Abstract. Regulated secretion from pancreatic acinar cells occurs by exocytosis of zymogen granules (ZG) at the apical plasmalemma. ZGs originate from the TGN and undergo prolonged maturation and condensation. After exocytosis, the zymogen granule membrane (ZGM) is retrieved from the plasma membrane and ultimately reaches the TGN. In this study, we analyzed

Late in gestation, the prenatal fetal alveolar epithelium switches from fluid secretion to resorption of salt and water via apical sodium channels and basal Na-K-ATPase. The amounts of lung sodium pump activity protein and mRNA increase in the lung just before birth. Because maternal glucocorticoids (GC) may promote maturation of the alveolar epithelium and augment fetal surfactant apoprotein levels, we hypothesized that GC increase the fetal lung Na-K-ATPase alpha- and beta-subunit gene expression in development. Timed-pregnant Sprague-Dawley rats were injected daily with intraperitoneal dexamethasone (1 mg/kg) or saline for 1, 3, or 5 days before death at fetal day (FD) 17 or 19. Maternal GC treatment altered the fetal lung wet to dry weight, decreasing it at FD17 and increasing it at FD19. Northern analysis of total lung RNA for the alpha1- and beta1-pump subunits demonstrated differential regulation of the mRNA in response to GC. At FD17, beta1-mRNA increased after 1 (FD16) or 3 days (FD14-FD16) of GC treatment, whereas alpha1-mRNA was not altered. There were accompanying increases in beta1-, but not alpha1-, protein. At FD19, GC treatment for 5 days (FD14-FD18) increased beta1- and decreased alpha1-mRNA levels, but treatment for 1 (FD18) or 3 days (FD16-FD18) had no effect. In all groups, the alpha1-Na-K-ATPase protein was predominantly on the basolateral surface of airspace epithelium by immunofluorescence. In summary, maternal dexamethasone differentially affected the fetal lung mRNA levels of the two sodium pump subunits in a complex manner, with increased beta1-mRNA levels dependent on duration of treatment and fetal age.

We undertook studies to determine whether secretagogue action on the exocrine pancreas and parotid is accompanied by phosphorylation of proteins in intact cells. For this purpose, rat pancreatic, and parotid Iobules were preincubated with 32p~ for 45 min at 37°C, washed, and then incubated at 37°C in the presence or absence of secretagogues that effect discharge through different second messengers. Among a variety of polypeptides exhibiting enhanced phosphorylation in pancreatic Iobules upon a 30-s incubation in the presence of the secretagogues carbamylcholine, cholecystokinin octapeptide, or secretin, one species with an Mr of 29,000 was especially notable for three reasons: (a) its enhanced level of phosphorylation was dependent on the dose of secretagogue used and was still apparent after incubation for 30 min at 37°C; (b) an analogous phosphorylated polypeptide was observed in isoproterenolstimulated parotid Iobules; and (c) in both tissues its selective dephosphorylation was observed upon termination of stimulation by administration of atropine to carbamylcholine-stimulated pancreatic Iobules and propranolol to isoproterenol-stimulated parotid Iobules. These results suggest that the phosphorylation of one protein with an Mr of 29,000 is closely correlated both temporally and in a dose-dependent fashion with secretagogue action in both the exocrine pancreas and parotid.The mechanism whereby secretagogues are able to elicit a characteristic biological response in their respective target ceils is unclear at the moment. There is now increasing evidence that protein phosphorylation is involved in mediating the effects of a variety of the actions of hormones in many diverse enzymological and physiological processes (1). To gain further insight into the mechanism of secretagogue action in exocrine glands, we examined the relationship between protein phosphorylation and hormone action in the intact cell to determine whether this covalent modification mediates or modulates any of the biological actions of secretagogues.The acinar cells of the exocrine pancreas and parotid offer good systems for studying secretagogue effects on endogenous protein phosphorylation since homogeneous cell preparations can be prepared, several different secretagogues exist which elicit discharge through different intracellular messengers, and the effects of these hormones on calcium and cyclic nucleotide levels during secretion are well characterized (2, 3, 4). Since cAMP, cGMP, and Ca 2+ have been implicated in the secretion of exportable proteins from the exocrine pancreas, protein kinases are plausible targets for these putative secretagogue mediators and, in fact, both cAMP and cGMP-dependent protein kinases have been partially purified from homogenates of rat pancreas (5, 6).We examined the relationship between endogenous protein phosphorylation and secretagogue action in situ in gland lobules under physiological conditions using the rat exocrine pancreas and parotid as model systems. A preliminary note on this research has...

Late in gestation lung epithelium changes from net chloride and fluid secretion to sodium and fluid absorption. Fluid resorption is required for postnatal gas exchange and occurs by combined action of epithelial sodium channels and Na, K-ATPase. We hypothesized that alveolar epithelial Na, K-ATPase increases perinatally. Immunofluorescence (IF) and immunoelectron microscopy (IEM) with a monoclonal anti-alpha subunit antibody demonstrated that Na, K-ATPase was present on the basolateral surfaces of columnar epithelial cells at fetal day (FD) 17 and on type II cells throughout development. However, type I epithelial cells did not have detectable Na,K-ATPase. The steady-state levels of both the alpha 1 isoform and the beta-subunit mRNAs were maximal at FD20-neonatal day (ND) 1, with consistent increases from FD17 level. Na, K-ATPase alpha-subunit protein also increased from FD17 to FD20-22 and then decreased in the early postnatal period. The ouabain-inhibitable sodium pump activity per milligram membrane protein increased 2.6-fold from FD17 to FD22-ND1 (P < 0.05). The quantities of sodium pump mRNA, antigenic protein, and enzyme activity increase in late gestation in accord with a proposed role for Na, K-ATPase in resorption of alveolar sodium and fluid in preparation for birth.

Using a battery of seven lectin-ferritin conjugates as probes for cell surface glycoconjugates, we have studied the pattern of plasmalemmal differentiation of cells in the embryonic rat pancreas from day 15 in utero to the early postpartum stage. Our results indicate that differentiation of plasmalemmal glycoconjugates on acinar, endocrine, and centroacinar cells is temporally correlated with development and is unique for each cell type, as indicated by lectin-ferritin binding. Specifically, (a) expression of adult cell surface saccharide phenotype can be detected on presumptive acinar cells as early as 15 d in utero, as indicated by soybean agglutinin binding, and precedes development of intracellular organelles characteristic of mature acinar cells; (b) maturation of the plasmalemma of acinar cells is reached after intracellular cytodifferentiation is completed, as indicated by appearance of Con A and fucoselectin binding sites only at day 19 of development ; conversely, maturation of the endocrine cell plasmalemma is accompanied by "loss" (masking) of Ricinus communis II agglutinin receptors; and (c) binding sites for fucose lectins and for soybean agglutinin are absent on endocrine and centroacinar cells at all stages examined . We conclude that acinar, centroacinar, and endocrine cells develop from a common progenitor cell(s) whose plasmalemmal carbohydrate composition resembles most closely that of the adult centroacinar cell .Finally, appearance of acinar lumina beginning at -17 d in utero is accompanied by differentiation of apical and basolateral plasmalemmal domains of epithelial cells, as indicated by enhanced binding of several lectin-ferritin conjugates to the apical plasmalemmal, a pattern that persists from this stage through adult life .Our previous studies showed that the plasmalemmas of acinar, centroacinar, and endocrine cells of the mammalian pancreas each possess a distinctive glycoconjugate composition as revealed by differential binding of a battery of lectin-ferritin conjugates (15, 16). Whereas acinar cells display receptors for all lectins tested (Con A', specific for mannosyl and glucosyl residues ; RCA I, galactosyl residues ; RCA 11 and SBA. Nacetyl-galactosaminyl residues, WGA, N-acetyl-glucosaminyl residues; Ulex europeus and Lotus tetragonolobus lectins, fucosyl residues), endocrine and centroacinar cells are devoid of ' Abbreviations used in this paper: Con A, concanavalin A; RCA 1 and RCA 11, Ricinus communis agglutinins l and 11 ; SBA, soybean agglutinin; WGA, wheat germ agglutinin ; BSA, bovine serum albumin; KRB, Krebs-Ringer bicarbonate salt solution supplemented with amino acids and glucose; STI, soybean trypsin inhibitor. 96binding sites for fucose-specific lectins and for SBA, but can be distinguished from each other by the ability of centroacinar cells to bind RCA 11 . Because these three cell types arise from an endodermal evagination of the embryonic foregut that forms the pancreatic primordial tissue, it was of interest to examine the pattern of differentiatio...