Smooth muscle contraction induced by Indigofera dendroides leaf extracts may involve calcium mobilization via potential sensitive channels

The contractile effects of the aqueous extract of the leaves of Indigofera dendroides (ID) were studied on the gastrointestinal motility in mice and isolated smooth muscle preparations obtained from rats and guinea pigs. The contractile effects of 10(-6) M acetylcholine, 80 mM KCl and 1.6 mg/ml ID were measured on the rat ileal smooth muscle exposed to calcium-free buffer or physiological solution, to determine the calcium pools mobilized by extract for activation of contraction. Acute toxicity test (LD(50)) was also carried out in mice. The result showed that ID (0.05-3.2 mg/ml) produced a concentration-dependent contraction of the guinea pig and rat ileum. These responses were not blocked by mepyramine (2.49 x 10(-9) M), verapamil (8.14 x 10(-9) M), or pirenzepine (4.7 x 10(-7) M), but were blocked completely by atropine (2.92 x 10(-9) M). A significant increase in propulsion of gastrointestinal motility was observed. Acetylcholine, KCl and ID produced contractions in Ca(2+) free media. The phasic components of the contractile responses to Ach as well as the tonic component of K(+) and ID-induced contractions were relatively resistant to short periods of calcium-free exposure. Ach, K(+) and ID still caused contractions in the presence of verapamil. The data revealed that ID-induced contractions were not mediated by histaminergic receptors, calcium channels, M1 muscarinic receptors. It also suggests that Ach mobilize Ca from some tightly bound or intracellular pool, whereas high K(+) and ID may mobilize Ca from some superficial or loosely-bound pool.     

Uterotonic effects of hydromethanolic extract of Parquetina nigrescens (Periplocaceae) on spontaneous contractile activity in the isolated myometrium of pregnant rats

Parquetina nigrescens (Afz.) Bullock (Periplocaceae) hydromethanolic extract (PAR) in reference solution exerts a stimulating and spasmogenic action on the pregnant rat myometrium in vitro. This action is characterised by increasing amplitude of spontaneous isometric contractions and a slight elevation of muscular basic tonus. These effects on myometrium may be compared with those provoked by sulprostone (PGE₂) in the same experimental conditions. In calcium free solution, Parquetina nigrescens extract could restore partially spontaneous contractile activity. On the contrary, in presence of EDTA PAR induces a complete abolition of the activity. On the other hand, in calcium free solution, sulprostone is able to develop a clear elevation of basic tonus (contracture). These observations support the hypothesis that the extract has an oxytocin-like effect, which is characterised by an extracellular influx of calcium, responsible for the increase of the maximum isometric contraction amplitude. Sulprostone activates both extracellular and intracellular calcium responsible for the increase of contractile activity and development of the contracture of uterine smooth muscle.     

Activity of aqueous extract of the bark of Vitex doniana on uterine muscle response to drugs

The activity of Vitex doniana on the uterine muscle response was investigated. The bark of Vitex doniana was extracted in boiled water at 100 degrees C, and the extracted solution filtered and centrifuged with refrigeration. The extract prepared from the dry powder extract was tested on uterine muscle strip preparations. The bark extract of Vitex doniana was analysed elementally and found to contain much more potassium and phosphate than calcium, magnesium, zinc and iron. The presence of potassium ions in excess may also be partly responsible for the effect of the crude extract on uterine muscle activity. In another study, Vitex doniana extract induced graded uterine muscle contractions and also potentiated the contractile effects of prostaglandins, ergometrine and oxytocin. However, the potentiating effect was not significant on the contractile responses to acetylcholine and potassium chloride. The investigation therefore suggests that the effect of the Vitex doniana bark extract may be not only voltage operated but may act via uterotonic receptors. Therefore, the use of Vitex doniana to control postpartum bleeding after child birth may be justified.     

Antispasmodic and hypotensive effects of Ferula asafoetida gum extract

The effects of Ferula asafoetida gum extract on the contractile responses of the isolated guinea-pig ileum induced by acetylcholine, histamine and KCl, and on the mean arterial blood pressure of rat were investigated. In the presence of extract (3 mg/ml), the average amplitude of spontaneous contractions of the isolated guinea-pig ileum was decreased to 54 +/- 7% of control. Exposure of the precontracted ileum by acetylcholine (10 microM) to Ferula asafoetida gum extract caused relaxation in a concentration-dependent manner. Similar relaxatory effect of the extract was observed on the precontracted ileum by histamine (10 microM) and KCl (28 mM). However, when the preparations were preincubated with indomethacin (100 nM) and different antagonists, such as propranolol (1 microM), atropine (100 nM), chlorpheniramine (25 nM) then were contracted with KCl, exposure to the extract (3 mg/ml) did not cause any relaxation. Furthermore, Ferula asafoetida gum extract (0.3-2.2 mg/100g body weight) significantly reduced the mean arterial blood pressure in anaesthetised rats. It might be concluded that the relaxant compounds in Ferula asafoetida gum extract interfere with a variety of muscarinic, adrenergic and histaminic receptor activities or with the mobilisation of calcium ions required for smooth muscle contraction non-specificly.     

Pharmacological effects of Agapanthus africanus on the isolated rat uterus.

The Agapanthus africanus plant is used by South African traditional healers as a phytomedicine in herbal remedies to treat pregnancy-related ailments and to augment labour. It has already been shown that an aqueous extract of A. africanus causes smooth muscle contractions in the isolated uterus and ileum preparations. In the present study, the effects of an aqueous extract of A. africanus leaves was examined on receptor systems involved in contraction of the uterine smooth muscle in order to determine the mechanism of its pharmacological effect relevant to its ethnic use to augment labour. The extract was tested on the isolated rat uterus preparation. The aqueous extract of A. africanus leaves was found to exhibit agonist activity on uterine muscarinic receptors and to promote the synthesis of prostaglandins in the oestrogenized rat uterus. Some pharmacological justification for the ethnic use of A. africanus as a herbal oxytocic in prolonged labour has been provided.     

黄芩苷对大鼠离体小肠及胃平滑肌条收缩性的影响

目的研究黄芩苷对大鼠离体小肠及胃平滑肌条收缩性的影响。方法以大鼠离体小肠及胃平滑肌条收缩性为研究模型,考察黄芩苷在高钙、低钙及正常克氏液中对平滑肌条收缩作用的影响;在正常克氏液中分别观察黄芩苷对新斯的明、乙酰胆碱、组胺及阿托品存在下平滑肌收缩性的影响。结果在高钙、正常钙及低钙不同条件下,黄芩苷在6.25~100μmol/L范围内均剂量依赖性抑制小肠、胃平滑肌收缩,拮抗新斯的明、乙酰胆碱及组胺引起的收缩作用,且相加阿托品引起的抑制收缩作用。结论黄芩苷对小肠、胃平滑肌收缩具有明显抑制作用。     

Inhibitory effects of quercetin on guinea-pig ileum contractions

The inhibitory effect of quercetin on the guinea-pig ileum contractile responses was examined to further clarify its mechanism of action. Quercetin inhibited, from doses of 5 μM , the spontaneous phasic contractions of the guinea-pig ileum, as well as the phasic and tonic components of either acetylcholine- or KCl-elicited contractions, in a dose-dependent way. The former effect was reversible and was unaffected by nicotinic blockade. The tonic component of the contractions was more responsive to quercetin than the phasic component. The inhibition of the tonic contraction was, at least partially, due to the blockade of calcium channels by quercetin, since increasing calcium concentration in the tissue bath reversed the inhibition when the organ was contracted by KCl. Quercetin may also interfere with calcium release from intracellular stores, since it inhibited acetylcholine-elicited phasic contraction even in a calcium-free solution. Finally, quercetin shifted the concentration-response curves to carbachol downwards, without modifying the ED₅₀ of the agonist, whereas the concentration-response curves to CaCl₂ were shifted downwards and to the right. In conclusion, quercetin inhibits the contractile responses of guinea-pig ileum, decreasing the calcium concentration available for contractile machinery.     

Participation of extracellular calcium in α-hederin-induced contractions of rat isolated stomach strips

Ethnopharmacological relevance

The dry extract of Hedera helix leaves, due to its secretolytic and antispasmodic effects, is commonly used to produce pharmaceuticals applied in case of cough and other respiratory symptoms. The results of some in vitro studies as well as the clinical signs of poisoning caused by Hedera helix suggest however strong contractile effect on smooth muscle. In order to clarify the impact of α-hederin (the main active agent of ivy extract) on smooth muscle, the origin of activated calcium involved in α-hederin-induced contraction of gastric smooth muscle preparations was studied.

Materials and methods

The study was carried out on rat isolated stomach corpus and fundus strips, under isotonic conditions. The effect of α-hederin (100 μM) on smooth muscle preparations was measured before and after the treatment with verapamil during the incubation in modified Krebs–Henseleit solution (M K–HS). Besides, the effect of saponin was measured during the incubation of preparation in Ca²⁺-free modified Krebs–Henseleit solution or Ca²⁺-free EGTA-containing modified Krebs–Henseleit solution.

Results

The obtained results revealed that the application of verapamil significantly inhibited the reaction evoked by α-hederin. The incubation of stomach strips in calcium-free modified Krebs–Henseleit solution did not change the force of the observed contraction in comparison to the reaction of the preparations incubated in regular incubation solution (M K–HS). In contrary, the replacement of M K–HS by calcium-free chelator-containing solution inhibited totally the reaction to α-hederin.

Conclusions

The results indicated that α-hederin-induced contraction results from the influx of calcium which is located in intercellular spaces or bound to the outside of the cell membrane. The Ca²⁺ influx occurs predominantly through voltage-dependent calcium channels of L-type.     

Phytochemical screening and effect of aqueous extract of Ficus sycomorus L. (Moraceae) stembark on muscular activity in laboratory animals

The stembark of Ficus sycomorus was collected, dried and extracted, to screen for some chemical constituents and study its effect on muscle contraction. The duodena and recti abdominis of 10 guinea pigs weighing between 330 and 345 g and 10 frogs weighing between 180 and 201 g, respectively, were isolated and used for this study. The extract was tested to see its effect on acetylcholine (ACH) induced contraction on kymograph. The extract reduced the acetylcholine contractile responses of guinea pigs duodena and recti abdominis muscles of frogs significantly, thus showing inhibitory effect on muscle contraction. The extract showed the presence of gallic tannins, saponins, reducing sugars, alkaloids and flavone aglycones. It was concluded that the extract has inhibitory effect on both smooth and skeletal muscles contractions and contains important constituents for pharmacological activities.     

Effects on rat uterine and aorta strip smooth muscle of Thymus leptophyllus extract

The diethylether extract from Thymus leptophyllus was found to be more active on uterine smooth muscle than on aorta strips. Rat uterus experiments with and without extracellular calcium, yielded similar IC₅₀ values. A non-specific mechanism for the relaxant activity can therefore be postulated. In rat aorta and in the presence of extracellular calcium the extract inhibited the contractile response induced by K⁺ depolarizing solution and had a less inhibitory effect on noradrenaline (NA) contraction. In a Ca²⁺-free solution the extract strongly reduced the Ca²⁺-release induced by NA, but it did not affect the transient contraction caused by caffeine (CAF).     

Smooth muscle relaxant effect of kaurenoic acid,a diterpene from Copaifera langsdorffii on rat uterus in vitro

The effect of kaurenoic acid, a diterpene isolated from the oleo-resin of the popular medicinal plant Copaifera langsdorffii (Leguminaceae), was analysed on rat uterine muscle responsiveness to various drugs in vitro. Cumulative concentration-response curves to acetylcholine and oxytocin were obtained before and after incubation of uterine segments with up to 160 microm of kaurenoic acid. The maximal contractile response (E(max)) evoked by these agonists was inhibited by kaurenoic acid in a concentration-related manner; at 160 microm, kaurenoic acid depressed the E(max) of oxytocin and acetylcholine by 83% and 91%, respectively. The relaxation caused by kaurenoic acid on oxytocin-induced contraction was unaffected in the presence of tetraethyl ammonium, a compound that blocks the calcium activated potassium channels. It was partially reversed by glibenclamide (10(-5) m), an ATP-sensitive potassium channel blocker. Also, kaurenoic acid at 160 microm concentration was found to inhibit significantly the CaCl(2)-evoked contractile responses in a medium of high potassium and zero calcium. Furthermore, kaurenoic acid was found to relax the sustained tonic contraction induced by acetylcholine, oxytocin, BaCl(2) and KCl in a concentration-dependent way. However, KCl-induced tonic contraction was only weakly inhibited by kaurenoic acid. These data indicate that the diterpene, kaurenoic acid, exerts a uterine relaxant effect acting principally through calcium blockade and in part, by the opening of ATP-sensitive potassium channels.     

Pharmacological basis for the use of peach leaves in constipation

The aqueous crude extract (PPL.Cr) of peach leaves (Prunus persica) was studied for the possible presence of gut stimulatory constituent(s) to rationalize the folkloric use of the plant in constipation. PPL.Cr at the dose of 1-10 mg/ml caused a moderate degree of spasmogenic effect in isolated guinea-pig ileum. Pretreatment of the tissue with atropine (1 M) completely abolished the contractile effect of the plant extract similar to that of acetylcholine which is suggestive of a cholinergic mechanism. In isolated rabbit jejunum preparations, PPL.Cr produced a week spasmogenic effect followed by relaxation of the spontaneous contractions at higher doses. Bioassay-directed fractionation revealed that the spasmogenic activity was separated in the aqueous fraction, while the spasmolytic activity was concentrated in the ethyl acetate fraction. When tested against K(+)-induced contraction, both PPL.Cr and its ethyl acetate fraction (PPL.EtAc) caused a dose-dependent inhibition, suggesting calcium channel blockade (CCB). The presence of CCB in peach leaves was confirmed when pretreatment of the tissue with PPL.EtAc caused a dose-dependent rightward shift in the Ca(2+) dose-response curves, similar to that produced by verapamil. These data indicate that the plant contains spasmogenic (cholinomimetic) and spasmolytic (calcium antagonist) constituents, which are concentrated in the aqueous and ethyl acetate fractions, respectively. Furthermore, the laxative effect of the plant reported in the traditional system of medicine may be partially due to the cholinergic action, which was dominant over the spasmolytic component.     

氢溴酸槟榔碱对C57BL/6和W/W^v突变小鼠离体胃肠平滑肌条动力的作用研究

目的利用C57BL/6和W/W^v突变小鼠研究Cajal间质细胞(interstitial cells of Cajal,ICC)对小鼠胃肠各部位平滑肌条运动的影响,并通过胆碱能受体拮抗剂阿托品和钠通道阻断剂河豚毒素(tetrodotoxin,TTX)给药干预初步探讨氢溴酸槟榔碱(arecoline hydrobromide,Ah)促进小鼠平滑肌条动力作用的机制。方法制备C57BL/6和W/W^v突变小鼠胃肠各部位(胃体、胃窦、空肠和结肠)平滑肌环形肌条标本,分别观察和记录2种小鼠离体胃肠各部位平滑肌条的运动,比较2种小鼠胃肠各部位肌条收缩情况(收缩振幅、收缩频率和张力),并观察Ah对2种小鼠胃肠各部位肌条收缩的影响及阿托品、TTX对Ah促动力作用的干预。结果C57BL/6小鼠胃体部平滑肌环形肌条的收缩为具有较高振幅储存食物和低频率少量推进性的收缩波形,胃窦部为高振幅高张力较高频率推进性的收缩波形,空肠部为低振幅高频率快速推进性的收缩波形,结肠部为分散性中等振幅和频率的收缩波形;W/W^v突变小鼠胃体部环形肌条的收缩频率、收缩振幅都在较低水平,张力较其他部位偏高,胃窦部收缩频率、收缩振幅和张力都在较高水平,空肠部的收缩频率仍是最高,收缩振幅和张力却最低,结肠部的收缩频率最低,收缩振幅和张力处于中等水平;C57BL/6小鼠各部位肌条的收缩力均远大于W/W^v突变小鼠(P<0.05,P<0.01,P<0.001);Ah能明显增加C57BL/6小鼠各部位肌条的收缩力(P<0.05,P<0.01,P<0.001),对W/W^v突变小鼠各部位肌条的收缩力只有轻微增加(P<0.05,P<0.001);阿托品能抑制Ah的促动力作用,而TTX并不能阻断。结论ICCs在调控小鼠胃肠平滑肌条运动中具有重要作用,氢溴酸槟榔碱的促动力作用机制除了ICCs网络的调控外,还有一部分神经调控。另外,氢溴酸槟榔碱可能是经M型受体起作用的。     

Evaluation of Alstonia scholaris leaves for broncho-vasodilatory activity

The present study demonstrates that the ethanol extract of Alstonia scholaris (Apocynaceae) leaves induced pronounced bronchodilatory activity in anaesthetized rats with the probable involvement of prostaglandins. However, in vitro preparations of guinea-pig trachea did not confirm this property, indicating that bronchodilation is not due to the direct tracheal smooth muscle relaxation. The vasodilatory activity of the extract was independent of adrenergic or muscarinic receptors or prostaglandins but was mainly via endothelial-derived relaxing factor, nitric oxide. The extract inhibited the spontaneous movements of rabbit jejunum and contractile effects of acetylcholine and histamine on guinea-pig ileum. Additionally, the extract caused marked reduction of barium chloride-, potassium chloride- and calcium chloride-induced contraction on guinea-pig ileum and pulmonary artery, implying a direct interference of plant extract with the influx of calcium ions into cells. However, the extract has no detectable effect on mobilization of intracellular calcium. These results coupled with the in vivo effects of ethanol extract reveal that the Alstonia scholaris leaves possess broncho-vasodilatory activity mediated presumably by prostaglandins, calcium antagonism and endothelium-derived relaxing factor(s).     

侧柏叶乙酸乙酯提取物对豚鼠离体气管平滑肌的作用

用离体豚鼠气管条,探讨侧柏叶乙酸乙酯提取物对气管平滑肌作用。结果表明,侧柏叶乙酸乙酯提取物能抑制乙酰胆碱,氯化钾所致气管平滑肌收缩,而且能使乙酰胆碱收缩气管平没肌的量效曲线右移,并抑制最大效应,其作用为剂量依赖性心肌肥厚提示侧柏叶乙酸乙酯提取物松驰气管平滑肌作用机制可能与影响Ｃａ＾２＋的跨膜转运有关。     

槲皮素对平滑肌收缩性及肌球蛋白Mg~（2＋）-ATPase活性的影响

目的：研究槲皮素对平滑肌收缩性及肌球蛋白Mg2＋-ATPase活性的影响。方法：以大鼠离体小肠及胃平滑肌条为研究模型,首先考察槲皮素在高钙、低钙及正常克氏液环境中对平滑肌条收缩性的影响。然后在正常克氏液中分别观察槲皮素对新斯的明、乙酰胆碱及阿托品存在下平滑肌收缩性的影响。用孔雀绿法测定槲皮素对肌球蛋白Mg2＋-ATPase活性的影响。结果：在高钙、正常钙及低钙不同条件下,槲皮素在4～64μmol/L范围内均剂量依赖性抑制小肠、胃平滑肌收缩;槲皮素（32μmol/L）拮抗新斯的明、乙酰胆碱引起的收缩作用（P〈0.01）,且相加性协同阿托品引起的抑制收缩作用（P〈0.01）;槲皮素（25μmol/L）对不同磷酸化程度的肌球蛋白的Mg2＋-ATPase活性均具有抑制作用（P〈0.01）。结论：槲皮素对胃、肠平滑肌收缩性及平滑肌肌球蛋白的Mg2＋-ATPase活性具有明显抑制作用。     

Barakol suppresses norepinephrine-induced inhibition of spontaneous longitudinal smooth muscle contractions in isolated rat small intestine

The present study aimed to investigate the purgative effects of barakol, the purified extract of Cassia siamea Lam., on the longitudinal smooth muscle contractions of the rat ileum. The extract increased the force of spontaneous muscle contractions in a concentration-dependent manner (EC50=0.3 mM). Saxitoxin (0.3 microM) abolished the stimulatory effects of barakol, a result indicating a neural mechanism of action. In addition, atropine (10 microM) but not propanolol (10 microM) or phentolamine (10 microM), partially inhibited barakol-induced smooth muscle contractions suggesting that cholinergic nerves were involved. The motor effects of barakol were further examined in muscle strips treated with catecholamines to suppress spontaneous contractile activity and decrease muscle tone. Norepinephrine or dopamine (10 microM) decreased the amplitude of spontaneous contractions by 72% and 18%, respectively. Pretreatment of the tissues with barakol (1 mM) significantly decreased the inhibitory effect of norepinephrine by 60%, but not that of dopamine. Its ability to potentiate atropine- and saxitoxin-sensitive contractions and inhibit the antimotility actions of norepinephrine suggests that barakol may increase longitudinal smooth muscle contractions by decreasing the inhibitory effect of norepinephrine on excitatory cholinergic motor neurons. Barakol may produce a purgative action in small intestine which may be clinically important in patients with intestinal hypomotility disorders.     

Pharmacological analysis of the inhibitory effect of jatrophone,a diterpene isolated from Jatropha elliptica,on smooth and cardiac muscles

The present work was conducted to analyse the effect of the diterpene jatrophone on drug and electrically evoked contractions of cardiac and smooth muscle preparations. Pre-incubation of the isolated rat uterus with jatrophone (1–100 μM ), for 20 min caused a decrease in the maximal contractions and a rightward displacement of the dose-response curves to acetylcholine, bradykinin, and oxytocin. In the isolated longitudinal muscle from guinea-pig ileum, jatrophone (1.6–12.8 μM ) produced a concentration-dependent inhibitory effect of acetylcholine, bradykinin and histamine-induced contractions. In this tissue, jatrophone was about 5 to 8-fold more potent in inhibiting acetylcholine and bradykinin than in uterine muscle. In strips of guinea-pig ileum, guinea-pig urinary bladder, rat left atrium electrically stimulated, and dog ureter contractions, jatrophone (1–100 μM ) caused a concentration-dependent inhibitory effect. In preparations electrically stimulated, jatrophone was about 2.5 to 23-fold less potent compared to those values obtained against agonist-induced contractions in the guinea-pig ileum. These results demonstrate that jatrophone exerts direct inhibitory effect on cardiac and smooth muscle preparations and show that its inhibitory potency may vary according to the tissue and/or stimulus employed to induce muscle contractions.     

Phytochemical screening and effect of aqueous extract of Ficus sycomorus L. (Moraceae) stem bark on muscular activity in laboratory animals

The stembark of Ficus sycomorus was collected, dried and extracted to screen for some chemical constituents and study its effect on muscle contraction. The duodena and recti abdominis of 10 guinea pigs weighing between 330 and 34 g and 10 frogs weighing between 180 and 201 g, respectively were isolated and used for this study. The extract was tested to see its effect on acetylcholine-induced contraction on kymograph. The extract reduced the acetylcholine contractile responses of guinea pigs duodena and recti abdominis muscles of frogs significantly, thus showing inhibitory effect on muscle contraction. The extract showed the presence of gallic tannins, saponins, reducing sugars, alkaloids and flavone aglycones. It was concluded that the extract has inhibitory effect on both smooth and skeletal muscles contractions and contains important constituents for pharmacological activities.     

Effect of ambrein on smooth muscle responses to various agonists

The pharmacological effects of ambrein (isolated from ambergris) on the contractile responses induced by some agonists in smooth muscle preparations were investigated. Ambrein in the concentration range of 10, 50 and 250 μg/ml decreased the spontaneous contraction of the isolated rabbit jejunum, rat uterus and guinea-pig vas deferens. Ambrein-induced antagonism to acetylcholine (Ach) in the guinea-pig ileum was abolished when the concentration of calcium chloride in the Tyrode's solution was increased to 5 mM/l. Furthermore, ambrein did not antagonise nicotine-induced contractions in the isolated rabbit jejunum or serotonin-induced contractions in the isolated guinea-pig ileum and vas deferens or the rat uterus. However, ambrein in the concentration range of 10, 50 and 250 μg/ml antagonised prostaglandins (PGs) E₂, D₂, F_2α, and oxytocin-induced contractions in the rat uterus in vitro. Ambrein also antagonised (±) noradrenaline and (−) adrenaline-induced contractions in the isolated guinea-pig vas deferens. It is concluded that ambrein-induced non-selective dose-dependent antagonism to the effects of some agonists (Ach, adrenaline, noradrenaline, PGs and oxytocin) in some smooth muscles may be due to the ability of this compound to interfere with the mobilisation of extracellular Ca²⁺ required for muscular contractions induced by these agonists.