pig. In the isolated ileum PGE1 (2 X 10(-10) to 5 X 10(-8) M) potentiated the response to electrical stimulation of the cholinergic nerves. PGE1 (10(-7 M) caused an increase in tone followed by a period of transient inhibition of twitch height. Responses to simulation of the ileum with drugs were not potentiated by PGE1. Responses of atropinized or plexus-free muscle to electrical stimulation were also not potentiated by PGE1. Acetylsalicylic acid (2.5 X 10(-4) M) diminished the twitch response and the output of acetylcholine from the ileum. Both effects were reversed by PGE1. Qualitatively similar observations were made on the trachea. It is concluded that prostaglandins facilitate acetylcholine release in the ileum and trachea. PGE1 diminished the effect of vagal stimulation on the heart rate. The response to stimulation of the phrenic nerve was not affected.     

Improving Pharmacokinetic–Pharmacodynamic Models of Muscle Relaxants Using Potentiation Modelling

Repeated motor nerve stimulation performed during neuromuscular monitoring enhances the evoked mechanical response of the corresponding muscle resulting in an increased twitch response. This is known as twitch potentiation or the staircase phenomenon. For neuromuscular modelling research twitch stabilisation techniques are often used to reduce the visible effect of potentiation, but such techniques are not always effective. Our objective was to model pharmacokinetic-pharmacodynamic (PK–PD) and twitch potentiation and to estimate neuromuscular block (NMB) in the presence of twitch potentiation. We combined a standard PK–PD model with a model describing the degree of twitch potentiation. The combined model was used to predict mechanomyographic twitch measurements and estimate NMB and twitch potentiation during muscle relaxation monitoring. Model parameters and prediction accuracy were compared to the standard PK–PD model with and without linear baseline correction. The PK–PD-potentiation model allows NMB to be estimated in the presence of twitch potentiation. It also accurately predicts data from twitch stabilisation, which is ignored with the standard PK–PD model. Compared to the standard PK–PD model, estimated PD parameters ec50 and gamma were found to be higher using the PK–PD-potentiation model. Compared to linear baseline correction, estimated PD parameters ke0 and ec50 were found to be higher. A PK–PD-potentiation model can estimate the degree of twitch potentiation and the degree of NMB during neuromuscular monitoring. This model leads to different PD parameter estimations than the standard PK–PD model however the differences are small enough to be unlikely to cause great concern among researchers.     

金环蛇(Bungarus fasciatus)蛇毒的分离及其毒性组分的药理研究

金环蛇毒用羧甲基纤维素柱层析分离出17个蛋白组分,其中5个组分是毒性较大的致死成分。在这5个组分中,Ⅺ、Ⅻ、ⅩⅤ是心脏毒,ⅩⅢ、ⅩⅣ是神经毒。心脏毒组分Ⅺ、ⅫⅩⅤ使离体大鼠心脏挛缩,心跳停止于收缩期;使小鸡离体颈二腹肌挛缩,最后致痉挛性麻痹;对兔眼结合膜有局部刺激作用,使结合膜充血水肿。组分ⅩⅤ还具有直接溶血作用。神经毒组分ⅩⅢ阻断大鼠、小鸡、青蛙神经肌肉标本的突触传递,标本对乙酰胆碱的反应消失,对直接刺激以及对高浓度氯化钾的反应无减少,神经末梢乙酰胆碱的释放最无显著改变;使蛙腹直肌乙酰胆碱量效曲线平行右移,新斯的明可以对抗这个作用。组分ⅩⅣ的作用与组分ⅩⅢ相似。实验提示,组分ⅩⅢ、ⅩⅣ是作用于终板后膜的神经毒。     

Actions of the selective inhibitor of cholinesterase tetramonoisopropyl pyrophosphortetramide on the rat phrenic nerve-diaphragm preparation

1. Tetramonoisopropyl pyrophosphortetramide (iso-OMPA) added for 15 min to the rat isolated phrenic nerve-diaphragm in a concentration of 30 muM, produced a complete selective and stable inhibition of cholinesterase. A concentration of 3 muM produced near complete inhibition of cholinesterase, and a concentration of 300 muM also inhibited acetylcholinesterase marginally.2. Inhibition of cholinesterase was associated with a sustained increase in the neuromuscular blocking action of exogenous butyrylcholine but not of exogenous acetylcholine. Iso-OMPA, 300 muM, in addition caused transient increases in the sensitivity of the rat diaphragm to exogenous acetylcholine and butyrylcholine. In the same concentration, it had a curare-like action on the frog rectus abdominis muscle preparation.3. Iso-OMPA, 30 muM, caused reversible increases in the amplitude of the twitch response and tetanic responses, which were of a similar magnitude in the indirectly stimulated preparation and the directly stimulated curarized preparation. Caffeine had a similar effect on the twitch response and its effectiveness was increased by iso-OMPA, and vice-versa. Amongst anticholinesterases, octamethyl pyrophosphortetramide and tetraethylpyrophosphate also enhanced the amplitude of the tetanic response, but paraoxon, dyflos, and mipafox did not.4. It is concluded that iso-OMPA, in concentrations (3 and 30 muM) which in 15 min give near maximal or maximal selective inhibition of cholinesterase, has no effect on the transmission of nerve impulses at the neuromuscular junction, but enhances reversibly the amplitude of the contractile response to stimulation by a direct action upon the muscle fibre, which involves a mechanism related to but not identical with that by which caffeine potentiates twitch tension. In higher concentrations, iso-OMPA has a curare-like action at the neuromuscular junction.     

The contractile response to and the release of noradrenaline by transmural nerve stimulation in the guinea-pig vas deferens and a comparison with the response to noradrenaline.

The release of noradrenaline from the guinea-pig vas deferens by transmural stimulation was measured electrochemically after separation by h.p.l.c. Variations in release with different durations of stimulation were then compared with the mechanical response during stimulation. During long periods of 270 s of stimulation at 10 Hz the average release of noradrenaline in the final 250 s remained at about 50% of that released during the first 20 s yet the mechanical response throughout the later time declined to near zero. Superfusion with constant concentrations of noradrenaline produced mechanical responses similar to those from transmural nerve stimulation, an initial rapid contraction declining to a low level of maintained activity. During the later period, when the response to infused noradrenaline was low, the twitch response to transmural stimulation was potentiated. Phentolamine 10(-6)M reduced the potentiation of the twitch response to transmural stimulation by noradrenaline and propranolol 10(-6)M enhanced the potentiation. These results suggest that the decline in the adrenergic component of the response to prolonged transmural nerve stimulation is not the result of a decline in noradrenaline release but mainly due to desensitization to the contractile effect of noradrenaline. Since the similar desensitization produced by infused noradrenaline does not reduce the twitch response to transmural nerve stimulation it is unlikely that the transmitter responsible for that component is noradrenaline.