Neural Control of the Gallbladder

描述(申请人提供)：收缩功能减弱是胆囊病的一个特征，它与炎症和胆结石的形成有关。最近的报道表明，胆汁中胆固醇和疏水性盐类的升高导致了胆囊动力的下降，随后的炎症和胆囊排空障碍导致结石的形成。此外，越来越多的证据表明，亲水性胆盐可能通过恢复胆囊平滑肌(GBSM)的收缩能力和减少胆结石的发展和炎症而具有治疗潜力。然而，收缩能力降低、炎症和胆结石形成之间的相互关系尚不清楚，这反映了我们对导致胆囊病患者静息张力降低和餐后收缩的细胞事件缺乏了解。这种相互竞争的更新将解决我们知识中的这些根本差距。这项拨款申请的总体目标是：(1)阐明在胆结石疾病中GBSM收缩功能受损的细胞和离子机制；(2)解决疏水性胆盐与GBSM相互作用的机制；(3)阐明炎症在胆结石疾病中平滑肌功能障碍和相关胆汁淤积中的作用；以及(4)阐明亲水性胆盐通过预防或逆转这些紊乱在保护胆囊功能中的作用。将使用两个实验模型。在一个模型中，我们将使用喂养致石(形成胆结石)饮食的野生型和转基因小鼠来评估正常小鼠、没有发生胆囊炎的小鼠以及缺乏核或膜胆汁酸受体的小鼠在胆结石疾病进展过程中GBSM发生的功能变化。我们还将评估在用保护性亲水性胆盐、熊去氧胆酸或环氧合酶阻滞剂治疗的同时喂食促结石饮食的小鼠的GBSM功能。第二个模型包括评估胆固醇和疏水性胆盐在体外应用于GBSM制剂的效果，以及预先应用熊去氧胆酸或不应用熊去氧胆酸的效果。这种方法使我们能够在受控环境中单独检查这些化合物对GBSM的作用。这项资助提案涉及一种综合方法，使用最先进的技术来研究动物疾病发展过程中从单分子到完整组织的胆囊病理生理学。可使用的技术包括完整肌束中的GBSM细胞的钙成像，分别来自完整和分离的GBSM细胞的细胞内和膜片钳记录，以及肌张力测量。结合我们以前对平滑肌功能的研究，我们将深入了解胆汁淤积是如何导致胆囊运动功能障碍的，以及运动功能障碍与炎症之间的关系。此外，这些研究将阐明亲水性胆盐对GBSM的治疗潜力，以防止或逆转导致胆囊收缩能力下降的细胞变化，并揭示其作用的细胞机制。与公共卫生相关：胆石症是一种极其常见的疾病，涉及到胆汁收缩能力的降低和胆汁排入肠道，从而导致胆结石的形成。这项拨款申请的目的是了解胆结石疾病中胆囊肌被破坏的细胞机制，并确定亲水性胆汁盐是否以及如何保护胆囊免受这些变化的影响。

DESCRIPTION (provided by applicant): Decreased contractility is a hallmark of gallbladder disorders, and it is associated with inflammation and gallstone formation. Recent reports suggest that decreased gallbladder motility is caused by elevated cholesterol and hydrophobic salts in the bile, and that ensuing inflammation and impairments in gallbladder emptying lead to stone formation. Furthermore, mounting evidence suggests that hydrophilic bile salts may have therapeutic potential by restoring gallbladder smooth muscle (GBSM) contractility and reducing gallstone development and inflammation. Nevertheless, the interrelationships between decreased contractility, inflammation, and gallstone formation are not understood, and this reflects our lack of understanding of the cellular events that lead to decreased resting tone and postprandial contractions in gallbladder disease. This competing renewal will address these fundamental gaps in our knowledge. The overall objectives of this grant application are to: (1) elucidate the cellular and ionic mechanisms by which GBSM contractility is disrupted in gallstone disease, (2) resolve the mechanism by which hydrophobic bile salts interact with GBSM; (3) elucidate the role of inflammation in smooth muscle dysfunction and associated biliary stasis in gallstone disease; and (4) elucidate the role of hydrophilic bile salts in the protection of gallbladder function by preventing or reversing these disruptions. Two experimental models will be used. In one model, we will use wild type and transgenic mice fed a lithogenic (gallstone forming) diet to evaluate the functional changes that occur in GBSM during the progression of gallstone disease in normal mice, mice that do not develop cholecystitis, and mice lacking nuclear or membrane bile acid receptors. We will also evaluate GBSM function in mice fed a lithogenic diet while being treated with the protective hydrophilic bile salt, ursodeoxycholic acid, or a cyclooxygenase inhobitor. The second model involves evaluation of the effects of cholesterol and hydrophobic bile salts applied in vitro to GBSM preparations, with or without prior application of ursodeoxycholic acid. This approach allows us to examine the actions of these compounds individually on GBSM in a controlled environment. This grant proposal involves an integrated approach using state-of-the-art techniques to investigate gallbladder pathophysiology from single molecules to the intact tissue during the progression of disease in the animal. Techniques to be used include Ca2+ imaging of GBSM cells in intact muscle bundles, intracellular and patch clamp recording from intact and isolated GBSM cells, respectively, and muscularis tension measurements. Together with our previous studies on smooth muscle function, we will provide insights on how gallbladder motor function is disrupted leading to biliary stasis, and the relationship between disrupted motor function and inflammation. Furthermore, these studies will elucidate the therapeutic potential of hydrophilic bile salts on GBSM to prevent or reverse cellular changes that underlie decrease gallbladder contractility, and uncover the cellular mechanisms that mediate their actions. PUBLIC HEALTH RELEVANCE: Gallstone disease is an extremely common disorder that involves a decrease in the ability of the gallbladder to contract and empty bile into the intestine, thus leading to gallstone formation. The objectives of this grant application are to understand the cellular mechanisms by gallbladder muscle is disrupted in gallstone disease, and to determine whether and how hydrophilic bile salts can protect the gallbladder from these changes.