The principal objective of this proposal is to understand the mechanisms of fluid and electrolyte secretion, by the exocrine pancreas. Pancreatic fluid and electrolyte secretion has an acinar and ductal components which most probably occur by different mechanisms. In addition, the secretory mechanism is likely to differ along the ductal tree. There are significant species variability; the rat pancreas represents one extreme secreting fluid relatively rich in Cl- whereas the guinea pig pancreas secretes fluid poor in Cl- and rich in HCO3-. We therefore propose to make use of these naturally occurring variabilities to identify and characterize the key mechanisms responsible for pancreatic fluid and electrolyte secretion. Towards achieving this goal, we have developed the techniques required to: prepare isolated interlobular ducts with or without acini attached to them, microdissect and perfuse intralobular ducts, and perfuse main ducts; measure the intracellular concentrations of and Cl-, H+/HCO3- and Ca 2+ using photon counting from single cells or image acquisition and analysis of cells loaded with fluorescent dyes; measure ion fluxes mediated by specific transporters. We have also developed a new and simple technique for the simultaneous recording of intracellular ionic activities and cell volume of single cells with high time resolution. These techniques will be used to 1) Study and compare H+/HCO3- transport mechanisms in rat and guinea pig pancreatic ducts, their regulation by specific agonists and relation to cell volume regulation. These studies are aimed at discovering the active mechanisms responsible for HCO3- secretion to the duct lumen. 2) Determine Cl- transport mechanisms by the various ductal (and acinar) cells. It is hoped that we will be able to determine the mechanism of active transepithelial Cl- absorption by duct cells and the basis for cellular and species variations. 3) Localize acid base and Cl- transporters to the serosal and/or luminal ductal cell membranes using perfused ducts. The purpose of these studies is to develop and test physiologically and thermodynamically sound models for acinar and ductal fluid and electrolyte secretion. 4) Study mechanisms of cell volume regulation in single pancreatic duct (and acinar) cells to understand the cellular volume compensatory mechanism and interrelationships between the transporters during the overall process of fluid and electrolyte secretion. I hope that the proposed studies will extend the knowledge and determine the underlying mechanisms of fluid and electrolyte secretion by the exocrine pancreas.
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