Immunoreactive Secretin Research Articles

Role of secretin in negative feedback regulation of postprandial pancreatic secretion in dogs

Background: A negative feedback regulation of pancreatic exocrine secretion has been observed in fasting rats, pigs, and humans, but this phenomenon could not be found in fasting dogs. The aims of the present study were to investigate whether or not postprandial pancreatic secretion is regulated by a negative feedback mechanism and to determine if the mechanism is mediated by secretion and/or cholecystokinin (CCK) in dogs. Methods: In eight dogs prepared with gastric and Herrera's pancreatic cannulas, pancreatic juice was collected for 3 hours after feeding a mixed meal to measure volume, bicarbonate, and trypsin output. Peripheral venous blood was obtained to determine plasma immunoreactive secretin and CCK levels. Four groups of experiments were performed while pancreatic juice was diverted from the duodenum: (1) diversion of pancreatic juice alone, (2) intraduodenal infusion of fresh pancreatic juice while pancreatic juice was diverted, (3) intraduodenal infusion of 150 mg/h of trypsin and 300 mg/h of chymotrypsin, and (4) intraduodenal infusion of 0.1 mol/L NaHCO3. Results: Pancreatic secretion during diversion of pancreatic juice was significantly greater than that obtained while pancreatic juice was returned. Diversion-induced pancreatic hypersecretion was significantly suppressed by intraduodenal administration of pancreatic proteases, but it was not influenced significantly by 0.1 mol/L NaHCO3. The suppression by either pancreatic juice or proteases paralleled the decrease in plasma secretin response, whereas the CCK response remained unchanged. The inhibitory effect of pancreatic proteases on pancreatic secretion was blocked by a physiological dose of exogenous secretin, 0.06 clinical U · kg−1 · h−1. Conclusions: In dogs, postprandial pancreatic secretion is controlled by a negative feedback mechanism mediated mainly by the release of secretin, but not by CCK.

Quantitation of immunoreactive secretin (IS) in canine neonatal gut and breast milk

Quantitation of immunoreactive secretin (IS) in canine neonatal gut and breast milk

Cerulein-induced acute pancreatitis in the rat. Study of pancreatic secretion and plasma VIP and secretin levels.

A study was made with different doses of cerulein (2, 4, 10 and 20 micrograms/kg) administered subcutaneously to rats by four injections at intervals of 1 hr; the aim of this work was to study exocrine pancreatic secretion of the rat under cerulein-induced acute pancreatitis, analyzing enzyme and hydroelectrolyte secretion of pancreatic juice. A further aim was to study the relationship between the dose of cerulein and the plasma levels of peptides controlling hydroelectrolyte secretion of the pancreas, like secretin and vasoactive intestinal peptide (VIP). At the lowest dose schedule, the amounts of total protein and enzymes (amylase and trypsin) in pancreatic juice decreased significantly, plasma amylase increased, and the pancreas became edematous. Higher doses magnified these effects. By contrast, ductular function (flow and HCO3-) was well preserved in cerulein-treated rats, and this was probably due to the significant increase in plasma levels of immunoreactive secretin whereas VIP levels were unchanged. The secretin released by treatment with cerulein is able to palliate the lack of flow from acinar origin that is affected in the process of acute pancreatitis, being a beneficial response to the cerulein treatment.

Route of Access of Secretin into Rabbit Duodenal Lumen During Acid Perfusion

The route of entry of immunoreactive secretin into the duodenal lumen during acid perfusion is unknown. Possible sites include paracellular diffusion from the extracellular space, transapical secretion from S cells, or leaching from cells damaged by the perfusate. Antisecretin in the extracellular space and interstitium should reduce access due to paracellular diffusion from the interstitium without significantly affecting secretion or leaching. We therefore studied the effect of intravenous antisecretin antibody on luminal secretin output. Two groups of rabbits received either intravenous normal rabbit serum (controls) or antisecretin antibody, before perfusing a closed segment of duodenum with hydrochloric acid. Preliminary experiments established that duodenal perfusion with HCl concentrations below 0.025N produced no apparent mucosal damage on electron microscopy. HCl concentrations above this significantly damaged the mucosa. In the control group, perfusion with 0.01N, 0.0125N, and 0.025N hydrochloric acid resulted in a dose-dependent increase in secretin in the perfusate. Secretin output was markedly reduced in the group injected with secretin antibody. Antisecretin antibody significantly reduced bile flow at all levels of HCl concentration in the duodenal perfusate. These data suggest that luminal secretin is probably derived from the interstitium and travels to the lumen across narrow paracellular channels.

Effects of MCI-727, a New Anti-Ulcer Agent, on Plasma Secretin Concentration in Rats and Dogs and Pancreatic Exocrine Secretion in Rats.

In the present study, we investigated the effects of MCI-727, a new anti-ulcer agent, on plasma immunoreactive secretin concentration in rats and dogs using secretin specific RIA with the ethanol extraction method. Plasma secretin levels were increased dose-dependently 10 min after oral administration of MCI-727 in rats (control: 5.5 +/- 0.6; MCI-727, 10 mg/kg: 10.4 +/- 2.6; 30 mg/kg: 15.3 +/- 1.5; 100 mg/kg: 20.7 +/- 2.6 pg/ml, n = 6). Teprenone also caused a significant increase of plasma secretin at 10 min after oral administration at the doses of 30, 100 and 300 mg/kg. Under the same conditions, MCI-727 and teprenone did not alter the plasma immunoreactive gastrin concentration in rats. From the results of the time course study, the increasing effect of MCI-727 (30 mg/kg, p.o.) on plasma secretin remained for at least 240 min after administration. On the other hand, the increasing effect of teprenone (200 mg/kg, p.o.) was only observed at 30 and 60 min after administration. Furthermore, MCI-727 had increasing effects on plasma secretin concentration in dogs, but teprenone had no effects in this species. The volume of pancreatic secretion and the pancreatic bicarbonate output increased after intra-duodenal administration of MCI-727 at 30 and 100 mg/kg in rats. Similar effects were also observed with teprenone (30-300 mg/kg, i.d.) or secretin (Secrepan, 0.1-1.0 unit/kg, i.v.).

Fasting prevents acute pancreatitis induced by cerulein in rats

We examined the effect of fasting on the course of experimental acute pancreatitis induced in rats by four subcutaneous injections of 20 micrograms/kg body weight of cerulein at hourly intervals. Rats were either fasted from 24 hr before to 9 hr after the first cerulein injection or fed ad libitum throughout the experiment. Twenty-four hours of fasting reduced cerulein-induced increases in serum levels of amylase and anionic trypsin(ogen) to 50 and 70% of those in fed rats, respectively. Increases in pancreatic wet weight after cerulein injections were also less in fasted rats than in fed rats. Pancreatic content of trypsin was significantly decreased after a 24-hr fast, and no further changes were induced by cerulein injections. The histological signs of acute pancreatitis were greatly alleviated by fasting. However, 24 hr of fasting did not alter the sensitivity and responsiveness of the exocrine pancreas to cerulein in both in vivo and in vitro. Plasma CCK bioactivity and immunoreactive secretin concentration in 24-hr-fasted rats were significantly lower than those in fed rats. Administration of CCK receptor antagonist, loxiglumide, 12 hr prior to the induction of acute pancreatitis reduced the increase in serum amylase activity in fed rats to nearly the same levels as that in fasted rats and alleviated histological signs of pancreatitis to some extent. These present observations suggest that fasting lessens the severity of cerulein-induced acute pancreatitis by reducing endogenous CCK release.

To extract or not to extract in secretin radioimmunoassay?

The importance of an ethanol extraction procedure in the radioimmunoassay of plasma secretin was investigated. The extraction step led to a higher assay sensitivity of 0.35 fmol/ml, compared to 5.93 fmol/ml using unextracted samples. The rise of plasma secretin after infusion of a low dose of secretin (1 pmol.kg-1.h-1) in 10 healthy humans was only detected after sample extraction. Higher doses (3 and 9 pmol.kg-1.h-1) resulted in increments of plasma IRS (immunoreactive secretin), which could be recorded both with and without sample extraction. After a steak meal 7 of 10 subjects showed a significant increase of plasma secretin assaying extracted plasma samples. The secretin release occurred in spikes. The mean increase of plasma IRS in this group was 0.6 fmol/ml, the mean maximal secretin release above basal was 2.3 fmol/ml. Without sample extraction, plasma secretin was not significantly changed. We conclude that plasma samples should be extracted in order to detect physiological postprandial secretin release.

Cationic environment and secretin secretion in canine duodenal mucosa in vitro

We examined the effects of the divalent cation calcium (Ca 2+) and the monovalent cations potassium (K +) and sodium (Na +) and different modalities that affect the fluxes of these cations on immunoreactive secretin (IRS) secretion from canine duodenal mucosa in vitro. In the absence of extracellular Ca 2+, the basal IRS secretion was inhibited by 29%. Increased uptake of Ca 2+ by passive diffusion and facilitated uptake of Ca 2+ by ionophore A23187 had no effect on basal IRS secretion. However, Ca 2+ channel blocking agents, LaCl 3 and verapamil, inhibited basal IRS secretion by 18 and 33% respectively. High concentrations of extracellular K + caused a dose-dependent but delayed stimulation of IRS secretion. K +-stimulated IRS secretion was not neurally mediated. Similarly, 9-aminoacridine, which blocks K + exchange, mimicked the effects of K + Valinomycin (10 −6M) inhibited both K +-stimulated and 9-aminoacridine-stimulated IRS secretion. In the absence of extracellular Na +, there was a delayed inhibition of both basal and K +stimulated IRS secretion. These results suggest that changes in cationic environment are associated with alterations in the secretion of IRS. High extracellular K + concentration is conducive, and the absence of extracellular Na + and Ca 2+ is inhibitory to IRS secretion.

Effect of synthetic trypsin inhibitor on plasma immunoreactive cholecystokinin in rats.

A sensitive and specific radioimmunoassay system for rat plasma cholecystokinin (CCK) was employed to study the effect of trypsin inhibitor on plasma CCK levels. Feeding the trypsin inhibitor, FOY-305 200 mg/kg, to rats for 10 days stimulated the pancreatic growth. However, there was no significant difference in fasting plasma concentrations and duodenal contents of CCK and secretin. On the other hand, acute ingestion of same dosage of FOY-305 caused a marked (20-fold) and sustained elevation of plasma CCK levels. The principal form of circulating CCK was CCK-22 like on Sephadex G-50, in contrast to that of the cerebrum and duodenum where CCK-8 like form, or CCK-8 and CCK-22 like forms were main components, respectively. A significant rise in plasma levels of immunoreactive secretin was also found. These results suggest that the pancreatic growth observed by feeding the trypsin inhibitor to rats is caused by an excessive amount of CCK in conjunction with secretin.

Bile-stimulated secretin release in cats.

Duodenal perfusion with sodium taurocholate (TC), pH 10.5 (60 mM, made isotonic with NaCl, 30 ml/h for 90 min), increased the plasma immunoreactive secretin level (IRS) from 2.1 +/- 1.0 to 19.1 +/- 6.0 pM (p less than 0.025; n = 6) in anaesthetized cats. Dose-response studies with TC, pH 7.2 (0-60 mM, made isotonic with NaCl, 45 ml/h for 20 min), demonstrated a threshold concentration of TC for IRS release between 10 and 15 mM (p = 0.05; n = 6). Instillation of gallbladder bile from the same animal (1.5 +/- 0.1 ml, pH 7.0 +/- 0.1, over 12 min) increased the IRS concentration from 1.5 +/- 0.4 to 8.1 +/- 3.4 pM (p less than 0.025; n = 6). IRS concentrations also increased when gallbladder bile was mixed with either saline (from 3.4 +/- 1.7 to 11.6 +/- 2.5 pM; p less than 0.05; n = 5) or feline pure pancreatic juice (PPJ) (from 3.1 +/- 0.7 to 5.7 +/- 1.1 pM; p less than 0.025; n = 6). The peak value was significantly lower when bile was mixed with PPJ than with saline (p less than 0.05). We conclude that TC, the dominating bile salt in feline bile, can release IRS also above pH 7, that physiological concentrations of TC can elicit this response, that the response is also produced by gallbladder bile from the same animal, and that PPJ inhibits the bile-induced IRS release. Thus secretin release by bile is likely to be a physiological principle in the regulation of the pancreatic secretion.

Hepatic bile and pancreatic exocrine secretions evoked by gastrointestinal peptides in sheep

1. 1. The secretory response of hepatic bile and exocrine pancreas to gastrointestinal peptides has been studied in chronically cannulated sheep. 2. 2. Prancreatic juice flow and protein output were evoked dose dependently by intraportal injection of secretin, CCK-8, caerulein, VIP and neurotensin. However, biliary secretion was evoked by only secretin. 3. 3. Biliary and pancreatic exocrine secretions were enhanced by delivered gastric juice into the duodenum as followed by the increased plasma concentration of immunoreactive secretin (1RS). 4. 4. Results suggest that secretin is the major peptide that regulates pancreatic exocrine secretion and hepatic bile production in the sheep.

EARLYLIFE PATTERN OF PLASMA SECRETIN LEVEL IN CALVES

Newborn calves given colostrum showed an increase of plasma immunoreactive secretin (IRS), that did not occur in fasted animals. Thereafter, fasting plasma IRS levels were high in preruminant calves, with a clear fall 1 h after the morning meal. IRS levels were lower in animals given fish instead of milk protein. The postprandial decrease was no longer observed after weaning. Key words: Calf, secretin, preruminant, ruminant

An enzyme immunoassay for secretin

A sensitive and specific enzyme immunoassay for secretin was developed with the use of enzyme-labeled antigens. Synthetic procine secretin and its carboxy-terminal fragments (residues 11–27 and 18–27) were conjugated with β- d-galactosidase for use in the immunoassay, and the assay method with the latter fragment (residues 18–27) linked to β- d-galactosidase was found to be the most sensitive. The minimum amount of secretin detectable by this method was 1–2.5 pg/assay. Serum levels of secretin after intravenous injection of the peptide in rats were determined by both the enzyme immunoassay and a commercial radioimmunoassay kit. The correlation coefficient between the levels measured by the two methods was 0.984. The enzyme immunoassay could detect immunoreactive secretin levels in normal human sera, giving a value of 16.9 ± 2.2 pg/ml (mean ± SE of six human subjects).

Secretin release in coeliac disease. Plasma secretin concentration and bicarbonate output to the duodenum after intraduodenal acid infusion in coeliac patients before and after treatment.

Infusion of 40 ml 0.1 mol/l HCl into the duodenum in eight untreated coeliac patients was followed by an increase of the plasma immunoreactive secretin (IRS) concentration from 1.6 +/- 0.2 pmol/l to a peak level of 2.4 +/- 0.3 pmol/l (p less than 0.05). After treatment with a gluten-free diet, the same patients showed an increase from 1.4 +/- 0.3 pmol/l to a peak level of 5.5 +/- 0.9 pmol/l after intraduodenal acid infusion, which was significantly higher than before treatment (p less than 0.01). In control subjects, intraduodenal acid infusion was followed by an increase from 1.4 +/- 0.2 pmol/l to 6.7 +/- 1.1 pmol/l, which was significantly higher than in untreated coeliac disease (p less than 0.01) but did not differ from what was found in treated coeliac patients. Significant differences in pH, volume, or bicarbonate content of the duodenal aspirates or the basal IRS levels were not found.

Secretin-like immunoreactivity and biological activity in the antral mucosa.

Using immunohistocytochemical techniques, secretin cells are again demonstrated in the antral mucosae of both dogs and rats. Secretin-like immunoreactivity was found in the crude extracts of antral mucosae in 15 dogs [1.18 +/- 0.48 (+/- SE) ng/g wet wt of mucosae], and a similar amount of SLI was also found in 82 rat antral mucosae. Upon ion exchange chromatography, the extracts of dog antral mucosae exhibited a predominant species eluted by the same salt concentration as porcine secretin. The rat antral mucosal extract also produced a chromatogram exhibiting the same predominant species on the ion exchanger. The main immunoreactive secretin peak, when gel filtrated on a Sephadex G-50 (superfine) column, produced an elution profile identical to that of standard natural porcine secretin. These results indicated that antral mucosae of both animal species contain an immunoreactive secretin-like material of the same charge and size as natural porcine secretin. Intravenous injection of a preparation of partially purified secretin from the extracts of canine antral mucosae resulted in a significant increase in the pancreatic flow in anesthetized rats. We conclude that a small number of secretin cells are, therefore, present in the antral mucosae of dog and rat, and this observation is supported by the presence of an immunologically and biologically active secretin-like molecule with charge and size similar to those of porcine secretin in the canine mucosal extracts.

Effect of acid perfusion of the terminal ileum on plasma immunoreactive secretin and pancreatic secretion in the rat.

The distribution of immunoreactive secretin in acid extracts of the alimentary tract of the rat was determined. It was found to be contained in higher concentrations in the duodenum and in the terminal ileum than in other parts of the intestine. To elucidate the possible role of secretin contained in the terminal ileum, this part of the intestine was perfused with hydrochloric acid, a potent secretin secretagogue, and changes in plasma immunoreactive secretin levels and in pancreatic secretion were investigated in anaesthetized rats. Plasma immunoreactive secretin was increased from 65 +/- 5 pg/ml to 108 +/- 13 pg/ml during a 10 minutes perfusion with hydrochloric acid. This was significantly higher than that of saline perfusion but significantly lower than that of duodenal perfusion with hydrochloric acid. Acid perfusion of the terminal ileum also caused a three-fold increase in pancreatic flow rate and a twofold increase in bicarbonate concentration. These results suggest that the terminal ileum, through release of secretin, might play a role in regulating the pancreatic secretion of water and bicarbonate in response to changes in intraluminal fluids in the distal intestine in the rat.

Role of the kidneys in elimination of glucagon, insulin, secretin and somatostatin in the rat.

Immunoreactive plasma glucagon and secretin in the rat was elevated 48 hours after nephrectomy and ureteral ligation. Since kidneys obstructed by ureteral ligation were unable to remove glucagon and secretin from the blood, renal handling of glucagon and secretin must include glomerular filtration. Insulin and somatostatin levels were significantly elevated 48 hours after nephrectomy, but not after ureteral ligation, indicating partial uptake from peritubular capillaries.

Hormonal effects on the pylorus.

The control mechanisms of pyloric pressure responses have not been elucidated clearly. The purposes of this study were twofold: 1) to determine the dose-related pressure responses of the pylorus to exogenous glucagon, secretin, and cholecystokinin, and 2) to correlate changes in pyloric pressure with serum concentrations of these hormones. Pyloric pressures were measured by infusion manometry, and the serum concentrations of glucagon and secretin were quantitated using radioimmunoassays. Each of the hormones tested, glucagon, secretin, and cholecystokinin, increased the pyloric pressure significantly. The lowest active dosages tested for each of these peptide hormones were 2 micrograms.kg-1.h-1, 2 U.kg-1.h-1, and 1 U.kg-1.h-1, respectively. The maximal pyloric pressure responses recorded were 8.7 +/- 1.1 (P less than 0.05), 12.6 +/- 2.1 (P less than 0.02), and 14.8 +/- 1.7 (P less than 0.02) mmHg, respectively. The pyloric pressure responses to insulin hypoglycemia, duodenal acidification, and intraduodenal olive oil were 11.3 +/- 1.5, 13.4 +/- 1.4, and 11.3 +/- 1.4 mmHg, respectively. The serum concentrations of immunoreactive glucagon during infusion of the lowest active dosage of glucagon and insulin hypoglycemia were 801 +/- 55 and 322 +/- 12 pg/ml, respectively. The serum concentrations of immunoreactive secretin during infusion of the lowest active dosage of secretin and during duodenal acidification were 980 +/- 60 and 110 +/- 7.0 pg/ml, respectively. Although pyloric contraction can be induced by administration of exogenous glucagon, secretin, and cholecystokinin, these studies suggest that these effects may have no physiological relevance.

Distribution and metabolism of secretin and aprotinin in isolated perfused rat pancreas.

The distribution and metabolism of secretin and aprotinin were studied in isolated perfused rat pancreas, which were kept under physiological conditions. At 125I-[Tyr1]secretin perfusion study the radioactivity appeared in the effluent, rapidly attaining an equilibrium with the perfusate. While the intact secretin concentration decreased to 55% of the perfusate; the distribution space of secretin was estimated to be about 0.11 ml/g pancreas. The ratio of intact secretin to degradated products in the pancreas increased concomitantly with the perfusion of 82 nM secretin. When unlabeled secretin was perfused, immunoreactive secretin concentration in the portal effluent reached also a rapid equilibrium to about 60% of the perfusate. These results suggested that pancreas might clear about 40% of the entering secretin. In the case of 125I-aprotinin perfusion, intact aprotinin fraction was predominant both in the effluent and in the pancreas. The apparent distribution space was about 40% of the volume of pancrease. Meanwhile, when 125I-reduced aprotinin or 125I-[S-carboxamidomethyl]aprotinin was perfused, the decomposed products were significantly increased. These findings indicate that the disulfide bonds in aprotinin molecule play an important role for its stability as well as its antiproteolytic activity.

Recovery of secretin in acid small intestinal lumen perfusates in the rabbit.

Immunoreactive secretin concentrations were measured in femoral venous plasma and in intestinal perfusates during acid and neutral perfusion of small intestinal segments in anesthetized rabbits. Steady-state plasma and luminal secretin concentrations were less than 20 pg/ml. Duodenal perfusion with with 0.1 N hydrochloric acid stimulated peak plasma immunoreactive secretin concentrations of 200 - 300 pg/ml within five minutes. Acid stimulated plasma concentrations remained constant during 30 minutes of perfusion. Secretin output into the acid perfusate (2000 - 4000 pg/min) accompanied the release into plasma. Subsequent perfusion with a neutral solution resulted in a rapid return to steady-state plasma and luminal secretin concentrations. Sequential acid perfusion of the ileum, jejunum, and duodenum stimulated similar concentrations of secretin in plasma and luminal perfusates.