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

Abstract

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.

Figure 1

Immunohistochemistry for 5-HT. The distribution of 5-HT–positive cells in the guinea pig pancreas was examined with mouse anti-serotonin mAb. The majority of 5-HT–immunoreactive cells are present in the islet of Langerhans (a). They are also present in the epithelium of the main (b), larger (c), and smaller (d) interlobular ducts and in acini (a). Bar, 5 μm.

Figure 2

Effects of 5-HT on basal fluid secretion in interlobular duct segments isolated from guinea pig pancreas. The closed ducts were initially superfused with HCO₃^–-CO₂-free HEPES-buffered solution. After a 5-minute period, the bath solution was switched to the standard HCO₃^–-CO₂–buffered solution. After a further 10 minutes, 0.1 μM 5-HT was added to the bath in the presence of HCO₃^–-CO₂. The fluid secretory rates are shown as means ± SEM of four experiments.

Figure 3

Effects of 5-HT on secretin-stimulated fluid secretion. (a) Ducts were stimulated with 1 nM secretin in the presence of HCO₃^–-CO₂. 5-HT (0.1 μM) was added to the bath during stimulation with 1 nM secretin. (b) Concentration-response curve for the effects of 5-HT on secretin-stimulated fluid secretion. The fluid secretory rates are shown as means ± SEM of four experiments. Asterisks indicate significant (P < 0.01) differences.

Figure 4

Effects of 5-HT on ACh-stimulated fluid secretion. Ducts were stimulated with 1 μM ACh in the presence of HCO₃^–-CO₂. 5-HT (0.1 μM) was added to the bath during stimulation with 1 μM ACh. The fluid secretory rates are shown as means ± SEM of four experiments.

Figure 5

Effects of 5-HT receptor agonists on secretin-stimulated fluid secretion. (a) One micrommolar of 2-methyl-5-HT (5-HT₃ receptor agonist) was added to the bath during stimulation with 1 nM secretin. (b) Effects of 1 μM of 5-HT, 5-CT (5-HT₁ receptor agonist), α-methyl-5-HT (5-HT₂ receptor agonist), 2-methyl-5-HT (5-HT₃ receptor agonist), and 5-MeOT (5-HT₄ receptor agonist) on secretin-stimulated fluid secretion. Filled bars, the secretory rate before the application of 5-HT agonists. Open bars, the secretory rate in the presence of 5-HT agonists. The fluid secretory rates are shown as mean ± SEM of four experiments. Asterisks indicate significant (P < 0.01) differences.

Figure 6

Effects of luminal 5-HT on fluid secretion. The lumen of the cultured ducts was micropunctured and the luminal fluid was replaced by the HEPES-buffered solution with or without 5-HT (1 μM). The ducts were initially superfused with HEPES-buffered solution for 5 minutes. The bath solution was switched to the HCO₃^–-CO₂-buffered solution. After a 6-minute period, 1 nM secretin was added to the bath. (a) The fluid secretory rate in ducts injected with 5-HT (1 μM). Mean ± SEM of four experiments. (b) Spontaneous (HCO₃^–-CO₂–dependent) and secretin-stimulated (1 nM) fluid secretion in the absence (filled bars) and presence (open bars) of luminal 5-HT (1 μM). Mean ± SEM of four experiments.

Figure 7

Effect of 5-HT on intracellular pH. Ducts were superfused with the standard HCO₃^–-buffered solution in the presence of secretin (1 nM). (a) One micrometer 5-HT was applied to the bath. Representative of six experiments. (b) Duct cells were acid loaded by exposure to 2-minute pulses of 20 mM NH₄⁺ in the absence and presence of 5-HT (1 μM). Representative of five experiments.

Figure 8

Effects of intraductal pressure on secretin-stimulated fluid secretion in anesthetized guinea pigs. Pancreatic fluid secretion was stimulated with an intravenous infusion of secretin (1 μg/kg/h). A hydrostatic pressure of +3.0 cmH₂O was applied to the duct system by elevating the position of the outlet of the needle connected to the pancreatic duct without or with an intravenous infusion of granisetron (40 μg/kg/h), a 5-HT₃ receptor antagonist. Mean ± SEM of four experiments in four guinea pigs.

Figure 9

A hypothetical function of 5-HT: a local feedback control of pancreatic duct fluid secretion by the luminal pressure. EC cell–like cells in the duct epithelium are pressure-responsive sensory receptors. The increase in the luminal pressure stimulates the release of 5-HT into interstitium. 5-HT binds to its receptors on the basolateral membrane of the duct cells and inhibits fluid secretion.