We examined, by using a specific PGE receptor subtype EP4 agonist and antagonist, the involvement of EP4 receptors in duodenal HCO 3 Ϫ secretion induced by PGE2 and mucosal acidification in rats. Mucosal acidification was achieved by exposing a duodenal loop to 10 mM HCl for 10 min, and various EP agonists were given intravenously 10 min before the acidification. Secretion of HCO 3 Ϫ was dose-dependently stimulated by AE1-329 (EP4 agonist), the maximal response being equivalent to that induced by sulprostone (EP1/EP3 agonist) or PGE2. The stimulatory action of AE1-329 and PGE2 but not sulprostone was attenuated by AE3-208, a specific EP4 antagonist. This antagonist also significantly mitigated the acidinduced HCO 3 Ϫ secretion. Coadministration of sulprostone and AE1-329 caused a greater secretory response than either agent alone. IBMX potentiated the stimulatory action of both sulprostone and AE1-329, whereas verapamil mitigated the effect of sulprostone but not AE1-329. Chemical ablation of capsaicin-sensitive afferent neurons did not affect the response to any of the EP agonists used. We conclude that EP4 receptors are involved in the duodenal HCO 3 Ϫ response induced by PGE2 or acidification in addition to EP3 receptors. The process by which HCO 3 Ϫ is secreted through these receptors differs regarding secondmessenger coupling. Stimulation through EP4 receptors is mediated by cAMP, whereas that through EP3 receptors is regulated by both cAMP and Ca 2ϩ ; yet there is cooperation between the actions mediated by these two receptors. The neuronal reflex pathway is not involved in stimulatory actions of these prostanoids. prostaglandin E2; EP receptor subtype; EP4 receptor SECRETION OF HCO 3 Ϫ FROM surface epithelial cells is one of the mucosal defensive mechanisms and plays an important role in protecting the duodenal mucosa against acid injury (6,4,18). Although the physiological regulation of this secretion involves several factors such as PGs, peptides, and neuronal factors (4,10,19,20), endogenous PGs are particularly important in the local control of the process. We (23) previously investigated the relationship between PGE receptor (EP) subtypes and HCO 3 Ϫ secretion, using specific EP1, EP2, and EP3 agonists, and found that the secretion was stimulated by EP agonists having a potent affinity for EP3 receptors. Using EP3 receptor knockout mice, we (21) also demonstrated the involvement of EP3 receptors in the acid-induced secretion of HCO 3 Ϫ in the duodenum. At that time, however, the possibility remained that the stimulatory action of PGE 2 is mediated by EP4 receptors, because specific EP4 agonists and antagonists were not available. It is now known that the increase in duodenal HCO 3 Ϫ secretion is mediated by the stimulation of adenylate cyclase (AC) activity and an elevation in intracellular cAMP levels (8,16,20) and that the activation of EP4 receptors results in an increase of intracellular cAMP via G s protein (3). Thus it is possible that EP4 receptors play a role in the regulation of duodenal HCO...

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Background/Aim: We investigated the role of prostacyclin (PGI₂) IP receptors in the acid-induced secretion of HCO₃^– using IP receptor knockout [IP (–/–)] mice, in comparison with capsaicin-induced secretion. Methods: Male C57/BL6 mice, both wild-type [WT] and [IP (–/–)], fasted for 18 h were used. Under urethane anesthesia, a proximal duodenal loop was perfused with saline, and the secretion of HCO₃^– was measured at pH 7.0 using a pH-stat method and by adding 2 mM HCl. The secretion was stimulated by exposure of the loop to 10 mM HCl, capsaicin, PGE₂ or cicaprost (a PGI₂ agonist) for 10 min. Results: PGE₂ stimulated HCO₃^– secretion in both WT and IP (–/–) mice, while cicaprost increased it in WT but not IP (–/–) mice. Luminal acidification increased the mucosal level of PGE₂ as well as 6-keto-PGF_1α, yet stimulated HCO₃^–secretion in both WT and IP (–/–) mice, in an indomethacin-inhibitable and sensory neuron-dependent manners. Perfusion of the duodenum with 20 mM HCl for 4 h caused severe damage in WT mice pretreated with indomethacin, but not in control WT or IP (–/–) mice. Capsaicin increased duodenal HCO₃^–secretion in WT mice, in an indomethacin-sensitive manner, yet no such response was observed in the animals lacking IP receptors. Conclusion: The presence of IP receptors is not essential for acid-induced HCO₃^– secretion and mucosal defense against acid injury in the duodenum, although activation of IP receptors results in stimulation of HCO₃^–secretion. Although duodenal HCO₃^– secretion induced by both acid and capsaicin depends on afferent neurons, it seems that the mode of interaction with the afferent neurons differs regarding dependency on the PGI₂/IP receptors.

We compared the HCO3(-) secretory response to capsaicin and mucosal acidification in rat duodenums, especially the relation to vanilloid receptor type 1 (VR1). A proximal duodenal loop was perfused with saline, and the HCO3(-) secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. The secretion was stimulated by exposing the loop to capsaicin (0.03-0.3 mg/ml) or 10 mM HCl for 10 min. Indomethacin subcutaneously or ruthenium red intravenously, a nonspecific VR1 antagonist, was given 60 or 10 min, respectively, before exposure to capsaicin or acid, while L-NAME was given intravenously 3 hr before these treatments. Capsazepine, another VR1 antagonist, was coapplied to the loop for 10 min with capsaicin or acid. Luminal application of capsaicin increased the secretion of HCO3(-) in a dose-dependent manner; this effect was markedly attenuated by chemical ablation of capsaicin-sensitive afferent neurons (CSN) as well as pretreatment with ruthenium red or capsazepine, and significantly mitigated by indomethacin or L-NAME (in an L-arginine-sensitive manner). The HCO3(-) secretion was also stimulated by mucosal acidification, and this response was attenuated by both capsaicin pretreatment, indomethacin and L-NAME, but not ruthenium red or capsazepine. Mucosal application of capsaicin as well as acid increased the mucosal PGE2 content, and these effects were both significantly attenuated by indomethacin and L-NAME. These results suggest that both capsaicin and acid cause the CSN-dependent increase in duodenal HCO3(-) secretion mediated by NO and PG, yet the mode of their action differs in terms of the ruthenium red or capsazepine sensitivity. Although luminal H+ plays a modulatory role in duodenal HCO3(-) secretion, it is unlikely that the action results from the interaction of H+ with the ruthenium red- or capsazepine-sensitive site of VR1.

Lafutidine is a new type of antiulcer drug, possessing both an antisecretory effect, exerted via a blockade of the histamine H2 receptor, and gastroprotective activity, mediated by capsaicin-sensitive afferent nerves (CSN). In the present study, we examined the effect of lafutidine on gastric mucosal blood flow (GMBF) and duodenal HCO3- secretion (DAS) under basal and acid-stimulated conditions in rats. Under urethane anesthesia, GMBF was measured using a laser Doppler flowmeter in a chambered stomach before and after exposure to 20 mM taurocholate (TC) plus 50 mM HCl, while DAS was measured in a proximal duodenal loop before and after mucosal acidification (10 mM HCl for 10 min) by titrating the perfusate at pH 7.0 using a pH-stat method and by adding 10 mM HCl. Lafutidine given intraperitoneally affected neither GMBF nor DAS under basal conditions, but augmented an increase in both GMBF and DAS induced by mucosal acidification. Although the acid-induced GMBF and DAS responses were significantly mitigated by both indomethacin and sensory deafferentation but not by ruthenium red (RT), the vanilloid receptor (VR)-1 antagonist, the responses were preserved in lafutidine-treated animals, even in the presence of indomethacin. Both GMBF and DAS were significantly increased by local application of capsaicin, the responses being attenuated by indomethacin and RT as well as sensory deafferentation. Lafutidine augmented the GMBF and DAS responses to capsaicin and preserved the responses, even in the presence of indomethacin. Capsaicin evoked an increase in [Ca2+]i in rat VR1-transfected HEK293 cells, while lafutidine had no effect by itself on [Ca2+]i in these cells and did not affect the increase in [Ca2+]i evoked by capsaicin. In conclusion, these results suggest that lafutidine mimics endogenous effects of prostaglandins to augment the GMBF and DAS responses to acid or capsaicin, probably by sensitizing CSN through an unknown site other than VR1. The luminal H+ itself is not a ligand for the RT-sensitive site of VR1 but plays a modulator role in the CSN-mediated physiological responses.