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Effect of L-NMMA on proximal stomach function in healthy volunteers
April 2000
AGAA387
2035
2037
EFFECT OF K+ AND cr CHANNEL BLOCKERS ON MECHANICAL AND ELECTRICAL PROPERTIES OF OPOSSUM LOWER ESOPHAGEAL SPHINCTER (LES). Yong Zhang, David V. Miller, William G. Paterson, Acad Univ, Kingston, ON, Canada.
EFFECT OF L-NMMA ON PROXIMAL STOMACH FUNCTION IN HEALTHY VOLUNTEERS. Guy E. Boeckxstaens, Sjoerd Kuiken, Siem H. Heisterkarnp, Guido N. Tytgat, Acad Med Ctr, Amsterdam, Netherlands. Impaired fundic accommodation to a meal is increasingly recognised as an important pathophysiological mechanism of functional dyspepsia. Animal studies have suggested that meal-induced fundic relaxation is mediated by nitric oxide (NO). The role of NO in meal-induced fundic relaxation and postprandial symptoms in man however is unknown. Therefore, we investigated the effect of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA) on proximal stomach function in healthy volunteers (HV). Methods: 10 male HV (19-29 yr) were enrolled in a double blind placebocontrolled study evaluating the effect of L-NMMA (6.7 mg/kg/h i.v.) on basal and postprandial (during 1 h, following 200 ml of Nutridrink) fundic volume measured at MDP + 2 mmHg. Symptoms (nausea, fullness, satiety, pain) were scored every 5 min during the postprandial period using a VAS scale. One hour after the meal, a stepwise distension protocol (2 mmHg steps during 2 min) was performed. Results: Diastolic blood pressure was increased by L-NMMA from 73 ± 7 mmHg to 83 ± 9 mmHg (p
The resting membrane potential (RMP) of LES circular smooth muscle (--40 mv) is relatively depolarized compared to adjacent esophageal body muscle (--50mv), and this depolarized state is believed to contribute to myogenic LES tone. To explore the ionic mechanisms underlying LES RMP, and hence the generation of myogenic tone, the effect of various K+ and Clchannel blockers on resting tone and membrane potential of LES circular smooth muscle was determined using conventional isometric tension and intracellular electrical recordings, respectively. Strips were stretched to optimal length (140%) and equilibrated in 10 ml organ baths containing oxygenated Kreb's solution. TEA (2 mM) a large conductance Ca++-activated K+ channel blocker, and 4-AP (2 mM), a transient K+ channel blocker, enhanced resting tone to 32.9± 15.5%(mean±se) and 83.1 ±6.5%, respectively, over basal values (n=4,p<0.05). However, apamin (300 nM), a small conductance Ca++-activated K+ channel blocker, and glibenclamide, an ATP-sensitive K+ channel blocker, were without effect. DIDS (500 /LM) and NPPB (500 /LM), nonspecific CI' channel blockers, decreased basal tone to 75.0± 7.4% and 67.3± 10.8%, respectively, of control (n=4). Niflumic acid (NA), a putative selective Ca++-activated Clchannel blocker, relaxed LES muscle strips in a dosedependent manner, while tamoxifen (lO/LM), a volume-activated Clchannel blocker, had no affect. Basal LES tone was decreased to 64.6±2.9% of control values by the ICso dose of NA (1.2±0.3/LM, n=9). The application of KCl (60 mM) produced contraction in the presence of 3/LM NA, that was no different than control KCl-induced contractions, indicating that NA was not non-specifically inhibiting contraction via an effect on contractile proteins or voltage-sensitive Ca++ channels. Intracellular recordings revealed a RMP of -41.3±0.9 mV (n=24) with superimposed spontaneous spike-like action potentials (frequency; 43.0±2.7/min: amplitude, 19.9±1.0 mY). NA dose-dependently hyperpolarized RMP and decreased the amplitude of, and then abolished the spontaneous action potentials. On the other hand, both TEA and 4-AP significantly depolarized RMP and increased the frequency and amplitude of spontaneous action potentials. These data suggest that K+ and and Cl (Ca++ -activated) channels have important opposing roles in the genesis of LES tone. Supported by MRC Canada.
2036 MECHANISM OF ESOPHAGEAL LONGITUDINAL SMOOTH MUSCLE CONTRACTION INDUCED BY SODIUM NITROPRUSSIDE (SNP) AND SUBSTANCE P (SP). Yong Zhang, David V. Miller, Tavis Basford, William G. Paterson, Queens Univ, Kingston, ON, Canada; Acad Univ, Kingston, ON, Canada. Previous investigations have suggested that Nitric Oxide (NO) and SP can induce sustained contraction of esophageal longitudinal smooth muscle, and therefore may be involved in esophageal shortening that occurs in association with acid-induced esophagitis. To determine the ionic mechanisms underlying the NO- and SP-induced contraction, conventional isometric tension recordings were performed on longitudinal smooth muscle strips from the opossum esophagus. Strips were stretched to optimal length (140%) and equilibrated in 10 ml organ baths containing oxygenated Kreb's solution. Data were standardized to the maximal contraction induced by bath application of 60 mM KCL Both SNP and SP evoked sustained contractions in a dose-dependent manner, with an EC so of 239.7 ± 78.2 /LM (n= 10) (mean ± se) and 48.9 ± 15.6 nM (n=9), respectively. MEN 10,376, a specific NK-2 receptor antagonist, but not CP-99,994-01, a specific NK-I receptor antagonist, dose-dependently antagonized contractions produced by SP, with an ICso of 3.4 ± 2.3 nM (n=4). Contraction produced by SNP (300 liM) was reduced from 57.9 ± 6.7 % (of maximal KCL-induced contraction) to 3.6 ± 1.2 % (n=7) by niflumic acid (300 /LM), a specific Ca2+ -activated Cl channel blocker. Similarly, the contraction induced by SP (100 nM) was decreased from 57.5 ± 5.2% to 27.7 ± 2.3 % and 4.4 ± 0.9 % (n=5) in the presence of niflumic acid (100, 300 /LM, respectively). Nifedipine (I/LM) inhibited the contractions induced by SNP (300 /LM) to 1.70 ± 0.96 % (n=7) and by SP (100 nM) to 5.05 ± 1.28 % (n=5). The application of 60 mM KCI in the presence of niflumic acid (300 /LM) produced 82.6± 8.7 % (n=3) of control KCI-induced contractions, suggesting that niflumic acid was not nonspecifically inhibiting contraction via an effect on L-type Ca2+ channels or contractile proteins. The SNP-induced contraction was mimicked by the cGMP analogue 8-Bromo-cGMP (I mM), and was reduced to 4.9±4.3% (n=5) of control by ODQ (lO/LM), an inhibitor of soluble guanylate cyclase. This data suggests that NO and SP may induce contraction of opossum esophageal longitudinal smooth muscle by opening Ca2+ -activated CI' channels, which in turn leads to extra-cellular Ca2+ entry through L-type Cai channels. The SNP-induced contraction also involves a cGMP-dependent pathway.
2038 ROLE OF NITRIC OXIDE IN GASTRIC RELAXATION INDUCED BY INTRAVENOUS ADMINISTRATION OF LIPID IN CONSCIOUS DOGS. Hajime Haneji, Masazumi Okajima, Masahumi Kikkawa, Yasutomo Ojirna, Shintaro Nakashima, Satoshi Ikeda, Shinya Kodama, Yosuke Shimizu, Masahiro Nakahara, Katsuhumi Kawahori, Toshimasa Asahara, Hiroshima Univ Faculty of Medicine, Hiroshima, Japan. Background We have previously demonstrated that cholinergic nervous system, not nitric oxide (NO), mainly mediates a gastric relaxation induced by oral intake of lipid in the conscious dogs . Recent studies showed that gastric relaxation is mediated by NO via NANC inhibitory nerves in vitro. The aim of this study was to determine the effects of intravenous administration of lipid on canine proximal gastric tone and to investigate the role of NO in its effects. Materials and methods Experiments were performed on conscious dogs with chronic gastric fistulae (N = 6). A flaccid polyethylene bag introduced into the proximal stomach via the gastric fistulae was connected to a barostat with a double lumen tube. I) Maintained at 4 mmHg intragastric pressure, Intralipos (20% refined soybean oil) was administrated intravenously (0.1, 0.25, 0.5, 1.0 ml/kg). The volume change of the proximal stomach was measured by means of the barostat. 2) 10 minutes after intravenous administration of W-nitro-L-arginine (5mg/kg, L-NNA), lipid was given intravenously (1.0 ml/kg). Then the same procedure was performed. Results I) Lipid administration induced proximal gastric relaxation dose-dependently (0.1 ml/kg; 25.4± 1.4ml, 0.25 ml/kg; 60.8±37.2ml, 0.5 ml/kg; 104.9±74.0ml, 1.0 ml/kg; 161.0±60.1ml). 2) L-NNA reduced proximal gastric relaxation (55.0±31.9ml; P<0.OO5). Conclusion Our results indicate that intravenous administration of lipid increased the relaxatory capacity of the proximal stomach via a NOdependent mechanism in the conscious dogs.
AGAA387
2035
2037
EFFECT OF K+ AND cr CHANNEL BLOCKERS ON MECHANICAL AND ELECTRICAL PROPERTIES OF OPOSSUM LOWER ESOPHAGEAL SPHINCTER (LES). Yong Zhang, David V. Miller, William G. Paterson, Acad Univ, Kingston, ON, Canada.
EFFECT OF L-NMMA ON PROXIMAL STOMACH FUNCTION IN HEALTHY VOLUNTEERS. Guy E. Boeckxstaens, Sjoerd Kuiken, Siem H. Heisterkarnp, Guido N. Tytgat, Acad Med Ctr, Amsterdam, Netherlands. Impaired fundic accommodation to a meal is increasingly recognised as an important pathophysiological mechanism of functional dyspepsia. Animal studies have suggested that meal-induced fundic relaxation is mediated by nitric oxide (NO). The role of NO in meal-induced fundic relaxation and postprandial symptoms in man however is unknown. Therefore, we investigated the effect of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA) on proximal stomach function in healthy volunteers (HV). Methods: 10 male HV (19-29 yr) were enrolled in a double blind placebocontrolled study evaluating the effect of L-NMMA (6.7 mg/kg/h i.v.) on basal and postprandial (during 1 h, following 200 ml of Nutridrink) fundic volume measured at MDP + 2 mmHg. Symptoms (nausea, fullness, satiety, pain) were scored every 5 min during the postprandial period using a VAS scale. One hour after the meal, a stepwise distension protocol (2 mmHg steps during 2 min) was performed. Results: Diastolic blood pressure was increased by L-NMMA from 73 ± 7 mmHg to 83 ± 9 mmHg (p
The resting membrane potential (RMP) of LES circular smooth muscle (--40 mv) is relatively depolarized compared to adjacent esophageal body muscle (--50mv), and this depolarized state is believed to contribute to myogenic LES tone. To explore the ionic mechanisms underlying LES RMP, and hence the generation of myogenic tone, the effect of various K+ and Clchannel blockers on resting tone and membrane potential of LES circular smooth muscle was determined using conventional isometric tension and intracellular electrical recordings, respectively. Strips were stretched to optimal length (140%) and equilibrated in 10 ml organ baths containing oxygenated Kreb's solution. TEA (2 mM) a large conductance Ca++-activated K+ channel blocker, and 4-AP (2 mM), a transient K+ channel blocker, enhanced resting tone to 32.9± 15.5%(mean±se) and 83.1 ±6.5%, respectively, over basal values (n=4,p<0.05). However, apamin (300 nM), a small conductance Ca++-activated K+ channel blocker, and glibenclamide, an ATP-sensitive K+ channel blocker, were without effect. DIDS (500 /LM) and NPPB (500 /LM), nonspecific CI' channel blockers, decreased basal tone to 75.0± 7.4% and 67.3± 10.8%, respectively, of control (n=4). Niflumic acid (NA), a putative selective Ca++-activated Clchannel blocker, relaxed LES muscle strips in a dosedependent manner, while tamoxifen (lO/LM), a volume-activated Clchannel blocker, had no affect. Basal LES tone was decreased to 64.6±2.9% of control values by the ICso dose of NA (1.2±0.3/LM, n=9). The application of KCl (60 mM) produced contraction in the presence of 3/LM NA, that was no different than control KCl-induced contractions, indicating that NA was not non-specifically inhibiting contraction via an effect on contractile proteins or voltage-sensitive Ca++ channels. Intracellular recordings revealed a RMP of -41.3±0.9 mV (n=24) with superimposed spontaneous spike-like action potentials (frequency; 43.0±2.7/min: amplitude, 19.9±1.0 mY). NA dose-dependently hyperpolarized RMP and decreased the amplitude of, and then abolished the spontaneous action potentials. On the other hand, both TEA and 4-AP significantly depolarized RMP and increased the frequency and amplitude of spontaneous action potentials. These data suggest that K+ and and Cl (Ca++ -activated) channels have important opposing roles in the genesis of LES tone. Supported by MRC Canada.
2036 MECHANISM OF ESOPHAGEAL LONGITUDINAL SMOOTH MUSCLE CONTRACTION INDUCED BY SODIUM NITROPRUSSIDE (SNP) AND SUBSTANCE P (SP). Yong Zhang, David V. Miller, Tavis Basford, William G. Paterson, Queens Univ, Kingston, ON, Canada; Acad Univ, Kingston, ON, Canada. Previous investigations have suggested that Nitric Oxide (NO) and SP can induce sustained contraction of esophageal longitudinal smooth muscle, and therefore may be involved in esophageal shortening that occurs in association with acid-induced esophagitis. To determine the ionic mechanisms underlying the NO- and SP-induced contraction, conventional isometric tension recordings were performed on longitudinal smooth muscle strips from the opossum esophagus. Strips were stretched to optimal length (140%) and equilibrated in 10 ml organ baths containing oxygenated Kreb's solution. Data were standardized to the maximal contraction induced by bath application of 60 mM KCL Both SNP and SP evoked sustained contractions in a dose-dependent manner, with an EC so of 239.7 ± 78.2 /LM (n= 10) (mean ± se) and 48.9 ± 15.6 nM (n=9), respectively. MEN 10,376, a specific NK-2 receptor antagonist, but not CP-99,994-01, a specific NK-I receptor antagonist, dose-dependently antagonized contractions produced by SP, with an ICso of 3.4 ± 2.3 nM (n=4). Contraction produced by SNP (300 liM) was reduced from 57.9 ± 6.7 % (of maximal KCL-induced contraction) to 3.6 ± 1.2 % (n=7) by niflumic acid (300 /LM), a specific Ca2+ -activated Cl channel blocker. Similarly, the contraction induced by SP (100 nM) was decreased from 57.5 ± 5.2% to 27.7 ± 2.3 % and 4.4 ± 0.9 % (n=5) in the presence of niflumic acid (100, 300 /LM, respectively). Nifedipine (I/LM) inhibited the contractions induced by SNP (300 /LM) to 1.70 ± 0.96 % (n=7) and by SP (100 nM) to 5.05 ± 1.28 % (n=5). The application of 60 mM KCI in the presence of niflumic acid (300 /LM) produced 82.6± 8.7 % (n=3) of control KCI-induced contractions, suggesting that niflumic acid was not nonspecifically inhibiting contraction via an effect on L-type Ca2+ channels or contractile proteins. The SNP-induced contraction was mimicked by the cGMP analogue 8-Bromo-cGMP (I mM), and was reduced to 4.9±4.3% (n=5) of control by ODQ (lO/LM), an inhibitor of soluble guanylate cyclase. This data suggests that NO and SP may induce contraction of opossum esophageal longitudinal smooth muscle by opening Ca2+ -activated CI' channels, which in turn leads to extra-cellular Ca2+ entry through L-type Cai channels. The SNP-induced contraction also involves a cGMP-dependent pathway.
2038 ROLE OF NITRIC OXIDE IN GASTRIC RELAXATION INDUCED BY INTRAVENOUS ADMINISTRATION OF LIPID IN CONSCIOUS DOGS. Hajime Haneji, Masazumi Okajima, Masahumi Kikkawa, Yasutomo Ojirna, Shintaro Nakashima, Satoshi Ikeda, Shinya Kodama, Yosuke Shimizu, Masahiro Nakahara, Katsuhumi Kawahori, Toshimasa Asahara, Hiroshima Univ Faculty of Medicine, Hiroshima, Japan. Background We have previously demonstrated that cholinergic nervous system, not nitric oxide (NO), mainly mediates a gastric relaxation induced by oral intake of lipid in the conscious dogs . Recent studies showed that gastric relaxation is mediated by NO via NANC inhibitory nerves in vitro. The aim of this study was to determine the effects of intravenous administration of lipid on canine proximal gastric tone and to investigate the role of NO in its effects. Materials and methods Experiments were performed on conscious dogs with chronic gastric fistulae (N = 6). A flaccid polyethylene bag introduced into the proximal stomach via the gastric fistulae was connected to a barostat with a double lumen tube. I) Maintained at 4 mmHg intragastric pressure, Intralipos (20% refined soybean oil) was administrated intravenously (0.1, 0.25, 0.5, 1.0 ml/kg). The volume change of the proximal stomach was measured by means of the barostat. 2) 10 minutes after intravenous administration of W-nitro-L-arginine (5mg/kg, L-NNA), lipid was given intravenously (1.0 ml/kg). Then the same procedure was performed. Results I) Lipid administration induced proximal gastric relaxation dose-dependently (0.1 ml/kg; 25.4± 1.4ml, 0.25 ml/kg; 60.8±37.2ml, 0.5 ml/kg; 104.9±74.0ml, 1.0 ml/kg; 161.0±60.1ml). 2) L-NNA reduced proximal gastric relaxation (55.0±31.9ml; P<0.OO5). Conclusion Our results indicate that intravenous administration of lipid increased the relaxatory capacity of the proximal stomach via a NOdependent mechanism in the conscious dogs.