bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamics of a metabolic system: what single-action agents reveal about acid secretion
Authors: Licko V. , Ekblad E. B.
Published: December 2017

A simple single-state nonlinear mathematical model of an open metabolic system is shown to be an adequate representation of acid secretion in frog gastric mucosa. The parameters of the elemental model were estimated from data, and subsystems of augmented models were established for histamine, a nonconservative stimulus acting via binding to receptors, and for two inhibitors of acid secretion. The latter included metiamide, a nonconservative histamine antagonist, which affects the formation of acid by competitive binding to the histamine receptors, and nitrite, a conservative inhibitor, which affects the rate of acid translocation. For parameter estimation, two data sets were analyzed by a nonlinear least-squares procedure: a dynamic (time dependent) set consisting of individual acid secretion rate curves and an integral (time independent) set consisting of the curves of acid secreted and suppressed above or below baseline, respectively, as functions of agent exposure. Because the model is instrumental in the estimation of parameters of subsystems that are not accessible through direct observation, it can serve as a research tool in the investigation of the mechanism of gastric acid secretion under a variety of experimental conditions.
Regulation of gastrin alpha-amidation in the developing rat stomach
Authors: Marino L. R. , Muglia B. H. , Yamada T.
Published: December 2017

Because the gastrin molecule must be alpha-amidated to have maximum biological activity, rat pups from 1 to 6 wk of age were treated with dexamethasone (2 mg.kg-1.day-1) for 3 or 7 days, diethyldithiocarbamate (DDC; 400 mg.kg-1.day-1 x 3 days), dexamethasone and DDC, pentagastrin (750 micrograms.kg-1.day-1), or bombesin (40 micrograms.kg-1.day-1) for 3 days to determine the effects of these agents on alpha-amidation and gastrin and glycine extended gastrin (G-Gly) concentration in the stomach. Three day treatment with dexamethasone increased gastrin concentration by increasing amidation in pups before 5 wk of age and thereafter by enhancing preprogastrin synthesis or processing. Seven day dexamethasone treatment had no substantial effect on amidation. DDC universally inhibited amidation and affected a sustained increase in gastrin plus G-Gly concentration after the third week of life. Dexamethasone did not reverse the effects of DDC. Pentagastrin increased amidation in 1-, 3-, and 6-wk old rat pups but had no consistent effect on peptide concentration. Bombesin increased the sum of gastrin and G-Gly concentration in all but 1- and 5-wk old pups but had variable effects on alpha-amidation. We conclude that alterations in gastrin alpha-amidation have age-specific effects on tissue gastrin and G-Gly concentration and speculate that changes in tissue gastrin and G-Gly stores available for release might ultimately affect parietal cell and G-cell function during development.
Anion channels in rat liver canalicular plasma membranes reconstituted into planar lipid bilayers
Authors: Sellinger M. , Weinman S. A. , Henderson R. M. , Zweifach A. , Boyer J. L. , Graf J.
Published: December 2017

Previous studies from this laboratory have demonstrated a Cl(-)-HCO3- exchanger and have provided evidence for a Cl- conductance in rat liver canalicular plasma membrane vesicles. To further investigate the apical Cl- conductance, we performed single-channel analysis after incorporation of canalicular liver plasma membrane vesicles into planar lipid bilayers. This was necessary, because the canalicular membrane is not accessible for the patch-clamp technique. Two types of anion channels could be identified (30- and 90-pS conductance) corresponding to the class of small and intermediate channels, respectively. The kinetics of the small channel were found to be voltage dependent with a maximum for the open probability at -20 mV. In contrast, intermediate channel kinetics were voltage independent. The anion channels described above could allow electrogenic Cl- efflux, to compensate Cl- influx via the electroneutral Cl(-)-HCO3- exchanger. Further studies will be required to prove their functional importance in bile formation.
Hormonal regulation of paracellular permeability in isolated rat hepatocyte couplets
Authors: Nathanson M. H. , Gautam A. , Ng O. C. , Bruck R. , Boyer J. L.
Published: December 2017

Many hormones and drugs exert their effects on cells by increasing cytosolic Ca2+ (Cai2+) and activating protein kinase C (PKC). Each of these actions results in cholestasis in the isolated perfused rat liver, but the responsible mechanisms are unclear. We used isolated rat hepatocyte couplets to observe the direct effects of increased Cai2+ and PKC activation on permeability of the hepatocyte tight junction and canalicular volume, two possible determinants of hepatocyte bile secretion. Couplets were stimulated with the Ca2+ agonist vasopressin (10(-8) M) in the absence and presence of the Ca2+ influx antagonist Ni2+ (5 x 10(-3) M) or with the PKC activator phorbol dibutyrate (10(-6) M). Cai2+ was determined by ratio microspectrofluorometry of indo-1, permeability of the couplet tight junctions was assessed by exclusion of horseradish peroxidase from the canalicular space, and changes in canalicular volume over time were measured directly by optical planimetry. Canalicular volume increased by 1.6 +/- 2.5%/min (mean +/- SD) under basal conditions. In response to vasopressin, there was a rapid 15-fold increase in Cai2+, followed first by an increase in paracellular permeability, then by canalicular collapse (15.9 +/- 5.9%/min). Pretreatment with Ni2+ markedly decreased the vasopressin-induced increase in Cai2+ and abolished both the increase in paracellular permeability and the canalicular collapse. Phorbol dibutyrate also increased paracellular permeability but resulted in neither increased Cai2+ nor canalicular collapse. The PKC inhibitor H-7 reversed the effects of both vasopressin and phorbol dibutyrate on tight junction permeability. Bile secretory pressure, measured in isolated perfused rat liver preparations, was acutely increased by vasopressin, but the increase was augmented rather than inhibited by Ni2+.(ABSTRACT TRUNCATED AT 250 WORDS)
Altered regulation of regional sucrase-isomaltase expression in diabetic rat intestine
Authors: Hoffman L. R. , Chang E. B.
Published: December 2017

Increased sucrase-isomaltase (SI) expression is a prominent feature of adaptive changes observed in the small intestine of streptozocin-treated chronically diabetic (CD) rats. In this study, we examine the cellular and molecular basis of increased SI expression in CD rats by determining SI specific activities and mRNA abundance in sequentially isolated enterocytes along the villus-to-crypt axis of proximal jejunum and distal ileum. In all regions, two- to fourfold increases in sucrase activity in diabetic rat enterocytes were paralleled by increases in SI mRNA. However, analogous to nondiabetic rat intestine, no differences in SI mRNA abundance were observed between corresponding enterocyte fractions from ileum and jejunum of diabetic rat intestine. By nuclear run-on assays, differences in rates of SI gene transcription were not observed in diabetic and nondiabetic intestinal tissues. We conclude that diabetes induces increased total and specific activities and mRNA abundance of intestinal SI, largely through the stabilization of SI mRNA. Furthermore, analogous to nondiabetic small intestine, differences in proximal-to-distal SI expression appear to be determined at the translational or posttranslational level.
Hypoxic liver injury and the ameliorating effects of fructose: the "glucose paradox" revisited
Authors: Brass C. A. , Crawford J. M. , Narciso J. , Gollan J. L.
Published: December 2017

It has been independently postulated that nutritional status is a modulator of the hepatic injury response to hypoxia and that glucose may be a poor substrate for hepatocellular metabolism. This study provides data linking these two concepts within the framework of metabolic zonation of the liver. With the use of a hypoxically perfused isolated rat liver model, cellular injury, as reflected by aspartate aminotransferase (AST) release, was significantly greater in the liver of fasted (mean AST 489 U/g liver at 3 h) than fed (40 U/g) animals. The extent of injury during hypoxia was decreased to a comparable degree in fasted livers perfused with Wisconsin solution (27 U/g) or 20 mM fructose (51 U/g). Perfusion with (11.5 mM) glucose plus insulin provided no hepatoprotection (791 U/g); however, supraphysiological amounts of glucose (100 mM) with (310 U/g) or without (321 U/g) insulin (10 U) or dihydroxyacetone (220 U/g) provided a modest reduction in AST release. Cellular injury measured by trypan blue uptake showed a marked zonal pattern, with upstream regions incurring greater parenchymal and nonparenchymal injury than downstream areas. These data that indicate that exogenous glucose is poorly utilized as an energy substrate by the liver during hypoxia are consistent with data from the fasted-refed rat model, suggesting a "glucose paradox" in the liver. The findings also suggest that low levels of oxygen are an important factor mediating "hypoxic" liver injury.
Intestinal myoelectrical activity and transit time in chronic portal hypertension
Authors: Stewart J. J. , Battarbee H. D. , Farrar G. E. , Betzing K. W.
Published: December 2017

This study was designed to determine the effects of portal hypertension on intestinal myoelectrical activity and propulsion. In a single surgery, adult rats were implanted with a serosal electrode at each quarter of the small intestine, and portal hypertension was produced by calibrated constriction of the portal vein. To determine intestinal transit, portal vein-stenosed (PVS) and sham-operated animals were chronically implanted with a catheter in the proximal small intestine. Transit time was determined by measuring the progression of radioactive chromium along the bowel. Studies were conducted 6, 9, and 14 days after surgical preparation. Portal hypertension was associated with both transient and persistent changes in intestinal myoelectrical activity during the experimental period. Slow wave frequency was significantly reduced in the proximal small intestine on all test days and in the distal small intestine on day 14. Occurrence of the migrating myoelectric complex was reduced on days 6 and 9. Phase III amplitude was significantly reduced in the distal small intestine on all test days. Changes in intestinal myoelectrical activity in PVS animals were not associated with measurable changes in intestinal propulsion. The results suggest that both transient and persistent changes in intestinal myoelectrical activity occur during the 2-wk period after portal vein stenosis. The functional significance of the changes is unknown.
Protection against alkali injury to rabbit esophagus by CO2 inhalation
Authors: Meyers R. L. , Glenn L. , Orlando R. C.
Published: December 2017

Lye ingestion, a poisoning with no known effective treatment, frequently results in esophageal ulceration and healing by stricture formation. Tissue injury by lye is due to its alkalinity, and so therapy is logically directed at neutralization by acid. Here we describe a novel method, the inhalation of CO2, for the rapid delivery of (carbonic) acid capable of neutralizing tissue and luminal alkalinity. We also show that CO2 inhalation in anesthetized rabbits provides protection to the lye-exposed esophagus against transepithelial necrosis. This method has the potential to protect the human esophagus against lye injury, because it is relatively safe, rapidly effective, and can be administered in the field under emergency circumstances.
Comparison of receptors for Escherichia coli heat-stable enterotoxin: novel receptor present in IEC-6 cells
Authors: Mann E. A. , Cohen M. B. , Giannella R. A.
Published: December 2017

Enterotoxigenic Escherichia coli elaborate a heat-stable enterotoxin that causes diarrhea in humans and animals. The primary event in the diarrheal cascade is the binding of this enterotoxin to specific receptors on enterocytes and activation of guanylyl cyclase. Two intestinal cell lines, Caco-2 and IEC-6, were tested for the presence of these receptors. Although both cell lines exhibited specific binding, only the Caco-2 cell line responded to heat-stable enterotoxin with increased guanylyl cyclase activity. Cloning and expression studies confirmed that the receptor present in Caco-2 cells is a homologue of guanylyl cyclase C, a known transmembrane heat-stable enterotoxin receptor. Expression of the receptor in differentiating Caco-2 cells increases with cell maturation, indicating that these cells are a suitable model for future studies. However, Northern and polymerase chain reaction analyses demonstrated that guanylyl cyclase C is not expressed in IEC-6 cells, strongly suggesting the presence of a novel heat-stable enterotoxin receptor that is not coupled to guanylyl cyclase activity.
Hepatic taurocholate uptake is electrogenic and influenced by transmembrane potential difference
Authors: Lidofsky S. D. , Fitz J. G. , Weisiger R. A. , Scharschmidt B. F.
Published: December 2017

Uptake of the bile acid taurocholate by hepatocytes is coupled to Na+ influx. The stoichiometry of uptake, however, is uncertain, as is the influence of the transmembrane electrical potential difference (PD) on this process. In this study, we examined the relationship between taurocholate extraction and PD (measured using intracellular microelectrodes) in perfused liver, and we measured taurocholate-induced transport current in cultured hepatocytes using patch-clamp recording techniques. In the perfused liver under basal conditions, PD averaged -28.4 +/- 0.6 (SE) mV, and extraction of 1, 50, and 300 microM taurocholate was 0.95 +/- 0.02, 0.98 +/- 0.01, and 0.41 +/- 0.03, respectively. When the Na+ chemical gradient was decreased by replacing perfusate Na+ with choline, the membrane depolarized to -17.2 +/- 1.1 mV, and taurocholate extraction markedly decreased at all taurocholate concentrations (P < 0.01). When perfusate Na+ concentration was held constant at 137 mM, membrane depolarization induced by substitution of gluconate for perfusate Cl- (-17.9 +/- 0.6 mV) or Cl- for nitrate (-10.3 +/- 2.1 mV) significantly decreased extraction of 300 microM taurocholate. Abrupt exposure to taurocholate produced a concentration-dependent membrane depolarization in the presence of Na+, but not in its absence (P < 0.001). In cultured hepatocytes, exposure to 100 microM taurocholate produced an inward current of -0.056 +/- 0.016 pA/pF at a holding potential of -40 mV. This current was Na+ dependent, and it increased twofold as holding potential was changed from -20 to -50 mV.(ABSTRACT TRUNCATED AT 250 WORDS)
Evidence for indirect dietary regulation of cholecystokinin release in rats
Authors: Sharara A. I. , Bouras E. P. , Misukonis M. A. , Liddle R. A.
Published: December 2017

Food ingestion stimulates cholecystokinin (CCK) release from the proximal intestine, but the mechanisms involved are not well understood. To investigate this effect in vivo in intact rats, plasma CCK was measured after orogastric feeding of proteins, protein hydrolysates, amino acids, glucose, and starch. Intact proteins were the only nutrients to stimulate CCK release. The possibility of direct interaction between different dietary constituents and intestinal CCK-secreting endocrine cells was then examined using a perfusion system containing isolated mucosal cells from the rat duodenojejunum. The functional validity of this system was established by demonstrating that monitor peptide and bombesin both stimulated CCK release in a dose-dependent manner. The stimulatory effect of bombesin required extracellular calcium and was not inhibited by addition of tetrodotoxin. Perifusion of proteins, protein digests, and carbohydrates did not stimulate CCK release. These results indicate that proteins stimulate CCK release postprandially via an indirect mechanism, most likely related to inhibition of intraluminal trypsin. Perifusion of dispersed mucosal cells constitutes a reproducible model to investigate hormonal and peptidergic regulation of CCK release in vitro.
Orogastric EGF enhances c-neu and EGF receptor phosphorylation in suckling rat jejunum in vivo
Authors: Thompson J. F. , Lamprey R. M. , Stokkers P. C.
Published: December 2017

Epidermal growth factor (EGF) in rat milk stimulates intestinal growth. We examined the role of EGF-stimulated tyrosine kinase activity in vivo in this process using an affinity-purified anti-phosphotyrosine antibody in Western blot. Jejunal sacs from fasted 8-day-old rat pups were incubated with intraluminal EGF and were then assayed for phosphotyrosyl proteins (p-Tyr) by Western blot. A 170-kDa p-Tyr was present in EGF-treated sacs but not in control. A 190-kDa p-Tyr was constitutively present in control but increased in abundance in EGF-treated sacs. Dose-response experiments demonstrated that the increase in p-Tyr was present at 100 ng/ml EGF, which is within the physiological range. The 170- and 190-kDa p-Tyr was confirmed by immunoprecipitation to be the EGF receptor and c-neu, respectively. A similar response was also observed in the jejunum after orogastric gavage feeding of EGF. By use of indirect immunofluorescence, the EGF receptor was localized primarily to the basolateral membrane of both the crypt and villus epithelium. c-neu was localized primarily in the villus enterocyte basolateral membrane. These data demonstrate that intraluminal EGF stimulates rapid tyrosine phosphorylation of the EGF receptor in vivo and that c-neu is a major substrate of the EGF receptor in suckling jejunum. The basolateral membrane localization of the EGF receptor and c-neu suggests that EGF is rapidly transported across the intestinal epithelium in suckling rat jejunum.
Protein phosphatase inhibition and in vivo hepatotoxicity of microcystins
Authors: Runnegar M. T. , Kong S. , Berndt N.
Published: December 2017

Administration of microcystin (MCYST)-YM or -LR (peptide hepatotoxins produced by the cyanobacterium Microcystis aeruginosa) to mice resulted in the inhibition of liver protein phosphatase 1 and 2A activity. In all cases significant inhibition preceded or accompanied clinical changes due to MCYST intoxication. Fifteen minutes after intraperitoneal injection of lethal doses of MCYST-YM protein phosphatase activity was already decreased to 44% of controls, and by 60 min was further decreased to 22% of controls. The inhibition was dose dependent: intraperitoneal injection with 84 nmol/kg of MCYST-YM and 48 nmol/kg of MCYST-LR were the minimum doses required for significant inhibition at 60 min. Pretreatment of mice with 200 mumol/kg of rifamycin prevented the inhibition of liver protein phosphatase. The inhibition was tissue specific, with none detected in the kidneys, an organ that, unlike the liver, does not accumulate MCYST. In contrast to MCYST intoxication, lethal doses of phalloidin, a peptide hepatotoxin that produces clinical and pathological changes similar to MCYST, did not cause any inhibition of protein phosphatases.
Interaction between cholephilic anions and bile acid transport across basal membrane of human trophoblast
Authors: Bravo P. , el-Mir M. Y. , Serrano M. A. , Boyd R. , Marin J. J.
Published: December 2017

The sensitivity of radiolabeled bile acid (BA) binding and transport by basal plasma membrane (BPM) vesicles of human trophoblast to cholephilic organic anions (COAs) was studied by a rapid filtration technique. Glycocholate (GC) efflux from preloaded (15 microM GC) vesicles was investigated in the presence of 300 microM COAs at the trans-side of the membrane. Bilirubin (BR) diglucuronide and rose bengal induced a very strong transstimulating effect, whereas phalloidin and phenol red showed a negligible effect. This effect was from strong to moderate for indocyanine green > bromosulfophthalein (BSP) > or = fusidic acid > or = phenolphthalein > or = BR ditaurate > or = rifamycin SV > or = rifampicin. BSP-induced transstimulation was not additive to the "velocity effect" previously reported for bicarbonate. At the cis-side, BSP reduced the saturable component of taurocholate (TC) binding to BPM vesicles. BSP also induced a partial and mixed type of inhibition both in TC uptake [inhibitor constant (Ki) 227 microM] and efflux (Ki 209 microM). Two binding sites with overlapping specificity for BAs and other COAs are proposed in this carrier, the site for non-BA COA presumably corresponding to that for bicarbonate. In summary, the results indicate that several COAs can act as potential substrates for the BA carrier located at the BPM of human trophoblast. This stresses the "biliary-like" role of the placenta and suggests the possibility of developing new functional tests for this organ on the basis of fetal-maternal transfer of nontoxic cholephilic dyes.
Identification of a 5-hydroxytryptamine (5-HT2) receptor on guinea pig small intestinal crypt cells
Authors: Siriwardena A. K. , Smith E. H. , Borum E. H. , Kellum J. M.
Published: December 2017

Radioligand labeling of [3H]ketanserin was examined in suspensions of dispersed guinea pig small intestinal mucosal cells prepared by modification of the EDTA-chelation method described by M. M. Weiser (J. Biol. Chem. 248: 2536-2541, 1973). Preferential incorporation of [3H]thymidine was used to confirm that suspensions were enriched in crypt cells. At 25 degrees C, binding of [3H]ketanserin to dispersed enterocytes was rapid, maximal by 5 min, saturable (dissociation constant = 1.5 nM), 65 +/- 5% specific, stable, and reversible. The maximal number of binding sites per cell was 92,000 (range 86,000-105,500). Binding was temperature dependent, with maximal binding at 37 degrees C, and was inhibited by 5-hydroxytryptamine (5-HT) (half-maximal inhibition of [3H]ketanserin binding observed in response to 1 microM 5-HT) and ketanserin (half-maximal inhibition of [3H]ketanserin binding observed in response to 1 nM ketanserin) but not by the 5-HT1P antagonist N-acetyl-5-hydroxytryptophyl 5-hydroxytryptophan amide (5-HTP-DP) or the 5-HT3 antagonist 3-tropanyl-indole-3-carboxylate methiodide (ICS-205-930). The second messenger system coupled to the putative mucosal 5-HT2 receptor was examined. 5-HT stimulated a concentration-dependent production of inositol 1,4,5-trisphosphate (IP3) in the dispersed enterocytes. This was maximal at 1 min and was inhibited in a concentration-dependent manner by ketanserin. 5-HTP-DP and ICS-205-930 had no effect on 5-HT-stimulated production of IP3. These data provide evidence for the existence of a mucosal 5-HT2 receptor located on guinea pig small intestinal crypt cells.
Cell survival in rabbit gastric glands: effect of extracellular pH, osmolarity, and anoxia
Authors: Carter K. J. , Lee H. H. , Goddard P. J. , Yanaka A. , Paimela H. , Silen W.
Published: December 2017

Although backdiffusion of luminal acid is regarded as a common mechanism of gastric injury, the extracellular pH (pHo) at which cells are irreversibly injured is not well defined. Exclusion of the fluorescent dye propidium iodide was used to estimate cell survival in rabbit gastric glands incubated in buffers of pHo 8.0-2.0. Mean survival (+/- SE) for n = 6 experiments at 2 h in a HEPES buffer of 300 mosM at pHo 8.0, 7.0, 6.0, 4.0, and 2.0 was 80 +/- 3, 91 +/- 2, 90 +/- 2, 71 +/- 2, and 17 +/- 4%, respectively. Survival at acidic pHo was improved in a high KCl buffer: 78 +/- 3 and 38 +/- 7% at pHo 4.0 and 2.0, respectively. Survival in HCO3- buffers was 73 +/- 3, 88 +/- 2, and 92 +/- 3% at pHo 8.0, 7.4, and 6.0. Brief (5 min) exposure to pHo 4.0 followed by reexposure to pHo 7.4 had no effect on acid secretion as estimated by [14C]aminopyrine uptake or cellular viability over 4 h. The influence on cell survival of changes in pHo under conditions of chemical anoxia and in buffers of different osmolarity was investigated. Chemical anoxia was induced using 2.5 mM KCN and 2.0 mM iodoacetic acid (IAA) to inhibit oxidative phosphorylation and anaerobic glycolysis. Survival in glands exposed to KCN and IAA in HEPES buffer was 54 +/- 5, 82 +/- 3, and 87 +/- 2% at pHo 8.0, 7.0, and 6.0.(ABSTRACT TRUNCATED AT 250 WORDS)
Biphasic effect of SNP on opossum esophageal longitudinal muscle: involvement of cGMP and eicosanoids
Authors: Saha J. K. , Hirano I. , Goyal R. K.
Published: December 2017

Effects of nitric oxide (NO)-containing compounds on opossum esophageal longitudinal smooth muscle in vitro were examined. Sodium nitroprusside (SNP) and authentic NO produced a biphasic concentration-dependent relaxation-contraction sequence in the esophageal longitudinal muscle (ELM) but only a concentration-dependent relaxation of the lower esophageal sphincter (LES) and no effect in the esophageal circular muscle. A cell membrane-permeable analogue of guanosine 3',5'-cyclic monophosphate (cGMP), 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) also produced relaxation-contraction sequence in the ELM and relaxation of the LES. The guanylate cyclase inhibitors methylene blue (MB) and LY-83583 increased resting tone and had no significant effect on SNP-induced relaxation of ELM. However, they abolished the SNP- and NO-induced contraction of ELM. The cyclooxygenase inhibitor indomethacin had no effect on ELM relaxation and abolished the contractions due to SNP, NO, and 8-BrcGMP. These studies show that in the ELM 1) SNP, authentic NO, and 8-BrcGMP cause a biphasic relaxation-contraction sequence; 2) MB and LY-83583 blocked contraction but not the relaxation associated with SNP and NO; and 3) indomethacin blocked contractions but not the relaxation due to SNP, NO, and 8-BrcGMP. These results suggest that in the ELM, NO donors exert an inhibitory action that is largely cGMP independent and an excitatory action via a cGMP-dependent pathway involving endogenous eicosanoids of the cyclooxygenase pathway.
Intestinal lactase: maturational excess expression of mRNA over enzyme protein
Authors: Nudell D. M. , Santiago N. A. , Zhu J. S. , Cohen M. L. , Majuk Z. , Gray G. M.
Published: December 2017

The mechanism of decline of intestinal lactase during mammalian development remains uncertain. Despite a major loss of catalytic activity, lactase mRNA appears to persist at detectable concentrations in adult rats. We quantified lactase activity, total lactase protein, and lactase mRNA in rats aged 7, 11, 15, 18, 22, 30, and 60 days using the 7S ribosomal RNA as the developmental control. The active lactase fraction was 0.81 of total lactase for all age groups except 60-day-old animals, in which it declined to 0.60 (P = 0.004), indicating that conversion of active lactase to inactive species contributed to the lower activity in the adult. Northern blots revealed a single discrete 6.8-kb message at all ages. Although lactase activity and immunoprotein decreased coordinately to a minimum by day 30 (20% of the 7-day value), lactase mRNA doubled to a maximum at day 22 and was maintained at 7-day concentrations even in 60-day adults. The lactase mRNA-to-protein ratio was low at 7 days (0.19) but more than doubled (0.50) by 22 days, achieved a fivefold increase (1.0) by 30 days, and persisted at 0.77 in adults. The relative excess of lactase message during maturation suggests that translational or post-translational events may be paramount in the developmental regulation of lactase gene expression.
Elevation of intraluminal pressure and cyclooxygenase inhibitors increases duodenal alkaline secretion
Authors: Sababi M. , Nylander O.
Published: December 2017

Our aim was to study the effects of intraluminal hydrostatic pressure on duodenal mucosal alkaline secretion (DMAS) and permeability in anesthetized rats. A segment of proximal duodenum was perfused with saline and the rate of DMAS determined by backtitration. Mucosal permeability was assessed by measuring the clearance of 51Cr-EDTA (ED-C1) from blood to lumen. Raising the intraluminal hydrostatic pressure 6 mmHg above basal for 40 min induced a 75% increase in DMAS and a transient increase in ED-C1. This stimulation of DMAS was not affected by pretreatment with the muscarinergic receptor antagonist atropine (0.5 mg/kg i.v.), or the opioid receptor antagonist naloxone (15 micrograms/kg i.v.) but was strongly reduced by stripping the nerves around the carotid arteries and abolished by the ganglion blocker hexamethonium (10 mg/kg i.v.). When the distension was prolonged to 90 min, the net increase in DMAS decreased with time, indicating an adaptive mechanism. The cyclooxygenase inhibitors indomethacin (5 mg/kg i.v.) and meclofenamate (5 mg/kg i.v.) induced intermittent elevations of intraluminal pressure and increased DMAS by > 100%. This was associated with a transient increase in ED-C1. Indomethacin increased DMAS in a dose-related fashion, and the rise in both intraluminal pressure and DMAS was abolished by hexamethonium but not affected by stripping the nerves around the carotid arteries. Elevation of intraluminal pressure did not augment the rise in DMAS induced by indomethacin. It is concluded that luminal distension and cyclooxygenase inhibition increase DMAS by a similar, but not identical, neural mechanism involving nicotinergic receptors. It is speculated that the indomethacin-induced rise in DMAS is mediated via induction of duodenal motility
Transport of bovine serum albumin across rat jejunum: role of the enteric nervous system
Authors: Kimm M. H. , Curtis G. H. , Hardin J. A. , Gall D. G.
Published: December 2017

To assess the mechanisms for movement of antigenically intact macromolecules across small intestinal mucosa, transport kinetics of bovine serum albumin (BSA) uptake and the effect of neural and metabolic inhibition were examined in stripped short-circuited rat jejunum. The mucosa was exposed to BSA, and, after a 50-min equilibration, mucosal-to-serosal movement of immunologically intact BSA was determined by enzyme-linked immunosorbent assay and total BSA by radiolabeled 125I-BSA. Intact BSA uptake demonstrated saturable kinetics. Immunologically intact BSA crossed the intestinal mucosa as 4.5% of total 125I-BSA flux. Colchicine and 4 degrees C significantly reduced uptake of immunologically intact BSA. NaF significantly reduced uptake of immunologically intact BSA and 125I-BSA. Treatment with tetrodotoxin significantly reduced intact BSA uptake, but did not significantly alter total BSA uptake. The muscarinic cholinoceptor antagonist atropine also significantly inhibited transport of intact BSA, whereas the nicotinic cholinoceptor antagonist hexamethonium had no effect. These findings indicate that transport of intact macromolecules across small intestinal mucosa is a saturable energy-dependent process that utilizes the microtubular network and is regulated by the enteric nervous system primarily through cholinergic nerves acting on muscarinic receptors.
Multiple inhibitory effects of genistein on stimulus-secretion coupling in rat pancreatic acini
Authors: Duan R. D. , Wagner A. C. , Yule D. I. , Williams J. A.
Published: December 2017

Genistein, a tyrosine kinase inhibitor, inhibited cholecystokinin (CCK)-induced maximal amylase release from rat pancreatic acini by 18, 31, and 46% at concentrations of 100, 300, and 750 microM, respectively, after 30 min preincubation. Genistein similarly decreased amylase release stimulated by bombesin but not that stimulated by secretin or vasoactive intestinal peptide. The steps of stimulus-secretion coupling affected by genistein were further evaluated. We found genistein dose dependently suppressed CCK-as well as NaF-induced polyphosphoinositide hydrolysis with a 50% inhibitory concentration of 380 and 400 microM, respectively, but only slightly suppressed the increase of intracellular Ca2+ concentration in response to either low or high concentrations of CCK. Genistein at 300 microM also decreased incorporation of [3H]inositol into phosphatidylinositol 4,5-bisphosphate. Most strikingly, 300 microM genistein inhibited Ca(2+)-stimulated amylase release by 85% in streptolysin O-permeabilized acini and thapsigargin-stimulated amylase release by 69% in intact acini. Daidzein, the inactive analogue of genistein, had no effect on any of the above parameters. Genistein, up to 750 microM, did not affect amylase release in response to phorbol ester. To relate these inhibitory effects of genistein to its inhibition of tyrosine phosphorylation, Western blotting was performed with an anti-phosphotyrosine monoclonal antibody. Genistein at 100 microM partly and at 300 microM completely inhibited CCK-induced tyrosine phosphorylation. In conclusion, genistein inhibits amylase release at multiple stages of stimulus-secretion coupling. These effects most likely involve both tyrosine kinase-dependent and -independent mechanisms.
Phosphoramidon attenuates big endothelin-1-induced vasoconstriction in canine stomach
Authors: Wood J. G. , Yan Z. Y. , Davis J. M. , Cheung L. Y.
Published: December 2017

We compared the effects of endothelin-1 and its precursor, big endothelin-1, on vascular resistance of a blood-perfused ex vivo stomach segment of chloralose-anesthetized dogs. In separate groups of dogs, endothelin-1 or big endothelin-1 was infused intra-arterially directly to the gastric segment. Endothelin-1 caused statistically significant dose-related increases in gastric vascular resistance at final blood concentrations of 0.15-10 nM. Although each dose was given for only 5 min, endothelin-1 at concentrations > 0.6 nM caused sustained responses with vascular resistance remaining above control values for approximately 45-90 min. In contrast, however, big endothelin-1 caused a small but statistically significant vasoconstriction only at the highest concentration (10 nM). In other experiments, using 15-min peptide infusions, we found that pretreatment with phosphoramidon, an inhibitor of endothelin-converting enzyme, markedly reduced response to big endothelin-1 but not to endothelin-1. Our results demonstrate that endothelin-1, but not big endothelin-1, is a potent vasoconstrictor of the canine gastric microcirculation. In addition, it appears that big endothelin-1 is degraded to endothelin-1 in the stomach by a phosphoramidon-sensitive metalloproteinase.
Apical nonspecific cation conductances in rabbit cecum
Authors: Sellin J. H. , Dubinsky W. P.
Published: December 2017

Rabbit cecum exhibits electrogenic Na absorption in vitro. However, because this transport process is not inhibited by amiloride nor does it demonstrate saturation kinetics typical of the amiloride-inhibitable Na channel, we considered whether the cecal transporter represented one of a recently described family of nonselective cation conductances or channels (NSCC). Both transepithelial and vesicle studies demonstrated that K, Cs, and Rb were transported via an apical conductance. Electrogenic transport was inhibited by divalent cations including Ca, Mg, and Ba but was unaffected by either lanthanum or gadolinium. Parallel studies in distal colon did not exhibit a similar response to either K substitution or Ba inhibition. Phenamil, verapamil, and nicardipine significantly inhibited the short-circuit current (Isc). stimulated by nominal Ca- and Mg-free conditions. Flux studies demonstrated a correlation between changes in Isc and Na transport. Microelectrode impalement studies suggested that there may be both NSCC and K conductance in the apical membrane. Planar bilayer studies identified a 190-pS cation channel that may correlate with the macroscopic transport properties of this epithelium. These studies are consistent with a model of cecal Na absorption mediated by a NSCC in the apical membrane; this may be the mechanism underlying the distinct epithelial transport characteristics of this intestinal segment.
Selective lesioning of interstitial cells of Cajal by methylene blue and light leads to loss of slow waves
Authors: Liu L. W. , Thuneberg L. , Huizinga J. D.
Published: December 2017

Incubation with 50 microM methylene blue (MB) and subsequent intense illumination resulted in abolition of the slow-wave activity in the submuscular interstitial cells of Cajal-circular muscle (ICC-CM) preparations of canine colon. This was often accompanied by a decrease in resting membrane potential. Repolarization of cells back to -70 mV did not restore the slow-wave activity, indicating that MB plus light directly interrupted the generation mechanism of slow waves. After MB incubation, a 2-min illumination consistently changed the mitochondrial conformation in ICCs from very condensed to orthodox, without inducing any obvious changes in smooth muscle cells. After 4- to 10-min illumination, ICCs became progressively more damaged with swollen and ruptured mitochondria, loss of cytoplasmic contrast and detail, loss of caveolae, and rupture of the plasma membrane. No damage was seen in smooth muscle cells or nerves. Gap junctional ultrastructure was preserved. Intense illumination without preincubation with MB left the slow waves and the ultrastructure of ICC-CM preparations unaffected. In CM preparations, without the submuscular ICC-smooth-muscle network, MB plus light induced no changes in electrical activity. We conclude that the correlation between selective damage to the submuscular ICCs (relative to smooth muscle) and selective loss of the slow-wave activity (relative to other electrical activity of the CM) strongly indicates that the ICCs play an essential role in the generation of slow waves.
Age-dependent increase of neurotensin expression in the ileum
Authors: Shimoda I. , Evers B. M. , Sato K. , Ishizuka J. , Rajaraman S. , Chung D. H. , Uchida T. , Tsuchiya T. , Greeley G. H. , Townsend C. M. , et al.
Published: December 2017

The proliferative activity of gut mucosa is increased with aging. Neurotensin (NT), a tridecapeptide localized to the distal small bowel, stimulates growth of gut mucosa; the effects of aging on gut NT are not known. Young (4-mo-old), adult (1-yr-old), and aged (2-yr-old) male Fischer 344 rats were killed; the ileal mucosa was scraped, weighed, and extracted for measurement of NT mRNA by slot-blot and Northern hybridization; and the relative NT transcription rate was determined by a nuclear run-on assay. In addition, NT tissue content and plasma levels were determined by radioimmunoassay, and full-thickness sections of ileum were examined for age-dependent alterations of NT endocrine cell (N-cell) number using immunohistochemical staining. Slot-blot and Northern-blot analyses showed that the steady-state levels of NT mRNA were increased threefold in the adult group and eightfold in the aged rats. In addition, NT peptide content and plasma levels were significantly increased in the aged group. The dramatic increases in the abundance of NT mRNA were not associated with increases in either NT transcription or N-cell number. In summary, we have demonstrated an age-dependent increase in the constitutive levels of ileal NT mRNA that appears not to be due to concomitant increases in transcription, suggesting that NT mRNA is increased by mechanisms involving mainly a posttranscriptional process. In addition, we have shown corresponding increases in the levels of both tissue and plasma NT with aging, indicating that expression of NT does not remain the same throughout life but actually increases with aging in the rat.
L-glutamine and L-asparagine stimulate Na+ -H+ exchange in porcine jejunal enterocytes
Authors: Rhoads J. M. , Chen W. , Chu P. , Berschneider H. M. , Argenzio R. A. , Paradiso A. M.
Published: December 2017

L-Glutamine (Gln) is a major respiratory fuel and substrate for nucleic acid synthesis in mammalian intestinal cells. The structurally related amino acid, L-asparagine (Asn), stimulates the proliferative enzyme ornithine decarboxylase in colonocytes, an effect that is blocked by the Na+-H+ exchange inhibitor amiloride. In an epithelial cell line derived from newborn piglet jejunum (IPEC-J2 cells), we determined intracellular pH (pHi) by computer-assisted microfluorimetry in single cells loaded with pH-sensitive dye 2',7'-bis(2-carboxyethyl)5-(6)- carboxyfluorescein. Resting pHi in N-2-hydroxyethylpiperazine-N'-2- ethanesulfonic acid-buffered NaCl Ringer was 7.06 +/- 0.02. Removal of external Na+ caused reversible acidification; recovery of pHi from NH+4-induced acid load was Na+ dependent, amiloride inhibitable, and Cl-independent. Asn and Gln had no measurable effect on resting pHi, but pretreatment with Asn or Gln induced a consistent twofold increase in pHi recovery from an acid challenge that was not seen with L-proline, D-glutamine, or L-phenylalanine. Inhibition of Gln metabolism by aminooxyacetate abolished the stimulatory effect of Gln on the exchanger. The tumor promotor phorbol 12-myristate 13-acetate (PMA) stimulated recovery rate from acid load and also increased resting pHi. The effects of PMA and Gln on Na+-H+ exchange from acid load were additive. Stimulation of Na+-H+ exchange by PMA, but not by Gln, was inhibited by protein kinase C (PKC) inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpeperazine. We conclude that Gln metabolism stimulates Na+-H+ exchange of acid-loaded porcine enterocytes by a mechanism not requiring activation of PKC.
Cerebral metabolism in sheep: a comparative study of the adult, the lamb, and the fetus
Authors: Jones MD  , Burd LI , Makowski EL , Meschia G , Battaglia FC
Published: December 2017

The cerebral uptakes of glucose, oxygen, lactate, pyruvate, acetoacetate and beta-hydroxybutyrate were compared in the adult, the fetal, and the newborn sheep. Beginning 1-2 days after surgery, we withdrew samples simultaneously from an artery and from the sagittal sinus for up to 2 wk. At all ages glucose was the only substrate taken up in significant quantity relative to oxygen. The glucose-oxygen quotients were 0.98 less than 1.03 less than 1.08 in adults; 0.92 less than 0.99 less than 1.06 in lambs; 0.92 less than 0.98 less than 1.03 in fetuses. These quotients were not significantly different. There was a significant (P less than .05) arteriovenous difference for lactate in the adult (-0.031 mM), but not in the lamb or fetus. A significant (P less than .05) arteriovenous difference pyruvate was found in the adult (-0.013 mM) and in the fetus (-0.020 mM). The findings of a glucose-oxygen quotient of approximately 1 and the lack of significant lactate production in fetal sheep brain are consistent with the hypothesis that anerobic glycolysis is unimportant to the steady-state cerebral metabolism of the fetal sheep.
Monosodium glutamate metabolism in the neonatal monkey
Authors: Stegink LD , Reynolds WA , Filer LJ  , Pitkin RM , Boaz DP , Brummel MC
Published: December 2017

Monosodium glutamate (MSG) administered by gastric tube to 10 infant monkeys at doses of 1-4 g/kg produced rapid increases in plasma glutamate (17- to 33-fold) and aspartate (50- to 90-fold) levels. The degree of elevation was proportional to the dose administered. Levels of other amino acids were unaffected. Two of the monkeys exhibited high fasting glutamate levels and abnormal glutamate tolerance curves. Despite this apparent decreased ability to metabolize glutamate, neither these animals nor any of the others for whom morphologic studies have been previously reported demonstrated neurotoxicity. Studies using 14-C-labeled glutamated indicated conversion of administered glutamate to two ninhydrin-negative compounds identified as glucose and lactate, as well as to asparate.
Heart functional responses to pressure overload in exercised and sedentary rats
Authors: Dowell RT , Cutilletta AF , Rudnik MA , Sodt PC
Published: December 2017

Female rats that had been subjected to a moderate treadmill running program were compared with sedentary animals on the basis of heart weight, selected biochemical measurements, and heart function. Exercised animals maintained normal growth rate, and cardiac hypertrophy was not present. Left ventricular RNA, DNA, and cytochrome c levels were unchanged. Heart functional measurements obtained in situ were similar in sedentary and exercised animals under control conditions. When subjected to sustained (1-3 days) aortic constriction pressure overload, exercised animals maintained or increased myocardial contractility. Contractility was depressed in sedentary animals. Both sedentary and exercised animals increased left ventricular end diastolic pressure without changing contractility during acute (1-3 min) pressure overload. However, exercised animals were able to fully regain normal cardiac output when the acute overload was relieved. Cardiac output remained approximately 10% below control in sedentary animals. The improved ability of previously exercised animals to withstand pressure overload appears to be due to alterations in adaptation rather than preliminary augmentation of metabolism or function.
Antidiuretic peptide in mammalian choroid plexus
Authors: Rudman D , Chawla RK
Published: December 2017

Lyophilized bovine, porcine, and human choroid plexuses contain .02-.09 U of antidiuretic activity per milligram. The antidiuretic factor in bovine choroid plexus was concentrated 100 times by extraction with acetic acid, fractional precipitation with acetone and ethyl ether, gel filtration, and paper chromatography. Resulting choroid plexus fraction IIgammaB2 was eluted from Sephadex G-25 in position corresponding to molecular weight between 750 and 3,500; its antidiuretic activity was destroyed by trypsin, performic acid, and thioglycollic acid, but was not affected by leucine aminopeptidase, carboxypeptidase A or B, or cyanogen bromide. HgammaB2 possesses antidiuretic, pressor, and oxytocic potencies (measured in anesthetized-hydrated rat, anesthetized rat, and isolated rat uterus, respectively) of 1.9, 0.5, and 0.1 U/mg, respectively.
Response to parathyroid hormone in defined segments of proximal tubule
Authors: Hamburger RJ , Lawson NL , Schwartz JH
Published: December 2017

Previous investigations have suggested that there is a functional heterogeneity along the length of the proximal convoluted tubule. This study was designed to confirm and extend these suggestions by examining the intrinsic absorption of fluid and the effect of parathyroid hormone (PTH) on net fluid absorption in isolated, anatomically defined segments of rabbit superficial proximal tubules. The EPCT (early proximal convoluted tubule), LPCT (late proximal convoluted tubule), and PR (pars recta) segments were studied under controlled conditions by the isolated perfused tubule technique. In 23 EPCT, base-line fluid absorption was 1.38 +/- 0.04 (SE), a rate significantly higher than those of 11 LPCT (0.62 +/- 0.02; P less than 0.001) and 12 PR (0.52 +/- 0.03 nl mm-1 min-1, P less than 0.001) segments. In 10 EPCT, mean control fluid absorption was 1.31 +/- 0.04 nl mm-1 min-1; addition of PTH resulted in a decrease to 0.95 +/- 0.05 nl mm-1 min-1 (P less than 0.001); and, after removal of PTH, fluid absorption increased (P less than 0.001). Parathyroid hormone had no effect on either seven LPCT segments or six PR segments. These results demonstrate differences in intrinsic capacity to absorb fluid by anatomically defined segments of the rabbit proximal tubule. This functional heterogeneity is further supported by the observed differential response to PTH by the various anatomic segments of the proximal tubule.
Influence of fasting on lung oxygen consumption and respiratory quotient
Authors: Gregorio CA , Gail DB , Massaro D
Published: December 2017

We measured the oxygen consumption (QO2) of lung slices from rats and rabbits and the respiratory quotient (RQ) of lung slices from fed and fasted rats. The QO2 of lung slices is lowered within 24 h after the onset of food deprivation; this decrease in QO2 lasts during at least 2 additional days of fasting and is not eliminated by addition of glucose to the reaction medium. In fed rats the RQ of lung slices after 30 min of incubation without glucose is 0.75 +/- 0.01 (mean +/- SE) and 0.96 +/- 0.02 with glucose present. Fasting for 72 h lowers the RQ of lung slices after 30 min of incubation without glucose to 0.68 +/- 0.03; addition of glucose raises the RQ of lung slices from 72-h-fasted rats to 0.76 +/- 0.02. We conclude that fasting depresses lung oxidative metabolism. In the fed rat glucose is a major substrate for oxidative processes but in the fasting rat the oxidation of glucose is impaired and lipids are an important source of lung energy.