bluuebunny/crossref_metadata_2025 huggingface dataset:
Glucose absorption by a nectarivorous bird: the passive pathway is paramount
Authors: Karasov W. H. , Cork S. J.
Published: December 2017

We tested the hypothesis that most sugar absorption across the small intestine's brush border is normally by a mediated pathway, i.e., the Na(+)-glucose cotransporter. In nectar-eating rainbow lorikeets (Trichoglossus haematodus, 120 g), we measured mediated D-glucose uptake in vitro using the everted-sleeve technique. The apparent Michaelis constant (7.7 mM) was similar to that observed in hummingbirds and other birds in general. Maximal mediated D-glucose uptake summed along the entire length of intestine (48 cm) was not notably high (5.34 mumol/min) when compared with other avian species and was an order of magnitude too low to explain observed rates of glucose absorption in vivo (54.5 mumol/min). This implied that nonmediated glucose absorption predominated, and independent verification of that was sought. We applied a pharmacokinetic technique to measure in vivo absorption of L-glucose, the stereoisomer that does not interact with the Na(+)-glucose cotransporter. Eighty percent of L-glucose that was ingested was absorbed, confirming that nonmediated absorption can be substantial. We discuss how equating L-glucose absorption with passive D-glucose absorption depends on certain assumptions regarding the relative importance of diffusive and convective passive flux. In either case, the conclusion about the relative importance of passive absorption should still hold.
Effect of adrenergic and cholinergic agents on esophageal bicarbonate secretion in opossums
Authors: Hamilton B. H. , Tobey N. A. , Starnes M. C. , Schreiner V. J. , Orlando R. C.
Published: December 2017

The effects of cholinergic and adrenergic agonists and antagonists on esophageal bicarbonate secretion in the opossum were studied in vivo using a recirculated unbuffered saline solution and pH stat technique. The basal rate of secretion was 0.28 +/- 0.02 mumol.h-1.cm-2, and this increased in a dose-dependent manner by intravenous administration of carbachol (maximal increase 0.74 +/- 0.07 mumol.h-1.cm-2 at 6 micrograms/kg). Furthermore, like carbachol, the acetylcholinesterase inhibitor edrophonium also increased bicarbonate secretion, whereas atropine and pirenzepine, which had no effect on basal secretion, abolished the increase in secretion produced by carbachol. Intravenous administration of either adrenergic agonists or antagonists had no significant effect on secretion. These data indicate that basal bicarbonate secretion in the opossum esophagus is independent of intrinsic adrenergic or cholinergic activity, and so not under autonomic nervous system control, but that endogenous release of acetylcholine, presumably from parasympathetic nervous fibers, can stimulate bicarbonate secretion through activation of cholinergic M1 receptors.
Rapid reversible substrate regulation of fructose transporter expression in rat small intestine and kidney
Authors: Burant C. F. , Saxena M.
Published: December 2017

To understand the regulation of fructose transport in the small intestine and kidney, we provided rats with "control" diets (46% glucose as starch) and with diets enriched in fructose, glucose, or sucrose (60% each of simple carbohydrate) and measured the concentration of facilitative glucose transporter isoform (GLUT5) protein and mRNA in these tissues. The fructose-enriched diet resulted in a five- and eightfold increase in GLUT5 protein at 1 and 7 days, respectively, in the small intestine, which declined rapidly with reversion to control diet. No change in GLUT5 protein levels was seen after glucose- or sucrose-enriched diets. Glucose, and to a lesser extent fructose, feeding resulted in an increase in the basolateral GLUT2 protein. Feeding glucose to the rats caused a rise in sodium-dependent glucose transporter isoform (SGLT1) protein levels compared with the control diet. There was a transient increase in the small intestine GLUT5 mRNA 1 day after fructose feeding, which returned to normal by 7 days. In the kidney, both fructose and sucrose increased GLUT5 protein levels three- to fourfold, whereas glucose had no effect. Fructose-enriched diet did not increase the levels of GLUT5 protein or mRNA in a segment of small intestine that was isolated from the rest of the small intestine but continued to have mesenteric blood supply. The results suggest that the levels of GLUT5 protein are regulated by fructose, its in vivo substrate, in both the small intestine and kidney, and the regulation requires fructose to interact with the brush border of the small intestine, possibly stabilizing the protein.(ABSTRACT TRUNCATED AT 250 WORDS)
Differential expression of substance P, somatostatin, and VIP in neurons from cultured myenteric ganglia
Authors: Grider J. R. , Bonilla O. M.
Published: December 2017

The expression of three enteric neuropeptides was examined in freshly dispersed ganglia and in ganglia cultured for up to 28 days. During culture, glial cells grew into a flat sheet surrounding a cluster of neurons identified with neuron-specific enolase (13 +/- 2/ganglion), which remained constant throughout the period of culture. The neurons underwent a distinctive temporal change, resulting in overexpression of substance P (SP), normal expression of somatostatin, and virtual suppression of vasoactive intestinal peptide (VIP). Three weeks after the start of culture, the ganglia contained and released (in response to 55 nM KCl, 0.1 mM 1,1-dimethyl-4-phenylpiperazinium, or 1 microM gastrin-releasing peptide) twice as much SP as freshly dispersed ganglia, corresponding to a sevenfold increase per cultured neuron; content and release of somatostatin did not change. SP content and release declined to 1.5% of those found in control cultures when nonneuronal cells were suppressed with cytosine arabinoside but were partially restored (13-17% of control) by nerve growth factor. In marked contrast, VIP was minimally (< 1%) present in and released from ganglia after the third day in culture. Suppression of VIP could reflect a selective loss of VIP neurons and/or VIP expression.
Nitric oxide attenuates leukocyte-endothelial interaction via P-selectin in splanchnic ischemia-reperfusion
Authors: Gauthier T. W. , Davenpeck K. L. , Lefer A. M.
Published: December 2017

We studied the effects of exogenous nitric oxide (NO) on leukocyte-endothelial interaction after 60 min of splanchnic artery ischemia and 120 min of reperfusion (SAO/R) in pentobarbital sodium-anesthetized rats via intravital microscopy. Treatment with the NO donor S-nitroso-N-acetylpenicillamine (SNAP, 20 micrograms/kg bolus followed by infusion at 20 micrograms.kg-1.h-1), beginning 10 min before reperfusion, resulted in significantly decreased leukocyte-endothelial interaction. This was manifested by a significant decrease in leukocyte rolling and adherence in the postcapillary venules. Tissue protection was demonstrated by a significantly lower plasma free amino-nitrogen concentration in the SNAP-treated SAO/R rats compared with those receiving NO-depleted SNAP (P < 0.05). Immunohistochemical localization of P-selectin showed significantly decreased P-selectin expression on the venular endothelium after SAO/R in rats given SNAP 10 min before reperfusion (23.0 +/- 3.2% vs. 54.9 +/- 12.1% positive staining, respectively, P < 0.01). From these data, we conclude that the effects of exogenous NO on leukocyte-endothelial interaction after ischemia-reperfusion appear to be at least partially mediated through the endothelial adhesion molecule P-selectin.
Intestinal lipid inhibits gastric emptying via CCK and a vagal capsaicin-sensitive afferent pathway in rats
Authors: Holzer H. H. , Turkelson C. M. , Solomon T. E. , Raybould H. E.
Published: December 2017

The mechanism by which lipid in the duodenum inhibits gastric emptying was investigated in awake rats fitted with chronic gastric and duodenal cannulas. Perfusion of the duodenum with lipid (Intralipid, 5 and 10%; total amount 50 and 100 mg) caused a significant inhibition (26 and 78%, respectively) of gastric emptying of a nonnutrient liquid (0.9% saline). Functional ablation of the capsaicin-sensitive vagal, but not the spinal, sensory innervation to the upper gastrointestinal tract significantly attenuated by 57% lipid-induced inhibition of gastric emptying. In intact rats, administration of a specific cholecystokinin (CCK)-A receptor antagonist, devazepide, significantly attenuated by 66% the response to lipid. Administration of devazepide in perivagal capsaicin-treated rats did not further reduce the response to lipid. These results suggest that lipid in the duodenum inhibits gastric emptying via a mechanism involving an action of CCK at type A receptors and capsaicin-sensitive vagal afferents.
Role of Kupffer cells in the pathogenesis of hepatic reperfusion injury
Authors: Bremer C. , Bradford B. U. , Hunt K. J. , Knecht K. T. , Connor H. D. , Mason R. P. , Thurman R. G.
Published: December 2017

The purpose of this study was to evaluate the role of Kupffer cell activation in the pathogenesis of reperfusion injury. In a blood-free liver perfusion model, pericentral hypoxia and reperfusion injury occurred. Lactate dehydrogenase (LDH) and malondialdehyde (MDA) release, oxygen uptake, and trypan blue staining were assessed. Within the first 10 min of reflow, LDH and MDA release reached maximal values of 44 U.g-1.h-1 and 115 nmol.g-1.h-1, respectively. Trypan blue cell staining was confined to pericentral regions of the liver lobule. When Kupffer cells were inactivated with GdCl3, release of enzymes and MDA was reduced significantly by > 50%, and hepatic cell death was almost completely absent. Since increases in MDA suggested involvement of free radicals, livers were perfused with phenyl N-t-butylnitrone (5 mM), a spin-trapping agent. Analysis of liver tissue by electron paramagnetic resonance spectroscopy revealed a typical six-line spectrum, providing direct evidence that carbon-centered radicals were generated on reflow. GdCl3 treatment decreased radical adduct formation by approximately 50%. Collectively, these results strongly support the hypothesis that activation of Kupffer cells plays an important role in the pathogenesis of hepatic reperfusion injury.
Role of insulin and insulin-like growth factor receptors in regulation of T84 cell monolayer permeability
Authors: McRoberts J. A. , Riley N. E.
Published: December 2017

We previously showed that insulin and insulin-like growth factor (IGF)-I and IGF-II caused a dose-dependent increase in permeability through the paracellular pathway of T84 cell monolayers over 3-4 days. Here we have determined which cell surface receptors were involved in this response. Using radioligand binding studies and receptor cross-linking studies, we found that T84 cells possess insulin and IGF-I receptors. There were approximately 20 x 10(3) insulin receptors/cell with a dissociation constant (KD) of 0.5 nM and 29 x 10(3) IGF-I receptors with a KD of 0.6 nM for IGF-I. Cross-linking studies identified the alpha-subunit of insulin and IGF-I receptors with deduced molecular weights of 126 x 10(3) and 128 x 10(3), respectively. IGF-II bound to T84 cells with an apparent KD of approximately 2.0 nM. Radioreceptor cross-linking indicated that IGF-II interacted principally with the IGF-I receptor, although low levels of the IGF-II/mannose 6-phosphate receptor were also expressed on the cell surface. We then correlated the biological effect with the radioligand binding studies. It was first demonstrated that insulin and IGF-I were degraded in medium in the presence of cells. In addition, we showed that continuous exposure for 2-3 days to insulin or IGF-I was required to produce their biological effect on permeability. Taking into account the rate of degradation and the requirement for continuous exposure, we found a close correlation between radioligand binding and the half-maximal effective concentration for the hormonal effects on transepithelial permeability.(ABSTRACT TRUNCATED AT 250 WORDS)
Validation studies of Bilitec 2000: an ambulatory duodenogastric reflux monitoring system
Authors: Vaezi M. F. , Lacamera R. G. , Richter J. E.
Published: December 2017

Bilitec 2000, a fiber-optic spectrophotometer, detects duodenogastric (DG) reflux in an ambulatory setting independent of pH by measuring bilirubin in bile. Studies were performed for independent validation and better understanding of limitations of this system. Bilitec and a conventional spectrophotometer were used to assess the effect of gastric acidity on the spectral absorbance of refluxed DG contents, employing bilirubin ditaurate (BDT) and human bile. Total bile acid concentrations correlated with Bilitec absorption. BDT and bile had hypsochromic shift (400 nm) in spectrophotometric maximum wavelength (lambda max; 450 nm) in an acidic environment (pH < 3.5), corresponding to an approximately 30% decrease in absorption measured by Bilitec. Bile acid concentrations correlated strongly with Bilitec absorption for all 63 samples (R = 0.82) with pH > 3.5 (n = 31, R = 0.81), but correlation was less strong for 32 samples with pH < 3.5 (R = 0.70). As currently designed, Bilitec adequately measures DG reflux but underestimates it by at least 30% in acidic medium (pH < 3.5). Design modifications are suggested for improving the sensitivity of this equipment.
Regulation of riboflavin intestinal uptake by protein kinase A: studies with Caco-2 cells
Authors: Said H. M. , Ma T. Y. , Grant K.
Published: December 2017

Although the mechanism of riboflavin (RF) intestinal uptake has been the subject of many studies, virtually nothing is known about the cellular regulation of the uptake process. In the present study, we investigated the role of protein kinase A (PKA)- and C (PKC)-mediated pathways in the regulation of RF intestinal uptake using the confluent Caco-2 monolayers. Treatment of Caco-2 cells with 3-isobutyl-1-methylxanthine (IBMX), forskolin, cholera toxin, or dibutyryl adenosine 3',5'-cyclic monophosphate caused a significant inhibition in RF uptake. The inhibitory effect of IBMX was reversible and resulted from a significant decrease in the maximal velocity of the RF uptake process with no change in its apparent Michaelis constant. The IBMX-induced inhibition in RF uptake was not mediated via inhibition in the synthesis of the RF carrier protein or through inhibition in the recruitment of preexisting carrier protein into the plasma membrane. Calyculin A also inhibited RF uptake when added alone and further potentiated the inhibitory effect of IBMX when added together. Phorbol 12-myristate 13-acetate or chelerythrine, on the other hand, showed no significant effect on RF uptake. These results demonstrate for the first time that compounds that increase intracellular cAMP levels downregulate RF intestinal uptake and that this effect is mediated via a decrease in the activity of the RF uptake carrier. It is suggested that a PKA-mediated pathway(s) plays an important role in regulating RF intestinal uptake.
Effects of female sex hormones on mitochondria: possible role in acute fatty liver of pregnancy
Authors: Grimbert S. , Fisch C. , Deschamps D. , Berson A. , Fromenty B. , Feldmann G. , Pessayre D.
Published: December 2017

Acute fatty liver of pregnancy occurs in some women. As other cases of microvesicular steatosis are due to impaired mitochondrial oxidation of fatty acids, we investigated the effects of female sex hormones on liver mitochondria in female mice. Three hours after administration of both estradiol (36 mumol/kg) and progesterone (150 mumol/kg), the in vitro beta-oxidation of [U-14C]palmitic acid and the activity of the tricarboxylic acid cycle decreased 49 and 54%, whereas the in vivo oxidation of [U-14C]palmitic acid decreased 38%. One week of treatment with both sex hormones produced ultrastructural lesions of mitochondria, decreased the recovery of mitochondrial proteins by 34%, increased state 4 respiration by 54-77%, and decreased the activities per gram of liver of several enzymes involved in the activation, mitochondrial uptake, and oxidation of fatty acids by 34-54%. We conclude that female sex hormones have deleterious effects on liver mitochondria and suggest that these effects, together with other factors, may contribute to the development of acute fatty liver of pregnancy in some women.
Mucosal mast cells and developmental changes in gastric absorption
Authors: Catto-Smith A. G. , Ripper J. L.
Published: December 2017

We aimed to establish whether gastric mucosal mast cells undergo degranulation during normal postnatal development and to correlate this with gastric electrical parameters, paracellular permeability, and macromolecular absorption. Sprague-Dawley rats were studied between 10 and 30 days after birth. Gastric mucosal mast cell degranulation occurred and was maximal on days 15 and 17, measured by histology and gastric and serum levels of rat mast cell protease II. Short-circuit current, transepithelial conductance, and permeability of voltage-clamped glandular stomach were elevated in younger animals, falling with age except for a transient but significant increase in conductance and permeability at 17 days, closely correlated with maximal mast cell degranulation. Macromolecular uptake was significantly increased in animals aged 10-15 days. Concanavalin A and antigen-induced mast cell degranulation increased conductance and permeability in vitro in younger animals. We conclude that 1) gastric mucosal mast cells degranulate during development, 2) the neonatal stomach has increased permeability and uptake of macromolecules, and 3) gastric mucosal mast cell degranulation during development may affect mucosal permeability.
Regulation of goblet cell degranulation in isolated pancreatic ducts
Authors: Hootman S. R. , de Ondarza J.
Published: December 2017

Neurohumoral control of goblet cell degranulation in isolated segments of the guinea pig main pancreatic duct was examined using morphometric procedures. Goblet cells represent 25-30% of the epithelial cell population at the head of the main pancreatic duct, a percentage that decreases to 5-10% as the distance from the ampulla increases. Carbachol, bombesin, and vasoactive intestinal peptide (VIP) each stimulated degranulation of duct goblet cells, although cholecystokinin octapeptide, secretin, and histamine did not. The stimulatory effects of carbachol on goblet cell degranulation in isolated pancreatic ducts were blocked by atropine and enhanced by simultaneous exposure to VIP. These observations indicate that goblet cells in guinea pig pancreatic ducts express bombesin, VIP, and muscarinic cholinergic receptors and that multiple intracellular signaling pathways are involved in the regulation of goblet cell degranulation.
Do pancreatic proteases play a role in processing prolactase and/or in the postweaning decline of lactase?
Authors: Keller P. , Poiree J. C. , Giudicelli J. , Semenza G.
Published: December 2017

To assess the role of pancreatic proteases in the proteolytic processing and in the postweaning decline of lactase-phlorizin hydrolase (LPH), we have determined lactase activity and the different LPH forms in postweaned rats in which a jejunal loop was excluded from contact with pancreatic secretions by a jejunal bypass procedure. As a control for the absence of pancreatic proteases, pro-sucrase-isomaltase (proSI), which is known to be split by pancreatic proteases into heterodimeric SI, was used. Nearly all proLPH was processed to mature LPH, indistinguishable from LPH isolated from control animals. SI was found only in the unsplit pro form, whereas it was normally processed to the heterodimeric SI in the control tissues. There were no significant differences in lactase and sucrase activities in operated and in sham-operated control animals. We conclude that pancreatic secretions are not essential for the processing of proLPH to LPH or in the postweaning decline of LPH.
Organic cation transport by rat liver lysosomes
Authors: Moseley R. H. , Van Dyke R. W.
Published: December 2017

Hepatic organic cation transport has been characterized in rat liver plasma membrane vesicles, using the quaternary amine tetraethylammonium (TEA) as a model substrate. Sinusoidal TEA uptake is stimulated by an inside-negative membrane potential; TEA transport across the canalicular membrane is mediated by electroneutral organic cation-H+ exchange. Substrates for these transport processes include procainamide ethobromide (PAEB) and vecuronium, cationic drugs that undergo biliary excretion. Given the apparent absence of sinusoidal transport mechanisms able to generate high hepatocyte-to-blood organic cation concentration ratios, intracellular transport of organic cations may involve sequestration and concentration within acidified organelles. Therefore, the characteristics of TEA uptake were examined in isolated rat liver lysosomes that are acidified by a well-described H(+)-adenosinetriphosphatase (ATPase). Lysosomal uptake of [14C]TEA was a time- and ATP-dependent process, reaching steady state after 30-60 min. Steady-state [14C]TEA uptake was significantly reduced by omission of ATP and by addition of monensin, conditions that alter lysosomal pH and membrane potential gradients, and by the H(+)-ATPase inhibitors, N-ethylmaleimide and bafilomycin A. ATP-dependent lysosomal [14C]TEA uptake was significantly inhibited by PAEB, vecuronium, and other organic cationic substrates of canalicular TEA/H+ exchange. These findings demonstrate that rat liver lysosomes sequester certain organic cationic drugs, most likely via organic cation/H+ exchange driven by H(+)-ATPase. Canalicular organic cation/H+ exchange may reflect, in part, the exocytic insertion of this transporter from an intracellular compartment to this membrane domain.
Ecto-adenosinetriphosphatase in rat small intestinal brush-border membranes
Authors: Schweickhardt C. , Sabolic I. , Brown D. , Burckhardt G.
Published: December 2017

Antibodies against the holo ecto-adenosinetriphosphatase (ATPase) of rat liver and antibodies against COOH-terminal peptides of the long isoform of this enzyme reacted in Western blots with a 105-kDa band from small intestinal brush-border membranes. Indirect immunofluorescence revealed reactive proteins predominantly at the apical surface of enterocytes with some staining of basolateral membranes and of vascular endothelium. Similar results were obtained with monoclonal antibodies against HA4, a protein from rat liver closely related to the ecto-ATPase. Since these results suggested the presence of an ecto-ATPase, ATP hydrolysis was studied in intact, right-side-out brush-border membrane vesicles. Nearly half of ATP hydrolysis was caused by alkaline phosphatase (AP). Besides purine and pyrimidine trinucleotides, AP also hydrolyzed ADP, AMP, pyrophosphate, and 4-nitrophenylphosphate. Inactivation of AP by cleavage of its membrane anchor and by removal of the Zn2+ necessary for its function left the ecto-ATPase that was activated by Ca2+ and Mg2+ and hydrolyzed purine and pyrimidine trinucleotides and dinucleotides, but not AMP, pyrophosphate, and 4-nitrophenylphosphate. These features are characteristic of an ATP diphosphohydrolase (EC 3.6.1.5, also called apyrase). The physiological role of the small intestinal ecto-ATPase may be the degradation of nutrient nucleotides.
Kinetics and uptake in vivo of oxidatively modified lymph chylomicrons
Authors: Umeda Y. , Redgrave T. G. , Mortimer B. C. , Mamo J. C.
Published: December 2017

The metabolism of oxidized chylomicrons (ox-CMs) was investigated in vivo. CMs from rats fed corn, linseed, or fish oil were oxidized by incubation with 2,2'-azobis(2-amidinopropane)hydrochloride (AAPH) or sodium hypochlorite (NaOCl). Oxidized CMs had a rapid phase of clearance, followed by a slow phase. Clearance of ox-CMs was decreased for corn oil but increased for linseed and fish oil particles. Differences in rats of uptake between CM types or treatment were independent of the rate of remnant formation, but were instead a consequence of decreased clearance. A greater triglyceride-to-cholesteryl ester ratio in liver suggested that there was less lipolysis of ox-CM triglyceride prior to uptake. Hepatic uptake of ox-CMs was decreased, whereas there was increased uptake in spleen. However, the uptake by Kupffer cells of ox-CMs was 43% of total liver uptake after AAPH treatment and 59% after NaOCl treatment, compared with 21% for control CMs. Collectively, our data show that oxidation can have differential effects on the rate of clearance of CMs and that ox-CMs are preferentially cleared by the reticuloendothelial system.
Uptake and interconversion of plasma unesterified n-3 polyunsaturated fatty acids by the GI tract of rats
Authors: Nilsson A. , Becker W.
Published: December 2017

The origin of the linoleic [18:2(n-6)], arachidonic [20:4(n-6)], and docosahexaenoic acid [22:6(n-3)] of the mucosal phospholipids in the gastrointestinal (GI) tract is not known. This study examines whether stomach, small intestine, and colon take up and desaturate-elongate unesterified polyunsaturated fatty acids (PUFA) from blood. Albumin-bound unesterified alpha-[14C]linolenate [18:3(n-3)] and [3H]eicosapentaenoate [20:5(n-3)] were injected intravenously. After 10 min, 1 h, and 18 h, radioactivity of tissue lipids and the degree of interconversion of the 3H-labeled and 14C-labeled fatty acids were determined. After 10 min, the lipids of the gastrointestinal tract contained 3.6% of the 14C (0.5% in stomach, 2.7% in small intestine, and 0.4% in colon) and 4.9% of the 3H (0.6% in stomach, 3.6% in small intestine, and 0.7% in colon). Both fatty acids were acylated mainly into phospholipids, both in liver and gastrointestinal tract. Although the proportions of radioactivity found in desaturation-elongation products increased with time, 15% of the 14C in the liver, 6% in the stomach, 8% in the small intestine, and 11% in the colon were in 20:5 already after 10 min. A rapid interconversion of 18:3 thus occurred both in the liver and in the gastrointestinal tract. In all tissues examined, interconversion of [3H]20:5 to [3H]22:5 also occurred. Interconversion of unesterified PUFA taken up from blood may be an important source of eicosanoid precursors in the gastrointestinal tract.
Somatostatin receptor subtype 3 mediates the inhibitory action of somatostatin on gastric smooth muscle cells
Authors: Gu Z. F. , Corleto V. D. , Mantey S. A. , Coy D. H. , Maton P. N. , Jensi R. T.
Published: December 2017

Previous functional studies show that somatostatin (SS) interacts with specific receptors to inhibit relaxation in gastric smooth muscle cells. There are no ligand binding studies, and it is unknown which of the five subtypes of SS receptors mediates the action. Dispersed gastric smooth muscle cells from guinea pig bound both 125I-labeled SS-14 and 125I-D-Phe-Cys-Tyr-D-Trp-Lys-Thr-Cys-Nal-NH2 (where Nal indicates N-naphthylalanine) (cyclo-SS-8), a synthetic peptidase-resistant octapeptide SS analogue. SS-28 and SS-14, cyclo-SS-8, and SS analogue D-Phe-Cys-Phe-D-Trp-Lys-Thr-Cys-Thr-ol [SMS-(201-995) (octreotide)] inhibited 125I-cyclo-SS-8 binding with relative potencies of SS-28 = cyclo-SS-8 = SMS-(201-995) (octreotide), and the binding was not affected by the addition of protease inhibitors. SS-14 caused inhibition only in the presence of protease inhibitors. Ligand analysis demonstrated a two-binding-site model. Analysis of the relationship between biological function and binding suggested the high-affinity sites mediated the relaxant action of SS. 5'-Guanylylimidodiphosphate [Gpp-(NH)p] inhibited binding by reducing the affinity of the high-affinity site. Six SS-8 analogues that distinguish SS subtypes showed that 125I-SS-14 bound to somatostatin receptor subtype 3 (SSTR3). The results demonstrated that gastric smooth muscle cells possess distinct receptors for SS of the SSTR3 subtype. Occupation of these sites inhibits relaxation in gastric smooth muscle cells. Modulation between the high- and low- affinity binding states of SSTR3 is at least partially mediated by activation of guanine nucleotide regulatory proteins.
Platelet-activating factor stimulates gastric acid secretion in isolated rabbit gastric glands
Authors: Sobhani I. , Bado A. , Moizo L. , Laigneau J. P. , Tarrade T. , Braquet P. , Lewin M. J.
Published: December 2017

We examined the effect of platelet-activating factor (PAF) on gastric acid secretion by isolated rabbit gastric glands as determined by [14C]aminopyrine ([14C]AP) uptake. PAF, histamine, and carbachol time- and concentration-dependently stimulated [14C]AP uptake, with estimated half-maximal effective concentrations of 60 pM, 0.25 microM, and 0.1 microM, respectively. PAF-induced [14C]AP uptake was inhibited by the specific PAF antagonists BN-50727 and SR-27417 and by the proton pump inhibitors omeprazole and lansoprazole. However, the H2-receptor antagonist famotidine had no effect. Buffering extracellular Ca2+ by ethylene glycol-bis(beta-amino-ethyl ether)-N,N,N',N'-tetraacetic acid resulted in a shift to the right of the time-course effect of PAF without altering the maximal response, whereas buffering intracellular Ca2+ by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid 2-acetoxymethyl ester, as well as blocking Ca2+ channels by verapamil, inhibited PAF-induced [14C]AP uptake. Intracellular Ca2+ concentration in isolated rabbit gastric glands, as measured by fura 2-acetoxymethyl ester, concentration-dependently increased in response to PAF, to a maximum of 1.5-fold for 0.1 microM. These results suggest that PAF stimulates gastric acid secretion via specific receptors activating intracellular Ca2+ mobilization, which could be located on the parietal cells.
Somatostatin excites canine ileum ex vivo: role for nitric oxide?
Authors: Vergara P. , Woskowska Z. , Cipris S. , Fox-Threlkeld J. E. , Daniel E. E.
Published: December 2017

Isolated perfused segments of canine ileum have no spontaneous motor activity and release large quantities of vasoactive intestinal polypeptide (VIP) continuously. Somatostatin perfusion was shown to decrease VIP release, accompanied by increased contractions and amplification of responses to low-frequency electrical field stimulation. After perfusion of higher somatostatin concentrations, the VIP output did not recover but quiescence returned. The actions of somatostatin on motor activity were not modified by hexamethonium, slightly reduced by atropine, and markedly reduced by tetrodotoxin. Inhibition of VIP output was not the major determinant of motor activity in the ileum because 1) a second infusion of somatostatin had similar motor effects despite markedly reduced VIP output, 2) abolition of tonic VIP output did not prevent induction of motor activity by somatostatin, and 3) artificial restoration of VIP levels did not prevent or antagonize somatostatin-induced ileal contractions. In contrast, the increment in motor responses induced by somatostatin was not apparent after N omega-nitro-L-arginine methyl ester, an inhibitor of nitric oxide (NO) synthase, but recovered after reversal by L-arginine. We conclude that the mode of somatostatin activation of intestinal motor activity involves reduced NO output, enhanced excitatory mediator action or release, a direct action on smooth muscle, and possibly inhibition of VIP output. Of these, reduced NO output plays the most important role.
Hepatorenal baroreflex in cirrhotic rats
Authors: DiBona G. F. , Sawin L. L.
Published: December 2017

A hepatorenal baroreflex has been described in which increases in intrahepatic sinusoidal pressure stimulate an intrahepatic baroreceptor, resulting in increases in afferent hepatic (HNA) and efferent renal sympathetic nerve activity (RNA). Hepatic denervation prevents the increase in RNA. This baroreflex is postulated to contribute to the increase in RNA found in cirrhosis in which intrahepatic sinusoidal pressure is increased. However, the increased fibrosis in the cirrhotic liver may render the intrahepatic baroreceptor less sensitive to increases in intrahepatic sinusoidal pressure. By use of thoracic inferior vena caval constriction, intrahepatic sinusoidal pressure (i.e., inferior vena caval pressure, IVCP) was increased in control rats and rats with cirrhosis due to common bile duct ligation (CBDL) while HNA and RNA were measured. With increases in IVCP of 5 mmHg, increases in HNA (+38 +/- 2 and +44 +/- 3%) and RNA (+25 +/- 1 and +34 +/- 3%) were not different in control and CBDL rats, respectively. The slope gain, % delta HNA/delta IVCP, was +7.1 +/- 0.6 and +8.1 +/- 0.7%/mmHg in control and CBDL rats, respectively. Therefore the hepatorenal baroreflex is not desensitized in the CBDL rat, and the hepatorenal baroreflex is capable of contributing to the increase in RNA observed in cirrhosis.
Microscope laser light-scattering spectroscopy of vesicles within canaliculi of rat hepatocyte couplets
Authors: Mockel G. M. , Gorti S. , Tandon R. K. , Tanaka T. , Carey M. C.
Published: December 2017

Employing microscope laser light-scattering spectroscopy, we investigated "primary" bile secretion into canalicular spaces of rat hepatocyte couplets in monolayer culture. Time-dependent scattered light intensities were fitted by bi-exponential decays. The "slow" decay was attributed to an undulating canalicular membrane motion, whereas the "fast" decay was consistent with rapidly diffusing intracanalicular vesicles with mean hydrodynamic radii (+/- SD) of 479 +/- 53 A. After addition of micromolar concentrations of common bile salts, increases in the amplitude of the fast component facilitated a quantitative estimate of vesicle secretion rates. A dose-response relationship with 0.1-200 microM sodium taurocholate was characterized by an initial concentration-dependent increase and then a decrease in the amplitude of the fast canalicular component. Since these taurocholate concentrations are nontoxic to cultured hepatocytes, the maximum in vesicle-sized particles at 10 microM taurocholate suggested that its critical micellar concentration of approximately 5 mM was attained within the canalicular spaces. Sodium taurolithocholate resulted in time- and dose-dependent diminution in vesicle secretion rates, which after 2 h was followed by spontaneous canalicular recovery. This suggested that acute bicellular "cholestasis" was followed by oxidative metabolism and detoxification of the monohydroxy bile salt. Microscope laser light-scattering spectroscopy should facilitate further physical-chemical and pathophysiological studies of bile secretion at the cellular level.
Speed and pressure characteristics of external anal sphincter contractions
Authors: Herdmann J. , Enck P. , Zacchi-Deutschbein P. , Ostermann U.
Published: December 2017

The principle of isochronism reflects constant contraction time for varying strengths of muscle contraction. This principle was studied for the innervation of the pelvic floor in humans using motor-evoked potentials (MEPs) and evoked pressure curves (EPCs) from the external anal sphincter muscle (EAS). MEPs and EPCs were simultaneously recorded after transcranial magnetic stimulation of the motor cortex. Voluntary contractions were also studied. Contraction times of the EAS were significantly longer in voluntary contractions (mean, 237 ms) than in EPCs (mean, 90 ms). Depending on either mode of contraction, contraction times varied only slightly despite a wide range of contraction strengths. It is shown that the contractile behavior of the EAS is a function of slow- and fast-twitch muscle fiber distribution and that the principle of isochronism governs motor performance not only of limb muscles but also of the EAS. There exists a unique optimal working range of each muscle to meet its individual function. Disturbance of this principle results in a less efficient contraction with either inappropriate basic tone or disturbed reflex activation in the EAS. Both are possible causes of incontinence.
Ca2+ currents in human colonic smooth muscle cells
Authors: Xiong Z. , Sperelakis N. , Noffsinger A. , Fenoglio-Preiser C.
Published: December 2017

Voltage-gated Ca2+ currents were investigated in single smooth muscle cells freshly isolated from the circular layer of the human colon (ascending and descending portions) using the whole cell voltage-clamp technique. Tissue samples were obtained at the time of therapeutic surgery. In physiological salt solution (containing 2 mM Ca2+), an inward current was observed when the cell membrane was depolarized in the presence of tetrodotoxin. This current disappeared when Ca2+ was removed from the bath solution and was inhibited when Ca2+ channel blockers were applied, indicating that the inward current was a Ca2+ current (ICa). Changing the holding potential (HP) from -100 mV to more positive potentials (e.g., -60 and -40 mV) markedly decreased the amplitude of ICa. The voltage dependence of steady-state activation and inactivation was represented by Boltzmann distributions; there was a substantial amount of overlap (window current) between -60 and -10 mV. A fast-inactivating ICa component followed by a slow-inactivating ICa component was observed in some cells from both ascending and descending colons. The fast ICa component was observed only when cells were held at -80 or -100 mV, and had a more negative threshold potential (-70 to -60 mV). This component was sensitive to low concentrations of Ni2+ (30 microM) but was resistant to nifedipine (10-20 microM). In contrast, the slow (sustained) ICa component was observed at all HPs (-40 to -100 mV) and had a more positive threshold potential (about -40 mV). This component was insensitive to low concentration of Ni2+ but was sensitive to nifedipine and BAY K 8644.(ABSTRACT TRUNCATED AT 250 WORDS)
Ca(2+)-dependent amylase secretion from SLO-permeabilized rat pancreatic acini requires diffusible cytosolic proteins
Authors: Padfield P. J. , Panesar N.
Published: December 2017

Streptolysin O (SLO)-permeabilized pancreatic acini are now frequently used to study regulated exocytosis in the exocrine pancreas. In this paper we introduce alpha-toxin as a possible alternative permeabilization agent to SLO. Both alpha-toxin and SLO are bacterial cytolysins, but the membrane pores generated by SLO are approximately 5-10 times larger than those formed by alpha-toxin. The Ca2+ requirements for amylase secretion from both types of permeabilized acini were identical, maximal amylase secretion being obtained at 30 microM Ca2+ with an effective concentration of approximately 3-4 microM Ca2+ producing 50% of the maximal response. However, Ca(2+)-stimulated amylase secretion from the SLO-permeabilized acini stopped after 10-15 min, unlike secretion from the alpha-toxin-permeabilized cells, which continued for at least 50 min. The rapid cessation of secretion from the SLO-treated acini reflects the rapid decline in the responsiveness of the cells observed after permeabilization. This decline in Ca(2+)-dependent secretion appears to be due to the loss of cytosol, since addition of purified rat brain cytosol to nonresponsive SLO-permeabilized acini reconstituted regulated secretion. Because alpha-toxin-permeabilized acini maintained their responsiveness, the cytosolic factors lost from the SLO-permeabilized cells must be retained within the toxin-treated cells. The reconstitutive activity of the brain cytosol was nondialyzable but heat and trypsin sensitive, suggesting that the factors responsible are proteins. Of the cytosols screened (brain, liver, spleen, muscle, and lacrimal) only those prepared from brain or lacrimal gland reconstituted Ca(2+)-dependent amylase secretion.(ABSTRACT TRUNCATED AT 250 WORDS)
EGF inhibits secretagogue-induced cAMP production and amylase secretion by Gi proteins in pancreatic acini
Authors: Stryjek-Kaminska D. , Piiper A. , Zeuzem S.
Published: December 2017

In pancreatic acinar cells, the epidermal growth factor (EGF) receptor interacts with both cholera toxin- and pertussis toxin (PTX)-sensitive G proteins. In the present study, isolated rat pancreatic acini were used to investigate the effect of EGF on basal and secretagogue-induced adenosine 3',5'-cyclic monophosphate (cAMP) production and amylase release. EGF increased cAMP production and amylase release in pancreatic acini. However, cAMP accumulation and amylase release elicited by either vasoactive intestinal peptide (VIP) or forskolin were inhibited by EGF (17 nM). EGF inhibited the VIP-induced cAMP production and amylase release with a half-maximal effective concentration of 3 and 2 nM, respectively. EGF had no effect on the N6,2'-O-dibutyryladenosine-3',5'-monophosphate-stimulated amylase release, suggesting that the inhibitory effect of EGF on the VIP- and forskolin-induced cAMP production is due to inhibition of adenylyl cyclase. PTX pretreatment of the acini led to an increase of the basal, EGF-, and VIP-stimulated cAMP accumulation and amylase release, indicating that PTX-sensitive G proteins exert tonic inhibition of adenylyl cyclase even in the absence of agonist. In PTX-pretreated acini, the inhibitory effect of EGF on the VIP-induced cAMP production and amylase release was abolished. In conclusion, these results suggest that EGF inhibits secretagogue-induced cAMP production via activation of PTX-sensitive G proteins in rat pancreatic acini, whereas EGF-induced cAMP production and amylase release occurs via a PTX-insensitive pathway.
Role of gastric blood flow in impaired defense and repair of aged rat stomachs
Authors: Gronbech J. E. , Lacy E. R.
Published: December 2017

To study impaired gastric mucosal tolerance against noxious agents in aged rats, possible factors underlying this observation were compared in anesthetized Fisher 344 young and aged rats. The gastric mucosa was damaged by in situ exposure to 80% ethanol for 30-45 s and by 1 M NaCl for 10 min followed by saline (pH = 1.0) for 60 min in chambered stomachs. The lesion area was significantly larger and epithelial restitution was significantly slower in aged than in young rats after both types of injury. Changes in gastric blood flow were monitored by laser-Doppler velocimetry. Young, but not aged, rats showed a marked increase in gastric blood flow in response to 1 M NaCl, acid challenge, and 640 microM capsaicin for 60 min. Young rats showed a higher density of calcitonin gene-related peptide (CGRP)-staining nerve fibers around submucosal blood vessels and higher mucosal release of prostaglandin E2 and leukotriene C4 than did aged rats. These data suggest that impaired mucosal defense and reduced restitution in aged rats is related to lack of hyperemic response caused by mucosal injury and H+ back-diffusion, which is probably due to decreased density of CGRP-staining nerve fibers and prostaglandin biosynthetic capacity in the mucosa.
Transport and steady-state accumulation of putrescine in brush-border membrane vesicles of rabbit small intestine
Authors: Brachet P. , Debbabi H. , Tome D.
Published: December 2017

Absorption of polyamines from the lumen is essential for cell proliferation in small intestine but also in other rapidly growing body tissues and tumors. Intestinal uptake of polyamines is thought to involve one or more transport systems, but the characteristics of these systems have not yet been clearly elucidated. Because high levels of putrescine have been identified in intestinal lumen, we explored kinetic, physiochemical, and structural features of uptake of this diamine across rabbit intestinal brush-border membrane vesicles (IBBMV) prepared by CaCl2 or MgCl2 precipitation procedure. Initial rates of putrescine influx were measured during 5-min incubations at 25 or 37 degrees C (optimal temperature) for concentrations of 0.45-145 microM. At both temperatures, kinetics of putrescine transport fitted a model with a single Michaelis-Menten uptake component plus a nonsaturable uptake component. At 37 degrees C, the kinetic parameters for the saturable component of putrescine uptake, Km,app and Vmax,app, were 16.8 +/- 4.7 microM and 19.9 +/- 2.8 pmol.mg protein-1.min-1, respectively. The value of the constant for the nonsaturable component of putrescine uptake (P = 0.45 +/- 0.06 x 10(-8) l.mg protein-1.s-1) suggested this component represented essentially nonspecific binding of putrescine to IBBMV. Cadaverine, spermidine, and spermine were competitive inhibitors of putrescine transport, with inhibition constants equal to 47, 117, and 219 microM, respectively. When effects of a variety of alkyldiamines and structural analogues of polyamines (1 mM) on influx of 5.6 microM putrescine were compared, cadaverine, methylglyoxal bis(guanylhydrazone) (MGBG), and cyclic derivatives of MGBG were found to exhibit the highest inhibitory potencies.(ABSTRACT TRUNCATED AT 250 WORDS)
Extracellular matrix incorporation of normal and NEM-alkylated fibronectin: liver and spleen deposition
Authors: Rebres R. A. , McKeown-Longo P. J. , Vincent P. A. , Cho E. , Saba T. M.
Published: December 2017

The incorporation of plasma fibronectin (pFn) into the extracellular matrix (ECM) is believed to influence tissue integrity, wound repair, and vascular permeability. In vitro, matrix assembly of Fn requires the binding of soluble Fn to cell-associated matrix assembly sites. Alkylation of human pFn (HFn) with N-ethylmaleimide (NEM) prevents the initial binding of Fn to matrix assembly sites as well as its in vitro incorporation into the ECM as reflected by detergent-insoluble 125I-labeled Fn (pool II Fn). We determined the kinetics of Fn matrix incorporation in tissue and whether NEM treatment of rat pFn (NEM-RFn) would limit its in vivo incorporation into ECM by analysis of pool I [deoxycholate (DOC) soluble] and pool II (DOC insoluble) 125I-Fn in tissues after its intravenous injection into rats. After intravenous injection, tissue incorporation of normal rat 125I-pFn was especially intense in liver and spleen, in agreement with the large amount of endogenous Fn detected in the matrices of these organs. Tissue deposition of plasma-derived 125I-RFn in liver and spleen peaked by 4 h, with significant (P < 0.01) loss over 24 h, indicating turnover of matrix Fn. Tissue localization of normal 125I-RFn in liver, lung, spleen, heart, and intestine was greater (P < 0.05) than 125I-NEM-RFn at 4 h. Normal HFn, but not NEM-HFn, was incorporated into tissues and colocalized with endogenous Fn in the matrix. To identify the cells mediating the intense incorporation of pFn into liver ECM, we compared matrix assembly of 125I-HFn by cultured fibroblasts, hepatocytes, and hepatic Kupffer cells. With fibroblasts, 125I-HFn in pool I reached steady state by 3 h, whereas 125I-HFn in pool II exceeded that in pool I by 6 h and continued to increase over 24 h. With hepatocytes, pool I 125I-HFn reached steady state by 1 h, and a progressive increase (P < 0.05) of 125I-HFn in pool II was observed over 24 h. Kupffer cells were not able to incorporate significant amounts of 125I-HFn into matrix. NEM-HFn displayed limited incorporation into ECM by both fibroblast and hepatocyte cultures. These novel observations suggest that the interaction of soluble pFn with matrix assembly sites is necessary to its in vivo incorporation into the ECM.