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The impact of extracellular and intracellular Ca2+ on ethanol-induced smooth muscle contraction.

Döndaş NY, Kaplan M, Kaya D, Singirik E - Acta Pharmacol. Sin. (2009)

Bottom Line: Although lidocaine (50 and 100 micromol/L), a local anesthetic agent, and hexamethonium (100 and 500 micromol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 micromol/L) and nifedipine (1-50 micromol/L), selective blockers of L-type Ca2+ channels, significantly inhibited the contractile responses of ethanol.Ryanodine (1-50 micromol/L) and ruthenium red (10-100 micromol/L), selective blockers of intracellular Ca2+ channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 mumol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca2+ stores, significantly inhibited the contractile responses induced by ethanol.This inhibition was significantly different from those associated with caffeine, ryanodine or CPA.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Faculty of Medicine, Cukurova University, 01330 Adana, Turkey. yakdas25@cu.edu.tr

ABSTRACT

Aim: To evaluate the impact of extracellular and intracellular Ca2+ on contractions induced by ethanol in smooth muscle.

Methods: Longitudinal smooth muscle strips were prepared from the gastric fundi of mice. The contractions of smooth muscle strips were recorded with an isometric force displacement transducer.

Results: Ethanol (164 mmol/L) produced reproducible contractions in isolated gastric fundal strips of mice. Although lidocaine (50 and 100 micromol/L), a local anesthetic agent, and hexamethonium (100 and 500 micromol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 micromol/L) and nifedipine (1-50 micromol/L), selective blockers of L-type Ca2+ channels, significantly inhibited the contractile responses of ethanol. Using a Ca(2+)-free medium nearly eliminated these contractions in the same tissue. Ryanodine (1-50 micromol/L) and ruthenium red (10-100 micromol/L), selective blockers of intracellular Ca2+ channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 mumol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca2+ stores, significantly inhibited the contractile responses induced by ethanol. In addition, the combination of caffeine (5 mmol/L) plus CPA (10 micromol/L), and ryanodine (10 micromol/L) plus CPA (10 micromol/L), caused further inhibition of contractions in response to ethanol. This inhibition was significantly different from those associated with caffeine, ryanodine or CPA. Furthermore the combination of caffeine (5 mmol/L), ryanodine (10 micromol/L) and CPA(10 micromol/L) eliminated the contractions induced by ethanol in isolated gastric fundal strips of mice.

Conclusion: Both extracellular and intracellular Ca2+ may have important roles in regulating contractions induced by ethanol in the mouse gastric fundus.

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(A) A representative trace showing the effect of cyclopiazonic acid (CPA; 10 μmol/L) on contractile responses to ethanol (164 mmol/L). EtOH: Ethanol, W: Wash, mN: milli Newton. (B) Effect of CPA (1, 5, 10 μmol/L) on contractions induced by ethanol (164 mmol/L) in isolated gastric fundal strips of mice. Control: 164 mmol/L ethanol includes DMSO (1:20000 dilution). For each concentration of the drug, the points are means±SEM of gastric fundal strips from 5 animals. Significant differences were evaluated by one-way analysis of variance (ANOVA). Post hoc: Bonferroni. bP<0.05 vs control.
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fig7: (A) A representative trace showing the effect of cyclopiazonic acid (CPA; 10 μmol/L) on contractile responses to ethanol (164 mmol/L). EtOH: Ethanol, W: Wash, mN: milli Newton. (B) Effect of CPA (1, 5, 10 μmol/L) on contractions induced by ethanol (164 mmol/L) in isolated gastric fundal strips of mice. Control: 164 mmol/L ethanol includes DMSO (1:20000 dilution). For each concentration of the drug, the points are means±SEM of gastric fundal strips from 5 animals. Significant differences were evaluated by one-way analysis of variance (ANOVA). Post hoc: Bonferroni. bP<0.05 vs control.

Mentions: The highest concentration (50 μmol/L) of ryanodine, a selective blocker of intracellular Ca2+ channels/ryanodine receptors, significantly inhibited (P<0.05) the contractile responses to ethanol (Figure 5A, 5B). The lower concentrations (1, 10 μmol/L) of this drug decreased the effect of ethanol (Figure 5B). This agent did not cause any significant alterations in the basal tones of the preparations (Figure 5A; wash-out response of this group: 25±3.1). The effect of this agent was irreversible. The other selective blocker of ryanodine receptors, ruthenium red (10, 50, 100 μmol/L), significantly inhibited (P<0.05) the contractile responses to ethanol at all concentrations used (Figure 6A, 6B). Like ryanodine, it did not cause any significant alterations in the basal tones of the preparations (Figure 6A; wash-out response of this group: 118.4±9.8). Cyclopiazonic acid (CPA; 1, 5, 10 μmol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca2+-ATPase, significantly inhibited (P<0.05) the ethanol-induced contractions at all concentrations used in this study (Figure 7A, 7B). At the beginning of the experiments, CPA only slightly and transiently increased the basal tone of the preparation (Figure 7A). This increment lasted about 5 min, after which the effects decreased. Between 40 and 50 min of the incubation time, which corresponds to the time of the second ethanol application in the normal experimental protocol, CPA slightly increased the basal tone (wash-out response of this group: 110.5±13.4). Similar to ryanodine, caffeine, a depleting agent of intracellular Ca2+ stores, decreased the effect of ethanol in all concentrations used in this study (0.5, 1, 5 mmol/L). Only the highest concentration of caffeine caused a significant inhibition of contractile responses to ethanol (Figure 8B). This agent did not cause any significant alteration in the basal tones of the preparations (Figure 8A; wash-out response of this group: 104.1±9.7). The combination of caffeine (5 mmol/L)+CPA (10 μmol/L) potentiated the inhibitory action of caffeine (5 mmol/L) and CPA (10 μmol/L) on ethanol-induced contractions. This inhibition was significantly different (P<0.05) from both caffeine (5 mmol/L) and CPA (10 μmol/L) results (Figure 9). In addition, the combination of ryanodine (10 μmol/L)+CPA (10 μmol/L) also potentiated the inhibitory action of ryanodine (10 μmol/L) and CPA (10 μmol/L) on ethanol-induced contractions. This inhibition was significantly different (P<0.05) from those of both ryanodine (10 μmol/L) and CPA (10 μmol/L) (Figure 9). Furthermore, the combination of caffeine (5 mmol/L)+ryanodine (10 μmol/L)+CPA (10 μmol/L) completely eliminated the contractile responses to ethanol in the gastric fundal strips of mice (Figure 9).


The impact of extracellular and intracellular Ca2+ on ethanol-induced smooth muscle contraction.

Döndaş NY, Kaplan M, Kaya D, Singirik E - Acta Pharmacol. Sin. (2009)

(A) A representative trace showing the effect of cyclopiazonic acid (CPA; 10 μmol/L) on contractile responses to ethanol (164 mmol/L). EtOH: Ethanol, W: Wash, mN: milli Newton. (B) Effect of CPA (1, 5, 10 μmol/L) on contractions induced by ethanol (164 mmol/L) in isolated gastric fundal strips of mice. Control: 164 mmol/L ethanol includes DMSO (1:20000 dilution). For each concentration of the drug, the points are means±SEM of gastric fundal strips from 5 animals. Significant differences were evaluated by one-way analysis of variance (ANOVA). Post hoc: Bonferroni. bP<0.05 vs control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4007325&req=5

fig7: (A) A representative trace showing the effect of cyclopiazonic acid (CPA; 10 μmol/L) on contractile responses to ethanol (164 mmol/L). EtOH: Ethanol, W: Wash, mN: milli Newton. (B) Effect of CPA (1, 5, 10 μmol/L) on contractions induced by ethanol (164 mmol/L) in isolated gastric fundal strips of mice. Control: 164 mmol/L ethanol includes DMSO (1:20000 dilution). For each concentration of the drug, the points are means±SEM of gastric fundal strips from 5 animals. Significant differences were evaluated by one-way analysis of variance (ANOVA). Post hoc: Bonferroni. bP<0.05 vs control.
Mentions: The highest concentration (50 μmol/L) of ryanodine, a selective blocker of intracellular Ca2+ channels/ryanodine receptors, significantly inhibited (P<0.05) the contractile responses to ethanol (Figure 5A, 5B). The lower concentrations (1, 10 μmol/L) of this drug decreased the effect of ethanol (Figure 5B). This agent did not cause any significant alterations in the basal tones of the preparations (Figure 5A; wash-out response of this group: 25±3.1). The effect of this agent was irreversible. The other selective blocker of ryanodine receptors, ruthenium red (10, 50, 100 μmol/L), significantly inhibited (P<0.05) the contractile responses to ethanol at all concentrations used (Figure 6A, 6B). Like ryanodine, it did not cause any significant alterations in the basal tones of the preparations (Figure 6A; wash-out response of this group: 118.4±9.8). Cyclopiazonic acid (CPA; 1, 5, 10 μmol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca2+-ATPase, significantly inhibited (P<0.05) the ethanol-induced contractions at all concentrations used in this study (Figure 7A, 7B). At the beginning of the experiments, CPA only slightly and transiently increased the basal tone of the preparation (Figure 7A). This increment lasted about 5 min, after which the effects decreased. Between 40 and 50 min of the incubation time, which corresponds to the time of the second ethanol application in the normal experimental protocol, CPA slightly increased the basal tone (wash-out response of this group: 110.5±13.4). Similar to ryanodine, caffeine, a depleting agent of intracellular Ca2+ stores, decreased the effect of ethanol in all concentrations used in this study (0.5, 1, 5 mmol/L). Only the highest concentration of caffeine caused a significant inhibition of contractile responses to ethanol (Figure 8B). This agent did not cause any significant alteration in the basal tones of the preparations (Figure 8A; wash-out response of this group: 104.1±9.7). The combination of caffeine (5 mmol/L)+CPA (10 μmol/L) potentiated the inhibitory action of caffeine (5 mmol/L) and CPA (10 μmol/L) on ethanol-induced contractions. This inhibition was significantly different (P<0.05) from both caffeine (5 mmol/L) and CPA (10 μmol/L) results (Figure 9). In addition, the combination of ryanodine (10 μmol/L)+CPA (10 μmol/L) also potentiated the inhibitory action of ryanodine (10 μmol/L) and CPA (10 μmol/L) on ethanol-induced contractions. This inhibition was significantly different (P<0.05) from those of both ryanodine (10 μmol/L) and CPA (10 μmol/L) (Figure 9). Furthermore, the combination of caffeine (5 mmol/L)+ryanodine (10 μmol/L)+CPA (10 μmol/L) completely eliminated the contractile responses to ethanol in the gastric fundal strips of mice (Figure 9).

Bottom Line: Although lidocaine (50 and 100 micromol/L), a local anesthetic agent, and hexamethonium (100 and 500 micromol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 micromol/L) and nifedipine (1-50 micromol/L), selective blockers of L-type Ca2+ channels, significantly inhibited the contractile responses of ethanol.Ryanodine (1-50 micromol/L) and ruthenium red (10-100 micromol/L), selective blockers of intracellular Ca2+ channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 mumol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca2+ stores, significantly inhibited the contractile responses induced by ethanol.This inhibition was significantly different from those associated with caffeine, ryanodine or CPA.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Faculty of Medicine, Cukurova University, 01330 Adana, Turkey. yakdas25@cu.edu.tr

ABSTRACT

Aim: To evaluate the impact of extracellular and intracellular Ca2+ on contractions induced by ethanol in smooth muscle.

Methods: Longitudinal smooth muscle strips were prepared from the gastric fundi of mice. The contractions of smooth muscle strips were recorded with an isometric force displacement transducer.

Results: Ethanol (164 mmol/L) produced reproducible contractions in isolated gastric fundal strips of mice. Although lidocaine (50 and 100 micromol/L), a local anesthetic agent, and hexamethonium (100 and 500 micromol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 micromol/L) and nifedipine (1-50 micromol/L), selective blockers of L-type Ca2+ channels, significantly inhibited the contractile responses of ethanol. Using a Ca(2+)-free medium nearly eliminated these contractions in the same tissue. Ryanodine (1-50 micromol/L) and ruthenium red (10-100 micromol/L), selective blockers of intracellular Ca2+ channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 mumol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca2+ stores, significantly inhibited the contractile responses induced by ethanol. In addition, the combination of caffeine (5 mmol/L) plus CPA (10 micromol/L), and ryanodine (10 micromol/L) plus CPA (10 micromol/L), caused further inhibition of contractions in response to ethanol. This inhibition was significantly different from those associated with caffeine, ryanodine or CPA. Furthermore the combination of caffeine (5 mmol/L), ryanodine (10 micromol/L) and CPA(10 micromol/L) eliminated the contractions induced by ethanol in isolated gastric fundal strips of mice.

Conclusion: Both extracellular and intracellular Ca2+ may have important roles in regulating contractions induced by ethanol in the mouse gastric fundus.

Show MeSH
Related in: MedlinePlus