Forskolin Changes the Relationship between Cytosolic Ca2+ and Contraction in Guinea Pig Ileum

This study was designed to clarify the mechanism of the inhibitory effect of forskolin on contraction, cytosolic Ca2+ level ([Ca2+]i), and Ca2+ sensitivity in guinea pig ileum. Forskolin (0.1 nM~10 micrometer) inhibited high K+ (25 mM and 40 mM)- or histamine (3 micrometer)-evoked contractions in a concentration-dependent manner. Histamine-evoked contractions were more sensitive to forskolin than high K+-evoked contractions. Spontaneous changes in [Ca2+]i and contractions were inhibited by forskolin (1 micrometer) without changing the resting [Ca2+]i. Forskoln (10 micrometer) inhibited muscle tension more strongly than [Ca2+]i stimulated by high K+, and thus shifted the [Ca2+]i-tension relationship to the lower-right. In histamine-stimulated contractions, forskolin (1 micrometer) inhibited both [Ca2+]i and muscle tension without changing the [Ca2+]i-tension relationship. In alpha-toxin-permeabilized tissues, forskolin (10 micrometer) inhibited the 0.3 micrometer Ca2+-evoked contractions in the presence of 0.1 mM GTP, but showed no effect on the Ca2+-tension relationship. We conclude that forskolin inhibits smooth muscle contractions by the following two mechanisms: a decrease in Ca2+ sensitivity of contractile elements in high K+-stimulated muscle and a decrease in [Ca2+]i in histamine-stimulated muscle.

Spasmolytic activity of lapachol and its derivatives, alpha and beta-lapachone, on the guinea-pig ileum involves blockade of voltage-gated calcium channels / Atividade espasmolítica do lapachol e seus derivados, alfa e beta-lapachona, em íleo de cobaia envolve bloqueio dos canais de cálcio dependentes de voltagem

O lapachol, alfa e beta-lapachona são o naftoquinonas obtidas de espécies de Tabebuia, apresentam propriedades antiinflamatória, antibacteriana, anticâncer e tripanossomicida. O objetivo deste trabalho foi investigar um possível efeito espasmolítico destas naftoquinonas em íleo de cobaia, uma vez que, outras naftoquinonas inibem a atividade contrátil de músculos lisos. O lapachol, alfa e beta-lapachona inibiram as contrações fásicas induzidas tanto por carbacol (CI50 = 1,5 ± 0,2 x 10-4; 7,3 ± 0,9 x 10-5 e 3,2 ± 0,5 x 10-5 M, respectivamente) quanto por histamina (CI50 = 3,6± 0,5; 3,6 ± 0,7 e 3,3 ± 0,6 x 10-5 M, respectivamente). Estes compostos também relaxaram o íleo pré-contraído com KCl (CE50 = 1,2 ± 0,4; 4,3 ± 0,8 e 2,7 ± 0,2 x 10-5 M, respectivamente); carbacol (CE50 = 2,6 ± 0,7; 3,5 ± 0,5 e 2,2 ± 0,7 x 10-5 M, respectivamente) ou histamina (CE50 = 3,0 ± 0,8; 1,1 ± 0,3 e 3,3 ± 0,6 x 10-5 M, respectivamente) de maneira dependente de concentração. Este efeito é provavelmente devido à inibição do influxo de Ca2+ através dos canais de Ca2+ dependentes de voltagem (CaV). Beta-lapachona antagonizou (pD'2 = 5,73 ± 0,12; "slope" = 1,51 ± 0,05) as contrações induzidas por CaCl2 em meio despolarizante nominalmente sem Ca2+. O achado de que a beta-lapachona inibiu as contrações tônicas induzidas por S-(-)-Bay K8644 (CE50 = 1,4 ± 0,1 x 10-5 M) é sugestivo que o CaV envolvido é o do tipo L. Em conclusão lapachol, alfa e beta-lapachona apresentam atividade espasmolítica não seletiva em íleo de cobaia, e beta-lapachona exerce este efeito pelo bloqueio dos canais CaV tipo L.

Lapachol, alpha and beta-lapachone are naphthoquinones extracted from species of Tabebuia that have shown antiinflammatory, antibacterial, anticancer and trypanosomicidal properties. The aim of this work was to investigate the spasmolytic effect of these naphthoquinones on the guinea-pig ileum, since other naphthoquinones are known to depress the contractile activity of smooth muscles. Lapachol, alpha and beta-lapachone inhibited the phasic contractions induced by both carbachol (IC50 = 1.5 ± 0.2 x 10-4; 7.3 ± 0.9 x 10-5 and 3.2 ± 0.5 x 10-5 M, respectively) and histamine (IC50 = 3.6± 0.5; 3.6 ± 0.7 and 3.3 ± 0.6 x 10-5 M, respectively). These compounds also relaxed the ileum pre-contracted with KCl (EC50 = 1.2 ± 0.4; 4.3 ± 0.8 and 2.7 ± 0.2 x 10-5 M, respectively); carbachol (EC50 = 2.6 ± 0.7; 3.5 ± 0.5 and 2.2 ± 0.7 x 10-5 M, respectively) or histamine (EC50 = 3.0 ± 0.8; 1.1 ± 0.3 and 3.3 ± 0.6 x 10-5 M, respectively) in a concentration-dependent manner. This effect is probably due to inhibition of calcium influx through voltage-gated calcium channels (Ca v). Beta-lapachone antagonized (pD'2 = 5.73 ± 0.12; slope = 1.51 ± 0.05) CaCl2-induced contractions in depolarizing medium nominally without Ca2+. The finding that Beta-lapachone inhibited the tonic contractions induced by S-(-)-Bay K8644 (EC50 = 1.4 ± 0.1 x 10-5 M) is suggestive that the L-type CaV is involved. In conclusion, lapachol, alpha and beta-lapachone showed non-selective spasmolytic activity in guinea-pig ileum, and beta-lapachone exerts this effect by to blockade of L-type CaV channels.

Pharmacological characterization of venoms obtained from Mexican toxoglossate gastropods on isolated guinea pig ileum

The protein-containing extracts prepared from the venom ducts of Conus austini, Conus spurius and Polystira albida caused a concentration-dependent inhibition of spontaneous contractions in guinea pig ileum. The most potent extract was obtained from P. albida venom ducts (IC50 equal 0.11 more or less 0.02 microg protein/mL). The three extracts produced a moderate inhibition of contractions elicited by acetylcholine (ACh 1 microM), suggesting the presence of anticholinergic compounds. The contractile response elicited by nicotine (10 microM) was significantly reduced by the extracts prepared from the ducts of C. austini and P. albida, which indicates that the venom produced by these species contains toxins that target neuronal nicotinic receptors. All three extracts significantly inhibited contractions evoked by histamine (0.5 miM), particularly those from C. spurius and P. albida. These findings reveal the presence of antihistaminergic compounds not previously described in any conoidean venom. Finally, we found that only the extract prepared from C. spurius ducts decreased KCl (60 mM)-induced contractions, indicating that the venom of this snail contains compounds that block voltage-dependent Ca2 more or Na more channels.

Efeito de Hypericum caprifoliatum Cham. & Schltdl. (Guttiferae) sobre contrações em íleo isolado de cobaio induzidas por diferentes agonistas / Effect of Hypericum caprifoliatum Cham & Schltdl (Guttiferae) on isolated guinea pig ileum serotonin response

Na última década, o gênero Hypericum ganhou repercussão mundial devido à utilização de Hypericum perforatum para obtenção de medicamentos antidepressivos. Por esta razão, a maioria dos estudos com outras espécies do gênero centra-se nesta atividade. Porém, um dos usos populares de espécies de Hypericum nativas do sul do Brasil é no tratamento de problemas gastrintestinais, inclusive como antiespasmódico. Neste trabalho, foi avaliado o efeito de uma das espécies de Hypericum nativas do Rio Grande do Sul, H. caprifoliatum, sobre as contrações induzidas por agonistas em íleo isolado de cobaio. Foi investigado o efeito de um extrato ciclo-hexano purificado (isento de clorofila e ceras), nas concentrações de 1, 3, 10 e 30 mg/mL, sobre curvas cumulativas de acetilcolina, histamina, potássio e serotonina (10-7 a 10-4 M). Na concentração de 30 mg/mL o extrato inibiu totalmente as contrações induzidas por todos os agonistas. Na concentração de 10 mg/mL, o extrato apresentou efeito antagonista não-competitivo de serotonina, reduzindo a contração máxima induzida por serotonina em cerca de 50 por cento. A resposta contrátil aos outros mediadores não foi alterada. Estes resultados indicam que espécies de Hypericum do sul do Brasil podem ser uma perspectiva interessante na busca de moléculas com atividade sobre a motilidade gastrintestinal.

In the last decade the genus Hypericum has achieved worldwide recognition due to the therapeutic value of H. perforatum as an antidepressant drug. Consequently this activity is the most investigated one. However, species native to Brazil have other folk uses such as for the treatment of digestive disorders, including cramps. In this study we evaluated the effect of a purified cyclohexane extract (chlorophyll and waxes free) (1,3,10 and 30 mg/mL) of H. caprifoliatum, a specie native to South Brazil, on isolated guinea pig ileum contractions induced by different mediators: serotonin, histamine, acetylcholine and potassium chloride (10-7 - 10-4 M). At 30 mg/mL all contractile responses were abolished. At 10 mg/mL only serotonin responses were altered: the extract reduced the maximal effect in 50 percent, which represents a non-competitive antagonism. At 1 and 3 mg/mL the extract was unable to modify all mediators response. These results point to native species of Hypericum as an interesting perspective in searching new molecules active on gastrointestinal motility.

Extratos metanólico e acetato de etila de Solanum megalonyx Sendtn. (Solanaceae) apresentam atividade espasmolítica em íleo isolado de cobaia: um estudo comparativo / Methanol and ethyl acetate extracts from Solanum megalonyx Sendtn. (Solanaceae) present spasmolytic activity in guinea-pig ileum: a comparative study

Solanum megalonyx Sendtn. (Solanaceae) é conhecida popularmente por "jurubeba" no Nordeste do Brasil e se apresenta na forma de arbusto. Várias espécies de Solanum apresentam efeito espasmolítico em órgãos isolados. Assim, objetivou-se investigar e comparar o efeito dos extratos metanólico (SM-MeOH) e acetato de etila (SM-AcOEt), obtidos das partes aéreas de S. megalonyx, em íleo isolado de cobaia. SM-MeOH e SM-AcOEt antagonizaram (n = 5) as contrações fásicas induzidas por 1 mM de acetilcolina (logCI50 = 3,2 ± 0,1 e 1,8 ± 0,6 mg/mL, respectivamente) ou de histamina (logCI50 = 2,8 ± 0,5 e 1,7 ± 0,3 mg/mL, respectivamente). SM-MeOH e SM-AcOEt também relaxaram (n = 5) o íleo pré-contraído por 40 mM de KCl (logCE50 = 1,9 ± 0,09 e 1,9 ± 0,1 mg/mL, respectivamente), por 1 mM de histamina (logCE50 = 1,9 ± 0,07 e 1,7 ± 0,08 mg/mL, respectivamente) ou de acetilcolina (logCE50 = 1,9 ± 0,02 e 1,7 ± 0,09 mg/mL, respectivamente) de maneira dependente de concentração e equipotente. Demonstra-se pela primeira vez que S. megalonyx apresenta efeito espasmolítico não seletivo em íleo isolado de cobaia, sugerindo que os extratos podem estar agindo em um passo comum da via de sinalização dos agentes contráteis testados.

Solanum megalonyx Sendtn. (Solanaceae) is known popularly as "jurubeba" in Northeastern Brazil where it can be found as a shrub. Several species of Solanum present spasmolytic effect in several tissues, thus this study was aimed to investigate and compare the effect of the methanol extract (SMMeOH) and ethyl acetate extract (SM-AcOEt), obtained from aerial parts of Solanum megalonyx Sendtn., in guinea-pig ileum. In this work, both SM-MeOH and SM-AcOEt antagonized the phasic contraction induced by acetylcholine 1 mM (logIC50 = 3.2 ± 0.1 and 1.8 ± 0.6 mg/mL) and histamine 1 mM (logIC50 = 2.8 ± 0.5 and 1.7 ± 0.3 mg/mL, respectively) (n = 5), without statistical differences between these values. In another set of experiments, SM-MeOH and SM-AcOEt also relaxed the isolated guinea-pig ileum pre-contracted by KCl 40 mM (logEC50 = 1.9 ± 0.09 and 1.9 ± 0.1 mg/mL, respectively), histamine 1 mM (logEC50 = 1.9 ± 0.07 mg/mL and 1.7 ± 0.08 mg/mL, respectively) or acetylcholine (logEC50 = 1.9 ± 0.02 mg/mL and 1.7 ± 0.09 mg/mL, respectively) (n = 5) in a concentration-dependent and equipotent manner. This study demonstrates for the first time that aerial parts of S. megalonyx present a non-selective spasmolytic effect in guinea-pig ileum, suggesting that the extracts could be acting in a common step of the pathway signaling that leads to contraction induced by the contractile agents tested.

Experiment study on antiallergic effects of norstemizole / 中国临床药理学与治疗学

AIM: To study the antiallerergic effects of norastemizole. METHODS: Antiallergic effects of norastemizole were study using rats and guinea pigs. RESULTS: Norastemizole(10 -9 -10 -8 mol?L -1 ) caused inhibition effect on the hisitamine induced contraction of isolated guinea pig ileum. Its IC 50 was 4.42 ?10 -9 mol?L -1 Norastemizole at oral doses of 0.1 , 0.5 , 1 mg?kg -1 , norastemizole caused a significant inhibition on the histamin phosphate induced shock in guinea pigs (P

Modulation of outward potassium currents by nitric oxide in longitudinal smooth muscle cells of guinea-pig ileum

To investigate the possible involvement of outward potassium (K+) currents in nitric oxide-induced relaxation in intestinal smooth muscle, we used whole-cell patch clamp technique in freshly dispersed guinea-pig ileum longitudinal smooth muscle cells. When cells were held at -60 mV and depolarized from - 40 mV to + 50 mV in 10 mV increments, sustained outward K+ currents were evoked. The outward K+ currents were markedly increased by the addition of 10 muM sodium nitroprusside (SNP). 10 muM S-nitroso-N-acetylpenicillamine (SNAP) and 1 mM 8-Bromo-cyclic GMP (8-Br-cGMP) also showed a similar effect to that of SNP. 1 mM tetraethylammonium (TEA) significantly reduced depolarization-activated outward K+ currents. SNP-enhanced outward K+ currents were blocked by the application of TEA. High EGTA containing pipette solution (10 mM) reduced the control currents and also inhibited the SNP-enhanced outward K+ currents. 5 mM 4-aminopyridine (4-AP) significantly reduced the control currents but showed no effect on SNP-enhanced outward K+ currents. 0.3 muM apamin and 10 muM glibenclamide showed no effect on SNP-enhanced outward K+ currents. 1 muM 1H-(1,2,4)oxadiazolo (4,3-a)quinoxaline-1-one (ODQ), a specific inhibitor of soluble guanylate cyclase, significantly blocked SNP-enhanced K+ currents. We conclude that NO donors activate the Ca2+-activated K+ channels in guinea-pig ileal smooth muscle via activation of guanylate cyclase.

Action mechanisms of NANC neurotransmitters in smooth muscle of guinea pig ileum

The relaxation induced by stimulation of the inhibitory non-adrenergic, non-cholinergic (iNANC) nerve is mediated by the release of iNANC neurotransmitters such as nitric oxide (NO), vasoactive intestinal peptide (VIP) and adenosine triphosphate (ATP). The mechanisms of NO, VIP or ATP-induced relaxation have been partly determined in previous studies, but the detailed mechanism remains unknown. We tried to identify the nature of iNANC neurotransmitters in the smooth muscle of guinea pig ileum and to determine the mechanism of the inhibitory effect of nitric oxide. We measured the effect of NO-donors, VIP and ATP on the intracellular Ca2+ concentration((Ca2+)i), by means of a fluorescence dye (fura 2) and tension simultaneously in the isolated guinea pig ileal smooth muscle. Following are the results obtained. 1. Sodium nitropnisside (SNP: 10(-5) M) or S-nitro-N-acetyl-penicillamine (SNAP: 10(-5) M) decreased resting (Ca2+)i and tension of muscle. SNP or SNAP also inhibited rhythmic oscillation of (Ca2+)i and tension. In 40mM K+ solution or carbachol (CCh:10(-6) M)-induced precontracted muscle, SNP decreased muscle tension. VIP did not change (Ca2+)i and tension in the resting or precontracted muscle, but ATP increased resting (Ca2+)i and tension in the resting muscle. 2. 1H-(1,2,4)oxadiazol(4,3-a)quinoxalin-1-one (ODQ:1 muM), a specific inhibitor of soluble guanylate cyclase, limited the inhibitory effect of SNP. 3. Glibenclamide (10 muM), a blocker of KATP channel, and 4-aminopyridine (4-AP:5 mM), a blocker of delayed rectifier K channel, apamin (0.1 muM), a blocker of small conductance KCa. channel had no effect on the inhibitory effect of SNP. Iberiotoxin (0.1 muM), a blocker of large conductance KCa channel, significantly increased the resting (Ca2+)i, and tension, and limited the inhibitory effect of SNP. 4. Nifedipine (1 muM) or elimination of external Ca2+ decreased not only resting (Ca2+)i and tension but also oscillation of (Ca2+)i and tension. Ryanodine (5 muM) and cyclopiazonic acid (10 muM) decreased oscillation of (Ca2+)i and tension. 5. SNP decreased Ca2+ sensitivity of contractile protein. In conclusion, these results suggest that 1) NO is an inhibitory neurotransmitter in the guinea pig ileum, 2) the inhibitory effect of SNP on the (Ca2+)i and tension of the muscle is due to a decrease in (Ca2+)i by activation of the large conductance KCa channel and a decrease in the sensitivity of contractile elements to Ca2+ through activation of G-kinase.

The effect of sodium nitroprusside on muscle tension in guinea-pig ileum

Nitric oxide (NO) has been known as a mediator of nonadrenergic, noncholinergic inhibitory neurotransmitter in intestinal smooth muscles. It has been suggested that NO donor such as sodium nitroprusside (SNP) produces relaxation of smooth muscle via activation of guanylate cyclase and elevation of cGMP levels. We have therefore investigated the effects of NO, using SNP, on muscle tension in the longitudinal smooth muscle of guinea-pig ileum. The possible role of cGMP was also investigated as well as the involvement of K+ channel on SNP-induced inhibitory effect. The results are summarized as follows; high KCl-or CCh-activated contractions were inhibited by SNP in a concentration-dependent manner. 8-Br-cGMP also showed a similar effect in that of SNP. TEA (1 mM) significantly reduced the SNP-induced inhibitory effect. SNP-induced effect was further reduced by the presence of 10 mM TEA. On the other hand, 4-AP (0.1 mM), glibenclamide (10 muM) and apamin (0.1 muM) showed little effects on SNP-induced relaxation. Zaprinast significantly potentiated the SNP-induced inhibitory effect in all ranges. ODQ also significantly decreased the SNP-induced inhibitory effect. Pretreatment with CPA (10 muM) slightly reduced the SNP-induced inhibitory effect. From the above results, both effect mediated by NO and cGMP might be responsible for the activation of Ca2+/-activated K+ channel by SNP in guinea-pig ileum. And this K+ channel activation by SNP also contributes to the SNP-induced membrane hyperpolarization and relaxation.

Effects of ondansetron on morphine physical dependence in mice / 中国药理学通报

AIM To investigate the effects of ondansetron, a selective 5 Hydroxytryptamine3 (5 HT 3) receptor antagonist, on morphine physical dependence. METHODS The morphine dependent models in mice and in isolated Guinea pig ileum were used. RESULTS Pretreatment of ondansetron for 12 days significantly reduced morphine withdrawal symptoms in mice ,such as body weight loss(Groups 2～100 ?g?kg -1 ?d -1 ) or reduced both body weight loss and jumping times (Group 100 ?g?kg -1 ?d -1 ). In addition, concomitant treatment with ondansetron(1～20 ?mol?L -1 ) dose dependently suppressed the contraction induced by naloxone in Guinea pig ileum. CONCLUSION The chronic pretreatment of ondansetron can prevent morphine physical dependence to some extent.

Effect of nitric oxide (NO) on dependence and tolerance of normorphine (NM) in the guinea-pig ileum in vitro / 中国药理学通报

0.05. The ratio of NM (200 nmol? L-l) plus NOArg (0.1 mmol. L-1 ) is 0.47 ? 0.05, compared with control (n = 6, P 0 .05). CONCLUSIONS These results suggest that NO has important role on dependence and tolerance of NM in the guinea Pig ileum.