Christophe Dupont

Cyclic AMP production in isolated colonic epithelial crypts: a highly sensitive model for the evaluation of vasoactive intestinal peptide action in human intestine

Abstract. Epithelial cells of human colon were isolated in the form of entire epithelial crypts devoid of vasoactive intestinal peptide (VIP) as measured by radioimmunoassay. This preparation allowed the demonstration, in any conditions tested, of the considerable sensitivity of cyclic AMP accumulation in colonic epithelium to VIP. The magnitude of cyclic AMP accumulation in this preparation was dependent on temperature, time of incubation and concentration of phosphodiesterase inhibitor. At 37 C, in the presence 0–2 mM 3‐isobutyl‐l‐methylxanthine, a 4‐fold increase of cyclic AMP above basal was observed 5 min after addition of 10‐10 M VIP and a 16‐fold increase (maximal response) was obtained after 20 min by 10‐8 M VIP. The dose‐response curve was studied in conditions of equilibrium, i.e. a 60 min incubation at 15CC in the presence of 0–2 mM 3‐isobutyl‐l‐methylxanthine, and appeared to be monophasic with a half‐maximal stimulation at l‐2 times 10‐9 M VIP. Synthetic secretin, porcine intestinal histidine isoleucine amide (PIHI) and the other active substances (prostaglandins E and isoproterenol) were 2500, 600, and at least 100 times less potent than VIP, respectively. Adenylate cyclase of membranes prepared from epithelial crypts was stimulated by concentrations of VIP identical to those acting on cyclic AMP level in intact crypts. Plasmas from patients with the watery diarrhoea syndrome were able to induce a significant rise of cyclic AMP production in human colonic epithelial crypts. This effect was suppressed by previous exposure of the plasmas to a specific anti‐VIP immunserum.

Characterization of a vasoactive intestinal peptide-sensitive adenylate cyclase in rat intestinal epithelial cell membranes.

A vasoactive intestinal peptide-sensitive adenylate cyclase in intestinal epithelial cell membranes was characterized. Stimulation of adenylate cyclase activity was a function of vasoactive intestinal peptide concentration over a range of 1 . 10(-10)-1 . 10(-7) M and was increased six-times by a maximally stimulating concentration of vasoactive intestinal peptide. Half-maximal stimulation was observed with 4.1 +/- 0.7 nM vasoactive intestinal peptide. Fluoride ion stimulated adenylate cyclase activity to a higher extent than did vasoactive intestinal peptide. Under standard assay conditions, basal, vasoactive intestinal peptide- and fluoride-stimulated adenylate cyclase activities were proportional to time of incubation up to 15 min and to membrane concentration up to 60 microgram protein per assay. The vasoactive intestinal peptide-sensitive enzyme required 5-10 mM Mg2+ and was inhibited by 1 . 10(-5) M Ca2+. At sufficiently high concentrations, both ATP (3 mM) and Mg2+ (40 mM) inhibited the enzyme. Secretin also stimulated the adenylate cyclase activity from intestinal epithelial cell membranes but its effectiveness was 1/1000 that of vasoactive intestinal peptide. Prostaglandins E1 and E2 at 1 . 10(-5) M induced a two-fold increase of cyclic AMP production. Vasoactive intestinal peptide was the most potent stimulator of adenylate cyclase activity, suggesting an important physiological role of this peptide in the cyclic AMP-dependent regulation of the intestinal epithelial cell function.

Selective inhibition by somatostatin of cyclic AMP production in rat gastric glands: Demonstration of a direct effect on the parietal cell function

Somatostatin, originally isolated from the hypothalamus as an inhibitor of growth hormone secretion [I], is released from somatostatin~ontaining D cells of the stomach [2] and pancreas [2,3] during the gastric phase of a meal [4]. In vivo, somatostatin is reported to inhibit the pepsinogen secretion induced by a meal [S] and the gastric acid secretion stimulated by histamine [6,7]. The presence of specific receptors for somatostatin on isolated gastric cells from the rat fundus [8] strongly suggests that somatostatin exerts a direct effect on these exocrine secretory processes: We have shown that secretin which stimulates pepsinogen [9] and mucous [lo] secretions is highly potent in stimulating cyclic AMP production in gastric glands isolated from the rat fundus and antrum, while histamine is only effective on the acid-secreting fundic preparation [ 1 l]. Here we have examined therefore the ability of somatostat~ to inhibit both secretin and histamine-induced cyclic AMP formation in our system. The effect of cimetidine, a specific Hz-receptor antagonist [ 121 was also investigated since it has been shown that histamine stimulates gastric acid secretion by interacting with an Hz-receptor cyclic AMP system [ 131 present in parietal cells ]14]. Our data demonstrate that somatostatin (1 O-‘low6 M) reduces considerably the effect of histamine on the cyclic AMP production in a non-competitive way, while cimetidine (1 O-*-l O4 M) inhibits competitively the histamine stimulation. In contrast, no effect of somatostatin was found on the cyclic AMP

Evidence for a cyclic AMP system highly sensitive to secretin in gastric glands isolated from the rat fundus and antrum

The effects of secretin and vasointestinal peptide (VIP) on the production of cyclic AMP have been studied in gastric glands isolated by means of EDTA from rat fundic and antral mucosa. (1) In gastric fundus, secretin and VIP caused a time- and temperature-dependent stimulation of cyclic AMP production that was maximal when the test agents were incubated for 60 min at 20 degrees C in the presence of 0.5 mM 3-isobutyl-1-methylxanthine as a phosphodiesterase inhibitor. The dose-response curve was monophasic for both peptides, the production of cyclic AMP being sensitive to 10(-10) M secretin and to 5 . 10(-8) M VIP. Half-maximal stimulation was obtained with 2.9 10(-9) M secretin or 2 . 10(-7) M VIP and the maximal stimulation represented a 21-fold and a 19-fold increase above control for secretin and VIP, respectively. Histamine also stimulated cyclic AMP production, with a Km of about 5 . 10(-4) M. No additive effect on cyclic AMP production was oberved when secretin and VIP were simultaneously added at maximally active concentrations, while an additive effect was observed when secretin and histamine were added together. (2) In gastric antrum, the characteristics of the secretin- and VIP-stimulated cyclic AMP production were similar to those observed in gastric fundus. Histamine nevertheless failed to stimulate the formation of cyclic AMP in antral mucosa. (3) These data demonstrate the existence of a cyclic AMP system highly sensitive to secretin in gastric glands isolated from the rat fundus and antrum and suggest that VIP operates through this system. (4) It is proposed that the pepsinogen- and/or mucous-secreting cells are implicated in the regulation of cyclic AMP production by secretin in gastric glands of the rat.

Regulation by vasoactive intestinal peptide of cyclic amp accumulation in gastric epithelial glands: A characteristic of human stomach

Cyclic AMP production in gastric epithelium is regulated by numerous substances. In the different species studied, histamine stimulated the production of cyclic AMP only in the fundic part of gastric epithelium ( [ 1,2], C.G. et al., in preparation). This effect was shown to be related to the regulation by histamine of the acid secretion by parietal cells [3,4]. In the other cells of the stomach, namely the pepsinogen and mucous-secreting cells, which occur both in fundus and in antrum [5], secretin was described as the sole peptide stimulating cyclic AMP production at low physiological doses ([4], C.G. et al., in preparation). When a stimulating effect of vasoactive intestinal peptide (VIP) was found, concentrations of VIP 200-times higher were needed to exert the same effect as secretin (C.G. et al., in preparation), strongly suggesting that VIP, a natural analog of secretin was a positive agonist of this peptide. This study was carried out in humans, using an original preparation of pure gastric epithelial glands. We demonstrate that in the human, the peptide stimulating at physiological doses the cyclic AMP system of gastric epithelium is not secretin but VIP, that acted at as low as lo-” M both in fundus and in antrum epithelial glands.

Action of histamine and vasoactive intestinal peptide (VIP) on cyclic AMP in gastric glands isolated from human fetal stomach.

Histamine and VIP produce an elevation of cAMP production in gastric glands isolated from the human fetal stomach at 15 weeks of gestation. These effects were attributed to the activation of 2 distinct receptor-cAMP systems, one being sensitive to histamine in parietal cells, and the other being sensitive to VIP in muco-peptic cell populations. The results suggest that histamine and VIP may play a role in inducing gastric secretion during fetal life in man.

Beta‐adrenergic regulation of cyclic adenosine 3‘,5’ monophosphate accumulation in human gastric epithelial glands. Inhibitory effect of somatostatin

Abstract. The action of catecholamines and somatostatin on cyclic adenosine 3′,5′ monophosphate (cyclic AMP) formation in human isolated gastric glands is reported.

Pancreatic lipomatosis and duodenal stenosis or atresia in children

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Vasoactive Intestinal Peptide (VIP) Control of Cyclic amp Accumulation in Digestive Epithelia: Involvement of Multiple Regulatory Processes

Abstract VIP is a 28-residue peptide structurally related to secretin, glucagon (Said and Mutt, 1972) and PHI (Tatemoto and Mutt, 1980) (Table I). It is a neuropeptide widely distributed in the peripheral nervous system of the digestive tract (reviewed in Said, 1980). Besides its relaxant effect on the smooth musculature, the most important known action of VIP in the digestive tract is to alter the transport of water, electrolytes and/or proteins in the epithelia: small and large intestine, gallbladder, stomach and exocrine pancreas (reviewed in Said, 1980. Laburthe and Dupont, 1981). All these tissues, as well as liver, exhibit specific binding sites for VIP (see Table II) that are, in most cases, linked to the stimulation of adenylate cyclase in plasma membranes and/or cyclic AMP accumulation in intact cells (reviewed in Laburthe and co-workers, 1977; Amiranoff and Rosselin, 1981). However, discrepancies occur in the patterns of VIP-induced cyclic AMP. They are related to at least three regulatory components or processes (see Fig.l): 1) The recognition site or receptor. 2) The coupling between receptors and adenylate cyclase which involves the guanine nucleotide regulatory component. 3) The phosphodiesterase activity within the cell.