| The role of nitric oxide in lower esophageal sphincterof achalasia model in dogAbstractAim: To develop an achalasia model in dog and to investigate the role of nitric oxide in lower esophageal sphincter and pathogenesis of achalasia.Methods: Benzyl-dimethyltetradecylammonium chloride ( BAG ) was endoscopically injected circumferentially into the lower esophageal sphincter(LES) in 10 adult dogs and saline injection in 10 adult dogs as control. Six weeks after injection, manometry was performed, and the frequency and degree of LES relaxation, clinical symptoms and esophageal barium test were observed. In vivo, pharmacological responses of the LES pressure to L-arginine, D-arginine , Nu-nitro~L-arginine (L-NNA, a competitive antagonist of NO synthase activity) and sodium nitroprusside (SNP) were observed respectively. In vitro, responses of LES strips to electric field stimulation ( EPS ) were recorded. By means of immunochemical method, NOS-positive myenteric ganglion cells were stained. The content of nitric oxide (NO) and the activity unit of nitric oxide synthase (NOS) in LES were determined.Results: (1) The LES pressures (LESP) in BAC-treated dogs were significantly higher than that in saline control (42. 43?.19mmHg vs. 22.71+5.19 mmHg, P<0. 01) . The frequency and degree of LES relaxation in BAC-treated dogs were lower (relaxation frequency: 40% vs. 100%; relaxation degree: 13.08 ?3.89 % vs. 94.33 + 3.36%, P<0.01 ) . Regurgitation and decreased food appetite were accordingly 60%, 80% andweightless was 1.1. 5kg in BAC-treated dogs. Compared with saline group the difference of symptoms was absolutely significant (P<0.01) . Esophageal barium retention was found in BAC-treated dogs but normal in saline control (rate of barium retention:81. 3 ?11. 4% vs 4. 2 ?.2%, P<0. 01) .These results showed that there were many similarities between BAC-treated dogs and achalasia of human in many aspects including clinical symptoms, manometry and esophageal barium test. (2) L-arginine significantly decreased LESP from 22. 71 + 5.19mmHg to 16. 70 + 3. 74mmHg and L-NNA increased LESP to 46. 90 + 8. 43mmHg in saline control . It was suggested that the L-arginine-nitric oxide pathway existed in LES and NO regulated the relaxation of LES. (3)L-arginine and L-NNA had no effects on BAC-treated dogs. It suggested that NOS may be decreased or disappeared in myenteric plexus of LES and the elevated LESP may be related to the loss of NO in BAC-treated dogs. (4) SNP reduced LESP in both BAC-treated and saline dogs (P>0. 05) . In vitro, responses of LES strips in two groups to carbachol and SNP were similar. It demonstrated that LES function in BAC-treated dogs was normal. L-arginine decreased LES tension and L-NAME antagonised its effect in saline control, but no effects on LES strips in BAC-treated dogs. It showed lack of NOS in LES strips of BAC-treated dogs. (5) With electrical field stimulation (EPS), the initially LES relaxing response appeared in saline control but lost in BAC-treated dogs. Only a contractile response of LES in BAC-treated group was similar to that in saline control. The LES relaxation in saline control was antagonised by L-NAME and increased by L-arginine partly. It unfolded that LES relaxation in saline group was caused by release of NO and lack of NO in LES of BAC-treated dogs. (6) With atropine, the contractile responses of LES strips in two groups disappeared ordecreased. It suggested that the contractile responses to EPS were caused by release of acetylcholine (Ach) and that Ach be present in LESof BAC-treated dogs. (7) Histological examination of BAC-treated LES verified the loss or decrease of myenteric ganglia. The content of NO and the activity unit of NOS in LES of BAG- treated dogs were markedly lower than that in saline control ( NO: 6. 058?. 067 umol/g vs. 1.797 ?.873 umol/g, NOS: 1.458 + 0.146 u/mgprot vs. 0.463+0.039 u/mgprot, P<0. 01). These results further demonstreated that decrease of No may be one of the most important causes. Conclusions:1. Injection o... |
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