Effects of clenbuterol on resting tension and contractile response in vesicourethral smooth muscle of rabbits

The effects of clenbuterol, a selective beta 2-agonist, on isolated smooth muscle preparations from the rabbit bladder body, bladder base and proximal urethra have been investigated. The inhibitory effects on resting tension and acetylcholine- and field stimulation-induced contractions in the bladder body were compared with those of flavoxate, atropine, and verapamil. Clenbuterol (10(-10)-10(-7) M) had a strong, concentration-dependent relaxant effect on resting tension of the bladder body, and the relaxant effect was antagonized by propranolol. However, clenbuterol had little effect on the bladder base or proximal urethra. Isoproterenol, a non-selective beta agonist, gave a similar result, but was less potent than clenbuterol. Flavoxate failed to reduce the resting tension, but rather enhanced the spontaneous rhythmic contraction in a concentration-dependent manner. Atropine had little effect. Verapamil produced a concentration-dependent relaxation in the bladder body. Acetylcholine-induced contraction in the bladder body was completely inhibited by pretreatment with atropine (10(-7) M). Clenbuterol, flavoxate, and verapamil concentration-dependently inhibited acetylcholine-induced contraction. Field stimulation-induced contraction in the bladder body was not completely inhibited by atropine. However, the residual contraction was completely inhibited by tetrodotoxin. Clenbuterol, flavoxate, and verapamil concentration-dependently inhibited field stimulation-induced contraction. The inhibitory effects of clenbuterol and verapamil were antagonized by an application of propranolol and an increase in external Ca, respectively. The data suggest that the selective relaxant effect of clenbuterol on the bladder body is due to beta 2-antagonistic action, resulting in the inhibition of the contractile response to acetylcholine or field stimulation. Also, its response was different from that of the other drugs used.     

Effects of prostaglandins on vesicourethral smooth muscle of rabbit therapeutic implications

The effects of PGF₂-alpha and PGE₂ on the vesicourethral smooth muscle of the rabbit were studied in vitro. PGF₂-alpha had potent contractile effects on the bladder body and comparatively less in the bladder base and the proximal urethra. PGE₂ contractile effects were two times greater than PGF₂-alpha on the bladder body but minimal or absent on the base and the urethra. The effects of PGF₂-alpha and PGE₂ seem to be mediated through a prostaglandin receptor as indicated by competitive antagonism of both prostaglandins by N-0164, a synthetic phenyl phosphonate. It also appears that the effects of PGF₂-alpha and PGE₂ may not be mediated through muscarinic, adrenergic, nicotinic, or histaminic receptors or direct smooth-muscle action. The therapeutic implications of PGE₂ in patients with problems of bladder emptying are discussed.     

粉防己碱对新西兰白兔阴茎海绵体平滑肌细胞内游离钙离子浓度的影响

目的探讨粉防己碱(Tet)松弛阴茎海绵体平滑肌的作用机制。方法体外培养的第3～4代新西兰白兔阴茎海绵体平滑肌细胞,经钙荧光指示剂Fluo-2/AM负载后,用荧光离子数字成像系统观察Tet对平滑肌细胞内[Ca~(2 )]i的影响。结果Tet对平滑肌细胞内静息[Ca~(2 )]i无明显影响(P>0.05)。在细胞外钙浓度为2.5 mmol/L时,1、10和100μmol/LTet对高钾和去氧肾上腺素(PE)引起的细胞内[ca~(2 )]i升高有浓度依赖性地抑制作用。对40 mmol/L KCl引起的细胞内[Ca~(2 )]i升高的抑制率分别为22.0%、41.0%和73.0%(P<0.05)。对10μmol/L PE引起的细胞内[ca~(2 )]i升高的抑制率分别为15.0%、26.4%和46.6%(P<0.05)。在无细胞外钙时,1μmol/L和10μmol/L Tet对PE引起的细胞内[Ca~(2 )]i升高,无明显影响(P>0.05);而100μmol/L Tet能明显抑制PE引起的细胞内[ca~(2 )]i升高,抑制率为28.2%(P<0.05)。结论Tet可能通过阻滞电压依赖性钙通道、α1受体依赖性钙通道和抑制细胞内钙库释放,降低阴茎海绵体平滑肌细胞内[Ca~(2 )]i水平,这是Tet松弛阴茎海绵体平滑肌的作用机制之一。     

Roles of prostaglandin on rabbit vesicourethral smooth muscle contraction]

The effects of prostaglandin (PG) E1, E2 and F2 alpha on isolated smooth muscles of rabbit bladder and urethra were studied by in vitro techniques for recording contractile activities. To examine the mechanism of PGs' effect, intracellular cyclic AMP content was also measured by radioimmunoassay. Spontaneous contractile force of muscle strips isolated from rabbit urinary bladder dome and base was increased dose-dependently by administration of PGE1, E2 or F2 alpha. Isolated muscle strips from bladder dome responded to PG more markedly than those from bladder base. The rank order of potency to induce contractile responses was PGF2 alpha greater than PGE2 greater than PGE1 in both dome and base muscles. Spontaneous contractile force of muscle strips isolated from rabbit urethra was increased dose-dependently by administration of PGF2 alpha, and, in contrast, was decreased dose-dependently by PGE1 or E2. These effects were not affected by pretreatment with atropine, phentolamine, propranolol and tetrodotoxin, but were significantly inhibited by pretreatment with verapamil, a Ca-antagonist. Cyclic AMP accumulation in urethral muscle strips significantly increased after administration of PGE1. These results demonstrated that contractile response of rabbit bladder smooth muscle to PG was mainly induced by Ca2+ influx and that cyclic AMP was related to the relaxation of rabbit urethral smooth muscle by PGE1.     

粉防己碱(Tet)对兔子阴茎海绵体平滑肌细胞质中自由钙离子浓度的影响(英文)

目的:探讨粉防己碱(tetrandrine,Tet)松弛阴茎海绵体平滑肌的作用机制。方法:体外培养新西兰白兔阴茎海绵体平滑肌细胞,经钙荧光指示剂 Fluo-2/AM 负载后,用荧光离子数字成像系统观察 Tet 对平滑肌细胞内[Ca~(2 )]_i 的影响。结果:Tet(1,10,100μmol/L)对平滑肌细胞内静息[Ca~(2 )]_i无明显影响(P>0.05)。当细胞外钙离子浓度为2.5 mmol/L 时,Tet(1μmol/L,10 μmol/L,100 μmol/L)抑制了高钾和去氧肾上腺素(PE)导致的细胞内[Ca~(2 )]_i 升高(P<0.05),这种抑制作用具有浓度依赖性。在无细胞外钙时,1 μmol/L和10μmol/L Tet 对 PE 引起的细胞内[Ca~(2 )]_i 升高无明显影响(P>0.05);而100 μmol/L Tet 能明显抑制 PE 引起的细胞内[Ca~(2 )]_i 升高(P<0.05)。结论:Tet 通过阻滞电压依赖性钙通道、α_1受体依赖性钙通道和抑制细胞内钙库释放,降低阴茎海绵体平滑肌细胞内[Ca~(2 )]_i 水平,这是 Tet 松弛阴茎海绵体平滑肌的作用机制之一。     

Effects of neferine on cytosolic free calcium concentration in corpus cavernosum smooth muscle cells of rabbits

To study the relaxation mechanisms of neferine (Nef) on the corpus cavernosum smooth muscle (CCSM), the CCSM cells from New Zealand White rabbits were cultured in vitro. [Ca(2+)](i) was measured by fluorescence ion digital imaging system (FIDIS), using Fluo-2/AM as a Ca(2+)-sensitive fluorescent indicator. Nef (0.1, 1 and 10 micromol l(-1)) had no effect on the resting [Ca(2+)](i) (P > 0.05). In the presence of extracellular Ca(2+) (2.5 mmol l(-1)), Nef (0.1, 1 and 10 micromol l(-1)) inhibited [Ca(2+)](i) elevation induced by high K(+) and phenylephrine (PE) in a concentration-dependent manner (P < 0.05). In calcium free solution containing egtaic acid (EGTA), Nef (0.1 micromol l(-1)) had no inhibitory effects on [Ca(2+)](i) elevation induced by PE (P > 0.05). However, Nef (1 and 10 micromol l(-1)) inhibited [Ca(2+)](i) elevation induced by PE (P < 0.05). These data suggest that Nef inhibited [Ca(2+)](i) in CCSM cells via blocking voltage-dependent Ca(2+) channel, alpha(1)-adrenoceptor-operated Ca(2+) channel and Ca(2+) release from intracellular Ca(2+) pool. This inhibitory action on [Ca(2+)](i) might be one of the relaxation mechanisms of Nef on the CCSM.     

Effects of verapamil on indirect muscle twitch responses

The effects on indirectly elicited muscle twitch amplitude associated with the calcium (slow) channel blocker, verapamil, with or without pancuronium were investigated using isolated bullfrog sciatic nerve-sartorius muscle preparations. Verapamil (2-8 mM) produced a dose-related depression of indirect muscle twitch height (P less than 0.05). Twitch response was depressed 11% below control by the lowest concentration employed and 86% by the highest concentration. Pancuronium (0.07 mM) depressed neuromuscular function 35% below control (P less than 0.05). The combination of 5 mM or 8 mM verapamil with 0.07 mM pancuronium caused significantly greater degrees of depression than either drug alone. Verapamil produced significant depression of twitch height in vitro in relatively high concentrations. The mechanism of action remains unknown. Verapamil possesses pharmacologic properties that may be unrelated to slow (calcium) channel inhibition. The reduction of muscle twitch height caused by verapamil alone (5 mM) could not be antagonized by neostigmine, calcium, or frequent washings.     

The mode of action of prostaglandin E2, F2 alpha and prostacyclin on vesicourethral smooth muscle

Interactions of prostaglandin E2 (PGE2), F2 alpha (PGF2 alpha) and prostacyclin (PGI2) with Ca2+ on the isometric contraction of rabbit detrusor muscle strips were studied using two types of Ca2+ antagonists of different mechanisms of action: verapamil and sodium nitroprusside (NP). The effects of PGI2 on vesicourethral smooth muscle and their relationship with cholinergic, adrenergic receptors and nervous activity were also investigated. PGE2 and F2 alpha (3 X 10(-8) to 3 X 10(-5) M) caused dose-dependent contraction of the strips. Pretreatment of the strips with verapamil (10(-7) to 10(-5)M) significantly inhibited PGs-induced contraction in a dose-dependent manner, whereas NP(10(-7) to 10(-5)M) failed to suppress the contraction. Relaxation of the preparations once contracted by PGE2 and F2 alpha (3 X 10(-6)M) was induced completely by addition of verapamil (10(-5)M), and incompletely by NP(10(-5) to 10(-3)M). Washing of the strips with Ca2+-free solution containing 0.01 mM EGTA completely eliminated spontaneous activity and diminished basal tension, but replenishment of Ca2+ (0.5 to 10 mM) to the medium caused dose-related contraction and spontaneous activity of the strips. Addition of PGE2 and F2 alpha to the Ca2+-free medium enhanced Ca2+-induced contraction and spontaneous activity during Ca2+ replenishment, which were significantly inhibited by pretreatment with verapamil (10(-7) to 10(-5)M) in a dose-dependent manner, but not affected by NP (10(-7) to 10(-5)M). In Ca2+-free medium containing 0.1 mM EGTA, PGE2 and F2 alpha caused a slight degree of tension increase of the strips dose-dependently at the higher concentration exceeding 3 X 10(-6)M. PGI2 (10(-9) to 3 X 10(-4)M) caused dose-dependent contraction of the strips from the bladder body, base and the urethra. The contractile action of PGI2 was greatest on the bladder body, less on the base and minimal on the urethra. The effect of PGI2 was less potent than those of PGE2 and F2 alpha. The PGI2-induced contraction was slow in onset, short lasting, and not affected by pretreatment with phentolamine, propranolol, atropine, hexamethonium, hemicholinium-3 and tetrodotoxin. The interactions of PGI2 with Ca2+ were similar to those of PGE2 and F2 alpha.(ABSTRACT TRUNCATED AT 400 WORDS)     

黄体酮对家兔输尿管平滑肌电活动和尿流量的影响

目的：探讨黄体酮对输尿管平滑肌电活动和尿流量的影响。方法：采用在体电活动记录观察家兔实验前后输尿管平滑肌电活动及尿流量的变化。结果：黄体酮能显著降低家兔输尿管电活动频率,并使尿流量增加,结论：黄体酮对家兔输尿管平滑肌产生一定负性影响。但对 尿流量却有增加作用。其尿流量增加的作用有助于验尿管结石的排出。     

In vitro study of antispasmodic effects of dicyclomine hydrochloride on vesicourethral smooth muscle of guinea pig and rabbit.

Dicyclomine inhibition of acetylcholine-induced and barium chloride-induced isotonic contractions of the smooth muscle from three segments of the lower urinary tract (bladder body, bladder base, and proximal urethra) of the guinea pig and the rabbit was studied in vitro. In the guinea pig dicyclomine caused competitive inhibition of acetylcholine-induced contraction of the bladder body (1 x 10(-7) M to 1 x 10(-5) M) and the bladder base (1 x 10(-6) M, 1 X 10(-5) M) and was less potent than atropine and propantheline. In the rabbit significant blockade of acetylcholine-induced contractions occurred at dicyclomine concentrations of 5 x 10(-6) M to 3 x 10(-5) M in the bladder body and at 1 x 10(-5) M and 3 x 10(-5) M in the bladder base. In both species dicyclomine inhibitory effects were most marked in the bladder body, moderate in the bladder base, and minimal in the proximal urethra. Dicyclomine failed to cause inhibition of the barium chloride-induced contractions in the guinea pig vesicourethral smooth muscle. In rabbits, however, significant antagonism P less than 0.01) of barium chloride-induced muscle contraction was observed with dicyclomine at concentration 1 x 10(-5) M in both bladder body and the bladder base. The clinical implication of such properties of dicyclomine are discussed.     

瑞芬太尼对人肠系膜小动脉平滑肌细胞L-型钙通道电流的影响

目的观察瑞芬太尼对人肠系膜小动脉平滑肌细胞(MASMCs)L-型钙通道电流的影响,探讨其扩张血管的机制。方法酶解法分离人肠系膜小动脉单个平滑肌细胞,采用全细胞膜片钳技术,以钡电流作为载流子,观察19.4nmol／L瑞芬太尼在不同钳制电压下以及不同浓度瑞芬太尼在最大激活电压下对MASMCs L-型钡电流(TBa-L)的影响。结果19．4 nmol／L瑞芬太尼可抑制TBa-L,使电流密度-电压曲线上移,最大激活电压及翻转电压均向膜的负电位方向移动(P<0．01)。瑞芬太尼浓度依赖性地抑制TBa-L,其半数最大抑制效应的浓度为(39±4)nmol／L。结论瑞芬太尼浓度依赖性地抑制人肠系膜小动脉平滑肌细胞L-型钙通道,从而产生扩张血管的作用。     

Effects of halothane on sarcoplasmic reticulum calcium release channels in porcine airway smooth muscle cells

BACKGROUND: Volatile anesthetics relax airway smooth muscle (ASM) by altering intracellular Ca2+ concentration ([Ca2+]i). The authors hypothesized that relaxation is produced by decreasing sarcoplasmic reticulum Ca2+ content via increased Ca2+ "leak" through both inositol trisphosphate (IP3) and ryanodine receptor channels. METHODS: Enzymatically dissociated porcine ASM cells were exposed to acetylcholine in the presence or absence of 2 minimum alveolar concentration (MAC) halothane, and IP3 levels were measured using radioimmunoreceptor assay. Other cells were loaded with the Ca2+ indicator fluo-3 and imaged using real-time confocal microscopy. RESULTS: Halothane increased IP3 concentrations in the presence and absence of acetylcholine. Inhibition of phospholipase C blunted the IP3 response to halothane. Exposure to 2 MAC halothane induced a transient [Ca2+]i response, suggesting depletion of sarcoplasmic reticulum Ca2+. Exposure to 20 microM Xestospongin D, a cell-permeant IP3 receptor antagonist, resulted in a 45+/-13% decrease in the [Ca2+]i response to halothane compared with halothane exposure alone. In permeabilized cells, Xestospongin D or 0.5 mg/ml heparin decreased the [Ca2+]i response to halothane by 65+/-13% and 68+/-22%, respectively, compared with halothane alone. In both intact and permeabilized cells, 20 microM ryanodine blunted the [Ca2+]i response to halothane by 32+/-13% and 39+/-21%, respectively, compared with halothane alone. Simultaneous exposure to Xestospongin D and ryanodine completely inhibited the [Ca2+]i response to halothane. CONCLUSIONS: The authors conclude that halothane reduces sarcoplasmic reticulum Ca2+ content in ASM cells via increased Ca2+ leak through both IP3 receptor and ryanodine receptor channels. Effects on IP3 receptor channels are both direct and indirect via elevation of IP3 levels.     

缺血对兔血管平滑肌的损伤作用

研究兔断肢血管平滑肌组织中钙、镁离子含量及线粒体形态的变化与术后血循环危象的关系。方法:用原子吸收光谱分光光度计测定血管平滑肌组织中钙、镁离子的含量。用图像分析仪定量分析线粒体的结构变化。结果:缺血时间超过8小时,血管平滑肌组织中钙离子含量明显升高(P<0.01);线粒体明显受损。结论:血管平滑肌组织中钙离子增高在血循危象的发生中起着重要作用。     

Cytosolic calcium dynamics and smooth muscle contraction.III Plasmalemmal calcium fluxes

Smooth muscle contractile activity depends upon cytosolic Ca2+, the Ca(2+)-sensitivity of actin-myosin interaction and several auxiliary mechanisms. This section presents the plasmalemmal Ca2+ fluxes in relation with the functional structure of their supportive proteins and the contractile impact. Summaries of classical data are accompanied by examples of recent advances. Ca2+ influx occurs mainly via the L and T types of voltage-operated Ca2+ channels, the store-operated Ca2+ channels and the non-selective cation channels (operated by membrane receptors or mechanical stimuli). The plasmalemmal Ca2+ ATPase and Na/Ca antiport function to limit increases in cytosolic Ca2+; Na/Ca effect is opposite when driven to operate in the reverse mode.     

Effects of streptozotocin-induced diabetes mellitus on intracellular calcium and contraction of longitudinal smooth muscle from rat urinary bladder

PURPOSE: To examine the effect of diabetes on [Ca2+]i and contractility in longitudinal smooth muscle of the urinary bladder. MATERIALS AND METHODS: Longitudinal smooth muscle strips were isolated from the urinary bladders of rats with STZ-induced diabetes as well as age matched controls. Force and [Ca2+]i were measured simultaneously in muscle strips loaded with the calcium indicator, fura-2. Contractions were initiated by electrical field stimulation (EFS) at various frequencies, as well as by high K+, carbachol (CCh) and cyclopiazonic acid (CPA) in the presence of varying concentrations of extracellular Ca2+. RESULTS: In unstimulated muscles, there was no significant difference in resting [Ca2+]i between the control and diabetic groups. However, the muscle strips from the diabetic animals produced higher force levels in response to EFS, high K+, CCh and CPA than those from control animals. The higher force development in the diabetic muscles was not associated with greater increases in [Ca2+]i, which in fact tended to be lower during stimulation in the diabetic tissues. When stimulated by CCh in the presence of nifedipine, both control and diabetic muscles exhibited a nifedipine-resistant component of contraction, however, this was significantly larger in the diabetic muscles. CONCLUSION: The results suggest that there are no major impairments in either intracellular calcium regulation or contractile function in bladder smooth muscle after 8-weeks of STZ-induced diabetes. However, a non-specific enhancement of force production was seen, which was not associated with increases in [Ca2+]i. These changes imply that the apparent sensitivity to [Ca2+]i is enhanced in bladder smooth muscle from diabetic rats.     

Effects of primary alcohols on airway smooth muscle

BACKGROUND: The investigation examined whether primary alcohols could be used as tools to explore the mechanism of anesthetic actions in airway smooth muscle (ASM). The hypothesis was that, like volatile anesthetics, the primary alcohols relax intact ASM by decreasing intracellular Ca2+ concentration ([Ca2+]i) and by inhibiting agonist-induced increases in the force developed for a given [Ca2+]i (Ca2+ sensitivity). METHOD: The effects of butanol, hexanol, and octanol on isometric force in canine tracheal smooth muscle were examined. The effects of hexanol on [Ca2+]i (measured with fura-2) and the relationship between force and [Ca2+]i were studied during membrane depolarization provided by KCl and during muscarinic stimulation provided by acetylcholine. RESULTS: The primary alcohols relaxed ASM contracted by KCl or acetylcholine in a concentration-dependent manner, with potency increasing as chain length increased. The alcohols could completely relax the strips, even during maximal stimulation with 10 microM acetylcholine (median effective concentrations of 28 +/- 12, 1.3 +/- 0.4, and 0.14 +/- 0.05 mM [mean +/- SD] for butanol, hexanol, and octanol, respectively). Hexanol decreased both [Ca2+]i and force in a concentration-dependent manner. Hexanol decreased Ca2+ sensitivity during muscarinic stimulation but had no effect on the force-[Ca2+]i relationship in its absence. CONCLUSIONS: Primary alcohols produce reversible, complete relaxation of ASM, with potency increasing as chain length increases, by decreasing [Ca2+]i and inhibiting increases in Ca2+ sensitivity produced by muscarinic receptor stimulation. These actions mimic those of volatile anesthetics on ASM, a circumstance suggesting that the primary alcohols may be useful tools for further exploring mechanisms of anesthetic effects on ASM.     

Effects of halothane and diltiazem on L-type calcium currents in single smooth muscle cells from rabbit portal veins

We have studied the effects of halothane and diltiazem on L-typevoltage-dependent calcium currents (I_Ca) in single smooth musclecells from rabbit portal veins using a whole cell voltage clamptechnique. The threshold of I_Ca was –30 mV and the peakcurrent was reached at 0 mV. Halothane (0.25, 0.5, 1.0, 1.5and 2.07%) decreased I_Ca in a concentration-dependent mannerand shifted the I_Ca activation threshold to the depolarizingside. Halothane 2.0% abolished Diltiazem 10^–8–10^–6mol litre^–1, a calcium channel antagonist, also depressedI_Ca in a concentration-dependent manner. Administration of both0.5% halothane and diltiazem 10 mol litre^–1 (concentrationslower than the clinical therapeutic range) abolished I_Ca however,halothane did not exhibit use-dependent inhibition of I_Ca whereasdiltiazem showed partial use-dependency. We conclude that thedecrease in I_Ca produced by halothane is associated with a directvasodilator effect of this anaesthetic, but is not explainedby block of Ca²⁺ channels similar to the action of diltiazem.Furthermore, administration of low concentrations of both halothaneand diltiazem decreased I_Ca and may reduce the contractilityof the vascular smooth muscle cells.