Ta-min Chang

Identification, characterization and distribution of motilin immunoreactivity in the rat central nervous system

Motilin-immunoreactivity was evaluated in rat brain using 15 different antisera and by combining gel filtration, high pressure gel filtration and reverse phase high pressure liquid chromatography with radioimmunoassay. Gel filtration chromatography demonstrated a high molecular weight and low molecular weight form of immunoreactive motilin. The high molecular weight form predominated in brain while the low molecular weight peptide was the predominant form of duodenum. The low molecular weight immunoreactive motilin was indistinguishable from synthetic porcine motilin by gel filtration and high molecular weight gel filtration. Low molecular weight rat motilin could, however, be distinguished from synthetic porcine motilin by high pressure liquid chromatography and certain antisera. Immunological results suggest that the slight structural difference may be in the N-terminal portion of the molecule. Immunoreactivity was measured in grossly and microdissected regions of the rat brain. The peptide had quite a unique distribution as highest concentrations are observed in the cerebellum. High concentrations were also observed in hypothalamic nuclei. Particularly high concentrations were noted in the organum vasculosum lamina terminalis. Lowest motilin concentrations in the rat brain were in the pons and the medulla. The distribution of motilin in rat brain suggests that it may have roles in regulating both neuroendocrine and neurological processes.

Pituitary adenylate cyclase-activating peptide stimulates rat pancreatic secretion via secretin and cholecystokinin releases☆☆☆

BACKGROUND & AIMS Pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates protein and/or amylase secretion from isolated rat pancreatic acini. The effect of PACAP on pancreatic secretion in vivo and its mechanism of action were studied. METHODS Rats were prepared with pancreatic duct cannulation, pyloric ligation, and bile diversion into duodenum, and 2.5, 5, and 10 nmol/kg PACAP-27 was administered intravenously while pancreatic juice was collected for 30 minutes. In other groups of rats, the effect of 10 nmol/kg PACAP-27 was studied under the influence of either atropine; loxiglumide, an antisecretin serum; a combination of both loxiglumide and the antiserum; or a PACAP antagonist (PACAP 6-38). Plasma secretin and cholecystokinin concentrations were measured by radioimmunoassay. RESULTS (1) PACAP dose-dependently increased pancreatic secretion of fluid, bicarbonate, and protein; (2) the increase in pancreatic secretion paralleled that of plasma secretin and cholecystokinin; (3) a combination of loxiglumide and antisecretin serum eliminated the PACAP-stimulated pancreatic secretion, whereas loxiglumide or antisecretin serum alone partially but significantly blocked pancreatic secretion; (4) atropine failed to influence PACAP-induced pancreatic secretion; and (5) PACAP antagonist profoundly suppressed the PACAP action. CONCLUSIONS PACAP-27 dose-dependently stimulates pancreatic secretion of fluid, bicarbonate, and protein in rats. This effect is mediated by release of both secretin and cholecystokinin and is independent of cholinergic tone.

Acid-independent Release of Secretin and Cholecystokinin by Intraduodenal Infusion of Fat in Humans

In order to clarify a possible role of fat content in the release of secretin and cholecystokinin by liquid nutritional supplements in humans, duodenal pH and plasma concentrations of secretin and cholecystokinin were studied during the intraduodenal infusion of Ensure, Vivonex, 10% Intralipid, and sodium oleate. Significant release of secretin was observed with Intralipid and sodium oleate, while significant release of cholecystokinin was observed with all four testing solutions. Duodenal pH was rarely below 4.5 during the infusion of Ensure, Intralipid, and sodium oleate. Duodenal pH was high, greater than 6.0, when plasma secretin and cholecystokinin levels were elevated during the administration of Ensure, Intralipid, and sodium oleate. We conclude that both secretin and cholecystokinin are released in response to fat solutions in the duodenum and that low duodenal pH was not responsible for either secretin or cholecystokinin release during intraduodenal infusions of Ensure, Intralipid, or sodium oleate.

Purification of two secretin-releasing peptides structurally related to phospholipase A2 from canine pancreatic juice.

We previously showed that canine pancreatic juice contains a secretin-releasing factor activity. In this study, we carried out isolation of two secretin-releasing peptides (SRPs) from canine pancreatic juice. Through ultrafiltration, anion and cation exchange, and reverse-phase high-performance liquid chromatography (HPLC) steps and an in vitro bioassay in STC-1 cells, two SRPs, SRP-1 and SRP-2, were isolated and purified to homogeneity. Both SRPs dose-dependently stimulated secretin release from STC-1 cells. The results of mass spectral analysis indicated that SRP-1 and SRP-2 had molecular masses of 14,061 Da and 14,053 Da, respectively. N-terminal amino acid sequence analysis indicated that SRP-1 was identical to canine pancreatic PLA2 in the 25 residues determined; whereas SRP-2 had 71% sequence homology to the enzyme in the first 21 residues. Commercially available porcine pancreatic PLA2 dose-dependently stimulated secretin release from STC-1 cells. Porcine pancreatic PLA2 also stimulated secretin release from a secretin-producing cells-enriched preparation isolated from rat duodenal mucosa. These results suggest that pancreatic PLA2 and its related peptide may participate in regulation of secretin secretion.

Secretin: historical perspective and current status.

Abstract This review describes the history of secretin discovery, identification, purification, and structural determination; cloning of secretin and its receptor; synthetic secretin; and highly specific and sensitive radioimmunoassay to define the characteristic physiological role on postprandial pancreatic fluid and bicarbonate secretion, which requires robust potentiation by cholecystokinin. Secretin plays a key role in the negative and positive regulatory mechanisms of exocrine pancreatic secretion. Secretin-releasing peptides were discovered in duodenal acid perfusates of both rat and dog and in canine pancreatic juice. The release and action of secretin and secretin-releasing peptides are in part mediated via vagovagal reflex mechanism involving afferent sensory neurons in proximal intestine and efferent cholinergic neurons in the pancreas. Besides acetylcholine, many neurotransmitters or neuromodulators influence release and action of secretin. The action of secretin in the pancreas depends on insulin, which also suppresses local release of somatostatin and pancreatic polypeptide. Thus, release and action of secretin are mediated via neurohormonal interaction. Clinical conditions with hypersecretinemia and hyposecretinemia are discussed. Synthetic human secretin is used for studies of exocrine pancreatic secretion, secretin-enhanced magnetic resonance cholangiopancreatography combined with exocrine pancreatic function test and diagnosis of gastrinoma syndrome. Therapeutic use of secretin is considered for the relief of severe pain in chronic pancreatitis.

Neurohormonal control of exocrine pancreas.

The exocrine pancreas is regulated by various hormonal factors derived from the gut through hormone-hormonal and neurohormonal interactions. Physiologic stimuli entering the upper small intestine elicit the release of intestinal hormones and activate sensory reflex mechanisms from the intestinal mucosa to stimulate or inhibit exocrine pancreatic secretion. In addition, the endocrine pancreas, intrapancreatic nerves, and some extrapancreatic neural pathways, with or without mediation by the vagus nerve, are known to participate in regulation of exocrine pancreatic secretion. It has been established that two key intestinal hormones, secretin and cholecystokinin (CCK), in physiologic doses, act through the vagal afferent pathway and interact with each other as well as with other gut hormones. The releases of these two hormones are mediated through the corresponding releasing peptides. In the past few years, the roles of secretin- and CCK-releasing peptides have become more clearly defined. The participation of several neurotransmitters and regulatory peptides in the regulation of exocrine pancreatic secretion has also been established. In addition, neurotransmitters and neuropeptides released from the central nervous system may participate in the regulation of pancreatic secretion. It is conceivable that a few neurotransmitters and neuropeptides are involved in each neural regulatory pathway. However, their roles and sites of action in each pathway remain to be determined.

Release of secretin by licorice extract in dogs

To determine the effect of a fraction of licorice extract, FM100, on the endogenous release of secretin and exocrine pancreatic secretion, five dogs were prepared with chronic pancreatic fistulas and gastric cannulas. In-traduodenal administration of licorice extract (pH 7.4) in three different doses (0.5, 1.0, and 2.0 g) resulted in significant increases of both plasma secretin concentrations and pancreatic bicarbonate secretion in a dose-related manner. The plasma secretin concentration and pancreatic bicarbonate output produced by licorice extract correlated well. Intragastric administration of licorice extract (2 g) in 5% liver extract meal (in which pH was maintained at 5.5 by the intragastric titration method) resulted in significant increases of both plasma secretin concentration and pancreatic bicarbonate output. The increase in pancreatic bicarbonate secretion was completely abolished by intravenous infusion of a rabbit antisecretin serum in the two dogs so studied. Thus, the present study indicates that the endogenous release of secretion is involved in a mechanism of an increase in exocrine pancreatic secretin induced by FM100.

CHAPTER 153 – Secretin

Secretin is the first peptide hormone ever discovered. Secretin is a 27-amino-acid peptide of secretin-glucagon-vasoactive intestinal polypeptide superfamily and is localized mainly in the upper small intestinal mucosa. The gene structures of secretin and its receptor have been determined. Compared with several bio-active prosecretins, secretin is the most active form. The physiological functions of secretin include the stimulation of pancreatic exocrine secretion of water and electrolytes, inhibition of gastric acid secretion, and motility. The physiological release and actions of secretin are subjected to hormone-hormonal and neural-hormonal regulations, among which the vagal afferent pathway plays a significant role. Secretin is used clinically to assess pancreatic function and test for pancreatic malignancy. Pathological states of both hyposecretinemia and hypersecretinemia have been well documented.

Synergistic stimulation of CCK secretion from STC-1 cells by sodium oleate and amino acids

A mixed meal is a potent stimulant for the release of gastrointestinal peptides. Fatty acids and amino acids (AA)are digested products of food that stimulate CCK secretion. The postprandial concentration of these nutrients in the intestinal lumen may not be comparable to those during intraduodenal infusion to demonstrate their secretagogue activity. Sodium oleate (SO) and AA have been shown to stimulate STC-l cells to secrete CCK. We hypothesized that these nutrients may act synergistically on CCK cells at subthreshold concentrations. Purpose: To investigate whether or not low doses of SO and AA (L-Trp and L-Phe)can act synergistically on CCK release from STC-l cells. Methods: STC-I cells were incubated without or with SO, L-Trp or L-Phe, or combination of SO with either AA in the absence or presence of different protein kinase inhibitors. The release of CCK in the medium was assessed by a specific RIA. The effect of SO and amino acid on intracellular Ca + concentration ([Ca +L) was determined spectrafluorometrically in Fura 2 AM-preloaded cells. Results: There was a significant synergistic stimulation of CCK release by SO and TrplPhe as demonstrated by the difference between their combinational stimulation and the sum of their individual effects shown in the Table below. The synergism was also timeand dose-dependent. There was no synergistic increase in [Ca+L level between SO (0.14 mM) and Phe (10 mM). The PKC inhibitor, staurosporine (10 nM)did not significantly inhibit each amino acid stimulated CCK release but significantly reduced their synergism with SO. On the other hand, the calmodulin antagonist, calmidazolium chloride did not significantly affect the synergistic effect between SO and amino acids. Conclusion: Combination of SO and TrplPhe synergistically stimulated CCK release from STC-I cells timeand dose-dependently. The synergism appears to be mediated by PKC rather than by Ca+{calmodulin-dependent protein kinase II nor the elevation of [Ca2+L known to mediate stimulation by SO. 599

ABSENCE OF IN-UTERO AMNIOTIC FLUID SWALLOWING IS ASSOCIATED WITH DECREASED CORD BLOOD TROPHIC GUT PEPTIDE LEVELS. † 701

The human fetus receives most of its nutrition parenterally via the placenta, but also swallows large volumes of amniotic fluid. The swallowed amniotic fluid provides nutrients, specific growth factors and mechanical distension for the fetal gut. These factors act in concert to stimulate gastrointestinal(GI) tract development. In several animal models, esophageal ligation causes decreased gastrointestinal weight and absorptive capacity, abnormal ultrastructural GI morphology and reduced blood levels of gut peptides. Similarly, human newborns who experience absent or reduced amniotic fluid swallowing(AFS) because of upper GI obstruction or neurologic disorders often present with GI immaturity and decreased absorptive capacity. We hypothesized that the absence of in-utero AFS would be associated with decreased umbilical cord blood levels of gut peptides shown to have trophic effects on the GI system.