Both acinar and duct cells in the mammalian exocrine pancreas participate information of pancreatic juice. Little is known, however, of the specific mechanisms involved in fluid and electrolyte secretion in either cell type, or how these mechanisms are modulated by hormones and neurotransmitters during stimulus-response coupling. Recent studies suggest that active transport of Na+ across both acinar and duct epithelia is essential to fluid production and that Na+ transport is dependent on activity of the plasma membrane enzyme Na+, K+ activated adenosine triphosphatase (Na+/K+ pump). In the proposed research, the role of the Na+/K+ pump in pancreatinc secretion will be explored. To accomplish this goal, suspensions of dispersed epithelial cells will be prepared from guinea pig pancreas and separated by gradient centrifugation into populations enriched in either acinar or duct cells. The numbers of Na+/K+ pumps in each cell type and their activity will be determined by analysis of ouabain-sensitive 86Rb+ uptake, ouabain-sensitive oxygen consumption and 3H-ouabain binding to the cells. The former two techniques allow direct estimation of Na+/K+ pump-mediated transport activity and its relationship to oxidative metabolism in the cells. The last technique not only measures pump activity, but also provides a means by which the number of Na+/K+ pump molecules in the membranes of each cell type can be determined. The effects of pancreatic secretagogues such as cholecystokinin, VIP, secretin, and acetylcholine on Na+/K+ pump activity will be assessed. Further studies will be carried out to establish the intracellular events that couple receptor activation to alterations in activity of the ion transport enzyme. These will include investigation of ion dependencies, intracellular messengers such as cyclic nucleotides and Ca++, and metabolic energy requirements. The studies proposed will provide information on the ways in which hormones and neurotransmitters modulate Na+/K+ pump activity in acinar and duct epithelia of the exocrine pancreas. This information will be used to construct a cellular model of pancreatic fluid and electrolyte secretion. These data also will establish a strong foundation for further studies probing the dysfunctions in fluid secretion that are characteristic of certain diseases of the pancreas such as pancreatitis and cystic fibrosis.
