bluuebunny/crossref_metadata_2025 huggingface dataset:
Bile secretory function of intrahepatic biliary epithelium in the rat
Authors: Alpini G. , Lenzi R. , Zhai W. R. , Slott P. A. , Liu M. H. , Sarkozi L. , Tavoloni N.
Published: December 2017

To shed light on ductular fluid secretion, hepatic histology and ultrastructure, cell proliferation and phenotypes, and several aspects of biliary physiology were studied in rats with ductular cell hyperplasia induced by either biliary obstruction (0-14 days) or 1-naphthylisothiocyanate (ANIT) feeding (0-28 days). In both groups of experimental animals, bile duct hyperplasia and spontaneous bile flow and secretin-induced choleresis increased with time of treatment in a linear fashion. Measurements of [14C]mannitol biliary entry and of biliary tree volume showed that the increase in both spontaneous and secretin-stimulated bile flow originated at the proliferated biliary structures. Ultrastructural examination, [3H]thymidine incorporation, and histochemical and immunohistochemical staining for various markers demonstrated that in both hyperplastic reactions the proliferated cells were the progeny of preexisting biliary epithelial cells and retained their characteristics. These results indicate that the increased bile secretory activity associated with either biliary obstruction or ANIT intoxication reflects a quantitative change due to the proliferation of biliary epithelial cells. Thus both models of bile ductular cell hyperplasia lend themselves to assessment of the transport function of intrahepatic biliary epithelium and its contribution to normal bile formation. In the present studies, we have estimated that net ductular secretion in the normal rat accounts for 10-13% of spontaneously secreted hepatic bile.
Differential action of somatostatin on peptide-induced release of acetylcholine
Authors: Kowal V. , Wiley J. W. , Owyang C.
Published: December 2017

In this study we examine the mechanism by which somatostatin (SRIF-14) inhibits cholecystokinin octapeptide- (CCK-8) but not substance P-mediated release of [3H]acetylcholine (ACh) from the guinea pig ileum. 2',5'-Dideoxyadenosine, an inhibitor of adenylate cyclase, antagonized the action of CCK-8 and forskolin but had no effect on substance-P-evoked release of [3H]ACh. Addition of theophylline enhanced the release of [3H]ACh stimulated by CCK-8 but not by substance P. These observations suggest that CCK-8, but not substance P, can stimulate cholinergic transmission via an adenosine 3',5'-cyclic monophosphate (cAMP)-dependent pathway. Somatostatin inhibited release of [3H]ACh evoked by CCK-8 and forskolin in a dose-related manner. CCK-8- and forskolin- but not substance P-evoked release of [3H]ACh were maximally inhibited in the presence of 10(-6) M somatostatin (49 +/- 5 and 48 +/- 7% of control, respectively). Pretreatment with pertussis toxin (inactivates inhibitory guanine nucleotide binding proteins) reversed the inhibitory effect of somatostatin on the release of [3H]ACh evoked by CCK-8. These observations suggest that CCK-8 but not substance P can stimulate [3H]ACh by a cAMP-dependent pathway. Somatostatin appears to inhibit the cAMP-dependent component of CCK-8-mediated cholinergic transmission via activation of a pertussis toxin-sensitive G protein.
Relationships between gastric emptying of solid and caloric liquid meals and alcohol absorption
Authors: Horowitz M. , Maddox A. , Bochner M. , Wishart J. , Bratasiuk R. , Collins P. , Shearman D.
Published: December 2017

The effects of three variations in meal composition (a solid and a liquid meal consumed together, a liquid meal consumed alone, and a liquid meal consumed 90 min after a solid meal) on the rates and patterns of solid and liquid gastric emptying were examined in 13 volunteers. By including alcohol (0.5 g/kg body wt) in the liquid meal, the relationship between alcohol absorption and gastric emptying was also assessed. The lag phase and the initial emptying phase of the solid meal were prolonged (P less than 0.001) when the liquid meal was consumed with the solid meal, compared with when the liquid meal was consumed 90 min after the solid meal. In this latter situation, consumption of the liquid meal caused the cessation of emptying of solid food, and this second lag phase was followed by a slower (P less than 0.001) than initial emptying phase. Gastric emptying of the liquid meal was slower (P less than 0.005) when solid food was present and was slowest (P less than 0.05) when liquid was consumed 90 min after the solid meal. Alcohol absorption was fastest (P less than 0.05) when the liquid meal was consumed alone and slower (P less than 0.01) when alcohol was consumed with or after the solid meal. For all three meals there was a close correlation (r greater than or equal to 0.91; P less than 0.001) between alcohol absorption and liquid emptying. We conclude that gastric emptying of liquid may be influenced by solid food and that the rate and pattern of solid emptying may be modified by the presence of liquid.(ABSTRACT TRUNCATED AT 250 WORDS)
pH regulation in hepatoma cells: roles for Na-H exchange, Cl-HCO3 exchange, and Na-HCO3 cotransport
Authors: Weintraub W. H. , Machen T. E.
Published: December 2017

Regulation of intracellular pH (pHi) was studied in Fu5, a rat hepatoma cell line that maintains a variety of differentiated functions. Microspectrofluorimetry of the pH-sensitive dye 2',7'-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF) was used to measure pHi in 10-15 cells growing on cover glasses that were mounted in a flow-through chamber on the stage of a microscope. In N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered solutions, pHi was 7.14, and intrinsic buffer capacity was inversely related to pHi. Amiloride (0.1 mM) caused pHi to decrease by 0.33 pH units in 4 min. Recovery from an acid load (using either NH4 prepulse technique or Na-free solutions) was completely blocked by amiloride. In HCO3-CO2-buffered solutions, pHi was 7.15, and buffer capacity was relatively insensitive to pHi between pHi of 6.6 and 7.2. Amiloride caused pHi to decrease by only 0.09 units. Recovery from an acid load was Na dependent, occurred in Cl-free solutions, and was totally blocked by the combination of amiloride plus 0.5 mM dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (H2DIDS); recovery occurred when either amiloride or H2DIDS was removed. Removal of external Cl caused a rapid, H2DIDS-blockable alkalinization that was faster in HCO3-CO2 than in HEPES. The apparent Km for Clout for relaxation of Cl-free alkalinization was 4.5 mM. Rate of HCO3 transport during Cl-free treatment increased at alkaline resting pHi. It is concluded that Fu5 cells have two Na-dependent base-loading mechanisms and an acid-loading Cl-HCO3 exchanger. In solutions containing HCO3-CO2, the Na-H exchanger accounts for approximately 40% of recovery from an acid load, and a Na-HCO3 cotransporter accounts for the remainder. Recovery from an alkaline load appears to occur through the activity of the Cl-HCO3 exchanger.
Gastric and pancreatic intrinsic factor-mediated absorption of cobalamin in the dog
Authors: Batt R. M. , Horadagoda N. U.
Published: December 2017

The effects of canine gastric and pancreatic intrinsic factors on uptake and subcellular localization of cobalamin have been investigated in vivo to determine whether these proteins could mediate the physiological absorption of cobalamin in the dog. Cyano [57Co]cobalamin was introduced into ileal loops in dogs under general anesthesia, either free (control) or bound to gastric or pancreatic intrinsic factor. At 2 h, total uptake of cobalamin by ileal mucosa was significantly enhanced after prior binding to either gastric or pancreatic intrinsic factor compared with controls. Displacement of receptor-bound cobalamin with EDTA showed that enhanced total uptake reflected increased internalization of cobalamin by both proteins. Findings after reorienting sucrose density gradient centrifugation of ileal mucosa from loops containing intrinsic factor-cobalamin complexes were consistent with a major lysosomal and perhaps endosomal localization of internalized cobalamin, in agreement with results after oral administration of cobalamin. In marked contrast, cobalamin was recovered predominantly in the soluble fractions and was not associated with particulate subcellular organelles in ileal mucosa from control loops. These findings suggest that both gastric and pancreatic intrinsic factors can promote the physiological absorption of cobalamin by receptor-mediated endocytosis in the dog.
Effect of aluminum on bidirectional calcium flux in rat everted intestinal sacs
Authors: Adler A. J. , Zara C. , Berlyne G. M.
Published: December 2017

The effect of aluminum on intestinal calcium absorption was determined in male Sprague-Dawley rats using an everted intestinal sac technique. Bidirectional calcium flux in the duodena and ilea of normal rats was assessed by means of dual calcium isotopes. Two micromolar aluminum significantly inhibited net calcium absorption (J net) in the duodenum through suppression of mucosato-serosa flux (J m----s). Jm----s was reduced from 2.21 +/- 0.50 mumol Ca.h-1.g wet wt-1 in controls to 0.93 +/- 0.35 mumol Ca.h-1.g-1 in aluminum exposed sacs, and Jnet was reduced from 1.88 +/- 0.14 mumol Ca.h-1.g-1 to 0.55 +/- 0.41 mumol Ca.h-1.g-1 (P less than 0.001). Serosa-to-mucosa calcium flux (Js----m) was not similarly influenced by aluminum. Inhibition of Jm----s occurred whether aluminum was initially present on the mucosal or serosal side of the duodenal sac and inhibition of Jnet calcium by 2 muM A1 occurred at all ambient concentrations of calcium studied. In the ileum, aluminum had no effect on any component of calcium flux. Aluminum did not induce any suppression of glucose transport in either the duodenum or ileum, suggesting that the effect on calcium transport is relatively specific. These results suggest that aluminum inhibits calcium absorption in the duodenum through an effect on active mucosa-to-serosa transport, but has no effect on ileal calcium absorption, which in the rat is not mediated by an active process.
Cholinergic control of smooth muscle peristalsis in the cat esophagus
Authors: Blank E. L. , Greenwood B. , Dodds W. J.
Published: December 2017

The aim of this study was to examine in detail the effects of selective cholinergic and other pharmacological antagonists on primary and secondary peristalsis in the smooth muscle of the cat esophagus in order to fully characterize the cholinergic contribution to peristalsis in this species. Primary and secondary peristalsis in the smooth muscle part of the feline esophagus was completely abolished by atropine, 4-dephenylacetoxy-N-methylpiperidine methiodide (4-DAMP) (a selective M2 muscarinic antagonist), hexamethonium, and high doses of nicotine. Pirenzepine (a selective M1 muscarinic antagonist), propranolol, and phentolamine were without effect, as were naloxone, methysergide, and pyrilamine. From these findings we conclude that primary and secondary peristalsis in feline esophageal smooth muscle involves nicotinic ganglionic neurotransmission as well as postganglionic release of acetylcholine that acts directly on muscarinic receptors located on the smooth muscle. Peristalsis in esophageal striated muscle does not involve either synaptic transmission or muscarinic receptors.
Effects of endotoxin on iron uptake by the hepatocyte
Authors: Potter B. J. , Blades B. , McHugh T. A. , Nunes R. M. , Beloqui O. , Slott P. A. , Rand J. H.
Published: December 2017

Administration of endotoxins is often followed within 12-24 h by marked hypoferremia. Because the hepatocyte is the major site of both iron storage and transferrin synthesis, we have investigated the effects of an Escherichia coli endotoxin (lipopolysaccharide, LPS) on these parameters on isolated hepatocytes from normal Wistar rats (ND), rats previously treated intraperitoneally with 2.5 mg/kg (LD) or 25 mg/kg (HD) LPS, and control rats injected intraperitoneally with sterile saline (CD). No effects were observed on iron uptake from transferrin by ND cells incubated in vitro with up to 350 micrograms LPS/10(7) hepatocytes. There was also no significant difference in iron uptake between CD, HD, and LD hepatocytes 1 h after LPS injection. However, hepatocytes isolated 24 h after LPS administration took up iron significantly faster than controls. The uptake of non-transferrin-bound iron was also increased in HD and LD hepatocytes at 24 h but only in HD cells at 1 h. Transferrin binding was not altered in LPS-treated cells from ND rats but was depressed in cells from LPS-treated rats both at 1 h and at 24 h after injection. Transferrin receptor recycling was significantly increased at 24 h in cells from both LD and HD rats. Transferrin and total protein synthesis were also depressed at 1 h in LPS-treated rats, returning to normal values at 24 h. Direct preincubation of ND cells, however, failed to increase synthesis except at the highest concentrations of LPS. We conclude that LPS has an immediate (although indirect) effect on protein synthesis by the hepatocyte but not on iron uptake.(ABSTRACT TRUNCATED AT 250 WORDS)
Effect of cholecystokinin on the accumulation of inositol phosphates in isolated pancreatic islets
Authors: Florholmen J. , Malm D. , Vonen B. , Burhol P. G.
Published: December 2017

Sulfated cholecystokinin octapeptide (CCK-8S) potentiated glucose-induced secretion in isolated pancreatic islets with a maximal effect at 12 mM glucose, whereas no effect was observed at 3.3 and 25 mM glucose. This effect of CCK-8S was maximal at 10(-7) M. Anion-exchange fast-protein liquid chromatography analysis of [3H]inositol phosphates derived from islets prelabeled with myo-[3H]inositol showed that glucose induced accumulation of the 1,4,5-isomer of inositol trisphosphate and of inositol tetrakisphosphate. At 3.3 mM glucose, CCK-8S stimulated accumulation of inositol trisphosphate and inositol tetrakisphosphate to levels induced by 25 mM glucose alone. The net effect of CCK-8S on the accumulation of the inositol phosphates was maximal at 12 mM glucose and decreased at higher glucose concentrations. At 12 mM glucose the accumulation of inositol phosphates increased gradually up to 10(-7) M CCK-8S. This study indicates that CCK-8S potentiates glucose-induced insulin secretion through a mechanism involving the hydrolysis of polyphosphoinositides and the generation of inositol phosphates. However, activation of the inositol cycle per se did not seem to induce insulin secretion.
Altered role of microtubules in asialoglycoprotein trafficking in developing liver
Authors: Kaufman S. S. , Blain P. L. , Park J. H. , Tuma D. J.
Published: December 2017

Efficient receptor-mediated endocytosis of asialoglycoprotein by mature liver requires participation of microtubules that convey newly internalized ligand to lysosomes for degradation and receptor back to plasma membrane to continue endocytosis. To ascertain whether microtubular participation in asialoglycoprotein endocytosis is altered during development, we compared endocytosis of 125I-labeled asialoorosomucoid (ASOR) in neonatal rat hepatocytes to that in adult cells, with and without microtubular disruption by colchicine. Control experiments demonstrated that 125I-ASOR degradation in neonatal hepatocytes occurred at 70% of the adult rate during continuous endocytosis, although neonatal surface receptors were only approximately 40% as numerous. Colchicine disruption of microtubules reduced 125I-ASOR degradation and steady-state intracellular ASOR more in adults during continuous endocytosis. Degradation of 125I-ASOR prebound to surface receptors was equally impaired by colchicine in the two groups. Continuous ASOR endocytosis by colchicine-treated adult hepatocytes progressively depleted their surface receptors but minimally in neonates. Unlike colchicine, the protonophore monensin markedly impaired receptor recycling as well as postinternalization ligand trafficking in both neonates and adults. Thus these experiments demonstrate that asialoglycoprotein processing proceeds as efficiently in neonatal as in adult hepatocytes despite a reduced surface receptor population. Microtubules are required to maintain receptors on cell surface as well as for postinternalization trafficking in adult cells. During development, only the latter process substantially requires microtubules, indicating that microtubular participation in protein trafficking is selectively, not uniformly, diminished at this time in life.
Role of neuropeptide Y in opossum internal anal sphincter
Authors: Nurko S. , Rattan S.
Published: December 2017

The present investigation was undertaken to examine the effects and role of neuropeptide Y (NPY) on the internal anal sphincter (IAS). The studies were performed in alpha-chloralose-anesthetized opossums. Resting pressure in the IAS (IASP) was recorded using low-compliant continuously perfused catheters. The effect of NPY administered close intra-arterially on the resting IASP was examined before and after different neurohumoral antagonists and the neurotoxin tetrodotoxin (TTX). To examine the influence of sympathetic nerves, the ventral roots of lumbar sympathetic nerves were isolated in some experiments and stimulated electrically. The influence of NPY on the IAS relaxation in response to rectoanal reflex mimicked by rectal balloon distension (RD) was also examined. NPY caused dose-dependent rise in the IASP, which was not modified by different neurohumoral antagonists and TTX. The rise in IASP by electrical stimulation of ventral lumbar nerve root (L3) was suppressed but not abolished by phentolamine. However, guanethidine (which depletes both norepinephrine and NPY) caused complete obliteration of this response. The other effect of NPY on the IAS was significant antagonism of IAS relaxation caused by RD. From these results, we conclude that NPY exerts two different actions on the IAS. First, it causes a rise in IASP by its action directly at the IAS smooth muscle. Sympathetic nerve stimulation may cause a rise in IASP by its action directly at the IAS smooth muscle partially through release of NPY. Second, it inhibits IAS relaxation produced by rectoanal reflex. These findings would suggest an important regulatory role of NPY in rectoanal physiology.
Modulation of feline esophageal contractions by bolus volume and outflow obstruction
Authors: Mittal R. K. , Ren J. , McCallum R. W. , Shaffer H. A. , Sluss J.
Published: December 2017

We studied the effect of bolus volume and esophageal obstruction on esophageal peristalsis by using synchronized video-fluoroscopic and manometric techniques in cats. A specially designed pressure cuff was surgically implanted around the distal esophagus to control the degrees of outflow obstruction. Secondary esophageal peristalsis was evoked by injecting bolus volumes of 3, 6, and 9 ml at cuff pressures of 0, 20, 40, and 60 mmHg. Increases in outflow obstruction reduced the velocity of peristalsis. The amplitude of esophageal contraction increased with increasing outflow obstruction at low bolus volumes but decreased with larger bolus volumes and larger outflow obstruction. In the absence of outflow obstruction, each esophageal contraction traversed the entire esophagus distal to its site of origin, but in the presence of outflow obstruction contractions only traversed part of the esophagus. The incidence and site of failure of propagation was directly related to cuff pressure and bolus volume. The relationship between the onset of manometric pressure complex at a given site in the esophagus to the passage of the bolus from that esophageal site was markedly affected by outflow obstruction. We conclude that esophageal peristalsis can be modulated by the bolus volume and outflow obstruction.
Multiple pathways for the regulation of ornithine decarboxylase in intestinal epithelial cells
Authors: Ginty D. D. , Marlowe M. , Pekala P. H. , Seidel E. R.
Published: December 2017

The regulation of ornithine decarboxylase (ODC) was examined in an intestinal epithelial crypt cell line (IEC-6). Addition of fetal bovine serum or growth factors to quiescent preconfluent cells resulted in a 20- to 30-fold increase in the specific activity of ODC, which was maximal at approximately 4 h. In contrast, ODC mRNA levels either did not change or increased only twofold over the time period examined. The increased enzymatic activity was blocked by cycloheximide, putrescine, and the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-napthalinesulfonamide (W-7). Cycloheximide alone increased mRNA levels and potentiated the induction in response to serum, suggesting that protein synthesis is not required for the increase in mRNA accumulation. In contrast to its effect on ODC activity, W-7 was without effect on the serum-stimulated increase in ODC or c-fos mRNA levels. Putrescine decreased ODC activity, but not mRNA content, in a dose-dependent manner with an IC50 between 0.1 and 1.0 microM. Also, serum stimulation resulted in a threefold increase in the stability of the enzyme in the presence of cycloheximide; this effect was blocked by pretreatment with W-7. Enzymatic activity was paralleled by ODC protein content as determined by [3H] difluoromethylornithine binding. Finally, the induction of enzyme activity was due entirely to an increase in Vmax as no detectable change in Km for ornithine was detected. These results suggest that ODC is regulated at multiple levels by independent signaling pathways in cultured intestinal epithelial cells. Increased levels of active ODC protein and enzymatic activity are sensitive to W-7 and putrescine.(ABSTRACT TRUNCATED AT 250 WORDS)
Effects of epidermal growth factor on signal transduction in rabbit parietal cells
Authors: Lewis J. J. , Goldenring J. R. , Asher V. A. , Modlin I. M.
Published: December 2017

The aim of this study was to elucidate the cellular mechanisms of action of epidermal growth factor (EGF) inhibition of parietal cell secretion. EGF effects on histamine- and carbachol-stimulated [14C]aminopyrine (AP) uptake and intrinsic factor (IF) secretion were evaluated in isolated rabbit parietal cells. EGF inhibited histamine-stimulated [14C]AP uptake and IF secretion through a reduction in stimulated adenosine 3',5'-cyclic monophosphate (cAMP) levels. EGF decreased the phosphorylation of a cytosolic 30-kDa, histamine-stimulated, cAMP-dependent protein kinase substrate. These effects on histamine-stimulated activation were reversed by pertussis toxin preincubation. EGF inhibited carbachol-stimulated [14C]AP uptake and IF secretion, but did not alter the carbachol-stimulated Ca2+ transient. These results indicate that EGF inhibits histamine-stimulated secretion through the inhibitory Gi guanosine 5'-triphosphate-binding protein and carbachol-stimulated secretion through a mechanism independent of the activation of an increase in intracellular Ca2+.
Galanin inhibits gastrin release from isolated rat gastric G-cells
Authors: Schepp W. , Prinz C. , Tatge C. , Hakanson R. , Schusdziarra V. , Classen M.
Published: December 2017

Enzymatically isolated rat gastric mucosal cells (0.25% G-cells) were separated by counterflow elutriation, yielding a fraction in which the G-cell content was relatively enriched to 1.4%. In this fraction, basal gastrin release (mean +/- SE) was 31.1 +/- 1.3 pg.10(6) cells-1.60 min-1 and was stimulated by 10(-8) M neuromedin C (222.3 +/- 18.1% of basal), 10(-4) M carbachol (227.5 +/- 25.9%), 10(-6) M 12-O-tetradecanoylphorbol-13-acetate (TPA) (196.3 +/- 14.7%), and 10(-3) M dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP) (193.9 +/- 6.8%), respectively. The neuropeptide galanin was tested at 10(-10) to 10(-7) M. Galanin had no effect on basal gastrin release but reduced the responses to neuromedin C, carbachol, TPA, and DBcAMP. IC50 ranged between 1 X 10(-10) and 8.6 X 10(-10) M galanin. Although in the relatively enriched G-cell fraction D-cells were not detectable by immunocytochemistry, a low rate of somatostatin release was still measured by radio-immunoassay (5.3 +/- 0.5 pg.10(6) cells-1.60 min-1). However, galanin failed to increase this rate under basal conditions or in response to any of the stimulants tested. These results favor the assumption that galanin might exert a direct inhibitory effect on rat gastric G-cells. Galanin seems to interfere at an intracellular mechanism(s), which is induced by neuromedin C and carbachol and which is commonly activated by protein kinase C- and cAMP-mediated stimulation.
Effect of swallowed bolus variables on oral and pharyngeal phases of swallowing
Authors: Dantas R. O. , Kern M. K. , Massey B. T. , Dodds W. J. , Kahrilas P. J. , Brasseur J. G. , Cook I. J. , Lang I. M.
Published: December 2017

In this investigation, we studied the effects of bolus volume and viscosity on the quantitative features of the oral and pharyngeal phases of swallowing. Concurrent videofluoroscopic and manometric studies were done in 10 healthy volunteers who were imaged in lateral projection. Videofluorography was done at 30 frames/s while concurrent manometry was done with 5 intraluminal transducers that straddled the pharynx and upper esophageal sphincter (UES). Submental electromyography was recorded also. Swallows of 2-20 ml were recorded for low-viscosity liquid barium and high-viscosity paste barium. Analysis indicated that the major effect of increases in bolus volume was an earlier onset of anterior tongue base movement, superior palatal movement, anterior laryngeal movement, and UES opening. These events provide receptive adaptation for receiving a swallowed bolus. Earlier UES opening was associated with an increase in the duration of sphincter opening and sphincter diameter. The major effects of high bolus viscosity, unrelated to bolus volume, were to delay oral and pharyngeal bolus transit, increase the duration of pharyngeal peristaltic waves, and prolong and increase UES opening. Thus the specific effect of bolus viscosity per se differs substantially from that of bolus volume. We conclude that 1) specific variables of swallowing are affected significantly by the variables of the swallowed bolus, such as volume and viscosity; 2) overall, bolus volume and viscosity affect swallowing in a different manner; and 3) the study findings have implications about the neural control mechanisms that govern swallowing as well as about the diagnosis and treatment of patients with abnormal oral-pharyngeal swallowing.
Characterization of mucous cell synthetic functions in a new primary canine gastric mucous cell culture system
Authors: Boland C. R. , Kraus E. R. , Scheiman J. M. , Black C. , Deshmukh G. D. , Dobbins W. O.
Published: December 2017

Mucin is a critical component of the protective layer secreted by gastrointestinal mucous cells. A detailed understanding of the molecular processing of gastric mucin and the physiology of its secretion has been limited by the lack of an adequate model for their study. We have developed a primary culture system of canine gastric mucous cells that has permitted us to study their synthetic and secretory functions. It was found that [3H]glucosamine used for metabolic labeling studies was incorporated into both mucin and lipid components of gastric mucus. To measure mucin with this model, a new immunoassay was developed to quantitate canine gastric mucin. Mucin was purified from the canine stomach, a polyclonal antibody was generated, and an enzyme-linked immunosorbent assay for gastric mucin was established. Mast cells were frequent contaminants of the gastric mucous cell preparation, and two methods were developed to limit their contamination. A new culture system has been developed for the study of gastric mucous cells. These cells synthesize and secrete both mucin and phospholipids. This system will permit us to study the molecular processing of mucin and the physiology of its production and release.
Inhibition of the alpha-amidation of gastrin: effects on gastric acid secretion
Authors: Dickinson C. J. , Marino L. , Yamada T.
Published: December 2017

Formation of biologically active amidated gastrin from glycine-extended progastrin processing intermediates (G-Gly) is achieved via the action of peptidyl-glycyl alpha-amidating monooxygenase. Since this enzyme requires copper for optimal activity, we examined the effects of a known copper chelator, diethyldithiocarbamate (DDC), on gastrin posttranslational processing and gastric acid secretion in vivo. DDC (400 mg.kg-1.day-1 ip X 3 days) administered to male Sprague-Dawley rats decreased antral amidated gastrin content, but increased antral G-Gly content. The ratio of amidated gastrin to G-Gly, which reflects in situ amidating activity, was decreased in DDC-treated rats. In contrast, tissue amidating potential, assayed directly under optimal copper concentrations in vitro, was increased in the antrum and unchanged in the pituitary. DDC markedly increased both basal and gastrin-stimulated gastric acid outputs despite the presence of normal serum amidated gastrin levels. These results suggest that copper chelation with DDC inhibits amidating activity in situ but selectively increases antral amidating enzyme synthesis. The marked increase in acid secretion despite normal circulating amidated gastrin concentrations, combined with the enhanced secretory response to exogenously administered gastrin, suggests the possibility that gastrin receptors are upregulated by the events precipitated via DDC administration.
Influence of Cl- on pH(i) in oxynticopeptic cells of in vitro frog gastric mucosa
Authors: Yanaka A. , Carter K. J. , Lee H. H. , Silen W.
Published: December 2017

The effect of Cl- on intracellular pH (pH(i)) was studied using sheets of frog (Rana catesbeiana) fundic mucosa in which oxynticopeptic cells were selectively loaded with the acetomethoxy ester form of the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF/AM). Before the measurement of pH(i), tissues were exposed to either 10(-5) M forskolin in the serosal solution (stimulated tissues) or 3 x 10(-4) omeprazole in the serosal solution (inhibited tissues). In HCO3- and N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) buffers, pH(i) increased significantly after removal of Cl- from serosal and luminal solution, both in stimulated and inhibited tissues. The presence of Cl- in the luminal solution prevented this rise in pHi, an effect abolished by serosal 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 3 x 10(-4) M) but not by serosal amiloride (10(-3)M). In the presence of serosal Cl-, pH(i) increased after exposure to serosal DIDS, more prominently in the stimulated than in the inhibited tissues. These results confirm the presence of a Cl(-)-HCO3-exchanger in the basolateral membrane of oxynticopeptic cells in intact sheets of mucosa and suggest that luminal Cl- contributes to the regulation of pH(i) in oxynticopeptic cells.
Effect of acid-base balance on biliary bicarbonate secretion in the isolated perfused guinea pig liver
Authors: Blot-Chabaud M. , Dumont M. , Corbic M. , Erlinger S.
Published: December 2017

Secretin-induced choleresis is of ductal origin and involves bicarbonate transport. Its mechanism is unknown. To determine the relative effects of systemic pH, PCO2, and bicarbonate concentration on secretin-stimulated bicarbonate transport, states of acute metabolic and respiratory acidosis or alkalosis were created in isolated perfused guinea pig livers with or without secretin infusion. During spontaneous secretion conditions, biliary bicarbonate secretion was not correlated with perfusate pH (7.19-7.62) or perfusate PCO2 (23.9-59.7) but was significantly correlated with perfusate bicarbonate concentration (17.5-37.9 mM). Under secretion infusion (25 mU/min), bile flow and biliary bicarbonate concentration increased significantly (109 and 51%, respectively). Biliary bicarbonate secretion was not correlated with perfusate pH (7.19-7.60) but was significantly correlated both with perfusate bicarbonate concentration (14.6-36.8 mM) and PCO2 (25.8-54.3 mmHg). Spontaneous and secretin-induced bile flow were correlated with biliary bicarbonate concentration. The correlation between biliary bicarbonate secretion and PCO2 during secretin-induced choleresis supports the hypothesis that secretin-induced biliary bicarbonate secretion could, at least in part, involve a transport of H+ (or OH-) rather than HCO3- itself and that intracellular pH could play a role in the regulation of this secretion. Amiloride (5 X 10(-4) M) did not influence secretin-induced biliary bicarbonate secretion. This result suggests that the Na(+)-H+ exchange is not involved in bicarbonate secretion by ductular cells.
Na(+)-Ca2+ exchange in cultured rat hepatocytes: evidence against a role in cytosolic Ca2+ regulation or signaling
Authors: Lidofsky S. D. , Xie M. H. , Scharschmidt B. F.
Published: December 2017

Plasma membrane Na(+)-Ca2+ exchange contributes importantly to the regulation of cytosolic Ca2+ concentration ([Ca2+]i) in excitable cells. Despite extensive study in excitable tissues, the role of this transporter in the regulation of [Ca2+]i in hepatocytes is unknown, and conflicting information has been reported regarding the presence of Na(+)-Ca2+ exchange in hepatocyte plasma membrane vesicles. We have therefore assessed the role of Na(+)-dependent Ca2+ transport in the regulation of [Ca2+]i in rat hepatocytes in primary culture under basal conditions and after exposure to vasopressin, a hormone that elevates [Ca2+]i. Ca2+ efflux, measured using 45Ca, did not differ in the presence or absence of extracellular Na+, either under basal conditions or in response to vasopressin. [Ca2+]i, measured using the Ca2(+)-sensitive dye fura-2, was not altered by transient or prolonged exposure to Na(+)-free media or by exposure to ouabain in concentrations sufficient to produce a five-fold elevation in intracellular Na+ concentration. The [Ca2+]i response to vasopressin was also unaffected by Na+ removal or ouabain. By contrast, in cultured rat cardiac myocytes, cells that possess Na(+)-Ca2+ exchange, transient or prolonged Na+ removal as well as ouabain exposure produced greater than fivefold increases in [Ca2+]i compared with controls. We conclude that Na(+)-Ca2+ exchange does not contribute to the regulation of [Ca2+]i in hepatocytes.
Regulated phosphorylation of secretory granule membrane proteins of the rat parotid gland
Authors: Marino C. R. , Castle J. D. , Gorelick F. S.
Published: December 2017

An antiserum raised against purified rat parotid secretory granule membrane proteins has been used to identify organelle-specific protein phosphorylation events following stimulation of intact cells from the rat parotid gland. After lobules were prelabeled with [32P]orthophosphate and exposed to secretagogues, phosphoproteins were immunoprecipitated with the granule membrane protein antiserum, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and visualized by autoradiography. Parallel studies of stimulated amylase release were performed. Isoproterenol treatment of parotid lobules resulted in an increase in the phosphate content of immunoprecipitable 60- and 72-kDa proteins that correlated with amylase release in a time-dependent manner. Forskolin addition mimicked these effects, but only the isoproterenol effects were reversed by propranolol treatment. To confirm the specificity of the antiserum to the secretory granule membrane fraction, subcellular isolation techniques were employed following in situ phosphorylation. The 60- and 72-kDa phosphoproteins were immunoprecipitated from both a particulate fraction and a purified secretory granule fraction. Furthermore, the extraction properties of both species suggest that they are integral membrane proteins. These findings support the possibility that stimulus-regulated secretion may involve phosphorylation of integral membrane proteins of the exocrine secretory granule.
Relationship between interdigestive duodenal motility and fluid transport in humans
Authors: Sjovall H. , Hagman I. , Abrahamsson H.
Published: December 2017

In 22 healthy volunteers distal duodenal fluid absorption was related to the interdigestive motility cycle. Fluid absorption was measured with a triple-lumen perfusion technique, and motility was registered with a low-compliance pneumohydraulic system. Pancreatic and biliary secretions were estimated by measurement of bilirubin and amylase release into the duodenal segment. Duodenal fluid absorption rate changed during the interdigestive motility cycle; the highest absorption rate was registered during phase I (low-motor activity) and absorption rate then decreased in parallel with increasing motor activity during phase II (r = -0.69, P less than 0.001). In late phase II a net fluid secretion was frequently registered, together with an increased release of bilirubin into the duodenal lumen. This pattern was seen during perfusion with both glucose-containing (30 mM) and glucose-free solutions. The results show that duodenal fluid absorption rate changes markedly during the interdigestive motility cycle. This effect may be a hydrodnamic phenomenon or may be due to activation of a neural secretory mechanism during phase II.
Identification of taurocholate binding sites in ileal plasma membrane
Authors: Simon F. R. , Sutherland J. , Sutherland E.
Published: December 2017

Intestinal absorption of bile salts occurs by passive processes throughout the length of the small intestine, whereas active carrier-mediated uptake is localized to the ileum. Although previous studies have extensively characterized brush-border transport of bile acids, their extrusion across the basolateral membrane is less well understood. Because previous reports had failed to show specific bile acid binding sites except with the use of photolabeled bile salt derivatives, we sought to identify and characterize the binding parameters of the physiological bile salt taurocholate in ileal and jejunal plasma membrane subfractions. Brush-border membrane (BBM) and basolateral membrane (BLM) fractions were rapidly and simultaneously isolated from the small intestinal mucosa. BBM fractions were isolated with enrichments of 50- to 54-fold for leucine aminopeptidase, whereas the basolateral membrane enrichment of Na(+)-K(+)-ATPase, its specific marker enzyme, was 22- to 25-fold. Contamination from intracellular organelles was minimal. Binding of [14C]taurocholate was demonstrated in both jejunal as well as ileal plasma membrane fractions. However, only ileal binding demonstrated saturation, reversibility, and susceptibility to proteolytic enzymes. [14C]taurocholate binding to BBM fractions also showed competition with bile acids but was not altered by pH or alkylating agents. In contrast, binding of taurocholate to the basolateral membrane showed optimal pH between 6.5 and 7.5 and was inhibited by thiol and alkylating agents. Kinetic analysis of specific ileal BBM and BLM binding showed the parameters for BBM as 288 +/- 70 microM and 2.4 +/- 0.6 nmol/mg protein and for BLM as 6.6 +/- 0.7 microM and 0.56 +/- 0.01 nmol/mg protein for dissociation constant and maximum binding capacity, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Amiloride and taurine inhibit cholate-induced HCO3(-)-rich choleresis in perfused rat livers
Authors: Anwer M. S. , Atkinson J. M. , Zimniak P.
Published: December 2017

Bile acid-induced HCO3(-)-rich choleresis may be due to primary activation of sinusoidal Na(+)-H+ exchange or to biliary reabsorption of unconjugated bile acid. To test these hypotheses, we studied the effect of cholate and taurocholate (TC) (infused at 10 mumol/min for 20 min) on net H+ efflux, biliary [HCO3-], and bile flow in perfused rat livers and on intracellular pH (pHi) in isolated hepatocytes. Cholate, but not TC, produced HCO3(-)-rich choleresis. Amiloride and taurine decreased cholate-induced choleresis and HCO3- excretion and biliary excretion of unconjugated cholate. Amiloride, but not taurine, decreased cholate-induced net H+ efflux. Both cholate and TC (200-750 microM) decreased pHi. Cholate was metabolized to a polar compound, most likely cholate glucuronide, in the presence of amiloride. These results are consistent with the hypothesis that the biliary reabsorption of unconjugated cholate may be involved in HCO3(-)-rich choleresis. Amiloride also inhibited net hepatic uptake and biliary excretion of cholate and TC without affecting hepatic content of bile acids. It is suggested that amiloride may decrease the maximal excretion rate of cholate and TC. Since cholate and TC induce amiloride-sensitive net H+ efflux and decrease pHi, it appears that cholate and TC activate Na(+)-H+ exchange indirectly by decreasing pHi.
Gastric ulceration induced by nonsteroidal anti-inflammatory drugs is a neutrophil-dependent process
Authors: Wallace J. L. , Keenan C. M. , Granger D. N.
Published: December 2017

The hypothesis that neutrophils play an important role in the pathogenesis of gastric ulceration induced by nonsteroidal anti-inflammatory drugs (NSAIDs) was tested in rats. Rats made neutropenic by prior treatment with an antibody to rat neutrophils raised in goat were found to be significantly more resistant to the gastric-damaging actions of indomethacin or naproxen than were control rats or rats pretreated with normal goat serum. The reduction of damage in neutropenic rats was not due to effects of the antineutrophil serum on either gastric acid secretion or the ability of indomethacin or naproxen to inhibit prostaglandin synthesis. Gastric cyclooxygenase activity was inhibited by greater than 95% in both normal and neutropenic rats that received indomethacin or naproxen. Reduction of circulating neutrophil numbers by treating rats with methotrexate also resulted in a significant reduction in the susceptibility to gastric damage induced by indomethacin. Since activation of circulating neutrophils appeared to be important in the development of gastric erosions after administration of indomethacin, and in the significant changes in vascular endothelial integrity (Monastral Blue staining) observed within 15 min of indomethacin administration, we investigated the possibility that leukotrienes (LTs) and platelet-activating factor (PAF) might be involved in the pathogenesis of indomethacin-induced ulceration. Changes in gastric LTB4 synthesis were not observed after indomethacin administration. Pretreatment with either an LTD4 antagonist or a PAF antagonist was without significant effect on the extent of gastric damage induced by indomethacin. These results suggest an important role for neutrophils in the pathogenesis of NSAID-induced gastric ulceration. Neutrophils may be important in the vascular injury that occurs early after administration of these compounds.
Bioavailability of dietary glutathione: effect on plasma concentration
Authors: Hagen T. M. , Wierzbicka G. T. , Sillau A. H. , Bowman B. B. , Jones D. P.
Published: December 2017

Plasma glutathione (GSH) concentration in rats increased from approximately 15 to 30 microM after administration of GSH either as a liquid bolus (30 mumol) or mixed (2.5-50 mg/g) in AIN-76 semisynthetic diet. GSH concentration was maximal at 90-120 min after GSH administration and remained high for over 3 h. Administration of the amino acid precursors of GSH had little or no effect on plasma GSH values, indicating that GSH catabolism and resynthesis do not account for the increased GSH concentration seen. Inhibition of GSH synthesis and degradation by L-buthionine-[S,R]-sulfoximine and acivicin showed that the increased plasma GSH came mostly from absorption of intact GSH instead of from its metabolism. Plasma protein-bound GSH also increased after GSH administration, with a time course similar to that observed for free plasma GSH. Thus dietary GSH can be absorbed intact and results in a substantial increase in blood plasma GSH. This indicates that oral supplementation may be useful to enhance tissue availability of GSH.
Ontogenetic development of monosaccharide and amino acid transporters in rabbit intestine
Authors: Buddington R. K. , Diamond J. M.
Published: December 2017

We measured brush-border uptakes of seven sugars and amino acids by rabbit intestine as a function of age from the day of birth to adulthood. Gut dimensions, especially those of the colon and cecum, increase more rapidly with body weight than would be true if rabbits maintained identical proportions as they grew. However, nominal small intestinal area increases in approximately direct proportion to the animal's basal metabolic rate. For all solutes except fructose, uptake per milligram of intestinal tissue is maximal at or near birth and declines to a level 2.5-5 times lower in the adult. Because of small intestinal growth, though, the total uptake capacity of the whole length of the small intestine increases in approximately direct proportion to metabolic rate. Fructose uptake per milligram is unique in increasing steeply at the time of weaning, correlated with the post-weaning first appearance of fructose in the natural diet. Age-related changes in uptake ratios among aldohexoses or amino acids suggest developmental sequences of related transporters. Correlated with the very high protein content of rabbit milk, the proline-to-glucose uptake ratio is higher in suckling rabbits than in other sucking mammals. Remarkably, the ratio for adult rabbits is higher than in other monogastric herbivores and is instead similar to values for carnivores. In explanation, although the transport capacity of the small intestine appears to account for proline absorption in rabbits of all ages and for sugar absorption in suckling rabbits, the hindgut may be a major site of carbohydrate digestion in adult rabbits.
Relationship between portal venous and hepatic arterial blood flows: spectrum of response
Authors: Kawasaki T. , Carmichael F. J. , Saldivia V. , Roldan L. , Orrego H.
Published: December 2017

The relationship between portal tributary blood flow (PBF) and hepatic arterial blood flow (HAF) was studied in awake, unrestrained rats with the radiolabeled microsphere technique. Six distinct patterns of response emerged. In group A (PBF+, HAF 0), ethanol, acetate, glucagon, prostacyclin, and a mixed diet increased PBF without a change in HAF; in group B (PBF+, HAF+), adenosine and histamine increased both PBF and HAF; in group C (PBF 0, HAF+), isoflurane and triiodothyronine did not change PBF but increased HAF; and in group D (PBF-, HAF+), halothane and vasopressin decreased PBF and increased HAF. Acute partial portal vein ligation decreased PBF (56%) and increased HAF (436%). Hypoxia (7.5% O2) decreased PBF (28%) and increased HAF (110%). In group E (PBF+, HAF-), acute hepatic artery ligation increased PBF (35%) and reduced HAF (74%), while in group F (PBF-, HAF-), thyroidectomy reduced PBF and HAF (36 and 47%, respectively). All blood flow responses were accompanied by the expected changes in both portal tributary and hepatic arterial vascular resistances. The data suggest that the portal and hepatic arterial vascular territories have regulatory mechanisms that allow for independent changes.