【博士論文】学術データベース

博士論文 / Motilin- and ghrelin-induced gastric contractions in different parts of Suncus stomach in vitro スンクスの胃各部位でのモチリン及びグレリンによる収縮調節機構

著者

書誌事項

タイトル

Motilin- and ghrelin-induced gastric contractions in different parts of Suncus stomach in vitro

タイトル別名

スンクスの胃各部位でのモチリン及びグレリンによる収縮調節機構

著者名

AMRITA DUDANI

学位授与大学

埼玉大学 (大学ID:0019) (CAT機関ID:KI00020X)

取得学位

博士(学術)

学位授与番号

甲第1066号

学位授与年月日

2017-03-22

注記・抄録

Background and Aim: Motilin and ghrelin are released in the interdigestive state to initiate and regulate phase III contractions of migrating motor complex (MMC). The MMC is responsible for emptying the stomach during the interdigestive period to prepare for the next meal. Gastric phase III contractions of the MMC originate in the stomach and propagate downward in the alimentary canal. Previously, I found that motilin and ghrelin synergistically induced gastric contractions both in vitro and in vivo. Motilin-induced contractions are regulated by a ghrelin-mediated GABAergic pathway. Therefore, I hypothesized that some regions of the stomach are more responsive to motilin and ghrelin and propagate strong contractions. The present study determined active responsive sites for motilin- and ghrelin-induced contractions in the stomach and elucidated mechanisms underlying the induction of these contractions.Methods: The stomachs of Suncus murinus or Asian house shrew, a small insectivorous mammal, were dissected, and the fundus, proximal corpus, distal corpus, and antrum were isolated to examine the effect of motilin- and/or ghrelin-induced contractions by using an organ bath system. The stomach segments were pretreated with tetrodotoxin, atropine, bicuculline, phaclofen, adenosine, and dopamine to determine the involvement of neural pathways. Quantitative PCR (qPCR) was performed to measure the mRNA expression of the motilin receptor GPR38. Distribution of ghrelin-immunopositive cells and mRNA expression of the GHSR in the different segments of the suncus stomach were examined by performing immunohistochemical analysis and RT-PCR, respectively.Results: Results of this in vitro study showed that treatment with 10-10 M motilin induced contractions only in the proximal corpus. In contrast, treatment with 10-9 M motilin induced strong contractions in the other segments of the suncus stomach. Motilin-induced contractions in each dissected stomach segment were inhibited by tetrodotoxin and atropine pretreatment, suggesting that these contractions were mediated by a cholinergic neural pathway in the myenteric plexus. Treatment with ghrelin (10-11-10-7 M) in the presence of low-dose motilin (10-10 M) induced gastric contractions in a dose-dependent manner in the fundus and proximal corpus but not in the distal corpus and antrum. In addition, pretreatment with ghrelin antagonist D-Lys3-GHRP6 blocked motilin-induced contractions in all the stomach segments. In contrast, treatment with GABA antagonists reversed this blockade in all the stomach segments. Treatment with adenosine A2A receptor and dopamine D2 receptor agonists also reversed ghrelin antagonist-induced inhibition of motilin-induced contractions. The mRNA expression of motilin receptor, GPR38 was highest in the proximal corpus and was the lowest in the antrum. The mRNA expression of GPR38 varied, with low expression in the mucosal layer and higher expression in the muscle layer. The mRNA expression of the ghrelin receptor, GHSR was detected in all the stomach segments. Density of ghrelin-immunopositive cells was significantly higher in the fundus and proximal corpus than in the other stomach segments.Conclusions: These results suggest that each gastric segment shows a different response toward motilin and/or ghrelin. The fundus and proximal corpus, including the cardia, are the most sensitive and responsive to motilin- and/or ghrelin-induced synergistic gastric contractions, suggesting that the proximal part of the stomach along with fundus is the first contractile site for MMC onset. In addition, present results indicate that adenosine(via A2A receptor) and dopamine (via D2 receptor) play vital roles in regulating motilin- and ghrelin-induced gastric contractions.

Declaration IThesis dedicated to IIAcknowledgement IIITable of contents VIList of figures XIList of tables XIIIList of appendix XIVAbbreviations XVAbstract XVIIIChapter 1: General Introduction and Objectives 11.1. Research background 11.1.1. The migrating motor complex (MMC) of the gastrointestinal (GI) motility 11.1.2. Motilin and its regulatory mechanism 21.1.3. Ghrelin and GI motility 31.1.4. Similar properties of motilin and ghrelin 41.1.5. Asian house shrew (Suncus murinus) 51.2. Hypothesis and objectives 7Chapter 2: The proximal gastric corpus is the most responsive site of motilin-induced contractions in the stomach of the Asian house shrew in vitro 82.1. Introduction 82.1.1. Motilin as an important regulator of MMC 82.1.2. Advantages of suncus for studying gastrointestinal physiology including gastric motility 92.2. Materials and Methods 112.2.1. Ethical Approval 112.2.2. Animals 112.2.3. Drugs used 112.2.4. Preparation of S. murinus isolated stomach 122.2.5. Gastric contractility study 132.2.6. GPR38 mRNA expression 142.2.7. Statistical analysis 152.3. Results 162.3.1. Spontaneous contractile pattern in the different segments of isolated stomach 162.3.2. Responses to motilin in different segments of stomach 162.3.3. The cholinergic pathway of the motilin-induced contraction 182.3.4. The myenteric plexus pathway of the motilin-induced contraction 182.3.5. GPR38 mRNA expression in the stomach 182.4. Discussion 202.4.1. Sensitivity towards motilin is region-specific 202.4.2. Physiological correlation of MMC and motilin 212.4.3. Regulatory mechanism of motilin-induced gastric contraction 212.4.4. Motilin receptor GPR38 mRNA expression 222.5. Summary24 Chapter 3: Synergistic effect of motilin and ghrelin induces different responses in different segments of the stomach of Suncus murinus in vitro 253.1. Introduction 253.1.1. Regulatory mechanism underlying motilin- and ghrelin-induced gastric contractions 253.2.2 Adenosine and dopamine as neurotransmitters and neuromodulators 263.2. Materials and Methods 283.2.1. Ethical approval 283.2.2. Animals 283.2.3. RT-PCR 293.2.4. Tissue preparation for morphological analysis 293.2.5. Immunohistochemical analysis 303.2.6. Morphometric analysis 313.2.7. Preparation of the isolated S. murinus stomach 313.2.8. Analysis of in vitro gastric contractions 323.2.9. Drugs used 333.2.10. Statistical analysis 333.3. Results 353.3.1. Synergistic effect of motilin and ghrelin on the different segments of the suncus stomach 353.3.2. Motilin and ghrelin synergistically induce gastric contractions through a cholinergic neural pathway 353.3.3. GHS-R mRNA expression in the different segments of the suncus stomach 353.3.4. Localization of ghrelin-ip cells in the suncus stomach 363.3.5. Effect of D-Lys3-GHRP6 and GABA receptor antagonists on motilin-induced gastric contractions in vitro 363.3.6. Effect of non-selective adenosine and dopamine receptor agonists on motilin-induced gastric contractions 373.3.7. Effect of adenosine A2A receptor agonist and dopamine D2 receptor agonist on motilin-induced gastric contractions 373.4. Discussion 393.4.1. Role of ghrelin in the MMC 393.4.2. Involvement of the GABAergic pathway in motilin-induced gastric contractions 403.4.3. Adenosine and dopamine are the key molecules for mediating motilin-induced gastric contractions 414. Summary and Conclusion 435. Figures, Tables and Appendix 456. References 78

主指導教員 : 坂井貴文

博士の専攻分野の名称 : 博士(学術)学位授与年月日 : 平成29年3月22日

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NII論文ID(NAID)

500001054908

NII著者ID(NRID)
  • 8000001169344
本文言語コード

eng

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機関リポジトリ / NDLデジタルコレクション

博士論文 / 埼玉大学 / 学術

博士論文 / 埼玉大学

博士論文 / 学術

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