Endogenous prostaglandins and microflora modulate DNA synthesis and neuroendocrine peptides in the rat gastrointestinal tract.

BACKGROUND: Previous studies suggest that E2 prostaglandins and the microflora may participate in the regulation of endocrine cells and of gastrointestinal cell kinetics. Our aim is to examine the actions of endogenous prostaglandins and of the microflora on gastrointestinal cell proliferation and tissue levels of neuroendocrine peptides. METHODS: Germfree and ex-germfree rats were treated with subcutaneous placebo or 1.5 mg/kg indomethacin for 3 days. All rats were labeled with 3H-methyl-thymidine, and biopsy specimens from different parts of the gastrointestinal tract were processed for autoradiography. DNA synthesis was estimated by the labeling index, except in the oxyntic mucosa, where the total number of labeled cells present in 7.5 mm mucosa was used. The concentration of neuroendocrine peptides was determined by radioimmunoassay. RESULTS: In the germfree rat, indomethacin reduced DNA synthesis in the fundus, duodenum, and proximal jejunum (P < 0.05) and the number of villous cells throughout the small intestine (P < 0.05). Exposure to microflora increased DNA synthesis in the proximal and distal jejunum, ileum, and colon (P < 0.05 versus germfree controls) and the number of crypt cells in the distal small intestine and colon (P < 0.05) and reduced the number of villous cells in the small intestine (P < 0.05) but did not affect tissue concentrations of neuroendocrine peptides. Indomethacin increased the concentration of somatostatin in the stomach, duodenum, and colon of germfree rats (P < 0.001), the concentration of calcitonin gene-related peptide (CGRP) and enteroglucagon in the proximal and distal jejunum and ileum (P < 0.001), and the concentration of glucagon in the colon (P < 0.05). The concentrations of somatostatin, CGRP, and glucagon were lower in indomethacin-treated ex-germfree rats than in indomethacin-treated germfree rats (P < 0.01). CONCLUSIONS: Indomethacin selectively reduced DNA synthesis in the upper gastrointestinal tract of germfree rats, indicating a basal stimulatory role for endogenous prostaglandins on cell proliferation. Endogenous prostaglandins modulate synthesis or release of gastrointestinal neuroendocrine peptides. Somatostatin may mediate indomethacin-induced reduction of DNA synthesis. The microflora stimulates cell proliferation and influences tissue levels of neuroendocrine peptides in a manner opposite to that of indomethacin.

HCl-stimulated duodenal HCO3- secretion in conscious rat. Interactions among VIP, nicotinic receptor mechanisms, and prostaglandins.

Using an isolated loop of the proximal duodenum of conscious rats, the role of vasoactive intestinal peptide (VIP) in the duodenal HCO3- response to HCl was examined, especially interactions with participating cholinoceptor mechanisms and prostaglandins. A 5-min perfusion with 150 mmol/liter HCl increased luminal VIP during 3 hr, with a peak output during and immediately after the acid challenge. The HCl-stimulated output was unaffected by atropine and hexamethonium, but was augmented by indomethacin from 13.6 (9.5-17.8) to 39 (20-85) fmol/cm/min. The HCO3- secretion in response to graded doses of intravenous VIP (0.00625-6 nmol/kg/30 min) was dose-dependent to maximally 33.5 +/- 10.5 mumol/cm/hr. The HCO3- secretion during a single intravenous infusion of VIP (12 nmol/kg/hr), 13.9 +/- 4.2 mumol/cm/hr, was unchanged by atropine, reduced to 10.0 +/- 3.5 mumol/cm/hr by hexamethonium, and augmented to 18.9 +/- 4.7 mumol/cm/hr by indomethacin. Exogenous VIP did not change the basal luminal output of PGE2; neither did exogenous PGE2 nor indomethacin affect the basal luminal output of VIP. HCl-induced increases in luminal outputs of VIP, substance P, and neurokinin A (the two latter with unknown roles) were differentially affected by atropine, hexamethonium, and indomethacin, indicating that the acid challenge released the peptides through controlled mechanisms. In conclusion, in the duodenal HCO3- response to luminal HCl, VIP may have a stimulatory role, which partially depends on nicotinic, but not on muscarinic cholinoceptor mechanisms, and which is negatively modulated by prostaglandins.

Aspects on the role of tachykinins and vasoactive intestinal polypeptide in control of secretion, motility and blood flow in the gut.

Both intrinsic and extrinsic neurons of the gut respond to mechanical and chemical stimuli by the release of neurotransmitters. We summarize here some of our recent work on the role of vasoactive intestinal polypeptide (VIP), substance P (SP) and neurokinin A (NKA) in the secretory, motor and vascular effects of hydrochloric acid stimulation in the isolated rat duodenal loop and electrical nerve stimulation and mechanical stimulation of the cat colon. Isolated duodenal loops of conscious rats were perfused with isotonic saline, and challenged at hourly intervals with brief exposures to increasing concentrations of HCL. The concentrations of bicarbonate and prostaglandin E2 (PGE2) released from the duodenal mucosa were significantly augmented already by pH 5.0 whereas VIP was significantly augmented at pH 3.0 and the tachykinins SP and NKA at pH 2.0. Continuous electric stimulation of the pelvic nerve in cats at 4 Hz during 1 s with 10 s rest produced a marked release of NKA-LI and SP-LI from the colon to blood. Reflex activation of the pelvic nervae by mechanical stimulation of the anus or rectal distension produced a less pronounced release of NKA-LI and SP-LI from the colon to blood. There was a simultaneous colonic contraction and vasodilation during each nerve stimulation. Close intraarterial infusions of NKA, neurokinin B, SP, neuropeptide K (NPK), eledoisin and physalemin at doses of 0.1-100 pmol/min induced dose-dependent proximal and distal colonic contractions and vasodilation, NKA being the most potent. The effects of the tachykinins were reduced after tetrodotoxin and atropine, but unchanged after treatment with hexamethonium.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholinergic mechanisms involved in release and effect of prostaglandin E2 in HCl-stimulated duodenal HCO-3 secretion in the conscious rat.

Using a proximal duodenal loop in conscious rats, we investigated interactions between prostaglandin E2 and nicotinic and muscarinic receptor mechanisms previously found to be involved in the duodenal HCO3- response to HCl. In previous studies using the same model, a 5-min perfusion of the duodenal loop with 150 mmol l-1 HCl produced a marked and sustained HCO3- response. In the present study, the identical challenge produced a rapid 20-fold increase in the luminal output of prostaglandin E2 during acid exposure, followed by a sustained more than twofold elevation above the basal level during the 45 min monitored. The prostaglandin synthesis inhibitor indomethacin (4 mg kg-1 i.p.) suppressed the output of prostaglandin E2 during the HCl challenge from 131 +/- 84 to 15.4 +/- 10.0 pmol cm-1 h-1, and in the post-stimulatory period from 17.3 +/- 9.1 to 4.4 +/- 2.2 pmol cm-1 h-1. The nicotinic receptor antagonist hexamethonium (20 mg kg-1 i.v.) had no effect on the output of prostaglandin E2. The muscarinic receptor antagonist atropine (0.5 mg kg-1 s.c.) had no effect on the output of prostaglandin E2 during HCl challenge, but reduced the post-stimulatory output to 7.7 +/- 4.1 pmol cm-1 h-1. Perfusion of the duodenal loop with 0.1 mmol l-1 prostaglandin E2 produced a HCO3- response that was abolished by hexamethonium (20 mg kg-1 i.v.), but not affected by atropine (0.5 mg kg-1 s.c.).(ABSTRACT TRUNCATED AT 250 WORDS)

Role of VIP in local control of secretion.

Vasoactive intestinal peptide (VIP) is a potent stimulant of duodenal HCO3- secretion and may, like prostaglandins, have a stimulatory role in the local duodenal HCO3- response to luminal HCl. Using a proximal duodenal loop in conscious rats, we examined the local luminal release of HCO3-, VIP and prostaglandin (PG) E2 in response to increasing concentrations of HCl (0.01-150 mmol l-1), perfused for 5 min at 60-min intervals. HCO3- and PGE2 were detected in all basal saline perfusate effluents, and were increased in a concentration-dependent manner by all acid concentrations tested. VIP was increased in a concentration dependent manner from pH 3. Exogenous VIP did not affect the basal luminal output of PGE2, or vice versa. Inhibition of prostaglandin synthesis by indomethacin augmented the HCl-stimulated luminal release of VIP, as well as the HCO3- response to exogenous VIP. The results are in agreement with previous studies, demonstrating that PGE2 is an important regulator of the duodenal HCO3- response to HCl in the rat. In addition, prostaglandins may negatively modulate the release of VIP from local VIPergic neurones, as well as the HCO3- secretagogue effect of VIP. Released VIP may contribute to the HCO3- response at pH less than or equal to 3.

Cholinergic influence on duodenal bicarbonate response to hydrochloric acid perfusion in the conscious rat.

Present information suggests that endogenous prostaglandins, hormonal factors, and neuropeptides participate in the regulation of duodenal bicarbonate secretion in response to luminal hydrochloric acid. The purpose of this study was to examine the effects of cholinergic antagonists and agonists on basal and acid-stimulated bicarbonate secretion in the proximal duodenum of the conscious rat. The basal bicarbonate secretion, 9 mumol X cm-1 X h-1, increased to 21 +/- 3 mumol X cm-1 X h-1 after 5 min of exposure to 150 mM HCl and remained significantly elevated for more than 3.5 h. The muscarinic blocker atropine, 0.5 mg X kg-1, reduced the acid-stimulated bicarbonate response by a third, and higher doses did not increase the inhibition. The ganglionic blocker hexamethonium, 10-20 mg X kg-1, suppressed in a dose-related manner the alkaline response by maximally one half. The opioid inhibitor naloxone was also an effective inhibitor. Hexamethonium, but not atropine, inhibited the basal duodenal bicarbonate secretion, which was unaffected by graded doses of the cholinergic agonists bethanechol and carbachol. We conclude that cholinergic nicotinic activity participates in maintaining basal duodenal bicarbonate secretion in the conscious rat. The bicarbonate response to luminal 150 mM HCl is partially dependent on cholinergic sites, but activation of such sites is not a single final step, since cholinergic agonists failed to elevate bicarbonate secretion from basal.

Stimulation of duodenal bicarbonate secretion by misoprostol.

Graded doses of topical misoprostol were examined for effects on duodenal bicarbonate output in fasted conscious rats and compared with perfusions of the H2-receptor blocker cimetidine. Chronic loops (2 cm) of the proximal duodenum were prepared in male Sprague-Dawley rats weighing 250 g. A thin perfusion tube was placed in the duodenal bulb, and a permanent cannula drained the distal end to the exterior. The stomach and remaining duodenum with biliary and pancreatic ducts were then reanastomosed. During tests, the loops were perfused with isotonic saline at 0.5 ml/min and samples of 15-min effluents analyzed for HCO3- using a modified back-titration method. Drugs and stimulants were added to the perfusion fluid. Misoprostol at 10(-10) M significantly (P less than 0.02) elevated the duodenal HCO3- from basal value (N = 10), and concentrations of 10(-10) to 4 X 10(-4) M produced dose-dependent increases of bicarbonate secretion, with a peak output of 49 +/- 7 mumol/cm/hr. A peak response of the same magnitude, 42 +/- 4 mumol/cm/hr, is observed after 5-min exposure of the loops to 150 mm HCl. The response to a single dose of 10(-4) M misoprostol was sustained, similar to the prolonged stimulatory effects of 5-min acid exposure. Perfusions of the loop with graded natural PGE2 result in similar increases of duodenal HCO3-, but the peak may be lower and the response to a single dose is less sustained. The H2-receptor blocker cimetidine, perfused at concentrations of 10(-7) to 10(-4) M failed to affect duodenal HCO3- output.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of graded doses of intraluminal H+, prostaglandin E2, and inhibition of endogenous prostaglandin synthesis on proximal duodenal bicarbonate secretion in unanesthetized rat.

The purpose of this study was to examine the effect of graded doses of intraluminal HCl and prostaglandin E2 on proximal duodenal bicarbonate secretion in unanesthetized rats. Basal bicarbonate secretion was approximately 14 mumol/cm . h and increased significantly in a dose-related manner to 50, 100, and 150 mM HCl as follows: 22.0 +/- 1.5, 29.2 +/- 4.0, and 42.0 +/- 4.3 mumol/cm . h, respectively (mean +/- SE). After a single 5-min infusion of 150 mM HCl, duodenal bicarbonate secretion remained significantly above basal for almost 3 h. The responses to 100 mM HCl and 10(-4) M prostaglandin E2 were similar (29.2 +/- 4.0 and 26.9 +/- 4.4 mumol/cm . h, respectively). Indomethacin (4 mg/kg, intraperitoneally) suppressed the bicarbonate response to 150 mM HCl; whereas the response to 10(-4) M prostaglandin E2 was unaltered. These studies in the in vivo rat indicate that (a) graded doses of intraluminal HCl produced graded increases in proximal duodenal bicarbonate secretion; (b) the bicarbonate output in response to a 5-min infusion of HCl persisted for almost 3 h; and (c) the bicarbonate response to HCl was essentially abolished by indomethacin pretreatment, suggesting that endogenous prostaglandins are at least in part involved in acid-induced proximal duodenal bicarbonate secretion.

Protection of the gastroduodenal mucosa by prostaglandins.

Like other nucleated cell populations in the body, the cells of the gastroduodenal mucosa are capable to metabolise arachidonic acid into prostaglandins, with prostaglandin E2 as the probable major metabolite. The production increases on demand and can be followed in the gastric lumen, where the output of prostaglandin E2 increases two to fourfold after exposure of the mucosa to hydrochloric acid. Exogenous prostaglandins, in particular of the E series, stimulate several identified mucosal defense factors in the upper gastrointestinal tract. Prostaglandins of the E series stimulate the transport of bicarbonate and the production and release of mucus glycoproteins from the gastroduodenal mucosa. They have trophic effects on gastrointestinal epithelia by increasing the survival time of mucosal cells and have cytoprotective properties. In addition, E2 prostaglandins suppress the gastric acid secretion and accelerate peptic ulcer healing. Non steroidal antiinflammatory drugs, which block the biosynthesis of prostaglandins, suppress the bicarbonate secretion, the production of mucus glycoproteins and cytoprotective properties. They interfere with the inhibitory feedback regulation of the gastric acid secretion and are ulcerogenic in experimental and clinical situations. These actions of PG biosynthesis blockers provide indirect information on the importance of local prostaglandin formation for maintenance of gastrointestinal mucosal integrity. It is hypothesised that biosynthesis of prostaglandins in the gastroduodenal mucosa is of importance and may be a key event in triggering the different components of the mucosal defense.

Secretin, VIP, and PHI stimulate rat proximal duodenal surface epithelial bicarbonate secretion in vivo.

The surface epithelial cells of the stomach and duodenum secrete bicarbonate at rest and in response to a number of agonists including the gastrointestinal hormones, glucagon, and GIP. Since those hormones with structural homology may have similar effects, the purpose of the present study was to examine the effect of graded doses (6, 24, and 96 nmol/kg) of pure porcine secretin, VIP, and PHI on bicarbonate secretion by the proximal duodenum containing Brunner's glands. Experiments were performed in vivo on unanesthetized Sprague-Dawley rats with chronic Thiry-Vella type loops of the proximal 2 cm of duodenum. The order of testing was random and only one hormone was tested on a single day. Compared to the saline control, each dose of VIP produced a significant increase in duodenal bicarbonate secretion in a dose-response manner. The two higher doses of secretin and only the 96 nmol/kg dose of PHI significantly increased bicarbonate output. The responses to 96 nmol/kg dose of secretin and VIP were similar, and each was significantly greater than observed with PHI. It is concluded that secretin and VIP stimulate proximal duodenal bicarbonate secretion and are more potent than PHI.