Pharmacological Characterization of the Autonomous Innervation of the Guinea Pig Tracheobronchial Smooth Muscle

Abstract: From the trachea, main bronchi and hilus bronchi of guinea pigs, preparations were isolated for registration of mechanical tension on electrical field stimulation and drugs. The trachea contracted on short trains of electrical stimulation. Usually these contractions were followed by a relaxant response. The contractions were abolished while the relaxations were potentiated by atropine. In the main bronchi field stimulation induced a contractile response which was abolished by atropine. In the latter preparation relaxant responses were never observed, even not after atropine. In the trachea and main bronchi neither the α₁ blocker prazosin nor the α₂ blocker yohimbine affected the contractile or relaxant responses when used in α-blocking concentrations. The relaxant response in trachea was neither affected by the “P1 blocker” theophylline nor by the “P2 blocker” quinidine but it was partially inhibited by guanethidine or β-blocking agents. The hilus bronchi contracted on field stimulation as well as by histamine. The contractile response on electrical stimulation was only slightly inhibited by atropine or guanethidine. In all three preparations responses on field stimulation were abolished by tetrodotoxin. It is suggested that in both trachea and main bronchi excitatory cholinergic innervation is present. Further, the trachea but not the main bronchi is innervated by both adrenergic and nonadrenergic inhibitory nerves. The hilus bronchi contains a non-adrenergic non-cholinergic excitatory nervous system.     

Ginsenoside Rb3 Inhibits Angiotensin II‐Induced Vascular Smooth Muscle Cells Proliferation

Abstract: This study was designed to examine the effect of ginsenoside Rb₃ on angiotensin (Ang) II‐induced proliferation of cultured rat vascular smooth muscle cells (VSMCs). VSMCs proliferation was evaluated by [³H]Thymidine incorporation. The cell cycle was examined by flow cytometry. The expression of mRNA of proto‐oncogene c‐myc, c‐fos and c‐jun was observed by RT‐PCR. Ginsenoside Rb₃ had no effects on VSMCs proliferation in physiological condition. Ang II significantly increased the proliferation of VSMCs and the expression of mRNA of proto‐oncogene c‐myc, c‐fos and c‐jun. Ginsenoside Rb₃ markedly inhibited Ang II‐induced VSMCs proliferation. Concomitantly, ginsenoside Rb₃ decreased cell cycle progression from G₀/G₁ to S phase. Furthermore, ginsenoside Rb₃ significantly attenuated the expression of mRNA of proto‐oncogene c‐myc, c‐fos and c‐jun. This study showed that ginsenoside Rb₃ inhibited Ang II‐induced VSMCs proliferation, at least in part by inhibiting Ang II‐induced G₀/G₁ to S phase transition and attenuating the expression of mRNA of c‐fos, c‐jun and c‐myc. The findings may explain the beneficial effects of ginsenoside Rb₃ in cardiovascular diseases, and it will be useful to develop prevention and therapeutics of cardiovascular diseases.     

大黄素增强结肠近端平滑肌细胞钙离子依赖氯离子通道的表达

目的:研究大黄素对结肠近端平滑肌细胞钙离子依赖氯离子通道(ClCa channel)的作用。方法:采用RM6200四道仪记录黄体酮对平滑肌的等长收缩活动作用,相对定量的单细胞RT-PCR法检测平滑肌细胞ClCa通道mRNA的表达。结果:大黄素显著增强离体结肠平滑肌肌条和单个平滑肌细胞的收缩,并可使ClCa电流显著增强,作用与剂量呈正相关。DIDS和NFA可阻滞大黄素的作用。豚鼠近端结肠平滑肌细胞有ClCa1和ClCa2基因的mRNA表达,ClCa1基因的mRNA表达强度是ClCa2基因的mR-NA的5.3±0.71(n=5,P<0.01)倍。50μM大黄素孵育可以使ClCa1基因的mRNA表达增强(n=5,P<0.01)。结论:大黄素可通过兴奋氯离子通道电流引起结肠平滑肌收缩,大黄素对氯离子通道兴奋作用可能与增强结肠平滑肌细胞ClCa1基因表达有关。     

Polyalthic Acid Isolated from Croton reflexifolius has Relaxing Effect in Guinea Pig Tracheal Smooth Muscle

The relaxing activity of Croton reflexifolius H.B.K (Euphorbiaceae) leaves was assessed in isolated guinea pig tracheal rings. The dichloromethane extract of C. reflexifolius was the most active relaxant (EC₅₀ = 118.98 ± 5.927 μ g/mL), and within this extract polyalthic acid was identified as the main active relaxing agent. Polyalthic acid showed a relaxing effect on tracheal rings precontracted with carbachol (EC₅₀ = 183.71 ± 3.28 μM), histamine (6.24 ± 0.28 μM), and KCl (195.79 ± 10.36 μM). The pretreatment with polyalthic acid did not affect the concentration response curve to histamine, and it reduced the Emax of carbachol without affecting its EC₅₀, thus suggesting that polyalthic acid produces a mild antimuscarinic activity. In addition, neither glibenclamide, l-NAME nor propranolol modified the effect of polyalthic acid, although the latter enabled isoproterenol activity. In conclusion, this study represents the first in which the relaxing effect of C. reflexifolius on tracheal rings of guinea pig was clearly demonstrated. Polyalthic acid, which was the main active agent in this relaxing action, produced a mild antimuscarinic activity in a noncompetitive manner. Its relaxing effect was independent of the contractile agent employed and was not related with β -adrenergic receptors, K channels, or nitric oxide. Further experiments are needed to clarify the mechanism of action of polyalthic acid.     

Endoluminal Norepinephrine Inhibits Smooth Muscle Activity of the Pig Pyeloureter by Stimulation of β‐Adrenoceptors without Side Effects

Abstract: It has been demonstrated in pigs that endoluminal administration of norepinephrine reduces the increase in renal pelvic pressure during perfusion. The purposes were to describe concentration–response relationship and receptor mechanism of the effect of norepinephrine on muscle function of pyeloureter and to reveal possible side effects on cardiovascular and renal functions. Renal pelvis was perfused, while pelvic pressure, cardiovascular and renal functional parameters were recorded. In group A, a pelvic pressure increase was examined during pressure flow studies with norepinephrine solutions (0, 1, 5, 50 and 100 µg/ml). In group B, pelvis was perfused with 6 ml/min. norepinephrine solutions (0, 0.001, 0.01, 0.1 and 1 µg/ml). In group C, pelvis was perfused with 6 ml/min. norepinephrine, norepinephrine + sotalol 10^?6 mol/l and norepinephrine + phentolamine 10^?6 mol/l. Norepinephrine solutions of 0, 10^?8, 10^?7, 10^?6, 10^?5 and 10^?4 mol/l were used. In group A, all norepinephrine solutions lowered the pelvic pressure increase significantly. Large increases in plasma and urine norepinephrine occurred with 50 and 100 µg/ml, but cardiovascular and renal functions remained unchanged. In group B, a significant diminishing pelvic pressure increase with all solutions was seen with a significant difference between all solutions. In group C, norepinephrine demonstrated a concentration–response curve with EC₅₀ between 10^?8 and 10^?7 mol/l (10^?7.27±0.40). Sotalol had a smooth muscle inhibitory effect on the pyeloureter and inhibited the effect of norepinephrine increasing EC₅₀ by about a factor 10 (10^?6.40±1.17). No convincing effect of phentolamine was observed. Endoluminal norepinephrine probably stimulates β‐adrenoceptors and inhibits a renal pelvis pressure increase to perfusion in a dose‐related way without side effects. Endoluminal norepinephrine is safe in pigs and may be useful under endoscopy of the pyeloureter.     

(‐)‐Epigallocatechin Gallate Inhibits Endothelin‐1‐Induced C‐Reactive Protein Production in Vascular Smooth Muscle Cells

Abstract: Numerous studies have shown that C‐reactive protein (CRP), a pro‐inflammation cytokine, makes a direct contribution to atherosclerosis, and that (–)‐epigallocatechin gallate (EGCG) is able to exert an anti‐atherosclerotic effect by anti‐oxidative and anti‐inflammatory activities. Based on our previous study, the effect of EGCG on endothelin‐1 (ET‐1)‐induced CRP production in rat vascular smooth muscle cells (VSMCs) and the possible mechanism were observed. The in vitro experiments showed that EGCG concentration‐dependently inhibited ET‐1‐stimulated expression of CRP both in protein and mRNA levels in VSMCs as determined by the immunocytochemical staining, the enzyme‐linked immunosorbent assay and the real‐time quantitative polymerase chain reaction (RT‐qPCR). The in vivo investigation with the double‐labelled immunofluorescence staining and RT‐qPCR displayed that EGCG also prevented ET‐1‐induced CRP expression in protein and mRNA levels in the aortic VSMCs of rats receiving the subchronic infusion of ET‐1. In addition, EGCG suppressed reactive oxygen species (ROS) generation evoked by ET‐1 in VSMCs as observed by the fluorescence probe. These demonstrate that EGCG may inhibit ET‐1‐stimulated generation of CRP in VSMCs so to relieve the inflammatory response and oxidative stress via blocking ROS signal, which provides new evidence for an anti‐atherosclerotic effect of EGCG.     

Methylglyoxal Inhibits Smooth Muscle Contraction in Isolated Blood Vessels

Methylglyoxal (MGO) is a metabolite of glucose. In addition to evidence that increased plasma MGO level is associated with diabetic vascular complications, recent studies demonstrated that MGO accumulated in vascular tissues of hypertensive animals. We hypothesized that MGO could directly affect vascular reactivity. To test the hypothesis, we examined effects of MGO on contraction of isolated blood vessels. Treatment of endothelium-denuded rat aorta with MGO (420 μM, 30 min) shifted the concentration–response curve for noradrenaline (NA: 1nM–1 μM) to the right. The inhibitory effect was concentration-dependent (MGO: 42 – 420 μM). Indomethacin (10 μM) and cimetidine (30 μM) could not prevent the inhibitory effect of MGO. However, a non-selective K⁺-channel inhibitor, tetramethylammonium (10 mM), prevented it. Glibenclamide (3 μM), an ATP-sensitive K⁺-channel inhibitor or apamin (1 μM), a small conductance Ca²⁺-activated K⁺-channel inhibitor was ineffective, but iberiotoxin (100 nM), a large conductance Ca²⁺-activated K⁺ (BK_Ca)-channel inhibitor significantly prevented the effect of MGO. MGO (420 μM, 30 min) also inhibited the NA (1 nM – 1 μM)-induced contraction in mesenteric artery. The present results indicate that MGO has an inhibitory effect on contractility of isolated blood vessel, which is mediated via opening smooth muscle BK_Ca channel.     

Potassium Currents in Isolated Deiters' Cells of Guinea Pig

Deiters'' cells are the supporting cells in organ of Corti and are suggested to play an important role in biochemical and mechanical modulation of outer hair cells. We successfully isolated functionally different K⁺ currents from Deiters'' cells of guinea pig using whole cell patch clamp technique. With high K⁺ pipette solution, depolarizing step pulses activated strongly outward rectifying currents which were dose-dependently blocked by clofilium, a class III anti-arrhythmic K⁺ channel blocker. The remaining outward current was transient in time course whereas the clofilium-sensitive outward current showed slow inactivation and delayed rectification. Addition of 5 mM tetraethylammonium (TEA) further blocked the remaining current leaving a very fast inactivating transient outward current. Therefore, at least three different types of K⁺ current were identified in Deiters'' cells, such as fast activating and fast inactivating current, fast activating slow inactivating current, and very fast inactivating transient outward current. Physiological role of them needs to be established.     

复方金钱草颗粒利胆及体外抗豚鼠胆囊平滑肌痉挛作用研究

目的探讨复方金钱草颗粒利胆作用及对胆囊平滑肌痉挛的影响。方法利用胆管引流法测定给复方金钱草颗粒前后大鼠胆汁的流量及成分;通过对豚鼠胆囊肌条的观察,探讨复方金钱草颗粒对乙酰胆碱所致胆囊平滑肌痉挛的影响。结果复方金钱草颗粒7．4g／kg剂量组可显著提高大鼠的胆汁流量,显著提高胆汁中胆红素浓度;62g／L剂量组可显著抑制由乙酰胆碱引起的豚鼠胆囊痉挛。结论复方金钱草颗粒具有利胆作用,可缓解乙酰胆碱引起的胆囊痉挛。     

Electrophysiological and Neurochemical Evidence for Voltage‐Dependent Ca2+ Channel Blockade by a Novel Neuroprotective Agent NS‐7

Abstract: The effects of a neuroprotective agent 4‐(4‐fluorophenyl)‐2‐methyl‐6‐(5‐piperidinopentyloxy) pyrimidine hydrochloride (NS‐7) on the Ca²⁺ currents in rat dorsal root ganglion neurones and on the depolarization‐evoked nitric oxide synthesis, which was estimated from cyclic GMP formation in slices of the rat cerebral cortex, were investigated, and its mode of action was compared with those of typical Ca²⁺ channel blockers. In rat dorsal root ganglion neurones, NS‐7 (0.3–100 μM) inhibited the whole‐cell Ba²⁺ currents (I_Ba) in a voltage‐dependent manner, in which the compound more potently blocked the I_Ba elicited from the holding potential of ?40 mV than that induced from ?80 mV. In slices of rat cerebral cortex, KCl‐evoked nitric oxide synthesis was markedly inhibited by ω‐conotoxin GVIA and ω‐agatoxin IVA, but only slightly attenuated by nifedipine, suggesting that the response is mediated predominantly through activation of N‐type and P/Q‐type Ca²⁺ channels. NS‐7 (1–100 μM) inhibited the KCl‐stimulated nitric oxide synthesis in a manner dependent on the intensity of the depolarizing stimuli. Moreover, weak but significant inhibitory effect of NS‐7 was observed even after wash‐out. Similar voltage‐dependent inhibition of the KCl response was observed by a limited concentration (10 μM) of verapamil. These findings indicate that NS‐7 in several concentrations blocks Ca²⁺ channel in a voltage‐dependent manner.     

Effects of Tartrazine on Different Contractile Stimuli in Guinea Pig Tracheal Muscle

Abstract: Tracheal smooth muscle obtained from ovalbumin-sensitized guinea pigs contracted with micromolar concentrations of tartrazine and indomethacin. The contractions were slightly reduced by clemastine, but were completely blocked by the SRS-A antagonist FPL 55712. Arylsulfatase B, which is an enzyme that inactivates SRS-A, also abolished the contractions induced by tartrazine. Contractions induced by carbacholine, histamine, egg albumin and arachidonic acid were slightly reduced by indomethacin and tartrazine. Tartrazine like indomethacin inhibited the generation of thromboxanes from human platelets. We suggest that SRS-A plays an important role in the immediate hypersensitivity reactions of tartrazine and indomethacin in guinea pig trachea.     

豚鼠气道平滑肌的急性分离及K_(ATP)通道单通道电流的记录

目的:建立急性分离豚鼠气道平滑肌的方法,并初步分析ATP敏感钾通道(KATP通道)单通道电流的性质。方法:急性分离出豚鼠3~4级支气管,并用链霉蛋白酶E分散气道平滑肌细胞,应用膜片钳技术的内面向外式记录方法,研究气道平滑肌KATP通道单通道电学性质。结果:成功记录到电导为112.4±5.14 pS的、可被优降糖所阻断的KATP通道单通道电流。钳制电压在0~-60 mV之间,通道电流无整流现象,且未见时间依赖性失活。结论:建立了急性分离豚鼠气道平滑肌的方法,并成功记录KATP通道单通道电流,为进一步研究气道平滑肌KATP通道在呼吸道疾病中的作用提供了基础。     

Icariin Inhibits Hydrogen Peroxide‐Mediated Cytotoxicity by Up‐regulating Sirtuin Type 1‐Dependent Catalase and Peroxiredoxin

Abstract: Previous studies suggest that flavonol icariin protects against neuron injury after oxygen and glucose deprivation by increasing SIRT1. This study demonstrates that icariin can inhibit H₂O₂‐induced neurotoxicity. The neuroprotection of icariin enhances the antioxidant capacity through both a direct scavenging effect on over‐produced free radicals and an indirect stimulating effect on the expression and activity of cellular antioxidant enzymes including catalase (CAT) and peroxiredoxin 1 (Prx1). The mechanism may be partially involved in the up‐regulation of SIRT1. The SIRT1 antagonist can partly block this neuroprotection and the enhancement of CAT/Prx1 by icariin. These results indicate that the effect of icariin on H₂O₂‐induced neurotoxicity is dependent on increasing SIRT1 and provides a potentially novel pharmacological strategy for stroke prevention and/or treatment.     

IQ_(23)对豚鼠心室肌细胞复极和延迟整流钾电流两种成分的作用(英)

采用膜片钳全细胞记录技术,研究了具有抗心律失常作用的苄基异喹啉衍化物IQ23对豚鼠心室肌单细胞动作电位（AP）和钾电流的作用。结果表明：IQ_23在10,30,100μmol·L~-皇呈浓度依赖性的减慢AP复极,APD_90分别延长15％,28％和31％,此效应不依赖细胞外Ca~2 。电压钳制下记录延迟整流钾电流,IQ_23对其两种成分,即I_ks和I_kr,都有阻断作用,30和100mol·L-1IQ_23阻断I_ks达21％和26％,对Ikr为67％和86％。即使在100mol·L-1,IQ_23也不影响内向整流钾电流（Ikl）。本实验表明,IQ23能浓度依赖性的延长心室肌细胞动作电位时程（APD）,此效应与胞外Ca2 无关。1Q23对Iks和Ikr两种成分都有阻断作用,无明显的选择性。     

On the Pharmacological and Physiological Role of Glibenclamide-Sensitive Potassium Channels in Colonic Smooth Muscle

Abstract: Actions of activators of glibenclamide sensitive K⁺ channels on canine colonic circular muscle were investigated. Cromakalim as well as its (-) enantiomer lemakalim caused inhibition of spontaneous phasic contractile activity (EC₅₀'s 4.4 ± 0.1 × 10^-7 M and 2.3 ± 0.4 × 10^-7 M, respectively) and of carbachol induced activity (EC₅₀'s: 9.4 ± 5.1 × 10^-7 M and 4.3 ± 1.4 × 10^-7 M, respectively). Cromakalim and lemakalim effects were completely inhibited by glibenclamide. Additive effects between K⁺ channel activators and other drugs relaxing colonic muscle (the L-type calcium channel blocker D600 and forskolin) were seen. A physiological role for specific glibenclamide sensitive K⁺ channels, if existing, remains unresolved. The present study indicates that the non-adrenergic inhibitory nerves do not act through these channels, neither does stimulation of muscarinic or β-adrenergic receptors.     

Quercetin Glucuronide Inhibits Cell Migration and Proliferation by Platelet-Derived Growth Factor in Vascular Smooth Muscle Cells

Many epidemiologic studies have reported that dietary flavonoids provide protection against cardiovascular disease. Quercetin, a member of the bioflavonoids family, has been proposed to have anti-inflammatory, anti-atherogenic, and anti-hypertensive properties leading to the beneficial effects against cardiovascular diseases. Recent studies demonstrated that orally administered quercetin appeared in plasma as glucuronide-conjugated forms in rats and humans. Therefore, we examined the effect of chemically synthesized quercetin glucuronide on platelet-derived growth factor (PDGF)-induced cell migration and kinase activation in cultured rat aortic smooth muscle cells (RASMCs). PDGF-induced RASMC migration was inhibited by quercetin 3-O-β-D-glucuronide (Q3GA). Q3GA also attenuated PDGF-induced cell proliferation in RASMCs. PDGF activated extracellular-signal regulated kinase (ERK) 1/2, c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein (MAP) kinase, and Akt in RASMCs. PDGF-induced JNK and Akt activations were suppressed by Q3GA, whereas ERK1/2 and p38 MAP kinase activations were not affected. We also confirmed that PDGF-induced JNK and Akt activations were inhibited by antioxidants, N-acetylcysteine and diphenyleneiodonium chloride, in RASMCs. These findings suggest Q3GA would be an active metabolite of quercetin in plasma and may possess preventing effects for cardiovascular diseases relevant to vascular smooth muscle cell disorders.     

葡萄糖对豚鼠心室肌细胞跨膜离子电流的影响

目的观察葡萄糖对豚鼠心室肌细胞膜APD,IK1,IK,ICa-L的影响.方法采用全细胞膜片技术记录单个豚鼠心室肌细胞膜的动作电位时程和离子电流.比较0、10和20mmol·L-1葡萄糖对心室肌细胞跨膜离子电流的作用.结果(1)与10 mmol·L-1葡萄糖相比,0和20mmol·L-1均可使豚鼠心室肌细胞的APD缩短(P＜0.05);(2)在细胞外葡萄糖浓度为10 mmol·L-1时,内向整流钾电流IK1的电流密度最大,标准化I-V曲线显示0和20mmol·L-1葡萄糖均可抑制IK1,并使I-V曲线左移,其翻转电位从-72.4移至-64.6 mV;(3)与mmol·L-1葡萄糖相比,0和20mmol·L-1葡萄糖均可增加ICa-L的电流幅度和电流密度.当钳制电位为10 mV,细胞外葡萄糖浓度为0 mmol·L-1时,ICa-L的电流密度为(-8.03±0.82)pA/pF(n=8),而10mmol·L-1和20mmol·L-1葡萄糖分别使电流密度增加为(-5.45±0.67)pA/pF和(-6.50±0.56)pA/pF;(4)当钳制电压为 70mV,细胞外葡萄糖浓度为0、10和20 mmol·L-1时,IK的电流密度分别为(18.96±2.86)pA/pF,(8.66±1.87)pA/pF,(15.32±3.12)pA/pF.结论细胞外不同浓度的葡萄糖可使豚鼠心室肌细胞APD,IK1,IK,ICa-L发生改变.细胞外葡萄糖浓度为0和20mmol·L-1对细胞膜离子电流产生相似的影响.     

Olibanum Extract Inhibits Vascular Smooth Muscle Cell Migration and Proliferation in Response to Platelet-Derived Growth Factor

Olibanum (Boswellia serrata) has been shown to have anti-inflammatory, anti-arthritic and anti-cancer effects. This study determined the role of a water extract of olibanum in platelet-derived growth factor (PDGF)-stimulated proliferation and migration of rat aortic smooth muscle cells (RASMCs). PDGF-BB induced the migration and proliferation of RASMCs that were inhibited by olibanum extract in a dose-dependent manner. The PDGF-BB-increased phosphorylation of p38 mitogen-activated protein kinase (MAPK); the heat shock protein (Hsp) 27 was significantly inhibited by the olibanum extract. The effects of PDGF-BB-induced extracellular signal-regulated kinase1/2 was not altered by the olibanum extract. Treatment with olibanum extract inhibited PDGF-BB-stimulated sprout out growth of aortic rings. These results suggest that the water extract of olibanum inhibits PDGF-BB-stimulated migration and proliferation in RASMCs as well as sprout out growth, which may be mediated by the inhibition of the p38 MAPK and Hsp27 pathways.     

Cerebral Vascular Smooth Muscle Potassium Channels and their Possible Role in the Management of Vasospasm

Abstract: One of the promising therapeutic uses of the potassium channel openers is in the management of cerebral vasospasm, a prolonged vasoconstriction of major cerebral arteries which follows aneurysmal subarachnoid haemorrhage. In this review, we first summarize the properties of potassium channels in cerebral vascular smooth muscle. Calcium-activated and ATP-dependent potassium channels are the major potassium channels identified in the cerebrovascular smooth muscle and both are believed to play a role in the regulation of cerebrovascular smooth muscle tone. The calcium-activated potassium channels can be activated by depolarization, by elevation of internal calcium and by some vasodilators. Some neuropeptides and potassium channel openers open the ATP-dependent potassium channels and produce vasodilation. We then review the effects of both synthetic and endogenous potassium channel openers in the cerebrovascular system, discuss their efficacy in the management of models of cerebrovascular spasm, and outline the clinical promise of these agents.