REGULATION OF SECRETION BY CALCIUM WAVES IN THE PANCREAS

胰腺中钙波的分泌调节

• 批准号: 6263275
• 负责人: MICHAEL H NATHANSON
• 金额: $ 4.03万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-06-01 至 2004-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6263275
• 关键词: acinar cell; basolateral membrane; calcium channel; calcium flux; calcium ion; calcium metabolism; cell growth regulation; confocal scanning microscopy; cooperative study; epithelium; exocytosis; flash photolysis; gap junctions; laboratory rat; liver cells; liver function; microinjections; pancreas; second messengers; secretion

Cytosolic Ca2+ (Cai2+) regulates a wide range of cell functions, from secretion to metabolism to cell growth and death. It is unknown how Ca2+ simultaneously controls such diverse activities in an individual cell, although Cai2+ waves and other types of Ca2+ gradients may be responsible by allowing distinct Cai2+ signals to occur in different subcellular regions. The parent grant for this FIRCA proposal tests the hypothesis that Cai2+ + waves in hepatocytes result from sequential release of distinct subcellular Ca2+ pools, and that localized, subcellular increases in Cai2+ canregulate specific hepatocyte functions. The current proposal would extend the hypothesis of the parent grant in an important way, by testing whether subcellular and intercellular Cai2+ waves regulate secretion in pancreatic acinar cells, a well-established model of polarized epithelium. The specific aims of this project are: 1. To determine the mechanism by which Cai2+ + waves spread across individual acinar cells. We will test whether Cai2+ can be increased selectively in either the apical or basolateral region. Subcellular Cai2+ signals will be elicited by microinjection of caged second messengers, followed by localized (apical or basolateral) release of these agonists using flash photolysis. Also, we will inject agents to specifically enhance or antagonize the effects of these second messengers to determine which of these messenger molecules affect Cai2+ waves. Individual acinar cells within intact acini will be used, and Cai2+ signals will be detected by confocal microscopy. 2. To determine the mechanism by which Cai2+ waves spread from cell to cell in the intact pancreatic acinus. We will test which second messengers must cross gap junctions in order to coordinate the spread of Cai2+ wavesfrom cell to cell. Increases in Cai2+ in individual acinar cells will be induced by microinjection of caged second messengers followed by their release through flash photolysis in the presence or absence of gap junction blockers. The resulting Cai2+ waves will be measured throughout the entire acinus using confocal microscopy. 3. To determine the relationship between subcellular and intercellular Cai2+ increases and a Ca2+-mediated event, exocytosis. We will testif apical exocytosis requires only that Cai2+ increases in the apical region, and whether exocytosis is potentiated if such apical Cai2+ increases are coordinated among neighboring cells. Localized apical or basolateral Cai2+ signals will be elicited in acinar cells by flash photolysis. Cai + and exocytosis will be detected simultaneously using confocal microscopy.

胞质Ca2+（CAI2+）调节分泌的广泛细胞功能 代谢细胞生长和死亡。尚不清楚Ca2+如何同时 控制单个细胞中的这种不同活性，尽管CAI2+波 和其他类型的Ca2+梯度可能是通过允许不同的CAI2+负责的 在不同的亚细胞区域发生的信号。父母的赠款 FIRCA提案检验了以下假设：肝细胞中的CAI2 + +波 从不同的亚细胞Ca2+池的顺序释放，以及该位置的， CAI2+的亚细胞增加可以调节特定的肝细胞功能。这 当前的提议将扩大父母赠款的假设 重要方法，通过测试亚细胞和细胞间CAI2+波是否 调节胰腺腺泡细胞中的分泌，这是一个完善的模型 偏振上皮。该项目的具体目的是： 1。确定CAI2 + +波散布在个人的机制 腺泡细胞。我们将测试是否可以选择性地增加CAI2+ 顶端或基底外侧区域。亚细胞CAI2+信号将通过 笼中的第二使者的显微注射，然后是局部化（根尖或 基底外侧）使用闪光分解释放这些激动剂。另外，我们会的 注入特定增强或对抗这些第二的效果 信使确定哪些信信体分子会影响CAI2+波。 将使用完整acini中的单个腺泡细胞，并使用CAI2+信号 将通过共聚焦显微镜检测。 2。确定CAI2+波从细胞到细胞中cai2+波的机制 完整的胰腺刺激。我们将测试哪些第二使者必须越过 间隙连接以将CAI2+波的扩散与从细胞到 细胞。单个腺泡细胞中CAI2+的增加将由 对笼中的第二使者的显微注射，然后通过 在存在或不存在间隙连接器的情况下，闪光光解。这 最终的CAI2+波将在整个acinus中测量 共聚焦显微镜。 3。确定亚细胞和细胞间CAI2+之间的关系 增加和Ca2+介导的事件，胞吐作用。我们将作证 胞吞作用仅需要CAI2+在顶端区域增加，以及是否是否 如果这种顶端CAI2+增加在 相邻的细胞。局部顶端或基底外侧CAI2+信号将是 通过闪光照射在腺泡细胞中引起。 CAI +和胞吐作用将是 使用共聚焦显微镜同时检测到。