Guinea Pig Pancreatic Duct Cells Research Articles

Electrophysiological properties of anion exchangers in the luminal membrane of guinea pig pancreatic duct cells

The pancreatic duct epithelium secretes the HCO3−-rich pancreatic juice. The HCO3− transport across the luminal membrane has been proposed to be mediated by SLC26A Cl−–HCO3− exchangers. To examine the electrophysiological properties of Cl−–HCO3− exchangers, we directly measured HCO3− conductance in the luminal membrane of the interlobular pancreatic duct cells from guinea pigs using an inside-out patch-clamp technique. Intracellular HCO3− increased the HCO3− conductance with a half-maximal effective concentration value of approximately 30 mM. The selectivity sequence based on permeability ratios was SCN− (1.4) > Cl− (1.2) = gluconate (1.1) = I− (1.1) = HCO3− (1.0) > methanesulfonate (0.6). The sequence of the relative conductance was HCO3− (1.0) > SCN− (0.7) = I− (0.7) > Cl− (0.5) = gluconate (0.4) > methanesulfonate (0.2). The current dependent on intracellular HCO3− was reduced by replacement of extracellular Cl− with gluconate or by H2DIDS, an inhibitor of Cl−–HCO3− exchangers. RT-PCR analysis revealed that the interlobular and main ducts expressed all SLC26A family members except Slc26a5 and Slc26a8. SLC26A1, SLC26A4, SLC26A6, and SLC26A10 were found to be localized to the luminal membrane of the guinea pig pancreatic duct by immunohistochemistry. These results demonstrate that these SLC26A Cl−–HCO3− exchangers may mediate the electrogenic HCO3− transport through the luminal membrane and may be involved in pancreatic secretion in guinea pig ducts.

Effects of CFTR gene silencing by siRNA or the luminal application of a CFTR activator on fluid secretion from guinea-pig pancreatic duct cells

AimsThe cystic fibrosis transmembrane conductance regulator (CFTR) is a cyclic AMP regulated chloride channel expressed in the apical plasma membrane of pancreatic duct cells where it plays an important role in fluid secretion. The purpose of this study was to elucidate the role of the CFTR chloride channel on ion and fluid secretion from the guinea-pig pancreas by manipulating the expression of CFTR by RNA interference or by luminal application of a CFTR selective activator, MPB91, in isolated cultured pancreatic ducts. Materials and methodsUsing cDNA isolated from the guinea-pig small intestine, fragments of the CFTR gene were generated by polymerase chain reaction and directly sequenced. Two different RNA duplexes for small interference RNA (siRNA) were designed from the sequence obtained. Fluid secretion from the isolated guinea-pig pancreatic ducts was measured using video-microscopy. The amount of CFTR chloride channel or AQP1 water channel expressed in pancreatic ducts was examined by immunoblotting with antibodies against CFTR or AQP1, respectively. ResultsGuinea-pig CFTR consists of 1481 amino acid residues. An additional glutamine residue was found to be inserted between amino acid residues 403 and 404 of human CFTR. Forskolin-stimulated fluid secretion from intact pancreatic ducts was significantly higher in the presence of MPB91 compared to fluid secretion in the absence of MPB91. Both basal and forskolin-stimulated fluid secretion in pancreatic ducts transfected with CFTR specific siRNAs were reduced by ∼50% compared to fluid secretion from ducts transfected with scrambled negative control dsRNAs. The amount of CFTR and AQP1 proteins was reduced to 34% and 45% of control, respectively. ConclusionsThe activity of the CFTR chloride channel or the amount of CFTR protein expressed determines the rate of fluid secretion from the isolated guinea-pig pancreatic ducts.

Dynamic Regulation of CFTR Bicarbonate Permeability by [Cl−]i and Its Role in Pancreatic Bicarbonate Secretion

Pancreatic bicarbonate (HCO3-) secretion is important for a healthy pancreas as well as digestive physiology. However, how human pancreatic duct cells secrete copious amounts of HCO3- has long been a puzzle. Here, we report that a dynamic increase in the cystic fibrosis transmembrane conductance regulator (CFTR) HCO3- permeability by intracellular Cl- concentration ([Cl-]i)-sensitive mechanisms plays a pivotal role in pancreatic HCO3- secretion. The role of [Cl-]i-sensitive kinases in CFTR-mediated HCO3- transport was examined in heterologous expression systems, PANC1 human pancreatic duct cells, and human and guinea pig pancreatic tissues using an integrated molecular and physiologic approach. In human pancreatic tissues, CFTR-positive duct cells abundantly expressed with-no-lysine (WNK1) kinase, oxidative stress-responsive kinase 1 (OSR1), and sterile 20/SPS1-related proline/alanine-rich kinase (SPAK), which are known to be activated by low [Cl-]i. Interestingly, CFTR activation rapidly decreased [Cl-]i in response to luminal Cl- depletion in polarized PANC1 human pancreatic duct cells. Notably, the WNK1-mediated OSR1 and SPAK activation by low [Cl-]i strongly increased CFTR HCO3- permeability in CFTR-transfected HEK 293T, PANC1, and guinea pig pancreatic duct cells, making CFTR primarily an HCO3- channel, which is essential for the secretion of pancreatic juice containing HCO3- at a concentration greater than 140 mmol/L. In contrast, OSR1 and SPAK activation inhibited CFTR-dependent Cl-/HCO3- exchange activity that may reabsorb HCO3- from the high HCO3--containing pancreatic juice. These results indicate that the [Cl-]i-sensitive activation of the WNK1-OSR1/SPAK pathway is the molecular switch to generate HCO3--rich fluid in the human pancreatic duct.

CFTR Expression But Not Cl− Transport Is Involved in the Stimulatory Effect of Bile Acids on Apical Cl−/HCO3 − Exchange Activity in Human Pancreatic Duct Cells

Low doses of chenodeoxycholate (CDC) stimulate apical anion exchange and HCO3(-) secretion in guinea pig pancreatic duct cells (Gut. 2008;57:1102-1112). We examined the effects of CDC on intracellular pH (pHi), intracellular Ca(2+) concentration ([Ca(2+)]i), and apical Cl(-)/HCO3(-) exchange activity in human pancreatic duct cells and determined whether any effects were dependent on cystic fibrosis transmembrane conductance regulator (CFTR) expression and Cl(-) channel activity. Polarized CFPAC-1 cells (expressing F508del CFTR) were transduced with Sendai virus constructs containing complementary DNAs for either wild-type CFTR or beta-galactosidase. Microfluorimetry was used to record pHi and [Ca(2+)]i and apical Cl(-)/HCO3(-) exchange activity. Patch clamp experiments were performed on isolated guinea pig duct cells. Chenodeoxycholate induced a dose-dependent intracellular acidification and a marked increase in [Ca(2+)]i in CFPAC-1 cells. CFTR expression slightly reduced the rate of acidification but did not affect the [Ca(2+)]i changes. Luminal administration of 0.1 mmol/L of CDC significantly elevated apical Cl(-)/HCO3(-) exchange activity but only in cells that expressed CFTR. However, CDC did not activate CFTR Cl(-) conductance. Bile salts modulate pHi, [Ca(2+)]i, and apical anion exchange activity in human pancreatic duct cells. The stimulatory effect of CDC on anion exchangers requires CFTR expression but not CFTR channel activity.

Effect of bile acids on ion conductances in guinea pig pancreatic duct cells

Effect of bile acids on ion conductances in guinea pig pancreatic duct cells

THE EFFECT OF BILE ACIDS ON PANCREATIC DUCTAL EPITHELIA

Introduction: Exposure of the pancreas to bile acids is considered to be one of the possible causes of acute pancreatitis. However, no information is available on the effects of bile acids on intact pancreatic ductal epithelia. The aim of this study was to investigate the effect of various bile acids on pancreatic duct cells. Methods: Isolated guinea pig intra/interlobular pancreatic ducts were microperfused from the luminal and basolateral membranes. The effects of various bile acids /chenodeoxycholate, ursodeoxycholate and lithocholate/ on intracellular calcium concentration [Ca2+]i, and intracellular pH (pHi) were measured. Results: The exposure of intact ducts to all three primary bile acids induced a marked increase in [Ca2+]i and chenodeoxycholate caused the largest response. The effect of chenodeoxycholate from the luminal side was more pronounced than from the basolateral membrane. Furthermore, glycin conjugated bile acids, which are not membrane permeable, resulted in Ca2+ signals when administered from either side of the ducts. Calcium signals could still be triggered by bile acids in a Ca2+-free external solution, but preincubation of the ducts with an intracellular Ca2+-chelator, BAPTA-AM, totally abolished the effect of bile acids. Basolateral administration of chenodeoxycholate or glycochenodeoxycholate caused a reversible intracellular acidification. Conclusion: These results suggest the presence of bile acid transporters on guinea pig pancreatic duct cells. The transport of bile acids into duct cells leads to an increase in [Ca2+]i which is probably mediated via the release of calcium from intracellular stores. The bile acid-induced acidification of pancreatic ductal epithelia may play a role during acute pancreatitis. Supported by OTKA, MTA, OM.

Bile acids cause acidification and CA2+-signalling in guinea pig pancreatic duct cells

Bile acids cause acidification and CA2+-signalling in guinea pig pancreatic duct cells

Apical Bicarbonate Conductance in Guinea-pig Pancreatic Duct Cells

Apical Bicarbonate Conductance in Guinea-pig Pancreatic Duct Cells

Ethanol induces fluid hypersecretion from guinea-pig pancreatic duct cells

Graphical Abstract Abstract figure legend HCN channels play an evolutionarily conserved pacemaker role in renal pelvic smooth muscle (RPSM) of lower and higher order mammals. The function of hyperpolarization-activated cation (HCN) channels in smooth muscle pacemakers remains controversial. Renal pelvic smooth muscle pacemakers trigger smooth muscle contractions that expel waste from the kidney, and HCN channels have been localized to these pacemaker tissues. To date, however, the mechanisms underlying RPSM pacemaker activity remain elusive. RPSM pacemaker activity was investigated in both lower (top left) and higher order (bottom left) mammalian models, which exhibit divergent upper urinary tract anatomies. We performed morphological and functional studies from the single-molecule to the whole-organ level and showed that HCN channels drive RPSM pacemaker activity. RPSM pacemakers (boxed regions) integrated into the muscular syncytium, expressed HCN channels on their plasmalemma and exhibited the Ih ‘funny’ pacemaker current conducted by HCN channels. Critically, HCN channel block abolished electrical pacemaker activity and peristaltic smooth muscle contractions in both lower and higher order mammalian upper urinary tracts. Thus, HCN channels play an evolutionarily conserved pacemaker role in RPSM pacemakers.

Ethanol induces fluid hypersecretion from guinea-pig pancreatic duct cells

Ethanol is the leading cause of pancreatitis; however, its cellular effects are poorly understood. We examined the direct effects of ethanol in the concentration range 0.1-30 mM, i.e. relevant to usual levels of drinking, on fluid secretion from guinea-pig pancreatic duct cells. Fluid secretion was continuously measured by monitoring the luminal volume of interlobular duct segments isolated from the guinea-pig pancreas. [Ca2+]i was estimated by microfluorometry in duct cells loaded with fura-2. Ethanol at 0.3-30 mM significantly augmented fluid secretion stimulated by physiological (1 pM) or pharmacological (1 nM) concentrations of secretin. It augmented dibutyryl cAMP-stimulated fluid secretion but failed to affect spontaneous or acethylcholine-stimulated secretion. Ethanol at 1 mM shifted the secretin concentration-fluid secretion response curve upwards and raised the maximal secretory response significantly by 41%. In secretin-stimulated ducts, 1 mM ethanol induced a transient increase in [Ca2+]i that was dependent on the presence of extracellular Ca2+. Ethanol failed to augment secretin-stimulated secretion from ducts pretreated with an intracellular Ca2+ buffer (BAPTA) or a protein kinase A inhibitor (H89). In conclusion, low concentrations of ethanol directly augment pancreatic ductal fluid secretion stimulated by physiological and pharmacological concentrations of secretin, and this appears to be mediated by the activation of both the intracellular cAMP pathway and Ca2+ mobilization.

Chloride transport in microperfused interlobular ducts isolated from guinea-pig pancreas.

Isolated interlobular ducts from the guinea-pig pancreas secrete a HCO3--rich fluid in response to secretin. To determine the role of Cl- transporters in this process, intracellular Cl- concentration ([Cl-]i) was measured in ducts loaded with the Cl--sensitive fluoroprobe, 6-methoxy-N-ethylquinolinium chloride (MEQ). [Cl-]i decreased when the luminal Cl- concentration was reduced. This effect was stimulated by forskolin, was not dependent on HCO3- and was not inhibited by application of the anion channel/transporter inhibitor H2DIDS to the luminal membrane. It is therefore attributed to a cAMP-stimulated Cl- conductance, probably the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel. [Cl-]i also decreased when the basolateral Cl- concentration was reduced. This effect was not stimulated by forskolin, was largely dependent on HCO3- and was inhibited by basolateral H2DIDS. It is therefore mediated mainly by Cl-/HCO3- exchange. With high Cl- and low HCO3- concentrations in the lumen, steady-state [Cl-]i was 25-35 mM in unstimulated cells. Stimulation with forskolin caused [Cl-]i to increase by approximately 4 mM due to activation of the luminal anion exchanger. With low Cl- and high HCO3- concentrations in the lumen to simulate physiological conditions, steady-state [Cl-]i was 10-15 mM in unstimulated cells. Upon stimulation with forskolin, [Cl-]i fell to approximately 7 mM due to increased Cl- efflux via the luminal conductance. We conclude that, during stimulation under physiological conditions, [Cl-]i decreases to very low levels in guinea-pig pancreatic duct cells, largely as a result of the limited capacity of the basolateral transporters for Cl- uptake. The resulting lack of competition from intracellular Cl- may therefore favour HCO3- secretion via anion conductances in the luminal membrane, possibly CFTR.

5-hydroxytryptamine strongly inhibits fluid secretion in guinea pig pancreatic duct cells

5-hydroxytryptamine strongly inhibits fluid secretion in guinea pig pancreatic duct cells

5-hydroxytryptamine strongly inhibits fluid secretion in guinea pig pancreatic duct cells.

We studied the distribution of 5-hydroxytryptamine- (5-HT-) containing cells in the guinea pig pancreas and examined the effects of 5-HT on fluid secretion by interlobular pancreatic ducts. The 5-HT-immunoreactive cells with morphological characteristics of enterochromaffin (EC) cells were scattered throughout the duct system and were enriched in islets of Langerhans. The fluid secretory rate in the isolated interlobular ducts was measured by videomicroscopy. Basolateral applications of 5-HT strongly but reversibly reduced HCO(3)-dependent, as well as secretin- and acetylcholine- (ACh-) stimulated, fluid secretion, whereas 5-HT applied into the lumen had no such effects. Secretin-stimulated fluid secretion could be inhibited by a 5-HT(3) receptor agonist, but not by agonists of the 5-HT(1), 5-HT(2), or 5-HT(4) receptors. Under the stimulation with secretin, 5-HT decreased the intracellular pH (pH(i)) and reduced the rate of pH(i) recovery after acid loading with NH(4)(+), suggesting that 5-HT inhibits the intracellular accumulation of HCO3(-). The elevation of intraductal pressure in vivo reduced secretin-stimulated fluid secretion, an effect that could be attenuated by a 5-HT(3) receptor antagonist. Thus, 5-HT, acting through basolateral 5-HT(3) receptors, strongly inhibits spontaneous, secretin-, and ACh-stimulated fluid secretion by guinea pig pancreatic ducts. 5-HT released from pancreatic ductal EC cells on elevation of the intraductal pressure may regulate fluid secretion of neighboring duct cells in a paracrine fashion.

Cystic fibrosis transmembrane conductance regulator currents in guinea pig pancreatic duct cells: Inhibition by bicarbonate ions

Background & Aims: Cystic fibrosis transmembrane conductance regulator (CFTR) Cl − channels play an important role in HCO 3 − secretion by pancreatic duct cells (PDCs). Our aims were to characterize the CFTR conductance of guinea pig PDCs and to establish whether CFTR is regulated by HCO 3 −. Methods: PDCs were isolated from small intralobular and interlobular ducts, and their Cl − conductance was studied using the whole-cell patch clamp technique. Results: Activation of a typical CFTR conductance by adenosine 3',5'-cyclic monophosphate (cAMP) was observed in 114 of 204 cells (56%). A larger (10-fold), time- and voltage-dependent Cl − conductance was activated in 39 of 204 cells (19%). Secretin had a similar effect. Coexpression of both conductances in the same cell was observed, and both conductances had similar anion selectivity and pharmacology. Extracellular HCO 3 − caused a dose-dependent inhibition of both currents ( K i, ̃7 mmol/L), which was independent of intracellular and extracellular pH, and the PCO 2 and CO 3 2− content of the bathing solutions. Conclusions Two kinetically distinct Cl − conductances are activated by cAMP in guinea pig PDCs. Because these conductances are coexpressed and exhibit similar characteristics (anion selectivity, pharmacology, and HCO 3 − inhibition), we conclude that CFTR underlies them both. The inhibition of CFTR by HCO 3 − has implications for the current model of pancreatic ductal HCO 3 − secretion. GASTROENTEROLOGY 2000;118:1187-1196

Anion interactions with CFTR and consequences for HCO3- transport in secretory epithelia.

We have been studying CFTR channels in guinea pig pancreatic duct cells and rather surprisingly found that luminal HCO3- had a pronounced inhibitory effect on cAMP-activated CFTR chloride currents. The block produced by HCO3- was rapid, voltage-independent and occurred over a physiological range of extracellular HCO3- concentrations. I- and ClO4- were also found to inhibit CFTR currents, but both were less effective than HCO3-. Although we have not identified how HCO3- is able to block CFTR our data suggests that an external anion-binding site on the channel itself is involved. Overall, our results show that luminal HCO3- acts as a potent inhibitor of CFTR channels (and by inference CFTR-mediated secretion), under normal physiological conditions. These data have implications not only for current models of pancreatic duct cell HCO3- transport, but also for other bicarbonate-secreting tissues, such as the liver, GI tract and lungs.

Arginine vasopressin inhibits fluid secretion in guinea pig pancreatic duct cells.

The effects of arginine vasopressin (AVP) on pancreatic ductal secretion were studied in guinea pigs. In the isolated vascularly perfused pancreas, AVP reduced secretin-stimulated fluid secretion and increased the vascular resistance when the perfusion rate was held constant. In the isolated interlobular duct segments, AVP inhibited secretin-stimulated fluid secretion, indicating the direct inhibitory action of AVP on the duct cells. AVP affected neither the basal nor the secretin-induced cAMP productions, suggesting that AVP inhibits the fluid secretion at a point distal to the production of cAMP. AVP increased intracellular Ca(2+) concentration ([Ca(2+)](i)) in the absence of extracellular Ca(2+). When [Ca(2+)](i) was elevated by the application of thapsigargin, AVP caused a rapid decrease in [Ca(2+)](i). AVP seems to activate both Ca(2+) release from intracellular stores and Ca(2+) efflux across the plasma membrane, but its relation to the inhibition of fluid secretion remains to be clarified. It is concluded that AVP directly inhibits secretin-stimulated ductal fluid secretion in the guinea pig pancreas.

Bicarbonate secretion in interlobular ducts from guinea‐pig pancreas.

1. The transport of HCO3- across the luminal membrane of pancreatic duct cells was studied by monitoring the luminal pH of isolated guinea-pig interlobular ducts after microinjection of an extracellular fluoroprobe, the dextran conjugate of 2'7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF-dextran). Luminal Cl- concentration was also measured by microfluorometry following microinjection of the dextran conjugates of 6-methoxy-N-(4-aminoalkyl)quinolinium bromide (ABQ-dextran) and Cl-NERF (Cl-NERF-dextran). 2. When HCO3-/CO2 was admitted to the bath, a transient acidification of the duct lumen was observed, followed by a marked alkalinization. The latter was abolished when the luminal Cl- concentration was reduced to 25-35 mM by replacement with glucuronate and may, therefore, be attributed to Cl(-)-HCO3- exchange at the luminal membrane. 3. Secretin, forskolin and acetylcholine stimulated HCO3- secretion into the lumen even when the luminal Cl- concentration was reduced to approximately 7 mM. Furthermore, agonist-evoked HCO3- secretion was not inhibited by luminal glibenclamide, dihydro-4,4'-diisothiocyanostilbene-2,2'-disulphonic acid (H2DIDS) or 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB). These observations are not easily reconciled with HCO3- transport across the luminal membrane being mediated by Cl(-)-HCO3- exchange in parallel with a Cl- conductance. 4. Agonist-stimulated HCO3- secretion was blocked by omitting Na+ from the bath but not by addition of N-methyl-N-isobutylamiloride (MIA) or bafilomycin A1. This supports our previous conclusion that HCO3- entry into duct cells from the extracellular fluid requires Na+ but is not dependent on Na(+)-H+ exchange or vacuolar-type H(+)-ATPase activity. 5. The three actions of secretin on guinea-pig pancreatic duct cells described in this and the accompanying paper - stimulation of a relatively Cl(-)-insensitive luminal HCO3- efflux pathway, stimulation of basolateral Na(+)-HCO3- cotransport, and lack of effect on intracellular pH- require the current model of pancreatic HCO3- secretion to be modified.

Accumulation of intracellular HCO3- by Na(+)-HCO3- cotransport in interlobular ducts from guinea-pig pancreas.

1. Short segments of interlobular duct were microdissected from guinea-pig pancreas following enzymatic digestion. After overnight culture, intracellular pH (pH1) and Na+ concentration ([Na+]i) were measured by microfluorometry in duct cells loaded with either the pH-sensitive fluoroprobe 2'7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) or the sodium-binding benzofuran isophthalate (SBFI). 2. The transporters responsible for maintaining pHi above equilibrium were investigated by using the NH4Cl pulse technique to acid load the cells. In the absence of HCO3-/CO2, the recovery of pH1 was Na+ dependent, abolished by 0.2 mM amiloride and by 10 microM N-methyl-N-isobutylamiloride and was therefore attributed to Na(+)-H+ exchange. 3. In the presence of HCO3-/CO2, amiloride only partially inhibited the recovery from acid loading. The amiloride-insensitive component was abolished by 0.5 mM H2DIDS and unaffected by depletion of intracellular Cl- and was therefore attributed to Na(+)-HCO3- cotransport. 4. Stimulation with 10 nM secretin did not cause a significant change in pH1 despite a significant increase in HCO3- efflux. However, in the presence of secretin, addition of 0.5 mM H2DIDS caused a decline in pH1 that was three times more rapid than that obtained with 0.2 mM amiloride. 5. In secretin-stimulated ducts, Na+ uptake increased when HCO3-/CO2 was added to the bath and this increase was strongly inhibited by 0.5 mM H2DIDS. 6. We conclude that Na(+)-HCO3- cotransport contributes approximately 75% of the HCO3- taken up by guinea-pig pancreatic duct cells during stimulation with secretin. It is proposed that electrical coupling between HCO3- efflux at the luminal membrane and electrogenic Na(+)-HCO3- cotransport at the basolateral membrane explains why secretin causes little change in pH1.

Insulin, transforming growth factors, and substrates modulate growth of guinea pig pancreatic duct cells in vitro

Background & Aims : Little is known of the physiological mechanisms that control cellular renewal in the pancreatic excretory duct system. This study investigated the effects of potential regulatory substances on the growth of cultured guinea pig pancreatic duct epithelial monolayers. Methods : Pancreatic duct explants were cultured for 3 days on plastic and on permeable filters in the presence and absence of different substances. Growth of epithelial monolayers from these explants was measured by 5-bromo-2′-deoxyuridine incorporation and morphometric procedures. Results : Epidermal growth factor and insulin both enhanced monolayer growth and together had an additive effect. Transforming growth factor α enhanced and transforming growth factor β inhibited growth, whereas glucagon, somatostatin, pancreatic polypeptide, secretin, cerulein, bombesin, and dexamethasone had no significant effects. Monolayer growth on type I collagen-coated filters was enhanced when compared with that of monolayers grown on tissue culture plastic. Cell growth from explants on filters coated with type IV collagen and fibronectin was comparable with that on plastic, whereas growth on Matrigel- or laminin-coated filters was reduced. Conclusions : Insulin, transforming growth factors, and substrate components modulate growth of pancreatic duct epithelial cells in vitro, suggesting that they are important regulators of cell division in the excretory duct system of the intact pancreas.

Regulation of pancreatic duct epithelial growth in vitro

Effects of a number of possible trophic factors on growth of guinea pig pancreatic duct epithelial monolayers were investigated. Isolated fragments of main and interlobular ducts were prepared and explanted onto both tissue culture plastic and thick gels of type I collagen. Monolayers growing out from explants were first cultured in a basal medium for 3 or 4 days. Next, the medium was supplemented individually with bombesin, carbachol, caerulein, epidermal growth factor (EGF), secretin, 12-O-tetradecanoylphorbol 13-acetate (TPA), or vasoactive intestinal peptide (VIP). Cells were cultured in the absence or presence of these possible trophic factors, and monolayer areas were determined morphometrically at 0, 2, and 4 days. Rate of growth was determined from increase in area over each 2-day period. Monolayers grown in basal medium alone on plastic increased to 479% of initial area over the 4-day test period; those grown on collagen increased to 523%. Explants cultured in presence of bombesin, carbachol, caerulein, secretin, TPA, and VIP on either substrate grew at rates not significantly different from those cultured in basal medium. By contrast, duct monolayers grown on plastic or collagen in presence of 10 nM EGF expanded in area to 722 and 1,070%, respectively, of their initial areas. The EC50 for this trophic effect was approximately 1 nM. These results show that EGF exerts a potent trophic effect on guinea pig pancreatic duct cells in vitro but also indicate that cell division in the pancreatic main and interlobular ducts is not regulated by caerulein and related peptide hormones that have been reported to have growth-promoting effects on exocrine pancreas in vivo.