Serotonin reuptake inhibitors fluoxetine and citalopram relax intestinal smooth muscle

Selective serotonin reuptake inhibitor antidepressants (SSRIs) exert depressant effects on cardiac myocytes and vascular smooth muscle cells by inhibiting Ca2+ channels. We hypothesized that the SSRIs fluoxetine and citalopram affect the contractile activity of intestinal smooth muscle by interfering with Ca2+ entry and (or) signaling pathways. The effects of fluoxetine and citalopram on contractions of guinea-pig ileum longitudinal muscle-myenteric plexus preparations (LMMP) were compared with the effects of the voltage-operated Ca2+ channel inhibitors nifedipine and diltiazem. In a concentration-dependent manner, nifedipine, diltiazem, fluoxetine, and citalopram elicited relaxation of LMMPs contracted by electrical field stimulation (EC50 values of 4 x 10(-7) M, 1.4 x 10(-6) M, 1.4 x 10(-5), and 6.8 x 10(-6) M, respectively). Nifedipine, diltiazem, fluoxetine, and citalopram also relaxed LMMPs contracted with a depolarizing concentration of KCl (48 mM; EC50 values of 1.8 x 10(-8) M, 1.4 x 10(-7) M, 3.7 x 10(-6) M, and 6.3 x 10(-6), respectively), a response that could be reversed by increasing the extracellular Ca2+ concentration (2.5-30 mM). These data suggest that fluoxetine and citalopram elicit relaxation of intestinal smooth muscle, likely by inhibiting Ca2+ channel(s). This effect may be of clinical importance.     

Hypothyroid state reduces calcium channel function in 18-day pregnant rat uterus

Hypothyroidism significantly reduced the mean amplitude and increased the mean frequency of spontaneous rhythmic contractions in 18 day pregnant rat uterus. Nifedipine (10(-12)-10(-9) M) and diltiazem (10(-10)-10(-6) M) caused concentration related inhibition of the myogenic responses of the uterine strips obtained from both pregnant and hypothyroid state. However, nifedipine was less potent (IC50:2.11 x 10(-11) M) in pregnant hypothyroid state as compared to pregnant control (IC50: 3.1 x 10(-12) M). Similarly, diltiazem was less potent (IC50: 3.72 x 10(-9) M) in inhibiting the uterine spontaneous contractions in hypothyroid than in pregnant rat uterus (IC50:5.37 x 10(-10) M). A similar decrease in the sensitivity to nifedipine and diltiazem for reversal of K+ (100 mM)-induced tonic contraction and K(+)-stimulated 45Ca2+ influx was observed with these calcium channel antagonists in uterus obtained from hypothyroid pregnant rats compared to the controls. Nifedipine-sensitive influx of 45Ca(2+)-stimulated either by K+ (100 mM) or by Bay K8644 (1,4-dihydro-2,6-methyl-5-nitro-4-[2'-(trifluromethyl)phenyl]-3-pyridine carboxylic acid methyl ester) (10(-9) M) was significantly less in uterine strips from hypothyroid rats compared to controls. The results suggest that the inhibition of uterine rhythmic contractions may be attributable to a reduction in rat myometrial Ca2+ channel function in the hypothyroid state.     

Cellular mechanisms involved in iso-osmotic high K+ solutions-induced contraction of the estrogen-primed rat myometrium

The aim of the present study was to investigate the mechanisms involved in the contraction evoked by iso-osmotic high K+ solutions in the estrogen-primed rat uterus. In Ca2+-containing solution, iso-osmotic addition of KCl (30, 60 or 90 mM K+) induced a rapid, phasic contraction followed by a prolonged sustained plateau (tonic component) of smaller amplitude. The KCl (60 mM)-induced contraction was unaffected by tetrodotoxin (3 microM), omega-conotoxin MVIIC (1 microM), GF 109203X (1 microM) or calphostin C (3 microM) but was markedly reduced by tissue treatment with neomycin (1 mM), mepacrine (10 microM) or U-73122 (10 microM). Nifedipine (0.01-0.1 microM) was significantly more effective as an inhibitor of the tonic component than of the phasic component. After 60 min incubation in Ca2+-free solution containing 3 mM EGTA, iso-osmotic KCl did not cause any increase in tension but potentiated contractions evoked by oxytocin (1 microM), sodium orthovanadate (160 micrM) or okadaic acid (20 microM) in these experimental conditions. In freshly dispersed myometrial cells maintained in Ca2+-containing solution and loaded with indo 1, iso-osmotic KCl (60 mM) caused a biphasic increase in the intracellular Ca2+ concentration ([Ca2+]i). In cells superfused for 60 min in Ca2+-free solution containing EGTA (1 mM), KCl did not increase [Ca2+]i. In Ca2+-containing solution, KCl (60 mM) produced a 76.0 +/- 16.2% increase in total [3H]inositol phosphates above basal levels and increased the intracellular levels of free arachidonic acid. These results suggest that, in the estrogen-primed rat uterus, iso-osmotic high K+ solutions, in addition to their well known effect on Ca2+ influx, activate other cellular processes leading to an increase in the Ca2+ sensitivity of the contractile machinery by a mechanism independent of extracellular Ca2+.     

Vasodilator action of docosahexaenoic acid (DHA) in human coronary arteries in vitro

Coronary arterial tissues obtained from mammalian hearts are known to develop spontaneous phasic contractions. The aim of the present study was to investigate the vasodilatory effects of docosahexaenoic acid (DHA) on the rhythmic contractions of isolated human coronary arterial (HCA) preparations obtained from the recipient hearts of patients undergoing cardiac transplantation. Results from 8 hearts show that: (i) most HCA tissues displayed spontaneous rhythmic phasic contractions with a cycle length around 10 min in the absence or presence of PGF2alpha or elevated [K+]0 (20 mM); (ii) the rhythmic activity could be suppressed by a free fatty acid DHA (30 microM); (iii) high [K+]0 (20 and 80 mM) could induce sustained tonic contraction in addition to phasic contractions in HCA tissues, the tonic contraction could be antagonized by L-type Ca(2+) channel blockers or by DHA (depending on [K+]0); (iv) a digitalis substance ouabain also could induce tonic contraction and suppress phasic contraction; (v) in isolated HCA vascular smooth muscle cells, DHA increased the magnitude of outward voltage-gated K+ (IKV) currents and the inwardly rectifying IK1 currents. Enhancement of K+ currents could be related to vasorelaxation induced by DHA in HCA preparations. Further studies on the effects of DHA on various ionic currents and intracellular Ca(2+) transient are needed to clarify the Ca(2+)-dependent and the Ca(2+)-independent actions of DHA in HCA.     

铁对血管收缩活动的影响及其机制

动脉粥样硬化的发生和铁引起的氧化应激密切相关。铁对血管的直接效应及其对血管收缩功能的影响尚不明确。本文采用血管环灌流装置 ,观察铁对离体SD大鼠去内皮胸主动脉环的直接效应 ,及对去内皮主动脉环KCl和苯肾上腺素 (PE)引发的收缩效应的影响。结果显示 :( 1) 10 0 μmol/L枸橼酸铁 (FAC)引起大鼠血管环发生相位性收缩 ,最大收缩幅度可达KCl诱发的最大收缩的 2 4 0 2± 2 3 7%。当 [Ca2 +]o 增加 1倍时 ,铁所致的血管环收缩幅度明显增加 (P <0 0 1)。阻断L 型钙通道后 ,铁所致的血管环收缩幅度明显降低 (P <0 0 1)。在无钙液中 ,用佛波酯收缩血管环 ,待收缩稳定后给予FAC ,此时收缩幅度增加 49 18± 3 75 %。 ( 2 )铁孵育 3 0min后 ,KCl引起血管环收缩的幅度显著降低 (P <0 0 1)。铁孵育可使PE引起的收缩量 -效曲线右移 (P <0 0 5 )。 ( 3 )二甲基亚砜、过氧化氢酶和谷胱甘肽可明显降低铁对PE血管收缩反应的抑制作用 (P <0 0 5 )。从这些结果可得到以下结论 :铁可引起胸主动脉发生相位性收缩 ,其机制可能与L 型钙通道短暂开放导致钙离子内流 ,及平滑肌对钙的敏感性增加有关 ;较长时间与铁孵育后 ,可对血管收缩功能产生损伤 ,氧自由基的生成增加和细胞内GSH的水平降低可能参与铁对收缩功能的     

Studies on the stereoselective effects of a novel 5-HT2 receptor antagonist on contractile responses of rat aorta

The effect of the enantiomers of a novel 5-HT2 receptor antagonist, (+/-)-(1R,3S)-1-[2-[4-[3-(p-fluorophenyl)-1-indanyl]-piperazinyl] ethyl]-2-imidazolidinone, was studied on serotonin (5-HT), noradrenaline (NA), potassium (K+), and calcium (Ca2+)-induced contractions in isolated rat thoracic aorta. The enantiomers shifted the 5-HT, NA, K+, and Ca2+ concentration-response curves to the right in a concentration-dependent manner and depressed the maximal contractile responses. The (+)-enantiomer was a far more potent inhibitor of 5-HT-induced contractions than the (-)-enantiomer. The (+)-enantiomer and phentolamine, both at 10(-6) M, had equal inhibitory effects on NA-evoked contractions. The (+)-enantiomer was again more potent inhibiting NA-induced contractions than the (-)-enantiomer. Both enantiomers had an equieffective inhibitory effect on K+ and Ca2(+)-induced contractions. The results show that the 5-HT and alpha-adrenoceptor antagonism of the two enantiomers is stereoselective, the (+)-enantiomer being more potent than the (-)-enantiomer. In contrast the enantiomers had equal, nonstereoselective inhibitory effects on K+ and Ca2(+)-evoked contractions.     

The vasodilator effect of thiamylal in dog mesenteric artery: contribution of intracellular action.

The effects of thiamylal on contractions induced by various mechanisms were investigated in mesenteric arteries isolated from dogs. Thiamylal (10(-4) to 10(-3) M) significantly inhibited contractions induced by KCl (20 mM) in normal media, and those induced by norepinephrine (10(-5) M) in normal and Ca(2+)-free media. Caffeine-induced contraction was significantly inhibited by thiamylal in the concentrations greater than 3 x 10(-5) M in intact fibers and 10(-5) M in chemically skinned fibers. Chemically skinned fibers that were precontracted with Ca2+ were relaxed by thiamylal in concentrations lower than those required to relax intact fibers that were precontracted with KCl (20 mM); the ED50 was 1.52 x 10(-5) M in skinned fibers and 5.50 x 10(-4) M in intact fibers. These results suggest that intracellular mechanisms are involved in thiamylal-induced vasodilatation of dog mesenteric artery.     

Contribution of sarcolemmal sodium-calcium exchange and intracellular calcium release to force development in isolated canine ventricular muscle

The aim of this work was to determine the relationship between peak twitch amplitude and sarcoplasmic reticulum (SR) Ca2+ content during changes of stimulation frequency in isolated canine ventricle, and to estimate the extent to which these changes were dependent upon sarcolemmal Na(+)-Ca2+ exchange. In physiological [Na+]o, increased stimulation frequency in the 0.2-2-Hz range resulted in a positive inotropic effect characterized by an increase in peak twitch amplitude and a decrease in the duration of contraction, measured as changes in isometric force development or unloaded cell shortening in intact muscle and isolated single cells, respectively. Action potentials recorded from single cells indicated that the inotropic effect was associated with a progressive decrease of action potential duration and a marked reduction in average time spent by the cell near the resting potential during the stimulus train. The frequency-dependent increase of peak twitch force was correlated with an increase of Ca2+ uptake into and release from the SR. This was estimated indirectly using the phasic contractile response to rapid (less than 1 s) lowering of perfusate temperature from 37 degrees C to 0-2 degrees C and changes of twitch amplitude resulting from perturbations in the pattern of electrical stimulation. Lowering [Na+]o from 140 to 70 mM resulted in an increase of contractile strength, which was accompanied by a similar increase of apparent SR Ca2+ content, both of which could be abolished by exposure to ryanodine (1 x 10(-8) M), caffeine (3 x 10(-3) M), or nifedipine (2 x 10(-6) M). Increased stimulation frequency in 70 mM [Na+]o resulted in a negative contractile staircase, characterized by a graded decrease of peak isometric force development or unloaded cell shortening. SR Ca2+ content estimated under identical conditions remained unaltered. Rate constants derived from mechanical restitution studies implied that the depressant effect of increased stimulation frequency in 70 mM [Na+]o was not a consequence of a decreased rate of refilling of a releasable pool of Ca2+ within the cell. These results demonstrate that frequency-dependent changes of contractile strength and intracellular Ca2+ loading in 140 mM [Na+]o require the presence of a functional sarcolemmal Na(+)-Ca2+ exchange process. The possibility that the negative staircase in 70 mM [Na+]o is related to inhibition of Ca(2+)-induced release of Ca2+ from the SR by various cellular mechanisms is discussed.     

Spasmolytic action of the methanol extract and isojuripidine from Solanum asterophorum Mart. (Solanaceae) leaves in guinea-pig ileum

Solanum asterophorum Mart. (Solanaceae) is a shrub popularly known as "jurubeba-defogo" in the northeast of Brazil. In the present work, the methanol extract (SA-MeOH, 3750 microg/mL) and isojuripidine (10(-7) - 3 x 10(-4) M), a steroidal alkaloid obtained from S. asterophorum Mart. leaves, inhibited phasic contractions induced by both 1 microM histamine [IC50 = (225.8 +/- 47.4), g/mL and (3.5 +/- 0.8) x 10(-5) M] or 1 microm acetylcholine [IC50 = (112.5 +/- 20.6) microg/mL and (2.3 +/- 0.4) x 10(-5) M] in guinea-pig ileum, respectively. The extract and isojuripidine also relaxed the ileum (SA-MeOH, 1-750 microg/mL, and isojuripidine, 10(-9) - 3 x 10(-4) M) pre-contracted with 1 M histamine [EC50 = (101.1 +/- 17.4) microg/mL and (1.2 +/- 0.3) x 10(-6) M] or 1 microM acetylcholine [EC50 = (136.8 +/- 21.1) microg/mL and (1.9 +/- 0.4) x 10(-6) M] or 40 mm KCl [EC50 = (149.4 +/- 19.5) microg/mL and (1.8 +/- 0.7) x 10(-6) M], respectively, in an equipotent and concentration-dependent manner. This effect is probably due to inhibition of calcium influx through voltage-operated calcium (Ca(v)) channels. To confirm this hypothesis, we evaluated their effect on cumulative CaCl2 curves in depolarizing medium nominally without Ca2+. SA-MeOH (27, 243, 500, and 750 microg/mL) and isojuripidine (3 x 10(-8), 10(-6), 3 x 10(-5), and 3 x 10(-4) M) inhibited the contractions induced by CaCl2, in a concentration-dependent manner. The concentration-response curves to CaCl2, in the presence of SA-MeOH and isojuripidine, were shifted downward in relation to a control curve in a non-parallel manner resulting in reduction of the maximum effect [E(max) = (71.2 +/- 9.2); (57.4 +/- 9.2); (43.8 +/- 3.4); (41.5 +/- 2.4) and (90.6 +/- 4.8); (74.7 +/- 8.7); (66.4 +/- 3.9); (31.3 +/- 4.1)%, respectively]. SA-MeOH and isojuripidine present spasmolytic action in guinea-pig ileum due to a partially blockade of calcium influx through Ca(v) channels.     

Ryanodine Inhibits Caffeine-Evoked Ca2+ Mobilization and Catecholamine Secretion from Cultured Bovine Adrenal Chromaffin Cells

The effects of ryanodine, a selective inhibitor of the Ca(2+)-induced Ca2+ release mechanism, on caffeine-evoked changes in cytosolic Ca2+ concentration ([Ca2+]i) and catecholamine secretion were investigated using cultured bovine adrenal chromaffin cells. Caffeine (5-40 mM) caused a concentration-dependent transient rise in [Ca2+]i and catecholamine secretion in Ca2+/Mg(2+)-free medium containing 0.2 mM EGTA. Ryanodine (5 x 10(-5) M) alone had no effect on either [Ca2+]i or catecholamine secretion. Although the application of ryanodine plus caffeine caused the same increase in both [Ca2+]i and catecholamine secretion as those induced by caffeine alone, ryanodine (4 x 10(-7) - 5 x 10(-5) M) irreversibly prevented the increase in both [Ca2+]i and catecholamine secretion resulting from subsequent caffeine application over a range of concentrations. The secretory response to caffeine was markedly enhanced by replacement of Na+ with sucrose in Ca2+/Mg(2+)-free medium, and this enhanced response was also blocked by ryanodine. Caffeine was found to decrease the susceptibility of the secretory apparatus to Ca2+ in digitonin-permeabilized cells. These results indicate that caffeine mobilizes Ca2+ from intracellular stores, the function of which is irreversibly blocked by ryanodine, resulting in the increase in catecholamine secretion in the bovine adrenal chromaffin cell.     

Effect of different calcium modulators on motilin-induced contractions of the rabbit duodenum. Comparison with acetylcholine

Motilin and acetylcholine (ACh) have a direct contractile effect on rabbit small intestinal smooth muscle. To explore the role of calcium influx in these contractions, we studied the effect of extracellular calcium concentration and of calcium antagonists on the response of longitudinal muscle preparations from rabbit duodenum. Motilin- (10(-7) M) and ACh- (10(-4) M)-induced contractions were abolished in Ca2+-depleted medium. ACh (10(-4) M) or motilin (10(-8) and 10(-7) M) increased the contractile response to added Ca2+ to 130 +/- 6%, 129 +/- 10% and 145 +/- 5% of the maximal response to Ca2+ added alone (10 mM in a cumulative concentration response curve). The sensitivity to Ca2+ was greater in the presence of ACh and motilin (EC50 = 1.0 and 1.1 mM Ca2+) than in the absence of any agonist (1.7 mM). In cumulative concentration response (CCR) curves for motilin and ACh, pD2'-values were 7.0 and 6.6 for diltiazem, 8.4 and 7.8 for verapamil (two calcium entry blockers), 5.6 and 5.2 for TMB-8 (an inhibitor of intracellular calcium), 5.3 and 5.2 for TFP (a calmodulin-antagonist). All CCR-curves showed metactoid-like action of the antagonistic drugs. We conclude that ACh and motilin cause calcium to enter the smooth muscle cell. They are probably operating via separate channels, and use a mechanism which differs from K+-induced influx. Intracellular calcium stores appear to play a minor role in these contractions.     

Modifications evoked by piretanide in the mechanical activity of cardiovascular tissues

The effects of piretanide upon mechanical activity and pharmacological reactivity of vascular and myocardial tissues from normotensive rats were investigated. Magnitude of phasic contractions of isolated rat portal vein was diminished by the drug in a dose-related manner; contractile depression induced by piretanide (10(-4)M) was less in the presence of insulin (0.1 U/mL), glucose (22 mM) or pyruvate (5 mM). Responses to KCl (90 mM), or norepinephrine (2.5 X 10(-5)M) were also reduced. Contractile activity of atria and ventricle strips was diminished only when piretanide reached 10(-4)M. Results support direct actions of piretanide upon cardiac and vascular tissues. Possible mechanisms of action are discussed.     

Role of Dendritic Dopamine of the Substantia Nigra in the Modulation of Nigrocollicular γ-Aminobutyric Acid Release: In Vivo Studies in the Rat

The release of gamma-[3H]aminobutyric acid ([3H]GABA) newly synthesized from [3H]glutamine was estimated in the superior colliculus of ketamine-anesthetized rats superfused via a push-pull cannula. A significant amount of [3H]GABA was spontaneously released in the superior colliculus (582 +/- 49 pCi/10 min). A major part of the large K(+)-evoked increase of the [3H]GABA release was Ca2+ dependent. When neuronal activity of the substantia nigra was enhanced by nigral application of K+ (30 mM) or bicuculline (10(-4) M), a persistent increase of the collicular [3H]GABA release was observed (60 and 80%, respectively). Conversely, when nigral activity was reduced by nigral application of GABA (10(-4) M) or superfusion with a Ca(2+)-free medium, a sustained decrease of the collicular [3H]GABA release was observed (-30 and -40%, respectively). Following the nigral application of a selective D2-receptor agonist. RU 24926 (10(-6) M), for 30 min in 6-hydroxydopamine-lesioned rats, a phasic increase (60%) of the collicular [3H]GABA release was detected. This effect could result from an activation of nigrocollicular GABAergic neurons by D2-receptor stimulation, because nigral activity and collicular release of [3H]GABA changed in a parallel direction.     

Primary role of sarcoplasmic reticulum in phasic contractile activation of cardiac myocytes with shunted myolemma

Homogeneous populations of single myocytes showing good preservation of ultrastructure were obtained by enzymatic digestion of rabbit and rat hearts, and maintained in a relaxed state in the presence of free Ca2+ concentrations less than 10(-7) M. Ultrastructural details such as a cytoskeleton of 100-A filaments connected to the sarcolemma at the Z lines were demonstrated especially well in these preparations. In spite of seemingly normal structure, electron probe analysis of cryosections reveals similar concentrations of electrolytes in the medium and in the cytoplasm, indicating the presence of electrochemical shunting across the external membrane. The dissociated myocytes display Ca uptake and phasic contractions that are apparently dependent on mitochondrial respiration, but are not affected by mitochondrial uncouplers when ATP and phosphocreatine are added. The uptake is augmented by oxalate and, based on identification of calcium oxalate crystals by electron microscopy and electron probe analysis, is localized to the sarcoplasmic reticulum (SR). An advantageous feature of the dissociated myocytes is that they are suitable for experiments using large numbers of cells in suspension. Thereby, velocities of calcium transport were measured directly by isotopic tracer and filtration methods. It was then found that the lowest CA2+ concentrations (5 x 10(-7) M for the rabbit and 1 x 10(-7) M for the rat) sustaining Ca transport also induce phasic contractile activity in all myocytes, even though the external membrane is electrochemically shunted. A stepwise rise in the Ca2+ concentration of up to one order of magnitude, increases transport velocities in parallel with the rates of phasic contractions. Both these parameters are affected by Mg2+, temperature, cyclic-AMP, and methylxanthines, even though the Ca2+ concentration is maintained constant in the medium. Therefore, Ca transport by SR is a requirement and a rate limiting factor for the occurrence of phasic contractile activation in dissociated cardiac cells retaining an electrochemically shunted external membrane. It is suggested that transient Ca release required for phasic contractile activation is due to equilibrium oscillations across the SR membrane. The sequential pattern of sarcomere activation is consistent with a self propagating mechanism of calcium release. SR in dissociated skeletal muscle cells sustains a greater Ca transport activity than in dissociated heart cells. However, the heart cells display a much higher phasic contractile activity, indicating that cardiac SR has a greater tendency to release accumulated calcium. If free Ca2+ in the medium is raised above 10(-6) M, both cardiac and skeletal myocytes undergo contractures and degenerative phenomena, accompanied by Ca, Mg, and phosphate accumulation in cardiac mitochondria.     

NO(+) but not NO radical relaxes airway smooth muscle via cGMP-independent release of internal Ca(2+)

We compared the effects of two redox forms of nitric oxide, NO(+) [liberated by S-nitroso-N-acetyl-penicillamine (SNAP)] and NO. [liberated by 3-morpholinosydnonimine (SIN-1) in the presence of superoxide dismutase], on cytosolic concentration of Ca(2+) ([Ca(2+)](i); single cells) and tone (intact strips) obtained from human main stem bronchi and canine trachealis. SNAP evoked a rise in [Ca(2+)](i) that was unaffected by removing external Ca(2+) but was markedly reduced by depleting the internal Ca(2+) pool using cyclopiazonic acid (10(-5) M). Dithiothreitol (1 mM) also antagonized the Ca(2+) transient as well as the accompanying relaxation. SNAP attenuated responses to 15 and 30 mM KCl but not those to 60 mM KCl, suggesting the involvement of an electromechanical coupling mechanism rather than a direct effect on the contractile apparatus or on Ca(2+) channels. SNAP relaxations were sensitive to charybdotoxin (10(-7) M) or tetraethylammonium (30 mM) but not to 4-aminopyridine (1 mM). Neither SIN-1 nor 8-bromoguanosine 3',5'-cyclic monophosphate had any significant effect on resting [Ca(2+)](i), although both of these agents were able to completely reverse tone evoked by carbachol (10(-7) M). We conclude that NO(+) causes release of internal Ca(2+) in a cGMP-independent fashion, leading to activation of Ca(2+)-dependent K(+) channels and relaxation, whereas NO. relaxes the airways through a cGMP-dependent, Ca(2+)-independent pathway.     

Inhibitor effect of omeprazole in isolated human myometrial smooth muscle.

The aim of the present study was to investigate the effect of omeprazole, an H+-K+-ATPase inhibitor, in myometrial smooth muscle strips from women undergoing elective caesarean section at term. Isolated myometrial strips taken with informed consent were obtained from eight pregnant women undergoing elective caesarean section at term (not in labour) and mounted in organ baths for recording of isometric tension. We recorded the effect of increasing concentrations of omeprazole on spontaneous and Ca2+-induced contractions of myometrial smooth muscle and on contractions of myometrial smooth muscle pretreated with indomethacin (3 x 10(-6) M) and L-NAME (3 x 10(-5) M). Omeprazole (10(-4)-10(-3) M) decreased the amplitude and frequency of spontaneous contractions in a time- and concentration-dependent manner in all myometrial smooth muscle isolated from pregnant women. The decrease in amplitude of contractions in myometrial smooth muscle reached statistical significance beginning from the concentration of 3 x 10(-4) M. Addition of indomethacin (3 x 10(-6) M) and L-NAME (3 x 10(-5) M) in to the organ baths 30 min before did not change relaxation responses to omeprazole. When 8 mM Ca2+-precontracted in Ca2+-free medium myometrial smooth muscle were exposed to increasing concentrations of omeprazole (10(-5)-10(-3) M), omeprazole produced relaxation responses in a time- and concentration-dependent manner, reaching statistical significance at 10(-4) M. These results show: (1) omeprazole time- and concentration-dependently decreased spontaneous contractile activity in myometrial smooth muscle isolated from pregnant women, (2) omeprazole-induced relaxations was not influenced by indomethacin and N(G)-nitro-L-arginine methyl ester (L-NAME), suggesting that it is not mediated by cyclooxygenase products and nitric oxide, and (3) omeprazole brought about time- and concentration-dependently relaxation of myometrial smooth muscle precontracted by 8 mM Ca2+ in Ca2+-free medium. This effect of omeprazole may be due to blockade of the calcium channels.     

Effects of palmatine on isometric force and intracellular calcium levels of arterial smooth muscle

The effects of palmatine on isometric force and intracellular free calcium levels ([Ca2+]i) were determined in isolated rat arterial strips. Palmatine dose-dependently relaxed the contractile responses stimulated by phenylephrine (PE) in aortic strips. In contrast, it only partially relaxed aortic strips contracted by 51 mM KCl. Pretreatment with palmatine shifted the dose-response curves of PE both rightwards and downwards in a dose-dependent manner. When Ca2+-free solution and re-addition of Ca2+ were applied to assess PE-induced phasic and tonic contractions, palmatine was found to be effective in inhibiting both contractions. The effects of palmatine on intracellular calcium levels were measured with the bioluminescent calcium indicator aequorin in rat tail artery strips. Palmatine caused a concomitant, dose-dependent decrease in PE-activated isometric force and [Ca2+]i, resulting in small changes in the [Ca2+]i-force relationship. These results suggest that vasodilatory effect of palmatine was mediated by reducing [Ca2+]i as well as affecting [Ca2+]i sensitivity of the contractile apparatus. Palmatine-induced [Ca2+]i decreases appeared to involve decreases in both Ca2+ release from intracellular stores and Ca2+ influx through calcium channels.     

Calcium pools mobilized during excitation-contraction coupling in flounder intestinal smooth muscle

The effects of Ca2+ free solutions (0 mM Ca2+/5mM EGTA) and low Ca2+ media (no added Ca2+) on the 100 mM K+ and 10(-5) M ACh contractions of the flounder intestine were examined. Ca2+-free solutions abolished the K+-contractions and reduced the normal ACh response to a smaller transient event. Low Ca2+ media blocked the prolonged tonic phase of the K+-contractions more readily than the initial phasic component. It was concluded that both ACh and K+ stimulate Ca2+ entry into the cell and that the phasic component of the K+-contraction relies on a more tightly bound extracellular Ca2+-fraction than that utilized during the tonic phase. ACh can also mobilize an intracellular Ca2+-store. Ultrastructural studies suggested that this ACh-releasable intracellular Ca2+-store may reside on the inner surface of the plasma membrane or within the peripheral S.R.     

Mechanism of cGMP contribution to the vasodilator response to NO in rat middle cerebral arteries

This study examined the mechanism by which cGMP contributes to the vasodilator response to nitric oxide (NO) in rat middle cerebral arteries (MCA). Administration of a NO donor, diethylaminodiazen-1-ium-1,2-dioate (DEA-NONOate), or 8-bromo-cGMP (8-BrcGMP) increased the diameter of serotonin-preconstricted MCA by 79 +/- 3%. The response to DEA-NONOate, but not 8-BrcGMP, was attenuated by iberiotoxin (10(-7) M) or a 80 mM high-K(+) media, suggesting that activation of K(+) channels contributes to the vasodilator response to NO but not 8-BrcGMP. The effects of NO and cGMP on the vasoconstrictor response to Ca(2+) were also studied in MCA that were permeabilized with alpha-toxin and ionomycin. Elevations in bath Ca(2+) from 10(-8) to 10(-5) M decreased the diameter of permeabilized MCA by 76 +/- 5%. DEA-NONOate (10(-6) M) and 8-BrcGMP (10(-4) M) blunted this response by 60%. Inhibition of guanylyl cyclase with 1H-[1,2,4]oxadiazole[4,3-a] quinoxalin-1-one (10(-5) M) blocked the inhibitory effect of the NO donor, but not 8-BrcGMP, on Ca(2+)-induced vasoconstriction. 8-BrcGMP (10(-4) M) had no effect on intracellular Ca(2+) concentration ([Ca(2+)](i)) in control, serotonin-stimulated, or alpha-toxin- and ionomycin-permeabilized vascular smooth muscle cells isolated from the MCA. These results indicate that the vasodilator response to NO in rat MCA is mediated by activation of Ca(2+)-activated K(+) channels via a cGMP-independent pathway and that cGMP also contributes to the vasodilator response to NO by decreasing the contractile response to elevations in [Ca(2+)](i).     

The electrophysiological and mechanical effects of atrial natriuretic peptide and acetylcholine on guinea pig ventricular papillary muscle

The direct effects of atrial natriuretic factor (ANF) and acetylcholine (ACh) on isolated guinea pig ventricular papillary muscle were studied. ANF (3 x 10(-9) - 3 x 10(-7) M), a cardiogenic hormone, had no significant electrical or mechanical effects on guinea pig papillary muscle driven at a frequency of 60 beats/min in normal (4 mM) and high [K]0 (27 mM) Tyrode solutions. On the other hand, ACh (3 x 10(-8) - 3 x 10(-7) M) caused a significant shortening of action potential duration and the contractile force showed no change or a slight decrease. At high concentration (5 microM), ACh reduced action potential durations at 50% and 90% repolarization (APD50 and APD90) by 10.5 +/- 2.1% and 12.4 +/- 1.8%, respectively, but the contractile force was slightly increased by 9.8 +/- 1.2%. In eleven of twenty-six preparations, spontaneous activity occurred and intermingled with driven activity. The ectopic rhythms were suppressed by ACh (1-5 microM). The changes in electrical but not mechanic activity induced by ACh were suppressed in the presence of five micromolar atropine. These results reveal that, in guinea pig papillary muscle, ANF had no direct chronotropic or inotropic effect. ACh may reduce APD and spontaneous discharges through an activation of muscarinic receptors but enhance twitch tension through other mechanisms.