bluuebunny/crossref_metadata_2025 huggingface dataset:
Somatostatin receptors on canine fundic D-cells: evidence for autocrine regulation of gastric somatostatin
Authors: Park J. , Chiba T. , Yokotani K. , DelValle J. , Yamada T.
Published: December 2017

Somatostatin is known to have inhibitory effects on the release and action of a wide variety of gut peptides. Previous studies in vivo have suggested a potential inhibitory role for somatostatin even on its own secretion. To determine whether this autoregulatory effect is the result of a direct action of the peptide on the cell that is responsible for its secretion, we examined the effect of a non-immunoreactive but biologically active analogue of somatostatin ([Leu8-D-Trp22-Tyr25]S28) on release of somatostatin-like immunoreactivity (SLI) from isolated canine fundic D-cells. We identified somatostatin binding sites with dissociation constants of 1.2 X 10(-9) and 3.8 X 10(-8) M that coenriched with D-cells. Somatostatin 14, somatostatin 28, and [Leu8-D-Trp22-Tyr25]S28 were equivalent in displacing 125I-[Leu8-D-Trp22-Tyr25]S28 from the binding sites. [Leu8-D-Trp22-Tyr25]S28 inhibited SLI release from D-cells stimulated with (DBcAMP), and pentagastrin. Pertussis toxin pretreatment prevented the inhibitory effects of [Leu8-D-Trp22-Tyr25]S28 on both SLI secretion and cAMP accumulation by on both SLI secretion and cAMP accumulation by D-cells stimulated with epinephrine and forskolin. In contrast, [Leu8-D-Trp22-Tyr25]S28 inhibition of SLI release induced by DBcAMP and pentagastrin was not altered by pertussis toxin. Our data suggest that somatostatin autoregulates its own secretion via specific receptors on D-cells. This inhibitory effect is mediated by mechanisms that are both dependent on and independent of pertussis toxin-sensitive inhibitory guanine nucleotide binding proteins.
Aging: altered responsiveness of gastric mucosa to epidermal growth factor
Authors: Majumdar A. P. , Arlow F. L.
Published: December 2017

The present investigation examines the responsiveness of the gastric mucosa to the growth-promoting action of epidermal growth factor (EGF) during advancing age. Two sets of experiments were performed. In the first set of experiments, groups of 4-, 8-, 16-, and 24-mo-old Fischer 344 rats were injected subcutaneously at 12-h intervals for 2 days with either EGF (10 micrograms/kg) in gelatin or the vehicle only (controls). The animals were killed 16-18 h after the last injection. The oxyntic gland mucosa was assayed for thymidine kinase and the rate of DNA synthesis in vitro (indicators of proliferative activity) as well as for tyrosine kinase (Tyr-k) activity. In control rats, the rate of DNA synthesis and thymidine kinase activity rose steadily between 4 and 24 mo of age. However, whereas Tyr-k activity in the gastric mucosal cytosol changed only marginally with age, activity of the enzyme in the membrane fraction rose steadily between 4 and 16 mo and then increased abruptly. EGF stimulated gastric mucosal DNA synthesis and thymidine kinase activity in 4- to 16-mo-old rats compared with the corresponding controls, but in the 24-mo-old animals, it caused a significant 40-50% inhibition. EGF had no demonstrable effect on Tyr-k activity in either cytosolic or membrane fraction. We postulated that Tyr-k activity might have returned to basal level 16-18 h after the last EGF injection.(ABSTRACT TRUNCATED AT 250 WORDS)
A new CCK analogue differentiates two functionally distinct CCK receptors in rat and mouse pancreatic acini
Authors: Matozaki T. , Martinez J. , Williams J. A.
Published: December 2017

Analysis of the competitive inhibition of 125I-labeled cholecystokinin octapeptide (CCK-8) binding to isolated rat or mouse pancreatic acini showed that in both species CCK-8 interacts with two different affinity sites. A newly synthesized CCK analogue modified at the COOH-terminal phenylalanine residue totally inhibited 125I-CCK binding. This interaction occurred with sites of a single affinity in rat acini but with two different affinity sites in mouse acini. When acini were incubated with increasing concentrations of CCK-8, a biphasic stimulation of amylase release was observed. By use of rat acini, the analogs stimulated amylase release but caused no inhibition at supramaximal concentrations. By contrast, in mouse pancreatic acini, analogues showed a biphasic stimulation of amylase release similar to CCK-8. Both CCK-8 and the analogue stimulated [3H]leucine incorporation into protein at low concentrations in rat pancreatic acini. Higher concentrations of CCK-8 profoundly inhibited [3H]leucine incorporation, whereas the analogue had no inhibitory effect. Moreover, the analogue at higher concentrations blocked the inhibition of [3H]leucine incorporation caused by CCK-8 but did not affect carbamylcholine-induced inhibition. In mouse acini, however, the CCK analogue inhibited [3H]leucine incorporation similar to the effect of CCK-8. The results support the concept that occupancy of distinct affinity sites or states of the CCK receptor is associated with specific biological actions. A model of the CCK receptor is proposed in which two interchangeable affinity states exist. By occupying all the receptors in only one state, the new CCK analogues serve as partial agonists of some and antagonists of other actions of CCK.
Prolonged ambulant monitoring of human colonic motility
Authors: Soffer E. E. , Scalabrini P. , Wingate D. L.
Published: December 2017

The study of human colonic motility under physiological conditions has proved to be an elusive goal. We have used a two-stage pernasal technique to position sensors in the human colon for the prolonged monitoring of motility in freely ambulant subjects. Nine healthy volunteers were studied for a total recording time of 263 h, each study lasting between 13 and 48 (mean 29) h. Motor activity in all regions of the large bowel was characterized by scant and irregular contractions with infrequent bursts that did not conform to any pattern. No motor coordination was apparent between different regions of the large bowel. Contractile activity throughout the large bowel was reduced to a minimum during sleep and was enhanced on waking. Meals were an inconsistent stimulus to motor activity. The technique obviates the need for colonic preparation and allows complete freedom of the subjects throughout the study. In demonstrating the practical feasibility of this mode of studying the colon, these preliminary data highlight a requirement for the availability of appropriate equipment but raise questions about the design and use of such equipment and methods of data analysis.
New insights into lactase and glycosylceramidase activities of rat lactase-phlorizin hydrolase
Authors: Buller H. A. , Van Wassenaer A. G. , Raghavan S. , Montgomery R. K. , Sybicki M. A. , Grand R. J.
Published: December 2017

Lactase-phlorizin hydrolase, a small intestinal disaccharidase, has been considered mainly an enzyme important only for the hydrolysis of lactose. After weaning in most mammals lactase-specific activity falls markedly, and, functionally, adult mammals are considered to be lactase deficient. However, the persistence of low levels of lactase activity in adulthood has never been explained. In addition, it has been suggested that lactase-phlorizin hydrolase is associated with glycosylceramidase activity when the enzyme is prepared by column chromatography, but it is unclear whether this represents copurified activities or two catalytic sites on one peptide. The developmental patterns of lactase-phlorizin hydrolase and other disaccharidases were investigated in homogenates of total rat small intestine; lactase and several glycosylceramidases were measured in immunoprecipitates from these homogenates using a monoclonal antibody. The developmental pattern of total lactase activity showed a steady 2.3-fold increase to adult levels (specific activity decreased eightfold), whereas total phlorizin-hydrolase activity increased 10.7-fold (specific activity decreased threefold). As expected, levels of both total and specific sucrase and maltase activities increased during development. In lactating rats total lactase activity showed a significant increase compared with adult males. The developmental pattern of the enzyme activities for the glycolipid substrates was similar to that found for lactase, and the immunoprecipitated enzyme showed a 40- to 55-fold higher affinity for the glycolipids than for lactose. Galactosyl- and lactosylceramide inhibited lactose hydrolysis by 38%, without a competitive pattern, suggesting two different active sites for lactose and glycolipid hydrolysis, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Delivery of high-density lipoprotein free and esterified cholesterol to bile by the perfused monkey liver
Authors: Scobey M. W. , Johnson F. L. , Rudel L. L.
Published: December 2017

The movement of cholesterol from high-density lipoproteins (HDL) into bile has been studied using perfused livers from cholesterol-fed African Green monkeys. Mass amounts of HDL were isolated from the plasma of African Green monkeys and were doubly labeled with either 125I-apolipoprotein and [3H]cholesteryl ester or with [3H]cholesteryl ester and [14C]cholesterol. For 3 h of perfusion HDL-free cholesterol was cleared from perfusate at a faster rate than HDL ester cholesterol which, in turn, was cleared at a faster rate than HDL protein. [14C]cholesterol from HDL appeared in biliary bile acids and cholesterol at a higher rate than [3H]esterified cholesterol from HDL. The specific activities of biliary [14C]cholesterol and HDL-free [14C]cholesterol had equilibrated by 60 min of perfusion, although the specific activity of whole liver free [14C]cholesterol was still only 46% of that in bile at 180 min of perfusion. In contrast, the specific activity of total liver free [3H]cholesterol was equal to that of biliary [3H]cholesterol by 180 min of perfusion. We conclude that, in this primate model, HDL-free cholesterol enters into a hepatic compartment that communicates with biliary cholesterol and bile acid precursor pools more efficiently than with other liver pools of cholesterol, whereas HDL-esterified cholesterol appears to mix with all liver pools with equal efficiency. Overall, these data support the concept of compartmentalization of cholesterol in the liver.
Gastric response to mucosal IgE-mediated reactions
Authors: Catto-Smith A. G. , Patrick M. K. , Scott R. B. , Davison J. S. , Gall D. G.
Published: December 2017

The effect of immunoglobulin E (IgE)-mediated anaphylaxis has been extensively studied in the small intestine, but little information is available on the response of the stomach to IgE-mediated mucosal reactions to food proteins. The effect of luminal antigenic challenge on gastric acid secretion, gastric emptying, and mucosal mast cell degranulation was examined in rats sensitized to egg albumin or in sham-treated controls. Intraluminal challenge of the stomach with egg albumin in sensitized animals significantly increased gastric acid secretion and delayed gastric emptying. The response was specific for the sensitizing antigen as challenge with bovine serum albumin was without effect. Sham-treated animals showed no response to egg albumin or bovine serum albumin. The increase in gastric acid secretion was reproduced by antigen challenge in naive animals passively transferred with hyperimmune serum. This effect was abolished by prior heat treatment of the serum. In sensitized animals challenged with egg albumin, there was histological evidence of mast cell degranulation in the stomach mucosa, increased intraluminal release of histamine, and increased serum levels of rat mast cell protease II, a marker specific for mucosal mast cell degranulation. The findings indicate that the stomach is a target organ for IgE-mediated reactions to food proteins. Antigen challenge in sensitized animals leads to increased gastric acid secretion and delayed emptying and evidence of mucosal mast cell activation.
Rates of triolein absorption in suckling and adult rats
Authors: Flores C. A. , Hing S. A. , Wells M. A. , Koldovsky O.
Published: December 2017

To determine the extent to which suckling animals differ from adults in their capacity to absorb fat, we compared the rate of absorption of orally administered [14C]triolein in 11- to 12-day-old suckling rats with that of 10-wk-old adults by three distinct methods. In the first, the rate of [14C]triolein disappearance was determined by quantitating substrate remaining in the gastrointestinal tract after oral administration. In the second, 14CO2 expiration in breath was measured continuously for 6 h after an identical feeding. In the third, intestinal triglyceride output was estimated by the lipoprotein lipase inhibitor, Triton WR-1339. Triolein disappearance, 14CO2 excretion, and intestinal triglyceride output were two- to threefold higher in suckling rats compared with adults (P less than 0.01, P less than 0.0001, and P less than 0.01, respectively). There was also a highly significant linear relationship between 14CO2 excretion and both triolein disappearance and intestinal triglyceride output for both age groups (P less than 0.0001 and P less than 0.003, respectively). These data show that consistent with its high dietary intake, the suckling rat can absorb triolein at rates significantly higher than the adult.
Comparison of the effects of leukotrienes B4 and D4 on distal colonic motility in the rabbit in vivo
Authors: Burakoff R. , Nastos E. , Won S. , Percy W. H.
Published: December 2017

Leukotrienes (LTs) have been shown to contract smooth muscle of the small and large intestine in vitro, and an increased mucosal synthesis of leukotrienes has been reported to occur in ulcerative colitis. The purpose of this study was to evaluate the effects of LTs B4 and D4 on the myoelectric and mechanical activity of the rabbit distal colon in vivo and to determine how these effects were modified by indomethacin. LTB4 had a weak excitatory action on both electrical and mechanical activity but this was not statistically significant; LTD4 on the other hand caused a significant dose-related increase in spike potential frequency and contractile activity. Indomethacin alone (1 mg/kg) had no significant effect on the electrical or mechanical activity of the colon but blocked the effects of subsequently administered LTD4. It is concluded from these data that at least part of the excitatory action of LTD4 results from its causing the synthesis and release of excitatory prostaglandins. However, whereas leukotrienes have the potential to play an important role in modulating colonic motility, it seems most likely that this effect would occur after pathological stimulus rather than in the normal healthy colon.
Development and tissue distribution of sucrase-isomaltase mRNA in rats
Authors: Leeper L. L. , Henning S. J.
Published: December 2017

Previous studies of sucrase-isomaltase (SI) activities have shown this complex to be absent in the suckling rat and to appear during the weaning period. We describe here the cloning of a heterologous SI cDNA and its use for the quantitation of SI mRNA as a first step toward understanding the molecular basis of SI development. A survey of RNA from 12 tissues of mature rats by Northern blot analysis showed a 6-kb band of SI mRNA only in the small intestine. Within the latter, both sucrase activity and SI mRNA peaked in the jejunum. Assay of jejunal tissue from developing rats showed sucrase activity and SI mRNA to be first detectable at 18 days, to rise in parallel through 24 days, and then to diverge a little (enzyme activity being lower) by 36 days. When glucocorticoid was administered to 10-day-old rats, neither sucrase activity nor SI mRNA was detectable 12 h later. Both parameters were readily detected 24 h postinjection, although the mRNA had risen relatively more than the enzyme activity. The two parameters increased in concert through 5 days postinjection and then plateaued. We conclude that, with respect to distribution along the intestine and to normal and precocious development, activities of SI in the rat are determined primarily by the abundance of its mRNA.
Interaction of cholera toxin with cloned human goblet cells in monolayer culture
Authors: Lencer W. I. , Reinhart F. D. , Neutra M. R.
Published: December 2017

Exposure of the intestinal mucosa to Vibrio cholerae enterotoxin (CT) results in mucus secretion from intestinal goblet cells. On the other hand, there is evidence that elevation of intracellular adenosine 3',5'-cyclic monophosphate levels is not sufficient to induce rapid mucin secretion. To determine whether CT has direct effects on human goblet cells and whether CT alone can elicit rapid exocytosis of apical mucin granules, purified CT was applied to monolayer cultures of well-differentiated HT-29-18 N2 cells, a goblet cell subclone derived from the human colon carcinoma line HT-29. CT bound with high affinity (dissociation constant = 10.5 +/- 1.9 nM) to receptors on these cells (approximately 60,000/apical membrane). Binding of radiolabeled CT was inhibited by excess CT or B subunit but not by A subunit. Preincubation of goblet cells with CT blocked the subsequent CT-specific ribosylation of a 45-kDa protein in membrane fractions of the cells and increased the activity of adenylate cyclase by 2- and 10-fold after 1 and 20 h, respectively. Light micrographs revealed that goblet cells incubated with CT, like control cells, contained abundant apical mucin granules. In contrast, goblet cells incubated with Ca2+ inophore A23187 and phorbol ester were rapidly depleted of mucin granules. Thus CT has direct physiological effects on HT-29 goblet cells, but these do not lead to rapid mucin secretion. These results raise the possibility that CT may accelerate mucin secretion in intact mucosa by an indirect mechanism perhaps mediated by mucosal nerves or other cell types.
An important role of endogenous insulin on exocrine pancreatic secretion in rats
Authors: Lee K. Y. , Zhou L. , Ren X. S. , Chang T. M. , Chey W. Y.
Published: December 2017

We have investigated a physiological role of endogenous insulin on exocrine pancreatic secretion stimulated by a liquid meal as well as exogenous secretin and cholecystokinin octapeptide (CCK-8) in conscious rats. Each rat was prepared with a chronic pancreatic fistula and an indwelling catheter in a jugular vein. Oral ingestion of a liquid meal (5 ml) resulted in significant increases in pancreatic secretion, including volume, bicarbonate, and amylase output, in these rats. A rabbit anti-insulin serum (1.0 ml) given intravenously completely blocked the postprandial exocrine pancreatic secretion, whereas a normal rabbit serum did not influence the pancreatic secretion in the same rats. When pancreatic secretion was stimulated by intravenous administration of both secretin and CCK-8 in three different doses, including 0.015, 0.03, and 0.06 clinical unit and microgram.kg-1.h-1, respectively, volume, bicarbonate, and amylase output increased significantly in a dose-dependent manner. This increase in pancreatic secretion was also completely blocked by a rabbit anti-insulin serum, whereas it was not influenced by a normal rabbit serum. The amount of the antiserum employed abolished the postprandial increases in plasma insulin concentration. We conclude that endogenous insulin plays an important role on the regulation of postprandial pancreatic secretion in rats. Furthermore, for the stimulatory action of the two intestinal hormones secretin and CCK-8 on the pancreatic exocrine secretion, endogenous insulin is need.
Small intestinal amyogenesia and dysmyogenesia induced by morphine and loperamide
Authors: Sarna S. K. , Otterson M. F.
Published: December 2017

We studied the effects of nitric oxide (NO) solution and acidified sodium nitrite (NaNO2), which produces NO, in splanchnic artery occlusion (SAO) shock in cats. NO is thought to be endothelium-derived relaxing factor (EDRF), a labile substance having several potentially valuable biological effects. Anesthetized cats subjected to total occlusion of the celiac, superior mesenteric, and inferior mesenteric arteries for 120 min, followed by reperfusion, usually died approximately 60 min after reperfusion. NO infusion significantly improved survival time in SAO-shock cats compared with those receiving vehicle (P less than 0.005). Administration of NO also attenuated the increase in plasma activities of the lysosomal hydrolase cathepsin D (P less than 0.05), total amino-nitrogen (P less than 0.001), and of the cardiotoxic peptide, myocardial depressant factor (MDF) (P less than 0.001). SAO-shock cats treated with NaNO2 at pH 2.0 also exhibited lower plasma cathepsin D (P less than 0.001), amino-nitrogen (P less than 0.05), and MDF activities (P less than 0.01), and survival time was also significantly improved (P less than 0.02). The same dose of NaNO2 infused at pH 7.4 failed to exert any significant protective effect. These results indicate that NO exerts beneficial effects in SAO shock in cats and suggest that exogenously administered NO may be a potentially useful therapeutic agent in splanchnic ischemic shock, probably via a cytoprotective rather than a vasodilator effect.
Effects of osmotic stresses on isolated rat hepatocytes. I. Ionic mechanisms of cell volume regulation
Authors: Corasanti J. G. , Gleeson D. , Boyer J. L.
Published: December 2017

Isolated hepatocyte suspensions were exposed to hypotonic and hypertonic stresses and serial cell volume measurements were made with an electronic particle size analyzer. With the exposure to hypotonic (160 mosM) buffer, hepatocytes swelled within 30-60 s as osomometers [relative volume (RV) = 1.44 +/- 0.08] and subsequently underwent regulatory volume decrease (RVD) back toward the resting (isotonic) level (1.16 +/- 0.05). This volume recovery was blocked by 65 mM extracellular K+ concentration and inhibited by barium (1 mM) and quinine (0.5 mM) but not by bumetanide (0.1 mM). Chloride depletion inhibited RVD by approximately 40% while 0.5 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) blocked the recovery by almost 90%. Calcium deprivation had no effect on RVD, nor did ouabain, amiloride, or sodium replacement. When exposed to buffer made hypertonic by addition of 200 mM sucrose, cells shrunk as osmometers (RV = 0.74 +/- 0.02) but did not exhibit regulatory volume increase (RVI). However, when cells that had first undergone RVD were reexposed to isotonic medium (relative hypertonic stress) RVI could be demonstrated from RV 0.77 +/- 0.17 to 0.91 +/- 0.20. This response was dependent on sodium, partially dependent on bicarbonate and chloride, and inhibited by the Na(+)-H+ exchange inhibitor amiloride (1 mM) but not by DIDS. Our findings suggest that RVD in rat hepatocytes is mediated by quinine- and barium-sensitive K+ conductance and DIDS-sensitive anion conductance, which is partly accounted for by Cl-; RVI is mediated by activation of Na(+)-H+ exchange coupled with a bicarbonate- and chloride-dependent but DIDS-insensitive process.
Effects of transmural field stimulation in isolated muscle strips from human esophagus
Authors: Tottrup A. , Forman A. , Funch-Jensen P. , Raundahl U. , Andersson K. E.
Published: December 2017

Smooth muscle strips representing longitudinal and circular muscle layers of the esophagogastric junction (EGJ) and esophageal body (EB) of the human esophagus were prepared. The strips were mounted in organ baths and isometric tension was recorded. Square wave stimulation was applied through platinum electrodes. Only responses abolished by tetrodotoxin (TTX) were considered neurogenic. Strips taken from longitudinal muscle layers of the EB and EGJ contracted during field stimulation. The responses evoked were abolished by atropine, and optimal frequency of stimulation was 40 Hz. In strips taken from the circular muscle layer of the EB, a contraction occurred after cessation of the stimulus. Atropine inhibited 90% of this response; the optimal stimulation frequency was 40 Hz. When a tone was induced in strips from this layer, a TTX-sensitive relaxation was seen during field stimulation. During stimulation of strips from the EGJ circular muscle layer, which was the only preparation developing spontaneous active tone, a relaxation was seen. A small contraction followed after termination of the stimulus. The relaxation, which was nonadrenergic, noncholinergic, reached maximum at 10 Hz. Atropine inhibited 40% of the contraction. The results suggest that in the longitudinal muscle layer of the human lower esophagus field stimulation causes postganglionic nerves to release transmitter(s) acting on muscarinic receptors. The responses of circular muscle layers seem to be mediated through release of at least two transmitters.
Gastric emptying in rats: role of afferent neurons and cholecystokinin
Authors: Forster E. R. , Green T. , Elliot M. , Bremner A. , Dockray G. J.
Published: December 2017

Peptone, acid, and hyperosmolal saline delay gastric emptying in conscious gastric fistula rats. We have now studied the emptying of these solutions in animals pretreated with capsaicin to lesion small diameter primary afferents and in rats with both a gastric and duodenal cannula. In capsaicin-treated rats, hyperosmolal saline did not significantly inhibit gastric emptying, whereas the inhibitory action of acid and peptone was reversed but not abolished. In control rats, the action of peptone was inhibited by the selective cholecystokinin antagonist L364,718, but in capsaicin-treated rats, L364,718 enhanced the action of peptone in delaying gastric emptying. In rats with a duodenal cannula approximately 5 cm from the pylorus, intragastric peptone or hyperosmolal solutions only delayed emptying when the duodenal cannula was closed; in contrast, intragastric acid inhibited gastric emptying when the duodenal cannula was open or closed. The results suggest 1) that all three test meals delay emptying by mechanisms depending at least in part on afferent neurons; 2) peptone delays emptying by at least two mechanisms: one is mediated by cholecystokinin A-type receptors and afferent neurons, and the other requires neither these receptors nor small diameter afferents; and 3) acid, but not peptone or hyperosmolal saline, regulates emptying by an action localized to the stomach or proximal duodenum. The results suggest that there are several different reflex pathways by which liquid test meals act to delay gastric emptying.
Effect of neurotensin on regional cerebral glucose utilization in cold water-restrained rats
Authors: Xing L. P. , King J. C. , Bryan R. M. , Kauffman G. L.
Published: December 2017

Neurotensin (NT), given intracerebroventricularly (icv), attenuates cold water restraint (CWR)-induced gastric mucosal injury; however, it is not clear which brain nuclear group or groups are involved. These studies tested the hypothesis that neuronal function, as measured by regional cerebral metabolic rate for glucose (rCMRGlc), is altered by icv NT with or without CWR. CWR resulted in a reduced global glucose utilization of 72 and 65% in control and NT-treated rats, respectively. NT, given icv, protected against ulcer formation induced by CWR. In those rats given NT icv, rCMRGlc was elevated significantly in amygdala, nucleus accumbens, substantia nigra, tuberculum olfactrium, hypothalamus, and cerebellum compared with CWR rats without NT pretreatment. This rCMRGlc increase was observed in both unstressed and stressed rats given NT icv in the nucleus accumbens and amygdala. These observations suggest that the nucleus accumbens and amygdala, both components of the mesolimbic dopamine system, are involved in the central action of NT on the gastric mucosa.
Isolation and characterization of the canalicular membrane bile acid transport protein of rat liver
Authors: Sippel C. J. , Ananthanarayanan M. , Suchy F. J.
Published: December 2017

The aim of this study was to isolate the Na(+)-independent bile acid transporter from rat canalicular plasma membranes by affinity chromatography. The affinity matrix used consisted of lysylcholic acid covalently linked to CH-Sepharose 4B, resulting in an anionic ligand essentially identical to glycocholic acid. The protein fraction, adsorbed and eluted from the affinity column, was markedly enriched in a 100-kDa band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) compared with the total membrane and membrane extract. The 100-kDa band, further purified by preparative SDS-PAGE, was electroeluted from excised gel fragments and used as an immunogen for antibody production in rabbits. The immune serum, but not preimmune serum, specifically recognized a single, 100-kDa polypeptide on one- and two-dimensional immunoblots of canalicular membranes. In contrast, no reactivity was observed with proteins in liver basolateral or ileal brush-border membranes. The 125I-labeled protein was immunoprecipitated from membrane extracts solubilized in NP-40 and was found to migrate with a pI of 5.3 on two-dimensional electrophoresis. The apparent molecular weight of the protein was reduced by 50% after deglycosylation with N-glycanase. The 100-kDa protein was localized specifically and exclusively by immunocytochemical methods to the bile canalicular domain of the hepatocyte plasma membrane. Moreover, the immunoglobin G fraction prepared from the antiserum significantly inhibited taurocholate transport by canalicular membrane vesicles and decreased the covalent labeling of the 100-kDa protein by the anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. Thus the isolation of a single 100-kDa protein by bile acid-affinity chromatography, as well as the inhibitory effects of antibodies directed against this polypeptide, provide further support for its role in the canalicular transport of bile acids.
Zinc blocks apical membrane anion exchange in gallbladder epithelium
Authors: Kitchens D. L. , Dawson K. , Reuss L.
Published: December 2017

The effect of Zn2+ on Cl- transport across the apical membrane of Necturus gallbladder epithelium was studied with intracellular conventional and Cl(-)-selective microelectrodes and measurements of apparent base secretion. Most studies were done on tissues incubated in HEPES-buffered solutions; intracellular adenosine 3',5'-cyclic monophosphate (cAMP) levels were elevated by adding to the serosal bathing medium either theophylline or dibutyryl cAMP. Under these conditions, Zn2+ (added to mucosal solution) had no effect on membrane voltages, apparent cell membrane resistance ratio, or rapid depolarization induced by reducing mucosal solution [Cl-]. However, Zn2+ reduced the rate of cell membrane repolarization during exposure to the low-Cl- solution and decreased significantly the rate of fall of intracellular Cl- activity (alpha Cli) elicited by lowering mucosal solution [Cl-]. Both effects were time dependent, became significant after 10 min, and were slowly reversible. In tissues not stimulated by cAMP and incubated in a HCO3-CO2-buffered solution, Zn2+ also reduced the rate of fall of alpha Cli on lowering mucosal solution [Cl-]. Base secretion from cells to mucosal solution was assessed from changes in mucosal pH on stopping superfusion with a poorly buffered (1 mM HEPES) medium in the presence of 1 mM amiloride or a Na(+)-free medium, without cAMP stimulation. Exposure to Zn2+ reduced the alkalinization observed with both protocols. We conclude that Zn2+ has no effect on apical membrane Cl- conductance stimulated by cAMP and inhibits Cl(-)-HCO3- exchange. The slow onset and reversal of the effects suggests slow binding of Zn2+, a covalent modification of the exchanger, or an effect requiring Zn2+ transport to the cell interior.
Ontogenetic development of nutrient transporters in bullfrog intestine
Authors: Toloza E. M. , Diamond J. M.
Published: December 2017

The ratio of intestinal glucose (Glc) to amino acid (AA) transporter activity rises with age in animals of species in which the ratio of dietary carbohydrate to protein also rises. Does this shift function to match transporter activity to current dietary substrate levels or to the high AA requirements of young growing animals? We compared intestinal brush-border uptake of Glc and the AA proline (Pro) in adult and tadpole bullfrogs, since with age this species changes from an herbivore to a carnivore and hence its dietary carbohydrate-to-protein ratio decreases rather than increases. Like typical adult herbivores, tadpoles have a long, highly coiled, narrow-bore, thin-walled intestine with a long, heavy colon, whereas adult bullfrogs have a short, wide-bore, thick-walled intestine typical of carnivores. The ratio of Glc to Pro uptake capacity is essentially the same in adults as in larval tadpoles and is typical of carnivores but unprecedently low for herbivores. This suggests that both functions proposed for developmental shifts of Glc and AA transport in other species are significant and that their effects cancel each other in bullfrogs. Transporter kinetic constants vary predictably with developmental stage. Intestinal daily uptake capacity is comparable to actual daily intake for Glc in adults and for Pro in both tadpoles and adults but is much lower than intake for Glc in tadpoles, probably because most carbohydrate ingested by tadpoles is in the form of high molecular weight polymers that are not utilized.
Transit of solids through the human colon: regional quantification in the unprepared bowel
Authors: Proano M. , Camilleri M. , Phillips S. F. , Brown M. L. , Thomforde G. M.
Published: December 2017

We used a noninvasive method to label the solid phase of contents in the unprepared human colon. 111In-labeled Amberlite pellets (0.5-1.8 mm diam) were placed in a gelatin capsule that was then coated with a pH-sensitive polymer (methacrylate). In vitro, the capsules disintegrated in simulated small bowel contents within 1-2 h; when ingested by healthy subjects, capsules released radiolabel in the distal ileum or proximal colon in 13 of 15 subjects. Transit of 111In-pellets through the unprepared colon could then be quantitated radioscintigraphically. Segmental transit was defined in the ascending (AC), transverse (TC), descending (DC), and rectosigmoid (RS) colon. Radioactivity was also quantitated in stools. At 12 h, radioactivity was most obvious in the AC (59 +/- 11%, mean +/- SE) and the TC (21 +/- 6%); at 24 h, counts were distributed equally between AC, TC, and stools (P greater than 0.05); by 48 h, 56 +/- 11% counts had been excreted, although 30 +/- 10% remained in the TC. At 24 and 48 h, the amount in DC or RS was lower (P less than 0.05) than in the TC or in stools. Emptying of the AC was characterized by an initial lag period, when no counts emptied into the TC, followed by a period of emptying that was approximately linear. Thus this simple approach is able to label contents in the healthy human colon. The ascending and transverse colon appear to be sites of storage of solid residue, whereas the left colon and rectosigmoid function mainly as conduits.
Effects of antral vagotomy in dogs on gastrin and gastric secretion with various stimuli
Authors: Hirschowitz B. I. , Fong J.
Published: December 2017

In four gastric-fistula dogs, selective antral vagotomy markedly reduced the vagal stimulation of gastrin release, thereby defining both the vagal pathway for stimulation of gastrin and the anatomic source of such gastrin release. Despite loss of gastrin response, vagal excitation by 100 mg/kg 2-deoxy-D-glucose (2-DG) produced the same acid and pepsin responses after antral vagotomy as before, but there was an approximately 40% diminished fundic response to pentagastrin, histamine, and synthetic human gastrin, as well as to endogenous gastrin released by graded doses of bombesin. Bethanechol did not reverse the defect, ruling out inadvertent fundic vagal denervation, nor did raising serum gastrin by bombesin alter the response to vagal stimulation by 2-DG. Fundic response to bethanechol was increased by approximately 60%, and the output of gastrin increased at least fivefold after antral vagotomy. Gastrin responses to food were diminished and those to sham feeding were eliminated. Separation of the denervated antral pouch had no additional effect on acid secretion. Vagal stimulation of gastric secretion thus occurs almost exclusively through direct cholinergic effects on the fundus with little or no contribution from antral gastrin. Vagal denervation sensitizes the antrum to cholinergic stimulation.
Regulation of ornithine decarboxylase activity in LoVo cells
Authors: McCormack S. A. , Tague L. L. , Gragoe E. J. , Johnson L. R.
Published: December 2017

The role of Na+ and Na(+)-H+ exchange in the stimulation of ornithine decarboxylase (ODC) activity has been investigated in a human colon adenocarcinoma cell line, LoVo. Asparagine (Asn; 10 mM) or 10% fetal bovine serum (FBS) increased ODC activity from undetectable levels to greater than 500 pmol CO2.mg protein-1.h-1 in 4 h. This increase could be reduced 50% by concentrations of Na(+)-H+ exchange inhibitors that did not reduce protein synthesis. (approximately 0.2 mM for amiloride and 0.05 mM for hexamethyleneamiloride). Asn was able to double the uptake of 22Na+, whether an ionic (choline chloride) or nonionic (D-mannitol) substance was substituted for Na+, and the substitution of these compounds as well as N-methyl-glucamine for Na+ largely prevented the stimulation of ODC by Asn. Another factor influencing ODC activity was extracellular pH (pHo). When pHo was lowered, intracellular pH (pHi) also fell, and ODC activity was reduced. When pHo was raised, pHi also rose, and ODC activity increased. The well-known correlation between increased pHi and Na+ uptake with the stimulation of growth may be due to their influence on ODC activity.
Inhibition of GSH efflux from rat liver by methionine: effects of GSH synthesis in cells and perfused organ
Authors: Fernandez-Checa J. C. , Maddatu T. , Ookhtens M. , Kaplowitz N.
Published: December 2017

The inhibition of efflux of intracellular reduced glutathione (GSH) by methionine was determined in isolated rat hepatocytes suspended either in Krebs-Henseleit buffer or in modified Fisher's medium. Methionine (1 mM) added to Krebs-Henseleit suspensions of isolated rat hepatocytes inhibited GSH efflux, with greater retention of GSH in the cells compared with control. Results were similar with methionine and 0.3 mM propargylglycine cystathionase inhibitor), suggesting no net synthesis of GSH from methionine. In Fisher's medium, the inhibitory effect of methionine on GSH efflux was masked due to increasing cellular GSH; however, the inhibitory effect of methionine was unmasked by propargylglycine, which prevented the utilization of methionine for GSH synthesis. The addition of serine (0.1 mM) to methionine in Krebs-Henseleit buffer raised cellular GSH, overcoming the inhibition of GSH efflux. In the perfused liver, infusion of 1 and 5 mM methionine initially inhibited GSH efflux, but the inhibition was reversed with continued methionine infusion. After removal of methionine, GSH efflux increased immediately. The reversal and rebound were blocked by propargylglycine, revealing concentration-dependent inhibition of sinusoidal GSH efflux by methionine. Thus, when methionine is utilized to promote GSH synthesis, its inhibitory effect on GSH efflux tends to be overcome.
Synthesis and secretion of somatostatin-28 and -14 by fetal rat intestinal cells in culture
Authors: Brubaker P. L. , Drucker D. J. , Greenberg G. R.
Published: December 2017

A fetal rat intestinal cell (FRIC) culture model was established to investigate the factors controlling synthesis and secretion of somatostatin-28 (S-28) by the intestine. Immunohistochemical analysis demonstrated the presence of cells containing somatostatin-like immunoreactivity (SLI), many of which emitted long processes toward neighboring cells. Gel chromatography of SLI stored and secreted by nonstimulated FRIC cultures demonstrated a predominance of S-28 (59%), lesser amounts of S-14 (31%), and a minor peak of large molecular weight SLI (10%). Secretion of SLI was stimulated by treatment of cells for 2 h with 5 mM dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP; P less than 0.01) or 2 microM phorbol ester (P less than 0.05), but was unaffected by the calcium ionophore A23187 (2 micrograms/ml). Concomitant treatment with all three agents increased SLI secretion in an additive fashion to 254 +/- 25% of controls (P less than 0.0001). DBcAMP treatment did not alter the distribution of stored or secreted S-28 (59%) and S-14 (33%). After 24 h of exposure to DBcAMP, but not after treatment with phorbol ester, a threefold increment in prosomatostatin mRNA transcript levels was observed. FRIC cultures therefore synthesize and secrete a predominance of S-28 in a regulated manner, making them a promising model for future studies on the biosynthesis and secretion of intestinal somatostatin.
Impaired gastric acid secretion in mast cell-deficient mice
Authors: Stechschulte D. J. , Morris D. C. , Jilka R. L. , Stechschulte D. J. , Dileepan K. N.
Published: December 2017

Gastric acid secretion in normal (+/+) C57B1/6J mice and congeneic, mast cell-deficient (mi/mi) C57B1/6J mice was examined. The mast cell-deficient animals had approximately 50% of the normal quantity of gastric histamine and a blunted basal acid level and secretory response. These observations were noted despite the presence of parietal cells, which were normal in number and morphology. The H2-antagonist ranitidine inhibited basal acid secretion in both groups of animals. Exogenous histamine induced a significant secretory response in normal and mast cell-deficient groups, but only the secretory response in normal animals could be blocked by the H2-antagonist. Treatment of mast cell-deficient animals with histamine for seven consecutive days before stimulation did not restore the histamine response to the normal (+/+) levels. The normal animals demonstrated an acid secretory response to pentagastrin. Mast cell-deficient mice also responded to pentagastrin, but the response was less than that observed in the normal animals, and a significant difference was not evident in all experiments. Furthermore, simultaneous injection of mast cell-deficient animals with histamine and pentagastrin did not restore pentagastrin responsiveness to normal levels, although the histamine concentration used was sufficient to raise acid secretion to basal levels of normal mice. These results support the conclusion that non-mast cell histamine only partially contributes to basal gastric acid secretion and is insufficient to facilitate full parietal cell responsiveness. Furthermore, pentagastrin requires the presence of mast cells to elicit a maximal secretory response but can use non-mast cell histamine to activate the parietal cells for acid secretion.
Effects of hypovolemia on blood flow, arterial [HCO3-], and HCO3- output in the rat duodenum
Authors: Jonson C. , Holm L. , Jansson T. , Fandriks L.
Published: December 2017

The effects of bleeding-induced hypovolemia on duodenal blood flow (microsphere technique), arterial [HCO3-], and duodenal HCO3- secretion (in situ titration) were investigated in chloralose-anesthetized rats. A 10% decrease in blood volume reduced duodenal HCO3- secretion by 44%, duodenal blood flow by 31%, and arterial [HCO3-] by 11%. In a group with cervically cut vagal nerves, basal duodenal HCO3- secretion was greater than 50% lower compared with controls. Basal blood flow and arterial [HCO3-] were on similar levels as in nonvagotomized animals. Furthermore, bleeding failed to lower duodenal alkaline output in rats with cut vagal nerves, although blood flow and arterial [HCO3-] were reduced to a similar extent as in the vagally intact controls. In a yohimbine-treated group, a 10% bleeding reduced duodenal blood flow by 28% and arterial [HCO3-] by 7% without influencing duodenal HCO3- secretion. We suggest that the hypovolemia-induced inhibition of duodenal alkaline secretion is not caused by a decrease in blood and/or arterial [HCO3-]. Instead, other factors may be of importance, for example, neural effects on enteric secretomotor neurons or directly on the secreting epithelium.
Determinants of upper esophageal sphincter pressure in dogs
Authors: Jacob P. , Kahrilas P. J. , Herzon G. , McLaughlin B.
Published: December 2017

Chronic experiments were done on six dogs fitted with EMG electrodes on pharyngeal and esophageal musculature. Electromyographic activity of the cricopharyngeus was recorded in awake and sedated animals with and without manometric recordings as well as during esophageal distension. Intraluminal upper esophageal sphincter (UES) pressure had two distinct components; active contraction accompanied by cricopharyngeal EMG activity and passive elasticity that persisted in the absence of EMG activity. Between swallows, the cricopharyngeal EMG activity patterns observed were of either tonic activity, no activity, or phasic activity with inspiratory bursts. The activity level was markedly affected by anesthesia, phonating, whining, panting, level of alertness, or changes in head posture. A brisk UES contraction was elicited in response to passage of the manometric assembly and to intraesophageal balloon distension. Persistent EMG augmentation after stationing of the manometric sensor suggested that intraluminal manometry tends to exaggerate resting sphincter pressure. We conclude that electrical activity of the cricopharyngeus, and by inference UES pressure, is markedly affected by many variables that are difficult to control during clinical or experimental determinations of UES pressure.
Na(+)-H+ exchange in basolateral plasma membrane vesicles from neonatal rat liver
Authors: Goodrich A. L. , Suchy F. J.
Published: December 2017

Basolateral plasma membrane vesicles (BLMV) were used to determine the activity of the Na(+)-H+ exchanger in neonatal compared with adult rat liver. Imposition of an outwardly directed H+ gradient (pH 5.5in/7.5out) markedly stimulated the uptake of 1 mM 22Na+ by adult and neonatal BLMV above rates observed under pH-equilibrated conditions (pH 7.5in/7.5out) and resulted in a transient uphill accumulation ("overshoot") of Na+ to levels 1.5- to 2-fold higher than at equilibrium. 22Na+ uptake via Na(+)-H+ exchange (identified as the amiloride-sensitive component of total Na+ flux) was greater in BLMV from neonatal compared with adult liver under both pH-gradient and -equilibrated conditions. These age-related differences in Na+ uptake could not be explained by membrane potential effects or by differences in membrane permeability to H+. Kinetic analysis revealed a saturable process with a similar Km for Na+ (8.6 +/- 1.6 vs. 8.1 +/- 1.5 mM) but with a 2.5-fold higher Vmax (8.06 +/- 0.67 vs. 3.20 +/- 0.40 nmol.mg protein-1.min-1; P less than 0.01) in neonatal vs. adult vesicles. We conclude from these studies that basolateral membrane vesicles from neonatal rat liver exhibit enhanced Na(+)-H+ exchange as a result of an increased number or translocation rate of carriers in the plasma membrane.
Molecular forms of cholecystokinin in rat intestine
Authors: Turkelson C. M. , Solomon T. E.
Published: December 2017

The predominant immunoreactive cholecystokinin (CCK) forms in acid extracts of rat intestine eluted from reverse-phase high-performance liquid chromatography columns in the positions of CCK-8, CCK-33/39, and CCK-58. Control experiments indicated that smaller CCK forms did not arise from artifactual degradation of large CCK forms. Less than 10% of CCK-8 added to and extracted from intestine was recovered in acid; CCK-8 could not be recovered by subsequent alkaline extraction, but subsequent urea extraction yielded approximately 25% of the added peptide. This suggests that CCK binds to proteins during acid extraction and that the preponderance of large CCK forms in acid extracts is not due to inhibition of CCK degradation but results from poor extraction of small CCK forms. No evidence for a CCK-22-like peptide was found in acid or subsequent urea extracts of rat intestine, suggesting CCK posttranslational processing in adult rats is like that in humans and dogs.