Chicken Small Intestine Research Articles

Antagonistic properties of [Phe 3,Leu 13]porcine motilin

We describe the antagonistic properties due to the replacement of Pro 3 by phenylalanine in porcine motilin. The analogue, [Phe 3,Leu 13]porcine motilin (OHM-11526), displaces iodinated [Nle 13]porcine motilin bound to a homogenate of rabbit antral smooth muscle tissue. The dissociation constant (p K d) was 9.26 ± 0.04, versus 9.11 ± 0.01 for motilin and 8.24 ± 0.06 for ANQ-11125, the (1–14) fragment of OHM-11526. The Hill coefficient was close to one and Schild plot analysis confirmed the competitive nature of the interaction. In the tissue bath OHM-11526 was unable to induce contractions of segments of rabbit duodenum. At a concentration of 10 −6 M, OHM-11526 inhibited the effect of maximally effective doses of porcine motilin and of the erythromycin derivative, EM-523, but was without effect on contractions induced by acetylcholine, substance P and serotonin. Increasing doses of OHM-11526 shifted the dose-response curves of motilin and EM-523 to the right, but caused a depression of the maximal response as well. From the motilin curves, and assuming a dual competitive and non-competitive interaction, the pA 2 was 7.79 ± 0.08, the pD′ 2 6.91 ± 0.08. The EM-523 curves yielded comparable data (pA 2 = 8.10 ± 0.12 and pD′ 2 = 7.06 ± 0.13). OHM-11526 also blocked the motilin responses observed with smooth muscle strips from the rabbit and human antrum. However, in a preparation of the chicken small intestine, OHM-11526 was a full agonist with a potency (pD 2 = 6.84) comparable to that of porcine motilin (pD 2 = 6.71). Our data confirm the interaction of motilides with the motilin receptor. Due to its increased affinity for the motilin receptor, OHM-11526 will be a valuable tool for studying the physiology of motilin and the pharmacology of motilin and motilides.

K+‐H+ Exchange Activity in Brush‐Border Membrane Vesicles Isolated from Chick Small Intestine

The purpose of this study was to investigate the presence of a K(+)-H+ exchanger in brush-border membrane vesicles (BBMV) isolated from chick small intestine. 86Rb+, as a tracer for K+ transport, was used to probe for the exchange mechanism. An outwardly directed proton gradient (pH 5.5 inside, pH 7.5 outside) stimulated 86Rb+ uptake into voltage-clamped BBMV. H(+)-driven 86Rb+ uptake was only weakly inhibited by 5-(N-ethyl-N-isopropyl)amiloride, whereas this agent strongly inhibited H(+)-driven Na+ uptake. At initial rates, proton-driven 86Rb+ uptake was significantly reduced by external K+ but it was not significantly affected by external Na+. Conversely, extravesicular Na+ inhibited proton-driven Na+ uptake, whilst K+ had little effect. H(+)-driven K+ uptake tended to saturate with increasing external K+ concentrations and Lineweaver-Burk analysis of the data revealed a Km for external K+ of 2 mM. These findings are consistent with the presence of K(+)-H+ exchange activity in the chicken jejunal brush-border membrane.

K+-H+ Exchange Activity in Brush-Border Membrane Vesicles Isolated from Chick Small Intestine

K+-H+ Exchange Activity in Brush-Border Membrane Vesicles Isolated from Chick Small Intestine

Na +-H + exchange activity in brush-border membrane vesicles isolated from chick small intestine

This study was undertaken to investigate the presence of a Na + H + antiporter in brush-border membrane vesicles (BBMV) isolated from chick small intestine. An outwardly directed proton gradient (pH 5.5 inside, 7.5 outside) stimulated Na + uptake into BBMV and resulted in a transient accumulation. No accumulation was observed in the absence of a proton gradient. Voltage clamping the membrane with K + and valinomycin decreased the Na + overshoot. Amiloride inhibited pH gradient-driven Na + uptake in a dose-dependent manner with an IC 50 of 44 μM. The relationship between pH gradient-driven Na + uptake and external Na + concentration followed simple, saturating Michaelis-Menten kinetics. Eadie-Hofstee analysis of the pH gradient-driven Na + uptake indicated a single transport system with a V max of 33 nmol/mg protein per 15 s and a K m for Na + of 12 mM. The initial rate of pH-driven Na + uptake increased as the intravesicular pH decreased, with a Hill coefficient close to 1. These findings indicate that BBMV isolated from chicken small intestine posses a Na + H + exchanger. This exchanger does not appear to be the one involved in cell pH regulation.

Differential regulation of vitamin D receptor and intestinal calcium transport occurring during sexual maturation in the fowl ( Gallus domesticus)

Quantitative measurements of intestinal vitamin D receptor (VDR) and calcium transport taking place in individual villus-attached enterocytes have been compared during the onset of sexual maturity to test for a direct involvement of VDR in controlling calcium transport across the chick small intestine. Chickens of all ages showed VDR to be present in crypt and villus enterocytes, the amounts of VDR declining only slightly during enterocyte migration from the crypts to the tips of villi. Calcium transport corrected for initial adsorption was lowest in the crypt and highest in villus tip enterocytes. These results are consistent with VDR initiation and possible later maintenance of calcium transport across differentiating enterocytes. The total amount of VDR determined along the crypt-villus axis was significantly less in immature 11-week-old compared with 17-week-old sexually mature non-laying and 25-week-old laying chickens. Calcium transport was significantly greater in 25-week-old compared with 17- and 11-week-old birds. This unexpected up-regulation of VDR in 17-week-old birds was not affected by dietary restriction of calcium. Increased VDR expression in 17-week-old sexually mature birds is probably initiated by estrogen acting indirectly to increase 1,25 (OH) 2D 3 production in the kidney. Increased calcium transport in 25-week-old laying chickens could involve estrogen interacting with estrogen receptors as well as 1,25 (OH)2D, interacting with vitamin D receptors to promote gene transcription in the intestine.

Ontogenic and regional changes in kinetic constants of alpha-methyl-D-glucoside transport in chicken small intestine.

Ontogenic and regional changes in kinetic constants of alpha-methyl-D-glucoside transport in chicken small intestine.

Chloride transport in brush-border membrane vesicles from chick jejunum

This study sought to characterize the mechanism(s) of Cl- transport across brush-border membrane vesicles (BBMV) isolated from chick jejunum. An inwardly directed proton gradient stimulated chloride (36Cl-) uptake. This uptake was inhibited by SITS and H2-DIDS. pH-gradient-stimulated Cl- uptake was electroneutral, since it was only slightly decreased by voltage clamping the BBMV with K+ and valinomycin. An outwardly directed HCO3- gradient significantly increased chloride uptake in the presence of a pH gradient. pH-driven chloride uptake was reduced by the presence of several anions in the uptake buffer. The rank order of potency for inhibition of pH-driven Cl- uptake was Cl- > SCN- > HCO3- > I- > Glu- > HPO4(2-). In the absence of a pH gradient, chloride was less concentrated inside the vesicles than outside. Chloride uptake under these conditions was stimulated by a positive electrical potential inside the vesicles. This stimulation was inhibited by the addition of several anions outside the vesicles. The order of inhibitory potency was SCN- > I- > Cl- > HCO3- > Glu- > HPO4(2-). The results are consistent with the presence of a Cl-/base exchanger and a chloride conductance pathway in the brush-border membrane of chick small intestine.

The appearance of direct contacts between Auerbach's plexus and smooth muscle cells in the small intestine of the chicken during in ovo development.

Direct contacts between the neural cell bodies of Auerbach's ganglia and the longitudinal smooth muscle cells in the chicken small intestine were demonstrated by electron microscopy during the second half of embryonic development. The possible functional explanation of these contacts is discussed.

Characterization of chicken intestinal brush border membrane Ns/H exchange

1. 1. Na/H exchange is the major pathway for Na uptake in brush border membrane vesicles from chicken small intestine. Hanes-Woolf analysis demonstrated that Na and H competed at the same extravesicular site. The K Na for Na + at extravesicular pH 6.6 is 35 mM and at pH 7.4, 12 mM. 2. 2. Similar to mammalian intestinal cells, the Na/H exchanger does not appear to have an internal proton modifier site. Varying intravesicular pH from 6.1 to 7.8 stimulates uptake, but a sigmoidal relationship is not observed. 3. 3. The ability of several amiloride analogs to inhibit the exchanger was tested and the inhibitory profile was similar, but not identical to Na/H exchangers in mammalian tissues. The potency series (from most to least potent) is hexamethylamiloride ≈ ethylisopropylamiloride > methylisobutylamiloride > dimethyl-amiloride > amiloride.

Histological distribution and developmental changes of tropomyosin isoforms in three chicken digestive organs.

Histological localization of tropomyosin isoforms in three digestive organs from embryonic and adult chickens was performed by using rabbit antisera against chicken skeletal muscle tropomyosin and against low-Mr-type tropomyosin from chicken small intestine mucosa. The former antiserum (named TM-SH) reacted with alpha, beta, and high-Mr-type isoforms, and the latter (named TM-HL) reacted with alpha, beta, high-Mr-type and low-Mr-type isoforms, alpha and beta Isoforms were detected in muscle cells of the muscular layer and the muscularis mucosa. Low-Mr-type isoforms, however, were detected along the cell membrane and cytoplasm of almost all nonmuscle cells, especially in terminal webs of epithelial cells. Developmental changes of tropomyosin isoforms in digestive organs were studied by two-dimensional gel electrophoresis and image analysis. The relative amounts of alpha and beta isoforms increased in the course of development, but those of low-Mr-type and high-Mr-type isoforms decreased.

Effect of the cyanobacterial (blue-green algal) toxins from Microcystis aeruginosa on isolated enterocytes from the chicken small intestine

Livestock deaths, and clinical reports of human injury, follow the consumption of toxic blue-green algae. The experiments described show that isolated intestinal enterocytes from chicks are both deformed and killed by exposure to Microcystis toxins, in a dose-dependent and time-dependent manner. The enterocytes were protected from toxicity by deoxycholate, bromosulphothalein and rifampicin. It was concluded that the gastroenteritis clinically associated with accidental Microcystis ingestion is likely to reflect enterocyte injury by Microcystis toxins, and that the therapeutic use of bile acids or transport inhibitors may be of value in treatment.

Regional differences for the D-amino acid oxidase-catalysed oxidation of D-methionine in chicken small intestine.

1. Enzymic oxidation of D-[1-14C]methionine (D-met) to 2-keto-4-methylthiobutanoate (KMB) has been determined using 100,000 g supernatants prepared from chicken tissue homogenates. 2. The small intestinal mucosa contains substantial oxidative activity towards D-met, which represents about one-half and one-tenth the hepatic and renal activity, respectively. 3. KMB is poorly decarboxylated and rather transaminated to L-met. 4. The specific activity for D-met oxidation in the duodenal mucosa is 1.5- and 4.0-fold than in the jejunal and ileal mucosa, respectively. 5. The intestinal D-met-oxidizing activity is dramatically altered by the D-amino acid oxidase specific inhibitor benzoate.

Cl −/base exchange and cellular pH regulation in enterocytes isolated from chick small intestine

Intracellular pH (pH i) and Cl −/base exchange activity have been examined in isolated chicken enterocytes, both in the presence and absence of 25 mM HCO 3 −/5% CO 2. Intracellular pH was measured with BCECF, a pH-sensitive car☐ylfluorescein derivative. Under resting conditions pH i was 7.17 in Hepes and 7.12 in HCO 3 −-buffered solutions. Cells became more alkaline upon withdrawal of Cl −. Cells depleted of Cl − acidified upon reinstatement of Cl −. These changes were faster in the presence of HCO 3 − than in its absence. After an alkaline load (removal of HCO 3 − from the medium) pH i decreases towards base line in the presence of Cl −, but not in its absence. The Cl −-dependent pH i changes were prevented by H 2DIDS and were unaffected by Na +. The Cl −-induced recovery from an alkaline load exhibited simple saturation kinetics, with an apparent K m of 12.5 mM Cl − and maximum velocity of ≈ 0.20pH units min −1. The Cl −/base exchange is functional under resting conditions, as shown by cell alkalinization on exposure to 0.5 mM H 2DIDS, both in the presence and in the absence of HCO 3 −. It is concluded that Cl −/base exchange participates in setting the resting intracellular pH in isolated chicken enterocytes and helps recover from alkaline loads. The exchange operates both in the presence and in the absence of bicarbonate.

Low-frequency electromagnetic pulses enhance Ca 2+ uptake by chick small intestine in vitro

The effect of external EMF on Ca 2+ uptake by the chick small intestine was investigated in vitro by use of the accumulating preparations of epithelial tissue. The intestinal tissue preparations were exposed to EMF in specific zones of Helmholtz coils. The steady magnetic field (SMF) having an induction value B 0 = 1.14 mT, as well as pulsed EMF—constant pulse rate ( f = 80 Hz) and randomized pulse rate (in range of 40–120 Hz)—with amplitude B M = B 0 was used. The fall time of the pulse was changed stepwise in the range 0.15–0.55 ms. It was found that SMF, as well as pulsed EMF significantly enhanced the uptake of Ca 2+ by the small intestine. A SMF was found to enhance uptake by up to 40–50% for both normal (D) and rachitic chicks. The effects of the induced E rot depended essentially of vitamin D status of chicks due to the complex action of vitamin D-dependent factors. The effect of randomized pulse EMF was qualitatively similar to that of constant pulse EMF, but quantitatively it was less. Data indicate the contribution of the B-, as well as E-component to the modification of the Ca 2+ uptake.

Low-frequency electromagnetic pulses enhance Ca 2+ uptake by chick small intestine in vitro

The effect of external EMF on Ca 2+ uptake by the chick small intestine was investigated in vitro by use of the accumulating preparations of epithelial tissue. The intestinal tissue preparations were exposed to EMF in specific zones of Helmholtz coils. The steady magnetic field (SMF) having an induction value B 0 = 1.14 mT, as well as pulsed EMF—constant pulse rate ( f = 80 Hz) and randomized pulse rate (in range of 40–120 Hz)-with amplitude BM = B 0 was used. The fall time of the pulse was changed stepwise in the range 0.15–0.55 ms. It was found that SMF, as well as pulsed EMF significantly enhanced the uptake of Ca 2+ by the small intestine. A SMF was found to enhance uptake by up to 40–50% for both normal (D) and rachitic chicks. The effects of the induced E rot depended essentially of vitamin D status of chicks due to the complex action of vitamin D-dependent factors. The effect of randomized pulse EMF was qualitatively similar to that of constant pulse EMF, but quantitatively it was less. Data indicate the contribution of the B-, as well as E-component to the modification of the Ca 2+ uptake.

Developmental control over vitamin-D-induced calbindin gene expression during early differentiation of chicken jejunal enterocytes

Abstract In situ hybridization and immunocytochemical techniques have been used to examine the distribution of vitamin-D-induced calbindin mRNA and calbindin protein in enterocytes lining the crypts and villi of chicken small intestine. Basal villus enterocytes contained approximately twice as much calbindin but over three times as much calbindin mRNA compared to values found in basal crypt and upper villus enterocytes, all values being measured 2 days after vitamin D injection into D-deficient chickens. Virtually no calbindin mRNA was detected in tissues taken from control D-deficient birds. Direct proportionality found between calbindin mRNA and calbindin content in enterocytes of basal crypt, mid and upper villus suggests pre-translational control over calbindin synthesis. The implications of possible inefficient translation of calbindin mRNA in basal villus enterocytes are discussed. Present methods of analysis provide a novel way to study mechanisms controlling gene expression throughout the whole process of enterocyte differentiation.

Na +-H + exchange activity and cellular pH regulation in enterocytes isolated from chick small intestine

Intracellular pH (pH i and Na +/H + exchange activity have been examined in isolated chicken enterocytes using pH sensitive fluorescence dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF), and in a nominally bicarbonate-free buffer. Under resting conditions the pH i (7.18) was higher than that observed in the presence of the proton ionophore FCCP (6.98), indicating that [H +] is below the value predicted for electrochemical equilibrium across the plasma membrane, i.e., pH i is regulated. Removal of extracellular Na + lowered pH i by 0.28 units and subsequent addition of 80 mM Na + rapidly increased pH i towards the control value. The acidification induced by Na +-removal was prevented by 1 mM amiloride. After an intracellular acidification by exposure to 30 mM NH 4Cl during 5 min, the pH i decreased from approx. 7.18 to approx. 6.86. Subsequent alkalinization of cells back to control pH i was observed after addition of Na + or Li + but not TEA +. Na +-dependent recovery of pH i after an acid-load was unaffected by valinomycin, and was 82% reduced by 1 mM amiloride. The inhibitory action of amiloride was abolished by 10 μM monensin. The initial rate of pH i recovery from an acid-load following exposure to Na + exhibited simple saturation kinetics, with an apparent K m of 12.5 mM Na + and maximum velocity of alkalinization of approx. 0.2 pH units·min -1. The rate of pH i recovery was inversely proportional to pH i. The ‘set point’ for the exchanger is approx. 7.35. It is concluded that in chicken enterocytes the Na +/H + exchange system is not quiescent at resting pH i and, thus contributes to the maintenance of a steady-state pH i at neutral or slightly alkaline levels.

Interleukins 1 and 3 stimulate anion secretion in chicken intestine

Interleukin 1 or 3 added to the serosal side of chicken small intestine transiently increases short-circuit current. Replacement of bathing-medium Cl and HCO3 with gluconate and HEPES abolished the short-circuit current increase, consistent with these cytokines stimulating electrogenic anion secretion. Cytokine-stimulated short-circuit current changes were inhibited by preincubation with piroxicam (10−5 M), an inhibitor of arachidonic acid cyclooxygenase, suggesting prostaglandin formation as an intermediate step for cytokine stimulation of short-circuit current. In intact mucosal strips, interleukin 1 and 3 stimulated prostaglandin E2 release and elevated tissue 3′,5′-cyclic adenosine monophosphate concentration. When prostaglandin E2 release from epithelial and subepithelial fractions of the mucosa by interleukin 1 was determined, increases were found only from the subepithelium. The secretory actions of cytokines appear to be mediated by arachidonic acid metabolites most likely produced by cells of the lamina propria and submucosa and may play a role in inflammatory processes in which intestinal secretion is enhanced.

Triiodothyronine stimulates 2-deoxy-D-glucose uptake by organ cultured embryonic chick small intestine

The possible contribution of increased D-glucose absorption from the intestine to the impairment of oral glucose tolerance in hyperthyroidism was evaluated by investigating the influence of T3 on different pathways of D-glucose transport, utilizing an organ culture system of embryonic chick small intestinal explants. T3, when present in the culture medium at a concentration between 10(-10)-10(-8) mol/l, had no effect on uptake of alpha-methyl-D-glucoside, but stimulated uptake of 2-deoxy-D-glucose by the intestinal epithelium in a dose-dependent fashion. T3 thereby enhanced the maximal velocity of a saturable, cytochalasin B-sensitive but phloretin-insensitive 2-deoxy-D-glucose transport system with an apparent Km of 7 mmol/l. The combined data are consistent with the assumption that T3 can enhance D-glucose entry into the intestinal epithelium through stimulation of a low-affinity transport system at the brush-border membrane of enterocytes. Our findings provide a basis for the explanation of adaptive modulation of intestinal glucose absorption in hyperthyroidism.

ZO-1 and cingulin: tight junction proteins with distinct identities and localizations.

The relative localization of ZO-1 and cingulin, the only two known components of the tight junction, was compared in Madin-Darby canine kidney (MDCK) cells, chicken small intestine, rat kidney distal convoluted tubule, and a hepatoma cell line. Immunoblot analysis demonstrated that cingulin and ZO-1 are immunologically unrelated and that, in the colon, cingulin is a single polypeptide with a molecular mass of 140 kDa. Immunofluorescent localization of cingulin and ZO-1 in confluent monolayers of MDCK cells showed identical staining patterns. However, subconfluent MDCK cells showed distinct localizations of the two proteins. Both cingulin and ZO-1 were found at the plasma membrane only at areas of cell-cell contact, but cingulin was diffusely distributed within the cytoplasm, whereas ZO-1 showed a more clustered internal arrangement. Cingulin and ZO-1 were identically localized at the plasma membrane of hepatoma tissue culture (HTC) cells at sites of cell-cell contact. In chicken intestine examined at the ultrastructural level, immunogold particles associated with cingulin were found approximately three times farther from the junctional membrane than those affiliated with ZO-1.