Studies on the efficacy and depressant potential of muscle relaxants in mice

The present study was undertaken to confirm and extend previous observations on the use of morphine-induced Straub tail and rotarod performance in mice as a means of assessing the efficacy of a compound as a muscle relaxant and its potential for depressant side-effects, respectively. The ratio of the rotarod to Straub tail ED₅₀s was calculated as an index of specificity. Our results generally confirmed previous reports in the case of diazepam, chlordiazepoxide and dantrolene. Additionally, the recently introduced muscle relaxants, cyclobenzaprine, baclofen, and DS 103-282 were found effective in blocking Straub tail and their reported clinical specificity was reflected by the specificity observed in the mouse tests. DS 103–282 was found to be the most potent and specific. It is concluded that muscle relaxants with specificity ratios significantly greater than 1.0 appear to have comparatively few depressant effects at therapeutic doses. The relatively simple methods used in the present study appear to be useful in the initial assessment of compounds as potential muscle relaxants.     

Potency of nondepolarizing muscle relaxants on muscle-type acetylcholine receptors in denervated mouse skeletal muscle

Aim:

To investigate the changing resistance to nondepolarizing muscle relaxants (NDMRs) during the first month after denervation.

Methods:

The denervated and innervated skeletal muscle cells were examined on days 1, 4, 7, 14, 21, and 28 after denervation. Individual denervated and innervated cells were prepared from the flexor digitorum brevis of the surgically denervated and contralateral hind feet, respectively. Nicotinic acetylcholine receptors (nAChRs) in the cells were activated with 30 μmol/L acetylcholine, either alone or in combination with various concentrations of vecuronium. Currents were recorded using a whole-cell patch-clamp technique.

Results:

The concentrations of vecuronium resulting in half-maximal inhibitory responses (IC₅₀) increased 1.2- (P>0.05), 1.7-, 3.7-, 2.5-, 1.9-, and 1.8-fold (P<0.05) at Days 1, 4, 7, 14, 21, and 28 after denervation, respectively, compared to the innervated control. Resistance to vecuronium appeared at Day 4, peaked at Day 7, and declined at Day 14 after denervation. Nevertheless, IC₅₀ values at Day 28 remained significantly higher than those for the innervated control, suggesting that the resistance to vecuronium had not disappeared at Day 28.

Conclusion:

The NDMR doses required to achieve satisfactory clinical effects differ at different times after muscle denervation.     

Microcomputer use in measuring onset,duration, and recovery from nondepolarizing skeletal muscle relaxants in rabbits

This report describes a method which has been developed to use an Apple II Plus microcomputer in studies of the time course and dose response of potential skeletal muscle relaxant drugs. Rabbits are anesthetized with pentobarbital, tracheotomized, and the right common carotid artery is cannulated for direct measurement of blood pressure and heart rate. The ulnar nerve is stimulated at the elbow. Twitch, “train-of-four” (T₄), tetanus, and posttetanic potentiation (PTP) are monitored by a force displacement transducer. These analog physiological signals are led from a polygraph through an lsaac Cyborg analog-to-digital converter attached to the microcomputer where they are stored in digital form. Each animal receives the test drug twice. Spontaneous recovery is allowed following the first administration while the second dose is reversed with neostigmine. The rate of onset, time and magnitude of maximal effect, time and rate of recovery, and time and magnitude of cardiovascular changes are all automatically calculated for both the control and the neostigmine reversal experimental phases. The effect of the test dose on each parameter is rated to provide a composite score for each dose of the test compound as the dose-response series is completed. All parameters are plotted using a dot-matrix printer and data are stored to disk. When doses, in 0.1-log intervals, from the dose which produces no measurable effect to those doses producing unacceptably long durations, have been administrated, the computer uses the composite scores to report an optimal dose. Using regression analysis, the dose required to provide a 90% depression of twitch is calculated. This calculated dose is compared to equiefficacious doses of other test compounds. A tabular, graphic, and narrative report of the drug's effects is generated by the computer with comparison to other compounds of pctential interest. These programs and methods provide efficient data collection and data analysis. They provide for standard report generation. Finally, they allow for comparisons to be made among many pharmacologic parameters of many test compounds, thus providing understanding of the mechanisms of action of the test compounds.     

Different magnitude of resistance to nondepolarizing muscle relaxants in the denervated mouse skeletal muscle

Aim:

To test the hypothesis that different magnitude of resistance of denervated skeletal muscle to nondepolarizing muscle relaxants (NDMRs) is related to their varying potencies at ɛ-AChR and γ-AChR.

Methods:

Both innervated and denervated mouse muscle cells, and human embryonic kidney 293 (HEK293) cells expressing ɛ-AChR or γ-AChR were used. The effects of NDMRs on nAChR were explored using whole-cell patch clamp technique.

Results:

NDMRs vecuronium (VEC), atracurium (ATR) and rocuronium (ROC) produced reversible, dose-dependent inhibition on the currents induced by 30 μmol/L acetylcholine both in innervated and denervated skeletal muscle cells. Compared to those obtained in innervated skeletal muscle cells, denervation shifted the concentration-response curves rightward and significantly increased the 50% inhibitory concentration (IC₅₀) values (VEC: from 11.2 to 39.2 nmol/L, P<0.01; ATR: from 24.4 to 129.0 nmol/L, P<0.01; ROC: from 37.9 to 101.4 nmol/L, P<0.01). In HEK293 cell expression system, ATR was less potent at γ-AChR than ɛ-AChR (IC₅₀ values: 35.9 vs 22.3 nmol/L, P<0.01), VEC was equipotent at both receptor subtypes (IC₅₀ values: 9.9 vs 10.2 nmol/L, P>0.05), while ROC was more potent at γ-AChR than ɛ-AChR (IC₅₀ values: 22.3 vs 33.5 nmol/L, P<0.05).

Conclusion:

Magnitude differences of resistance to different NDMRs caused by denervation are associated with distinct potencies of NDMRs at nAChR subtypes.     

5-HT3 receptor antagonists inhibit the response of kappa opioid receptors in the morphine-induced Straub tail [corrected

We used the morphine-induced Straub tail to examine the actions of a 5-HT₃ receptor antagonist and κ opioid receptor agonist. The κ opioid receptor agonist, U-50,488H (4–16 mg/kg i.p.), produced a dose-related inhibition of the tail elevation induced by morphine (10 mg/kg s.c.) in ICR male mice. ICS-205-930 (3 and 10 mg/kg i.p.) and zacopride (0.3 and 1 mg/kg i.p.), 5-HT₃ receptor antagonists, attenuated the inhibitory effect of U-50,488H in a dose-dependent manner. ICS-205-930 and zacopride did not inhibit the morphine-induced Straub tail. These observations suggest that the actions of κ opioid receptors may be modulated by 5-HT³ receptors in the morphine-induced Straub tail.     

5-HT3 receptor antagonists inhibit the response of κ oploid receptors in the morphine-reduced straub tail

We used the morphine-induced Straub tail to examine the actions of a 5-HT₃ receptor antagonist and κ opioid receptor agonist. The κ opioid receptor agonist, U-50,488H (4–16 mg/kg i.p.), produced a dose-related inhibition of the tail elevation induced by morphine (10 mg/kg s.c.) in ICR male mice. ICS-205-930 (3 and 10 mg/kg i.p.) and zacopride (0.3 and 1 mg/kg i.p.), 5-HT₃ receptor antagonists, attenuated the inhibitory effect of U-50,488H in a dose-dependent manner. ICS-205-930 and zacopride did not inhibit the morphine-induced Straub tail. These observations suggest that the actions of κ opioid receptors may be modulated by 5-HT³ receptors in the morphine-induced Straub tail.     

PKC and PKA inhibitors reinstate morphine-induced behaviors in morphine tolerant mice

Male Swiss Webster mice exhibited antinociception, hypothermia and Straub tail 3h following a 75mg morphine pellet implantation. These signs disappeared by 72h, and the morphine-pelleted mice were indistinguishable from placebo-pelleted ones, although brain morphine concentrations ranged from 200 to 400ng/gm. We previously demonstrated that chemical inhibitors of protein kinase C (PKC) and A (PKA) are able to reverse morphine tolerance in acutely morphine-challenged mice. However, it was not known whether the reversal of tolerance was due to the interaction of kinase inhibitors with the morphine released from the pellet, the acutely injected morphine to challenge tolerant mice, or both. The present study aimed at determining the interaction between the PKC and PKA inhibitors and the morphine released "solely" from the pellet to reinstate the morphine-induced behavioral and physiological effects, 72h after implantation of morphine pellets. Placebo or 75mg morphine pellets were surgically implanted, and testing was conducted 72h later. Our results showed that the intracerebroventricular (i.c.v.) administration of the PKC inhibitors, bisindolylmaleimide I and G?-6976 as well as the PKA inhibitors, 4-cyano-3-methylisoquinoline and KT-5720, restored the morphine-induced behaviors of antinociception, Straub tail and hypothermia in morphine-pelleted mice to the same extent observed 3h following the pellet implantation. The tail withdrawal and the hot plate reaction time expressed as percent maximum possible effect (%MPE) was increased to 80-100 and 41-90%, respectively, in PKC and PKA inhibitor-treated morphine tolerant mice compared to 2-10% in non-treated mice. Similarly, a significant hypothermia (1.3-4.0 degrees C decrease in body temperature) was detected in PKC and PKA inhibitor-treated morphine tolerant mice compared to an euthermic state in non-treated morphine tolerant mice. Finally, the Straub tail score was increased to 1.1-1.6 in PKC and PKA inhibitor-treated tolerant mice, whereas it was totally absent in non-treated animals. It is noticeably that the kinase inhibitors used in the study had no effect in placebo-pelleted mice. Our results provide the first evidence on the ability of PKC and PKA inhibitors to reinstate the behavioral and physiological effects of morphine in non-challenged morphine-tolerant animals.     

糖尿病对肌松药肌松效应的影响探讨

糖尿病周围神经病变在围术期影响肌松药的肌松效应,导致糖尿病患者术中应用肌松药后术后呼吸功能不全的发生率显著增加,加之糖尿病本身就可损伤呼吸系统,严重影响糖尿病患者的预后。本文探讨糖尿病对肌松药肌松效应的影响机制,并就糖尿病对临床常用肌松药(维库溴铵、罗库溴铵、阿曲库铵及顺式阿曲库铵)的影响进行综述。     

[d-Pen2,d-Pen5]enkephalin,the standard delta opioid agonist,induces morphine-like behaviors in mice

[d-Pen²,d-Pen⁵]enkephalin (DPDPE; 3–30 µg) and morphine (10 µg) both caused Straub tails, increased locomotion, and circling after ICV administration to ICR mice. DPDPE-induced tail stiffening was reduced when mice were pretreated with naloxone (0.5 mg/kg SC) or-funaltrexamine (10 µg ICV), but not with ICI 174864 (2 mg/kg SC), the selective antagonist at delta opioid receptors. These results point to (a) mu receptors mediating the tail stiffening and (b) the loss of delta receptor selectivity after 10 and 30 µg DPDPE.     

无肌松药全身麻醉用于鼾症手术的临床研究

目的尝试在鼾症（OSAHS）手术全身麻醉维持中免用肌松药的可行性，探讨其临床价值。方法选择ASAI-Ⅱ级悬雍垂腭咽成形术（UPPP）鼾症患者180例，随机分为A、B、C三组（每组n=60），均以瑞芬太尼、丙泊酚及氯化琥珀胆碱静脉快诱导，经鼻明视或盲探气管插管，A组以瑞芬太尼、丙泊酚维持麻醉，B、C组分别在A组基础上加用阿曲库铵、维库溴铵维持麻醉。观察记录三组患者各时点循环和呼吸功能指标及术中肌松程度、体动次数、麻醉苏醒时间、拔管时间、不良反应及并发症等。结果三组气管插管成功率100％，麻醉维持平稳，无体动反应，均顺利完成手术；A组拔管前后清醒彻底，呼吸功能和肌力正常，无麻醉并发症发生，B、C组清醒后有不同程度肌松药残余，须使用新斯的明拮抗，拔管后分泌物增多、出血、心动过缓、低氧血症、舌后坠、喉痉挛、呼吸道梗阻、支气管痉挛发生率明显高于A组（P〈0．01）。结论瑞芬太尼一丙泊酚无肌松药全凭静脉麻醉维持用于鼾症手术安全有效，可明显缩短麻醉恢复时间，提高麻醉恢复质量，降低麻醉恢复期风险，减少并发症发生。     

A comparative study of dantrolene sodium and other skeletal muscle relaxants with the Straub tail mouse

Male mice treated subcutaneously with morphine sulphate, 15 mg/kg, develop a Straub tail reaction within 15 min. The elevated tail has been used as a model of contracted muscle for studying the effects of a variety of drugs which induce muscle relaxation. The effective Straub tail antagonists (dantrolene sodium, diazepam, chlordiazepoxide, neuromuscular blocking agents and local anaesthetics) were further compared for their ability to cause loss of righting reflex and acute toxicity. Effects on motor coordination were also determined for the two benzodiazepines and dantrolene sodium. Dantrolene sodium was more specific than diazepam > chlordiazepoxide > neuromuscular blocking agents > local anaesthetics in relaxing the Straub tail without causing acute toxicity, loss of righting reflex or motor incoordination.     

A comparative study of the effect of some centrally acting skeletal muscle relaxants in mice

The possibility of differentiating between sedative-hypnotics and centrally acting muscle relaxants has been investigated in mice. The ED50 values for loss of muscle tone and loss of righting reflex were determined for 2-(3î,6î-dioxaheptyl) aminomethyl-1,4-benzodioxane (ambenoxan), 2-(4î,7î-dioxaoctyl) aminomethyl-1,4-benzodioxane (WB 4123) and compared with mephenesin, meprobamate, methocarbamol and the sedative-hypnotic, sodium pentobarbitone. Comparison of the slopes of the regression lines for hypotonia and loss of the righting reflex for the compounds suggests that ambenoxan offers a greater separation between the two responses than clinically used muscle relaxants.     

Quantitative grip strength assessment as a means of evaluating muscle relaxation in mice

The effects of various centrally acting drugs and some peripherally acting agents on the forelimb grip strength of CD-1 mice were explored. Forelimb grip strength was assessed by use of a strain gauge to measure the lateral pull force, in grams, exerted by mice as an index of muscle relaxation. The muscle relaxants, diazepam, midazolam, baclofen, methocarbamol, dantrolene sodium and the neuromuscular blocking agents, succinylcholine and pancuronium bromide, dose-dependently reduced forelimb grip strength. 2-Amino-7-phosphonoheptanoic acid (AP7), which has also been shown to have muscle relaxant effects, also reduced grip strength. Pentobarbital, ethanol, phencyclidine, ketamine and chlorpromazine reduced grip strength at doses which produced behavioral impairments. Lithium chloride, a toxic compound used to induce taste aversions, and clonidine, at doses which affect blood pressure, body temperature and locomotor activity, did not affect grip strength. In addition, stimulant doses of amphetamine and caffeine, but not of morphine, increased grip strength in a dose-dependent manner. These results extend previous findings and suggest that this forelimb grip strength procedure may be a useful screening test for the identification of the potential muscle relaxant properties of drugs.     

N-palmitoylethanolamide, an endocannabinoid, exhibits antidepressant effects in the forced swim test and the tail suspension test in mice

The antidepressant-like effects of N-palmitoylethanolamide (PEA), a putative endocannabinoid, was investigated in mice using the tail suspension test (TST) and the forced swimming test (FST). In TST, PEA (10, 20, and 40 mg/kg) produced a statistically significant reduction in immobility (50, 32, and 34%, respectively, vs. the control group), whereas fluoxetine (20 mg/kg) reduced immobility by 38%. In FST, PEA (5, 10, and 20 mg/kg) produced a statistically significant reduction in immobility (15, 21, and 36%, respectively), whereas fluoxetine (20 mg/kg) reduced immobility by 18%. Moreover, PEA (20 mg/kg) did not significantly change motor activity in a spontaneous behavioral test. In conclusion, PEA (dose range of 5-40 mg/kg) administered orally reduced immobility in TST and FST, comparable to the antidepressant effect of fluoxetine, and had no effect on spontaneous activity in mice.     

Zinc deficiency induces behavioral alterations in the tail suspension test in mice. Effect of antidepressants

BackgroundRecently, experimental zinc deficiency has been correlated with depression-like alterations in rodents.MethodsIn the first part of present study, the time course of zinc deficient diet induced alterations in the tail suspension test (TST) in mice was investigated. In the second part, the effect of imipramine and escitalopram in control and zinc-deprived for 3 weeks mice was examined in the TST.ResultsA 4- and 10-week administration of a Zn-deficient diet enhanced the immobility time in the TST (by 20% and 57%, respectively). By contrast, a 2-week period of a zinc deficient diet effected the reduction (by 24%) of the immobility time. Moreover, a 2- and 4-week (but not 10-week) of a Zn-deficient diet resulted in the reduction of the body weight (by 37% and 18%, respectively). These results indicate the developing response to zinc deficiency induced by a zinc-deficient diet. The antidepressant-like effect (reduction in the immobility time) of both drugs was significantly reduced in zinc-deprived mice, which suggests treatment-resistance induced by zinc deprivation.ConclusionsZinc deprivation induces “pro-depressive” behavior and alters antidepressant efficacy.     

A double-blind trial of methocarbamol versus placebo in painful muscle spasm

Summary

A double-blind trial of methocarbamol versus placebo was carried out in 59 matched pairs of patients suffering from painful muscle spasm. Methocarbamol {1500?mg. q.d.s) was found to be effective in approximately 60 % of patients compared with 30 % of patients receiving placebo (p< 0.01). Side-effects were of almost equal incidence in the two groups.     

Effects of neuroleptics on morphine-induced tail erection in mice.

Morphine elicits dose-dependent tail erection in mice. Pretreatment of mice with atropine, phenoxybenzamine, propranolol, diphenhydramine, cyproheptadine or parachlorophenylalanine did not interfere with tail erection induced by morphone. Several neuroleptic drugs which are dopamine receptor blocking agents showed a clear antagonistic effect on morphine-induced tail erection (MITE). Haloperidol and penfluridol blocked MITE at doses which only produced a slight behavioral depression. Pimozide and chlorpromazine were less antagonistic than haloperidol and penfluridol and inhibited MITE only at doses which produced a marked behavioral depression. Results indicated that dopamine might be involved in tail erection induced by morphine. MITE in mice might be a useful model for the evaluation of neuroleptic drugs.     

Effects of serine/threonine protein phosphatase inhibitors on morphine-induced antinociception in the tail flick test in mice

The aim of this study was to evaluate the effects of serine/threonine protein phosphatase (PP) inhibitors on morphine-induced antinociception in the tail flick test in mice, and on [3H]naloxone binding to the forebrain crude synaptosome fraction. Neither okadaic acid nor cantharidin (1-10000 nM) displaced [3H]naloxone from its specific binding sites, which indicates that they do not interact at the opioid receptor level. The i.c.v. administration of very low doses of okadaic acid (0.001-1 pg/mouse) and cantharidin (0.001-1 ng/mouse), which inhibit PP2A, produced a dose-dependent antagonism of the antinociception induced by morphine (s.c.). However, L-nor-okadaone (0.001 pg/mouse-1 ng/mouse, i.c.v.), an analogue of okadaic acid lacking activity against protein phosphatases, did not affect the antinociceptive effect of morphine. On the other hand, high doses of okadaic acid (10 ng/mouse, i.c.v.) and cantharidin (1 microg/mouse, i.c.v.), which also block PP1, and calyculin-A (0.1 fg/mouse-1 ng/mouse, i.c.v.), which inhibits equally both PP1 and PP2A, did not modify the morphine-induced antinociception. These results suggest that the activation of type 2A serine/threonine protein phosphatases may play a role in the antinociceptive effect of morphine, and that PP1 might counterbalace this activity.     

On the mechanism of tolerance to morphine-induced Straub tail reaction in mice

The effect of 5-HT and opioid receptor antagonists on morphine-induced Straub tail was studied in mice. Straub tail behavior was induced by subcutaneous administration of different doses (20, 30, and 40 mg/kg) of morphine hydrochloride to mice. The effect of morphine was dose-dependent. Maximum response was obtained with 40 mg/kg of the drug. The response induced by morphine (20 and 40 mg/kg) was decreased by different doses of intraperitoneal injection of naloxone (1 and 2 mg/kg) or methysergide, mianserin, and ritanserin (1 and 2 mg/kg). The effect of morphine (40 mg/kg) was also reduced by intracerebroventricular injection of naloxone (0.4-0.8 microg/animal) or mianserin (2 and 4 microg/animal). Different groups of mice received one daily dose (50 mg/kg sc) of morphine sulfate for 3 days to develop tolerance to morphine. The Straub tail reaction induced by morphine hydrochloride (40 mg/kg) was tested on the fourth day. Naloxone injection (1 and 2 mg/kg ip) on Day 3 (1 h after morphine sulfate injection) or on Day 4 (1 h before test dose of morphine hydrochloride), decreased tolerance induced to morphine. Methysergide, mianserin, or ritanserin (intraperitoneal) on Days 2 and 3 (1 h after morphine sulfate injection) or on Day 4 (1 h before test dose of morphine hydrochloride), also decreased tolerance induced to morphine. Intracerebroventricular injection of either naloxone or mianserin also reduced tolerance to morphine. It is concluded that 5-HT(2) and opioid receptor mechanisms are involved in morphine-induced Straub tail reaction and tolerance induced to morphine also may be mediated through these receptors.     

Calpastatin overexpression in the skeletal muscle of mice prevents clenbuterol-induced muscle hypertrophy and phenotypic shift

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