bluuebunny/crossref_metadata_2025 huggingface dataset:
CNS vasopressin mediates emotional stress and CRH-induced colonic motor alterations in rats
Authors: Bueno L. , Gue M. , Delrio C.
Published: December 2017

The effects of emotional stress (ES) corresponding to conditioned fear on colonic motility and its antagonism by [deamino-Pen1, Val4, D-Arg8]vasopressin, a vasopressin antagonist, were investigated by electromyography in conscious fasted rats fitted with chronically implanted electrodes. A 117% increase (19.6 +/- 2.1 vs. 9.0 +/- 0.9 cycles/10 min during the control period) in the frequency of colonic spike bursts was observed when rats were placed for 30 min in a box in which they had previously received electric foot shocks. Intracerebroventricular (icv) administration of corticotropin-releasing hormone (CRH; 0.5 micrograms/kg) mimicked the effects of ES and increased the spike burst frequency of the colon by 88.6% from 5 to 15 min after its administration. At doses between 5 and 20 micrograms/kg the antagonist [deamino-Pen1, Val4, D-Arg8]vasopressin significantly reduced or abolished the effects of ES and CRH administration on colonic motility. Injected icv at doses of 2.5 and 5 ng/kg [Arg8]vasopressin dose dependently increased the frequency of colonic spike bursts. These effects were not reproduced by similar or higher (50 ng/kg) doses given intraperitoneally, and the effects were abolished after previous administration of vasopressin at a dose of 20 micrograms/kg. It is concluded that the effects of ES on colonic motility in rats previously shown to be linked to the central nervous system (CNS) release of CRH are in turn mediated through the central release of vasopressin.
Actions of somatostatins on gastric smooth muscle cells
Authors: Gu Z. F. , Pradhan T. , Coy D. H. , Mantey S. , Bunnett N. W. , Jensen R. T. , Maton P. N.
Published: December 2017

The effects of somatostatin-28, somatostatin-14, and a synthetic somatostatin octapeptide analogue, D-Phe-Cys-Tyr-D-Trp-Lys-Thr-Cys-Nal-NH2 (cyclo SS-8) were examined on contraction of dispersed gastric smooth muscle cells from guinea pigs. The somatostatins did not cause contraction of gastric smooth muscle cells, nor did they inhibit carbachol-stimulated contraction. However, they reversed vasoactive intestinal peptide (VIP)-induced inhibition (relaxation) of carbachol-stimulated contraction. Somatostatin-28 had a half-maximal effect (EC50) at 1.6 +/- 0.8 nM, cyclo SS-8 at 0.6 +/- 0.3 nM, but somatostatin-14 had no effect even when used in concentrations as high as 1 microM. Incubation of muscle cells with peptidase inhibitors phosphoramidon (1 microM) plus amastatin (10 microM) had no effect on the EC50 of somatostatin-28 or cyclo SS-8 but increased the potency of somatostatin-14 greater than 1,000-fold. When peptides were incubated with muscle cells and the products applied to high-performance liquid chromatography, cyclo SS-8 was not degraded, but somatostatin-14 was rapidly degraded when present alone, and the addition of peptidase inhibitors partially inhibited the degradation. Cyclo SS-8 had its maximal effect at 0.5-1 min and inhibited relaxation induced by VIP, isoproterenol, glucagon, or dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP). Cyclo SS-8 partially inhibited the increase in VIP-stimulated cAMP. Preincubation with pertussis toxin blocked the inhibitory action of cyclo SS-8 on VIP or DBcAMP-induced relaxation. These results indicate that gastric smooth muscle cells rapidly degrade somatostatin-14 and suggest that muscle cell peptidases could have a major effect on the actions of somatostatin-14.(ABSTRACT TRUNCATED AT 250 WORDS)
23Na- and 31P-NMR studies of perfused mouse liver during nitrogen hypoxia
Authors: Bowers J. L. , Lanir A. , Metz K. R. , Kruskal J. B. , Lee R. G. , Balschi J. , Federman M. , Khettry U. , Clouse M. E.
Published: December 2017

The effects of nitrogen hypoxia on isolated perfused mouse livers from fed mice were studied at 37 degrees C using 23Na and 31P nuclear magnetic resonance (NMR) spectroscopy. The paramagnetic shift reagent, dysprosium-triethylenetetraminehexaacetic acid, was used to distinguish intracellular from extracellular sodium. The area of the intracellular sodium resonance remained relatively constant over the first 30 min of hypoxia and then increased by a factor of approximately 2 relative to controls over the next 30 min. High-energy phosphate metabolites were measured using 31P-NMR. The beta-ATP resonance decreased to zero, and the intracellular pH decreased from 7.3 to 6.9 during 60 min of hypoxia. Liver enzyme activity in the effluent exiting the liver increased in direct proportion to the length of hypoxia up to 56 min. The sodium, ATP, and enzyme changes during hypoxia were correlated with histological and electron-microscopic findings. The morphology of liver specimens exposed to 30 min of hypoxia was close to normal, whereas extensive centrilobular and midlobular necrosis was seen in specimens subjected to 60 and 90 min of hypoxia. The effect of 30 min of reoxygenation after 15, 30, 45, 60, and 90 min of hypoxia was also studied. The level of beta-ATP recovery depended on the duration of hypoxia. For 60 min of hypoxia followed by reoxygenation, beta-ATP recovered to only 20% of control values. The morphology of hypoxic livers after 30 min of reoxygenation was similar to livers subjected to hypoxia alone.
Digestive modulation in a seasonal frugivore, the American robin (Turdus migratorius)
Authors: Levey D. J. , Karasov W. H.
Published: December 2017

American robins (Turdus migratorius) switch from eating fruits in the fall to insects in the spring. Our objective was to determine the physiological and morphological changes associated with such a switch. Three nonexclusive hypotheses addressed possible mechanisms operating on different levels. First, we hypothesized that birds on the two diets would differ in intestinal absorption rates of sugars and amino acids. We predicted that individuals on a high-protein low-carbohydrate insect diet would exhibit higher proline and lower glucose absorption than individuals on a low-protein high-carbohydrate fruit-based diet. Uptake rates of glucose and proline were measured in vitro, using an everted sleeve technique. We found no significant differences in uptake between the two groups and so rejected the hypothesis. The second hypothesis, that retention time of digesta in the gut changes with diet, was tested using an inert marker (polyethylene glycol) injected into insects or fruit and fed to birds. We predicted and found that the marker was excreted significantly faster in birds eating fruit rather than insects. Thus this hypothesis is supported. The third hypothesis focused on changes in gut morphology as a basis for shifts in digestive processing. We found no differences in gut length, nominal surface area, or volume and so rejected this hypothesis. Note that hypotheses two and three are closely related, because retention time is proportional to gut volume/digesta flow. Because gut volume did not differ between the two groups, the difference in retention time must have been due to a change in flow. Taken together, these results suggest that short retention times are likely an important adaptation to frugivory.
Ontogeny of intestinal phosphate absorption in rabbits
Authors: Borowitz S. M. , Granrud G. S.
Published: December 2017

The ontogeny of intestinal phosphate transport was examined in brush-border membrane vesicles prepared from 2-wk, 4-wk, 6-wk, and 3-mo-old rabbits. At all four ages, vesicles prepared from the duodenum demonstrated sodium-phosphate cotransport in that uptake in the presence of a sodium gradient was significantly greater than uptake in the presence of a potassium gradient. While sodium-independent phosphate uptake was comparable at all ages, sodium-phosphate cotransport was greatest in the youngest animals and declined with increasing age. Peak phosphate uptake was 10-fold the equilibrium value at 2 wk and declined to 3-fold in adults. Maximal transport capacity of sodium-phosphate cotransport fell from 2,292 +/- 161 at 2 wk to 286 +/- 12 pmol.mg-1.10 s-1 at 3 mo, whereas the Michaelis constant did not change with age, varying between 0.032 and 0.054 mM. At all ages, uptake was half-maximally stimulated between 30 and 50 mM sodium, and Hill coefficients were between 1.5 and 2. Generation of inside negative diffusion potentials did not significantly enhance phosphate uptake at any age. These data suggest that throughout development, the transport of each phosphate molecule across the rabbit duodenal brush-border membrane is coupled to the transport of two sodium molecules. There were distinct developmental changes in the distribution of the sodium-phosphate cotransport along the length of intestine. In 2-wk old animals, sodium-phosphate cotransport was present in the duodenum, proximal and distal jejunum, and proximal ileum. In 4-wk-old animals, sodium-phosphate cotransport was present throughout the duodenum and proximal and distal jejunum, and in 6-wk and 12-wk-old animals, sodium-phosphate cotransport was observed only in the duodenum and proximal jejunum.
Calcium and potassium currents in canine gastric smooth muscle cells
Authors: Sims S. M.
Published: December 2017

Membrane ionic currents were recorded in single smooth muscle cells dissociated from circular muscle of dog stomach (corpus region). When studied under voltage clamp with K+ in the patch electrode, depolarization to potentials more positive than -40 mV, from a holding potential of -70 or -80 mV, evoked transient inward current followed by outward current. Evidence that the outward current was due to K+ came from analysis of deactivation tail currents, which reversed direction close to the K+ equilibrium potential. In addition, the outward current was reduced by tetraethylammonium (TEA, 1-5 mM) applied to the external surface of cells. The Ca(2+)-channel blocker Cd2+ blocked the inward current and also reduced outward current, suggesting Ca(2+)-activated K+ current contributed to the outward current. The voltage-activated inward current was studied in isolation with Cs+ and TEA in the recording electrode to block K+ current. In standard bathing solution containing 2.5 mM Ca2+, the inward current activated between -50 and -40 mV, with peak inward current at +10 mV. The depolarization-activated inward current was blocked by nifedipine and enhanced by BAY K 8644, providing evidence that it was Ca2+ current. The Ca2+ current showed transient and sustained components, both of which showed similar voltage activation and inactivation ranges. The half-inactivation potential was approximately -37 mV. These results provide evidence that smooth muscle cells from the canine gastric corpus possess K+ and Ca2+ channels. Based on the voltage dependence of activation and inactivation and sensitivity to dihydropyridines, L-type Ca2+ channels predominate in canine gastric corpus smooth muscle.
Characterization of a membrane tyrosine phosphatase in AR42J cells: regulation by somatostatin
Authors: Tahiri-Jouti N. , Cambillau C. , Viguerie N. , Vidal C. , Buscail L. , Saint Laurent N. , Vaysse N. , Susini C.
Published: December 2017

A phosphotyrosyl protein phosphatase (PTPase) activity has been characterized in the plasma membranes of confluent AR42J pancreatic tumor cells using 32P-labeled poly(Glu, Tyr) as substrate. Membrane PTPase activity exhibited an apparent Michaelis constant of 3 microM and an apparent maximal velocity of 0.9 nmol.min-1.mg-1. It was inhibited by orthovanadate, zinc, poly(Glu,Tyr) and was stimulated by EDTA and dithiothreitol. Gel filtration of solubilized plasma membranes gave a peak of enzyme activity at a relative molecular weight of 70,000. Plasma membrane PTPase activity was changed during AR42J cell growth. At the beginning of culture, the control PTPase activity was minimal. Over the 5 days of culture, PTPase activity increased to reach a maximum (3.5-fold over control activity) preceding confluency by 2 days. Then the high level of PTPase activity was sustained until confluency. Incubation of the cells with the stable somatostatin analogue SMS 201-995 (SMS) resulted in a rapid and transient activation of crude membrane PTPase activity. Activation reached a maximum level within 5 min of addition and return to control levels within 20 min. The effect of SMS was dose dependent with half-maximal and maximal activation occurring at 6 pM and 0.1 nM SMS respectively.
Effect of osmotic swelling on K+ conductance in jejunal crypt epithelial cells
Authors: MacLeod R. J. , Lembessis P. , Hamilton J. R.
Published: December 2017

To further elucidate differences in ion transport properties between jejunal crypt and villus cells, we compared the responses of purified cell suspensions to hypotonic stress using electronic cell sizing to evaluate volume changes and 86Rb and 36Cl efflux. After hypotonic swelling, villus enterocytes undergo a regulatory volume decrease (RVD) due to the loss of K+ and Cl- through volume-activated conductances. After 0.6x isotonic challenge in Na(+)-free medium, crypt cells exhibited only partial RVD, with t1/2 congruent to 15 min. The addition of a cation ionophore, gramicidin (0.25 microM), to hypotonically swollen crypt cells caused an accelerated RVD, which was complete with t1/2 congruent to 5 min. Crypt epithelial cells showed no volume-activated 86Rb efflux, but villus enterocytes had an increased rate of 86Rb efflux after hypotonic dilution (P less than 0.001). Gramicidin added to hypotonically diluted crypt cells greatly increased the rate of 86Rb efflux compared with controls. Both villus (30 s; P less than 0.005) and crypt (2 min; P less than 0.001) cells exhibited volume-activated 36Cl efflux in absence of gramicidin. Cl- channel blockers anthracene-9-carboxylate (9-AC, 300 microM) and indanyloxyacetic acid (IAA-94, 100 microM) prevented crypt RVD (P less than 0.001) in the presence of gramicidin. Ouabain (P less than 0.001) or K(+)-free Na(+)-containing medium, but not Ba2+ (5 mM) or quinine (100 microM), prevented crypt partial RVD. We conclude that crypt cells lack volume-activated K+ conductance. The RVD exhibited by crypt cells, although partial, was due to Cl- loss through a volume-activated Cl- conductance and Na+ loss via Na(+)-K(+)-ATPase.
Polyamine transport systems in isolated rat hepatocytes derived from resting and regenerating livers
Authors: Minuk G. Y. , Bennaroch A. , Ding L. X.
Published: December 2017

Polyamines and their principal metabolite, gamma-aminobutyric acid (GABA), modulate eukaryotic cell growth. To determine whether the liver possesses specific polyamine transport sites and whether changes occur to these or GABA transport sites during hepatic regeneration, suspensions of rat hepatocytes derived from in situ collagenase perfusions of livers at times 0, 24, 48, and 72 h post-partial hepatectomy were incubated at 4, 20, and 37 degrees C with various concentrations of the following ligands: [3H]putrescine, [3H]spermidine, [14C]spermine, and [3H]GABA together with or without excess unlabeled ligand, KCN, ouabain, or digitoxigenin. Of the ligands studied, only [14C]spermine and [3H]GABA were associated with specific binding to hepatocytes derived from nonregenerating livers. Spermine binding correlated with the concentration of hepatocytes in the incubation mixture and reached equilibrium within 60 min. The approximate affinity constant (KD) was 5.5 x 10(-5) mol/10(6) hepatocytes, and maximum number of binding sites (Bmax) was 1.8 +/- 1.2 x 10(-7) mol.10(6) hepatocytes-1.min-1. Binding was neither temperature nor sodium dependent and was not inhibited by KCN, ouabain, digitoxigenin, other polyamines, or GABA. Aside from a 43% decrease in spermine binding at 24 h post-partial hepatectomy [5.1 +/- 1.1 vs. 8.9 +/- 3.1 x 10(3) disintegrations per minute (dpm)/10(6) hepatocytes at time 0, P less than 0.05] and a 39% decrease in GABA binding (3.4 +/- 1.3 vs. 5.5 +/- 1.9 x 10(3) dpm/10(6) hepatocytes, P less than 0.05), there were no significant changes in ligand binding during hepatic regeneration.(ABSTRACT TRUNCATED AT 250 WORDS)
Measurement of axial forces during emptying from the human stomach
Authors: Vassallo M. J. , Camilleri M. , Prather C. M. , Hanson R. B. , Thomforde G. M.
Published: December 2017

Our aim was to measure axial forces in the stomach and to evaluate their relation to circumferential contractions of the gastric walls and the emptying of gastric content. We used a combination of simultaneous radioscintigraphy, gastroduodenal manometry, and an axial force transducer with an inflatable 2-ml balloon fluoroscopically placed in the antrum. In vitro studies demonstrated that the axial force transducer records only antegrade forces along the longitudinal axis of this probe in an intensity-dependent manner. In vivo studies were performed in five healthy subjects for at least 3 h after ingestion of radiolabeled meals. When administered separately, the emptying of liquids or solids from the stomach is associated with generation of antral axial forces and coincident phasic pressure activity; however, almost 20% (average) of gastric axial forces during emptying of liquids or solids are unassociated with proximal or distal antral pressure activity ("isolated" forces). High amplitude antral axial forces and pressures occur during both lag and postlag emptying phases. During emptying of liquids, there is a trend for axial forces to be coincident more often with proximal than with distal antral pressure activity and vice versa for the emptying of solids (P = 0.015). These data suggest that when placed in the antrum, the transducer can semiquantitatively record axial forces during gastric emptying. By combining these observations with the data from in vitro studies, it appears that axial forces predominantly result from traction on the balloon by the longitudinal vector resulting from circumferential gastric contractions. The combination of radioscintigraphy and measurement of antral axial forces is a promising method to evaluate mechanical forces involved in the emptying of the human stomach.
Bombesin receptors interact with Gi and p21ras proteins in plasma membranes from rat pancreatic acinar cells
Authors: Profrock A. , Zimmermann P. , Schulz I.
Published: December 2017

This study investigates interaction of bombesin receptors with heterotrimeric guanosine triphosphate (GTP)-binding proteins (G proteins) and monomeric small molecular weight GTP-binding proteins (smg proteins), respectively, in plasma membranes (PM) of rat pancreatic acinar cells. Addition of bombesin to isolated PM stimulated the incorporation of the photoaffinity analogue [alpha-32P]GTP-gamma-azidoanilide into Gi proteins of 40-41 kDa and reduced the pertussis toxin-induced ADP ribosylation of three 40-41 kDa proteins, which had been previously identified as Gi1, Gi2, and Gi3 (30). In PM isolated from bombesin-prestimulated acinar cells, binding of [alpha-32P]-GTP to PM proteins of 21-22 kDa and of a monoclonal antibody against p21ras proteins was increased. Two-dimensional separation of PM proteins revealed the presence of 18 or 19 differently charged smg proteins. The p21ras proteins could be separated into two differently charged proteins with isoelectric points of 5.58 and 5.79. In microsomal membranes (MM), [alpha-32P]GTP binding to yet unidentified 21-22 kDa smg proteins was decreased compared with membranes from unstimulated acinar cells. The data suggest that Gi proteins as well as p21ras proteins are involved in bombesin receptor-mediated signal transduction in the PM. Furthermore, 21-22 kDa smg proteins in MM might play a role in bombesin-induced stimulation of intracellular pathways that lead to enzyme secretion from pancreatic acinar cells.
Two-dimensional coupling by gap junctions in cultured gastric smooth muscle monolayers
Authors: Larson D. M. , Gilbert R. J. , Beyer E. C.
Published: December 2017

We studied intercellular transfer in cultured rabbit gastric smooth muscle cell monolayers after microinjection of electrotonic current or the fluorescent probe Lucifer yellow CH. Because cultured gastric muscle cells proliferate in vitro and form regular arrays of parallel spindle-shaped cells, we sought to assess the role of cell shape and orientation in determining two-dimensional coupling properties. With the use of electron microscopy, gap junctions were identified between adjacent cells. Northern blot analyses using specific cDNA probes demonstrated expression of mRNA for the gap junction protein connexin43. Dye injection of Lucifer yellow resulted in 97% transfer to at least one adjacent cell, and 88% of adjacent cells received dye. Electrophysiological studies were performed using two intracellular microelectrodes to measure electrotonic current flow between cells at varying interelectrode distances. Current flow in the monolayers was modeled using a modified two-dimensional analysis. Initial assessment showed that the ratio of calculated space constants (longitudinal axis/perpendicular axis) was 4.4, indicating anisotropic conditions. However, when a geometric transform was used to normalize the spindle-shaped cells to regular hexagons, the space constants became statistically equivalent (200 microns longitudinal, 256 microns perpendicular). These results suggest that anisotropy of current flow in the monolayer of gastric smooth muscle cells was due primarily to the shape of the cells and not to intrinsic membrane properties or the distribution of gap junctions.
Regional differences in gut blood flow and mucosal damage in response to ischemia and reperfusion
Authors: Leung F. W. , Su K. C. , Passaro E. , Guth P. H.
Published: December 2017

Ischemia and reperfusion of the small intestine and colon in rats were produced by reversible occlusion (for 30 min and 1 or 3 h) of the superior mesenteric artery and the aorta above the inferior mesenteric artery. Despite a greater reduction of mucosal perfusion in the colon than in the small intestine with 30 min of ischemia, the depth of mucosal damage was significantly smaller in the former than in the latter. Thirty minutes of ischemia followed by 1 h of reperfusion induced an increase in polymorphonuclear leukocyte infiltration in both locations. Exacerbation of mucosal injury occurred only in the small intestine, suggesting that reperfusion injury is independent of polymorphonuclear leukocyte infiltration. Reperfusion after 1 or 3 h of ischemia did not exacerbate mucosal damage in either location. Allopurinol significantly diminished the exacerbation of injury after reperfusion in the small intestine. The protective effect of allopurinol, however, was neither associated with an improvement in perfusion nor a reduction in polymorphonuclear leukocyte infiltration. These data indicate that there is a window (30 min) of reperfusion injury in the small intestine, but there is no evidence of reperfusion injury in the colon.
Effects of nerve stimulation and zymosan on glycogenolysis in perfused livers from cold-exposed rats
Authors: Shiota M. , Kurano Y. , Mochizuki Y. , Kimura K. , Ohta M. , Sugano T.
Published: December 2017

The effects of sympathetic nerve stimulation and zymosan (cell wall particles from yeast) on glycogenolysis were studied in perfused livers from rats kept for 5 and 20 days at 4 degrees C. The rate of glycogenolysis induced by nerve stimulation decreased significantly without any decrease in norepinephrine outflow during cold exposure, and the rate induced by norepinephrine did not change. By contrast, the rate of zymosan-induced glycogenolysis increased markedly during cold exposure. The rats with denervated hepatic nerves did not show the increased response to zymosan. In cold-exposed rats, both mepacrine and ibuprofen inhibited the effects of zymosan and of nerve stimulation without any inhibition of the outflow of norepinephrine. Neither inhibitor had any effect on the effects of norepinephrine. The metabolic effects of nerve stimulation and zymosan were not additive in cold-exposed rats. These results suggest that cold exposure may modulate the metabolism of arachidonic acid in Kupffer cells via hepatic nerve and decrease the eicosanoid-dependent glycogenolysis by nerve stimulation.
A primary role for protein kinase A in smooth muscle relaxation induced by adrenergic agonists and neuropeptides
Authors: Gu Z. F. , Jensen R. T. , Maton P. N.
Published: December 2017

Many studies suggest that smooth muscle relaxation caused by beta-adrenergic agents and various neuropeptides occurs as a result of an increase in cellular adenosine 3',5'-cyclic monophosphate (cAMP). However, the evidence is indirect, and furthermore does not demonstrate that an increase in cAMP is essential for mediating relaxation. To define more clearly the role of cAMP in receptor-mediated smooth muscle relaxation, we used a specific competitive antagonist of the action of cAMP on protein kinase A, (R)-p-adenosine 3',5'-cyclic phosphorothioate [(R)-p-cAMPS], and its S isomer, (S)-p-cAMPS, which functions as a cAMP agonist. In gastric smooth muscle cells from guinea pig, (S)-p-cAMPS caused a dose-related relaxation [50% inhibitory concentration (IC50) 86 +/- 59 nM]. Vasoactive intestinal peptide (VIP) produced smooth muscle cell relaxation (IC50 2.3 +/- 0.8 nM) through occupation of specific VIP receptors. (R)-p-cAMPS inhibited VIP-induced relaxation, with a rightward shift in the VIP dose-response curve, suggesting competitive antagonism. Furthermore, (R)-p-cAMPS inhibited relaxation induced by other agents that increase cellular cAMP (isoproterenol, calcitonin gene-related peptide, and glucagon) but not that induced by ATP or sodium nitroprusside. (R)-p-cAMPS had no effect on contraction stimulated by carbachol, cholecystokinin, or substance P. These data demonstrate that activation of protein kinase A is primarily responsible for mediating gastrin smooth muscle relaxation produced by adrenergic agents and various neuropeptides.
Rebound elevation of fibronectin after tissue injury and ischemia: role of fibronectin synthesis
Authors: Thompson P. N. , Cho E. , Blumenstock F. A. , Shah D. M. , Saba T. M.
Published: December 2017

Plasma fibronectin (pFn) stimulates macrophage phagocytosis of tissue debris; pFn deposition in tissues may influence vascular integrity. Although the acute depletion of pFn after surgery and/or injury has been described, less attention has been given to the rebound hyperfibronectinemia presumably "triggered" by the early pFn depletion. Using a model that compartmentalized the site of tissue injury and thus attenuated the initial pFn depletion, we studied this rebound elevation of pFn in anesthetized rats (250-350 g) after the surgical trauma of groin dissection alone (sham group) or surgery coupled with 4 h of hindlimb ischemia (experimental group). Nonoperated control rats were also anesthetized. Shams had baseline (preoperative) 6-, 8-, and 22-h postoperative pFn levels of 573 +/- 61, 598 +/- 62, 695 +/- 57, and 929 +/- 87 micrograms/ml, respectively. In the surgery-ischemia group, pFn also elevated to 1,117 +/- 40 micrograms/ml at 22 h postsurgery. Nonoperated control rats (only anesthetized) had no elevation of pFn. Intravenous infusion of gelatin-coated lipid particles (50 mg/100 g) depleted pFn by 89.3% but was unable to prevent the rebound elevation of pFn. The blood clearance of 125I-labeled pFn was very similar in control, sham, and experimental rats. In contrast, pFn synthesis over the 22-h period was dramatically altered and equal to 2.12 +/- 0.16, 3.40 +/- 0.56, and 4.49 +/- 0.17 mg pFn synthesized/100 g body wt, in control, sham, and experimental rats respectively. Thus a rapid increase in pFn synthesis contributes to the rebound hyperfibronectinemia after sublethal surgical injury.(ABSTRACT TRUNCATED AT 250 WORDS)
Mouse hepatocyte membrane potential and chloride activity during osmotic stress
Authors: Wang K. , Wondergem R.
Published: December 2017

Hepatocyte transmembrane potential (Vm) during osmotic stress responds as an osmometer, in part because of changes in membrane K+ conductance. This may contribute to the electromotive force that drives transmembrane Cl- fluxes. To test this, double-barreled ion-sensitive microelectrodes were used to measure changes in steady-state intracellular Cl- activity (aiCl) during osmotic stress applied to mouse liver slices. Hyperosmotic and hyposmotic conditions were created by rapidly switching to a solution in which sucrose concentrations were increased or reduced, respectively. Hyperosmotic stress [1.4 x control osmolality (280 mosmol/kgH2O)] decreased hepatocyte Vm 46% from -39 +/- 1 to -21 +/- 1 mV (SE; n = 16 animals). Corresponding aiCl increased twofold from 19 +/- 2 to 38 +/- 3 mM. This shifted the Cl- equilibrium potential (ECl) 19 mV, from -38 +/- 0.3 to -19 +/- 2 mV. Hyposmotic stress [0.71 x control osmolality (290 mosmol/kgH2O)] increased hepatocyte Vm 64% from -28 +/- 1 to -46 +/- 1 mV (SE; n = 13 animals). Corresponding aiCl decreased 0.53-fold from 17 +/- 1 to 8 +/- 1 mM. This shifted the ECl 20 mV from -26 +/- 2 to -46 +/- 3 mV. Thus hepatocyte aiCl is in electrochemical equilibrium with Vm. The paired measurements above were repeated after addition of K(+)-channel blockers quinine or Ba2+. Ba2+ (2 mM) had no effect on either Vm or aiCl during hyperosmotic stress; however, Ba2+ significantly inhibited changes in Vm and aiCl during hyposmotic stress. Effects of quinine (0.5 mM) on Vm and aiCl during both hyperosmotic stress and hyposmotic stress were similar to those of Ba2+.(ABSTRACT TRUNCATED AT 250 WORDS)
Glutathione as a primary osmotic driving force in hepatic bile formation
Authors: Ballatori N. , Truong A. T.
Published: December 2017

Indirect evidence suggests that transport of glutathione (GSH) across the canalicular plasma membrane into bile contributes to the formation of the bile acid-independent fraction of bile flow. To directly test this hypothesis, the present study measured bile flow in isolated perfused rat livers whose biliary GSH excretion rate was selectively modulated by administration of GSH monoethyl ester (50, 100, and 200 mumol infused over a 20-min interval), a high dose of GSH itself (550 mumol over 20 min), and the three amino acid components of GSH (70 mumol each) with and without methionine (35 mumol). Animals were starved overnight to decrease hepatic GSH levels, and livers were pretreated with acivicin to inhibit gamma-glutamyl transferase. Livers perfused single pass with Krebs-Henseleit buffer excreted bile acids at a relatively low rate of 1-3 nmol.min-1 x g-1, and this rate was unaffected by agents used to alter biliary GSH efflux. In comparison, basal biliary GSH efflux rates were 8-13 nmol.min-1 x g-1. Administration of the GSH ester produced a dramatic dose-dependent choleresis, a stimulation of biliary GSH excretion, and resulted in the biliary excretion of intact GSH ester. Changes in total biliary GSH excretion and bile flow were temporally and quantitatively related. Infusion of GSH and amino acid supplementation also resulted in higher rates of bile flow and biliary GSH excretion, but their effects were more modest.(ABSTRACT TRUNCATED AT 250 WORDS)
Actions of 5-hydroxytryptamine on myenteric neurons in guinea pig gastric antrum
Authors: Tack J. F. , Janssens J. , Vantrappen G. , Wood J. D.
Published: December 2017

Intracellular recording methods were used to study the actions of 5-hydroxytryptamine (5-HT) on 257 myenteric neurons in the guinea pig gastric antrum. Application of 5-HT caused three types of postsynaptic responses. A fast-activating depolarizing response was accompanied by a decreased input resistance and desensitized quickly to repeated applications. It was mediated by a 5-HT3 receptor. A slowly activating depolarization, accompanied by an increase in the input resistance and enhancement of the excitability, was mainly observed in after hyperpolarizing/type 2 neurons. It was suppressed by the prokinetic benzamide compound renzapride, while classical 5-HT1-4 receptor antagonists had no effect, suggesting the involvement of a 5-HT1p receptor as described in small intestinal neurons. A long-lasting hyperpolarizing response, accompanied by a decreased input resistance, was observed in a small subset of neurons. This response seemed to be mediated by a 5-HT1a receptor. Superfusion of 5-HT caused a dose-dependent inhibition of the stimulus-evoked nicotinic cholinergic fast excitatory postsynaptic potential (EPSP), which was mediated by a presynaptic 5-HT1a receptor. 5-HT also presynaptically inhibited the slow EPSP.
Neuroimmune interactions: role for cholinergic neurons in intestinal anaphylaxis
Authors: Javed N. H. , Wang Y. Z. , Cooke H. J.
Published: December 2017

The role of cholinergic neurons in mediating chloride secretion in anaphylaxis was assessed in muscle-stripped segments of distal colon from guinea pigs immunized to bovine milk. beta-Lactoglobulin evoked a concentration-dependent increase in short-circuit current (Isc) in immune, but not nonimmune, tissues. The Isc response to beta-lactoglobulin was reduced by piroxicam, pyrilamine, and cimetidine. Tetrodotoxin and atropine reduced the Isc response to beta-lactoglobulin in immune animals, whereas mecamylamine and ICS 205-930 were ineffective. beta-Lactoglobulin evoked a concentration-dependent increase in acetylcholine (ACh) release in immune, but not nonimmune, animals. In immune tissues after challenge with beta-lactoglobulin, ACh release paralleled the change in Isc. Piroxicam, cimetidine plus pyrilamine, or a combination of piroxicam, cimetidine, and pyrilamine significantly reduced the release of ACh after beta-lactoglobulin challenge. Histamine, dimaprit, and prostaglandins E2 evoked an increase in ACh release. These results suggest that beta-lactoglobulin releases prostaglandins and histamine probably from mast cells. Secretory responses that occur when immune animals are challenged with beta-lactoglobulin result, in part, from activation of cholinergic neurons that utilize muscarinic synapses for transfer of signals to the epithelium.
Release of nitric oxide by activation of nonadrenergic noncholinergic neurons of internal anal sphincter
Authors: Chakder S. , Rattan S.
Published: December 2017

The purpose of the present study was to investigate the direct release of nitric oxide (NO) in response to the stimulation of nonadrenergic noncholinergic (NANC) nerves. The studies were performed on isolated smooth muscle strips of the opossum (Didelphis virginiana) internal anal sphincter (IAS). Electrical field stimulation (EFS) using the appropriate parameters caused a frequency-dependent fall in the resting tone of the IAS. The release of NO was measured directly by the chemiluminescence method. The stimulation of NANC neurons by EFS and the nicotinic stimulant 1,1-dimethyl-4-phenylpiperazinium (DMPP) caused IAS relaxation with an accompanying release of NO. The release of NO and the fall in the resting tension of IAS in response to lower frequencies of EFS and DMPP were abolished by pretreatment of the smooth muscles with the neurotoxin tetrodotoxin and the NO-synthase inhibitor NG-nitro-L-arginine (L-NNA). The obliteration of the release of NO and the IAS relaxation in the presence of L-NNA reversed to control levels by the addition of the NO precursor L-arginine. The effect of L-NNA and L-arginine on NO release and IAS relaxation was stereoselective, since D-NNA and D-arginine had no significant effect. Vasoactive intestinal polypeptide also caused release of NO from IAS smooth muscle strips, which was abolished by L-NNA. However, isoproterenol and atrial natriuretic factor caused IAS relaxation without any increase in NO release. In conclusion, these studies demonstrate the direct release of NO in response to the stimulation of NANC inhibitory neurons of the gut.
Ursodeoxycholate mobilizes intracellular Ca2+ and activates phosphorylase a in isolated hepatocytes
Authors: Bouscarel B. , Fromm H. , Nussbaum R.
Published: December 2017

In isolated hamster hepatocytes, ursodeoxycholic acid (UDCA) mobilized intracellular free calcium ([Ca2+]i) and activated phosphorylase a with a half-maximally effective concentration of 188 and 9 microM, respectively. Addition of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) did not affect the maximum [Ca2+]i mobilized by UDCA; however, [Ca2+]i returned to basal levels in 4-5 min compared with > 10 min in the absence of EGTA. Both UDCA and vasopressin activated phosphorylase a to the same extent in the presence and absence of extracellular Ca2+, and the effect of both agents was abolished when the cells were depleted in Ca2+. Vasopressin (100 nM) did not further mobilize [Ca2+]i or activate phosphorylase a when combined with 500 microM UDCA. However, unlike vasopressin, UDCA did not stimulate inositol 1,4,5-trisphosphate (IP3) formation. In contrast to taurine-conjugated UDCA (TUDCA), concentration < or = 500 microM of glycine-conjugated UDCA (GUDCA) did not affect either [Ca2+]i or phosphorylase a. Lithocholic acid and taurolithocholic acid (TLCA) displayed the highest affinity for Ca2+. In addition, TLCA, chenodeoxycholic acid, and NaF stimulated Ca2+ efflux at concentrations as low as 100 microM, 200 microM, and 5 mM, respectively. Conversely, UDCA, TUDCA, and GUDCA presented the lowest affinity for Ca2+ and had no effect on Ca2+ efflux. The 28% increase in Ca2+ release induced by TLCA alone was further augmented to approximately 60% when TLCA was combined with UDCA, TUDCA, or GUDCA. However, Ca2+ efflux induced by NaF was not further increased by UDCA and its conjugates.(ABSTRACT TRUNCATED AT 250 WORDS)
Role of calcium in chloride secretion mediated by cAMP pathway activation in rabbit distal colon mucosa
Authors: Calderaro V. , Chiosi E. , Greco R. , Spina A. M. , Giovane A. , Quagliuolo L. , Servillo L. , Balestrieri C. , Illiano G.
Published: December 2017

Effects of Ca2+ on adenosine 3',5'-cyclic monophosphate (cAMP)-mediated Cl- secretion were investigated in intact mucosa and isolated crypt cells of rabbit descending colon. Addition of 10 microM prostaglandin (PG)E2 or forskolin to tissues incubated in Ca(2+)-free medium increased the size of short-circuit current (Isc) and Cl- secretion as estimated by unidirectional 36Cl flux measurements (net flux = -2.31 +/- 0.24 vs. -1.22 +/- 0.10 mueq.h-1.cm-2, n = 4, P < 0.001). Addition of 10 microM PGE2 to tissues incubated in 1.2 mM Ca2+ Ringer induced a 7-fold increase in mean cAMP level, whereas it produced an 11-fold increase in tissues exposed to Ca(2+)-free medium. Membrane preparations from whole mucosa incubated in Ca(2+)-free medium displayed a cyclic nucleotide phosphodiesterase activity significantly lower than controls (18.76 +/- 0.54 vs. 31.20 +/- 0.39 pmol cAMP. mg protein-1.min-1, means +/- SE, n = 4, P < 0.001). Ca2+ removal also affected adenylate cyclase (AC) responsiveness to agonists; AC activity increased in controls by 54 and 226% after stimulation with 10 microM PGE2 and forskolin, respectively, but it increased more (77 and 325%, respectively) after incubation in Ca(2+)-free solutions.(ABSTRACT TRUNCATED AT 250 WORDS)
Role of nitric oxide in gastric hyperemia induced by central vagal stimulation
Authors: Tanaka T. , Guth P. , Tache Y.
Published: December 2017

The effects of N omega-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide (NO) synthesis, on the vagal cholinergic increase in gastric mucosal blood flow (GMBF) and acid secretion induced by intracisternal injection of the thyrotropin-releasing hormone (TRH) analogue, RX 77368, were studied. GMBF and acid secretion were measured simultaneously by the hydrogen gas clearance technique and titration of gastric perfusate in urethan-anesthetized rats. RX 77368 (30 ng) injected intracisternally stimulated gastric acid secretion and GMBF for 90 and 180 min respectively. GMBF was increased from basal 63 +/- 4 to 166 +/- 14 ml.min-1.100 g-1 at 60 min postinjection. L-NAME (3 mg/kg) injected intravenously 15 min before RX 77368 completely prevented the increase in GMBF induced by the TRH analogue, whereas the acid response was not modified. The effect of L-NAME was reversed by L-arginine but not by the stereoisomer D-arginine. These results show that the increase in GMBF, but not the stimulation of acid secretion, induced by central vagal activation is mediated through a product of L-arginine-NO pathway.
Site density of mouse intestinal glucose transporters declines with age
Authors: Ferraris R. P. , Hsiao J. , Hernandez R. , Hirayama B.
Published: December 2017

To evaluate the effect of age on nutrient transport, the absorption rates of D-glucose, D-fructose, L-alanine, L-aspartate, L-leucine, L-lysine, L-proline, folic acid, and nicotinamide were determined in isolated jejunal tissues of young (6.7 mo old) and aged (23.7 and 27.0 mo old) mice (COBS:SFW). D-Glucose and D-fructose uptakes per milligram tissue were approximately 20-120% higher in the proximal jejunum and 15-50% higher in the distal jejunum of young mice. Amino acid and vitamin uptakes per milligram were also higher in young mice, but differences were not statistically significant. The number of Na(+)-D-glucose transporters per milligram tissue as estimated by specific phlorizin binding decreased with age. There was no age-related change in passive L-glucose permeability, in Kd of specific phlorizin binding, in transporter turnover rate, and in the molecular weight of the Na(+)-D-glucose transporter. Thus a reduction in D-glucose transporter site density fully accounts for the age-related decline in D-glucose transport rate per milligram small intestine.
Role of carboxyl and sulfhydryl residues on rabbit small intestinal brush-border membrane Na(+)-glucose cotransporter
Authors: Peerce B. E. , Cedilote M. , Clarke R. D.
Published: December 2017

The role of sulfhydryl (SH) and carboxylic acid residues in Na(+)-dependent glucose uptake, Na(+)-dependent phlorizin binding, and substrate exchange by the rabbit small intestinal brush-border membrane (BBM) Na(+)-glucose cotransporter was examined in sodium dodecyl sulfate-BBM vesicles. The sulfhydryl reagent p-chloromercuribenzoate (PCMB) inhibited all three measures of cotransporter function in a dithiothreitol-sensitive manner with similar K0.5 values (concn of PCMB resulting in 50% inhibition). PCMB sulfonate had no effect on Na(+)-glucose cotransporter function < 250 microM. The carboxylic acid reagent 1-ethyl-3-(4-azonia-4,4-dimethylpentyl)carbodiimide no effect on Na(+)-glucose cotransporter function. N,N'-dicyclohexylcarbodiimide (DCCD) inhibited all three measures of cotransporter function with similar K0.5 values for inhibition. Inhibition by DCCD did not require addition of a nucleophile. In contrast, PCMB-pretreated cotransporter was insensitive to DCCD in the absence of added nucleophile with respect to substrate transport (Na(+)-dependent glucose uptake) but not Na(+)-dependent phlorizin binding. These results indicate an intravesicular or lipophilic environment for both the PCMB-reactive SH residue and the DCCD-reactive carboxylic acid residues, suggesting that a SH residue may act as an endogenous nucleophile for interaction of DCCD with the Na(+)-glucose cotransporter and suggesting that different carboxylic acid residues may be involved in Na(+)-dependent glucose uptake and Na(+)-dependent phlorizin binding.
Ontogeny of sucrase-isomaltase gene expression in rat intestine: responsiveness to glucocorticoids
Authors: Nanthakumar N. N. , Henning S. J.
Published: December 2017

The goal of our study was to determine how sucrase-isomaltase (SI) gene expression in the rat small intestine is modulated by glucocorticoids during the second and third postnatal weeks. SI mRNA was quantitated by Northern blot and was compared with sucrase activities in the same tissue samples. The role of endogenous glucocorticoids was assessed by measuring SI mRNA and enzyme activity in rat pups adrenalectomized (ADX) on postnatal day 9. ADX pups showed a retarded appearance of sucrase activity compared with sham-operated control pups, and the appearance of SI mRNA paralleled the enzyme activity. To determine the role of exogenous glucocorticoids, a saturating dose of dexamethasone (Dex) was administered daily in three series of experiments starting on days 10, 16, and 18. In the day 10 series, Dex caused precocious appearance of both SI mRNA and sucrase activity. In the day 16 series, Dex accelerated the rate of rise of the two parameters, whereas by day 18 there was no significant effect of Dex. To investigate whether the accelerated rise in the day 16 series was associated with changes in epithelial cell kinetics, the location of SI protein was assessed by immunofluorescence staining. The results indicated that the effect of Dex at this age was due to faster emergence of SI-bearing enterocytes from the intestinal crypts.(ABSTRACT TRUNCATED AT 250 WORDS)
Effects of PACAP on morphologically identified myenteric neurons in guinea pig small bowel
Authors: Christofi F. L. , Wood J. D.
Published: December 2017

Intracellular microelectrodes were used to examine the actions of pituitary adenylate cyclase-activating peptide (PACAP) on morphologically identified myenteric neurons and glial cells of the guinea pig small bowel. PACAP-27 and PACAP-38 evoked excitatory responses in 96% of after hyperpolarizing (AH)/type 2 neurons. The half-maximal concentration for PACAP-27 was 1.5 nM. The responses consisted of membrane depolarization in association with increased input resistance, suppression of hyperpolarizing afterpotentials, and repetitive spike discharge. Forskolin mimicked the action of PACAP in all AH/type 2 neurons. PACAP excited 36% of S/type 1 neurons. Most of the AH/type 2 neurons had Dogiel II morphology, whereas the S/type 1 neurons were uniaxonal with morphology characteristics of Dogiel I or filamentous neurons. No glial cells responded to PACAP. A selective A1 adenosine receptor agonist blocked the excitatory action of PACAP, and this was reversed by a selective A1 antagonist. The results suggest that excitatory PACAP receptors and inhibitory adenosine A1 receptors are linked to adenylate cyclase in AH/type 2 myenteric neurons.
Protective action of oral N-acetylcysteine against gastric injury: role of hypertonic sodium
Authors: Barreto J. C. , Smith G. S. , Tornwall M. S. , Miller T. A.
Published: December 2017

N-acetylcysteine (NAC), when administered orally as a 20% solution, is a potent protective agent against gastric injury in the rat stomach induced by absolute ethanol. The present study was undertaken to define the means by which this protection is mediated. The notion that NAC acts as a glutathione precursor was excluded when N-acetylserine (NAS) was noted to be equally protective against alcohol injury. The NAS molecule contains a hydroxyl moiety at the site where NAC contains a sulfhydryl. To orally administer 20% NAC at a neutral pH, NaOH is added to the free acid form to keep NAC in solution. We determined by titration that a sodium concentration of 1.2 M results. Thus it became apparent that the protective effect of NAC might be mediated through the sodium employed to titrate NAC. Accordingly, we examined various sodium salts and assessed their relative protective effects against alcohol injury. Both sodium acetate and sodium chloride in 1 M solutions were found to be equally effective in preventing alcohol injury with the same efficiency as 1 M sodium solutions of NAC and NAS, excluding the acetate portion of NAC and NAS as being of primary importance for protection to occur. Further study, using different concentrations of sodium chloride (i.e., 150-1,000 mM) revealed that the 1 M solution was most optimal in preventing alcohol injury. One molar sodium by itself and when administered as part of the NAC solution also prevented gastric injury by concentrated acid and base.(ABSTRACT TRUNCATED AT 250 WORDS)