SIGNAL TRANSDUCTION IN THE PANCREATIC DUCT SYSTEM

对于细胞内信号转导的分子机制知之甚少， 排泄管的杯状细胞和分泌碳酸氢盐的主细胞 外分泌胰腺的导管上皮，尽管它们在 胰液的加工 通过这些机制， 中枢神经系统调节 胃肠道激素（例如分泌素）尤其不清楚。 在 建议的研究，最近开发的孤立和 培养的豚鼠胰管上皮细胞将被用于 鉴定生理调节剂和细胞内信使， 控制导管粘蛋白和碳酸氢盐分泌，并确定细胞 识别的信号转导级联的目标。 在这些研究中 导管碳酸氢盐分泌的潜在激动剂对 测定细胞内游离Ca 2+和环核苷酸水平 以及环核苷酸和肌醇之间的潜在相互作用 磷脂介导的信号通路。 三维动力学 激动剂引起的钙变化将通过新的 视频速率激光扫描共聚焦显微技术的发展 （LSCM），使用荧光钙探针。 的细胞靶标 确定的细胞内信号分子将被定义， 特别注意阴离子和阳离子交换机制， 使用同位素通量测量和LSCM的阴离子通道。 的影响 假定的生理调节剂和细胞内信使， 细胞内pH将使用LSCM和pH敏感的 荧光染料。 激动剂诱导粘蛋白从杯状体释放的动力学 分离的胰管中的细胞将通过组合的 固相放射免疫分析、形态计量学和共聚焦显微镜 显微镜 这些实验将提供一个详细的图片， 副交感神经系统通过细胞内机制 和胃肠激素调节碳酸氢盐和粘蛋白分泌， 胰管上皮细胞，并提供必要的 这些信号通路的扰动的未来研究背景 发生在胰腺疾病如囊性纤维化和胰腺- 炎症。

Little is known of the molecular mechanisms of signal transduction in the goblet cells and bicarbonate-secreting principal cells of the excretory duct epithelium of the exocrine pancreas, despite their central role in elaboration of pancreatic juice. The mechanisms through which input from the central nervous system modulates the regulatory effects of gastrointestinal hormones such as secretin are particularly obscure. In the proposed research, the recently developed model of isolated and cultured guinea pig pancreatic duct epithelial cells will be exploited to identify physiological regulators and intracellular messengers that control ductal mucin and bicarbonate secretion and to define the cellular targets of the identified signal transduction cascades. In these studies the effects of potential agonists of ductal bicarbonate secretion on intracellular free Ca2+ and cyclic nucleotide levels will be determined as will potential interactions between cyclic nucleotide- and inositol phospholipid-mediated signaling pathways. The three dimensional kinetics of calcium changes induced by agonists will be examined through the newly developed technology of video rate laser scanning confocal microscopy (LSCM), using fluorescent calcium probes. The cellular targets of identified intracellular signaling molecules will be defined, with particular attention directed to anion and cation exchange mechanisms and anion channels using isotope flux measurements and LSCM. Effects of putative physiological regulators and intracellular messengers on intracellular pH will be examined using LSCM and pH-sensitive fluorochromes. The kinetics of agonist-induced mucin release from goblet cells in isolated pancreatic ducts will be examined through the combined approaches of solid phase radioimmune assay and morphometry and confocal microscopy. These experiments will provide a detailed picture of the intracellular mechanisms through which the parasympathetic nervous system and gastrointestinal hormones regulate bicarbonate and mucin secretion by the epithelium of the pancreatic ducts as well as providing a necessary context for future studies on perturbations of these signaling pathways that occur in pancreatic diseases such as cystic fibrosis and pancrea- titis.