Magnesium uptake by intestinal brush-border membranes of spontaneously hypertensive rats.

Magnesium uptake by intestinal brush-border membranes (BBM) was studied in duodenal and jejunal vesicles of the spontaneously hypertensive rat (SHR) and normotensive control, the Wistar-Kyoto (WKY) rat. In the duodenum, no statistical difference was evidenced between the two types of rats. By contrast, initial rates of magnesium uptake in jejunal vesicles were lower in SHR (5.4 +/- 2.1 nmol/mg protein x 10 sec) in comparison to WKY rats (11.0 +/- 2.5 nmol/mg protein x 10 sec) at a magnesium concentration of 1 mM (P less than 0.01). In jejunal BBM, kinetic analysis of magnesium uptake showed three components in WKY rats, with one being diffusional. In SHR, only two components were seen, with the diffusional one being absent. The two saturable components showed Vmax of 6.5 +/- 1.3 and 26.2 +/- 6.0 nmol/mg protein x 10 sec and apparent Km of 0.22 +/- 0.12 mM and 1.9 +/- 0.4 mM in WKY rats, and Vmax of 10.9 +/- 3.5 and 14.8 +/- 5.9 nmol/mg protein x 10 sec and apparent Km of 0.43 +/- 0.23 mM and 1.3 +/- 0.2 mM in SHR. Only the component with the lowest apparent affinity appeared statistically different in SHR as compared with WKY rats for both Vmax and apparent Km (P less than 0.05). Time course evolution of magnesium uptake in jejunal BBM indicated, by extrapolation at zero time, that 2.5 and 5.1 nmol magnesium/mg protein in SHR and WKY rats, respectively, would be in the bound state. The study of the influence of medium osmolarity on 60-min magnesium uptakes was also indicative of a smaller binding compartment in jejunal BBM of SHR (3.70 and 8.26 nmol/mg protein in SHR and WKY rats, respectively); at the four osmolarities assayed, the 60-min uptakes were significantly lower in SHR as compared with WKY rats (P less than 0.01). From 60-min glucose uptakes, a smaller volume of jejunal BBM vesicles was determined for SHR as compared with WKY rats (0.34 +/- 0.06 and 0.63 +/- 0.17 microliter/mg of protein in SHR and WKY rats respectively, P less than 0.05), this volume being significantly augmented by the presence of 1 mM MgCl2 (0.48 +/- 0.05 and 1.27 +/- 0.02 microliter/mg of protein in SHR and WKY rats respectively, P less than 0.01). These results suggest that magnesium uptake and binding by jejunal BBM are altered in SHR in comparison to WKY rats, implying a possible role of the small intestine in the abnormalities of magnesium metabolism in genetic hypertension.     

Increased Na(+)-dependent D-glucose transport and altered lipid composition in renal cortical brush-border membrane vesicles from bile duct-ligated rats.

Erythrocyte membranes of patients with liver disease are characteristically enriched in cholesterol, a change known to impair several carrier-mediated membrane transport functions. In the present study we have assessed whether experimental liver disease can affect the membrane lipid composition and transport function of kidney epithelial cells. Small (about 5%) but significant (P less than 0.01) increases were found in the cholesterol-to-phospholipid molar ratio (C/PL) of rat renal cortical brush-border membrane (BBM) vesicles 3, 8, and 15 days after bile duct ligation which correlated closely with increased fluorescence polarization, i.e., decreased membrane fluidity (r = 0.75, P less than 0.001; n = 27). A lipoprotein-mediated pathogenesis was suggested by the close relationship between BBM C/PL and plasma C/PL (r = 0.69, P less than 0.001). The mean high-affinity Na(+)-coupled D-glucose uptake by BBM vesicles was higher 1, 3, 8, and 15 days after ligation than in non-operated rats, significantly so at 3 and 8 days (611 +/- 37 and 593 +/- 22 vs. 507 +/- 21 pmol/mg protein per 4 sec; P less than 0.05), and was positively correlated with BBM C/PL (r = 0.58, P less than 0.01) and fluorescence polarization (r = 0.41, P less than 0.05). Brief incubation of BBM vesicles from normal rats with cholesterol-rich phospholipid liposomes simultaneously increased BBM C/PL and Na(+)-dependent D-glucose uptake. Stimulation of BBM Na(+)-glucose cotransport in ligated rats was not due to delayed dissipation of the Na+ gradient or to a more rapid development of membrane potential. High-affinity Na(+)-dependent D-glucose uptake kinetics in 3-day bile duct-ligated rats showed a lower Kt, without an alteration in maximum velocity, Vmax, compared to sham-operated animals (0.298 +/- 0.015 vs. 0.382 +/- 0.029 mM; P less than 0.05), whilst the binding dissociation constant, Kd of high-affinity phlorizin binding sites was reduced by ligation (0.453 +/- 0.013 vs. 0.560 +/- 0.015 microM; P less than 0.001). We conclude that an early effect of bile duct ligation is to enrich renal cortical brush-border membranes in cholesterol, thereby decreasing membrane fluidity and stimulating Na(+)-dependent D-glucose uptake by increasing the affinity of the carrier.     

Transport of a tripeptide, Gly-Pro-Hyp, across the porcine intestinal brush-border membrane.

The transcellular transport of oligopeptides across intestinal epithelial cells has attracted considerable interest in investigations into how biologically active peptides express diverse physiological functions in the body. It has been postulated that the tripeptide, Gly-Pro-Hyp, which is frequently found in collagen sequences, exhibits bioactivity. However, the mechanism of uptake of dietary di- and tripeptides by intestinal epithelial cells is not well understood. In this study, we used porcine brush-border membrane (BBM) vesicles to assess Gly-Pro-Hyp uptake, because these vesicles can structurally and functionally mimic in vivo conditions of human intestinal apical membranes. The present study demonstrated the time-dependent degradation of this tripeptide into the free-form Gly and a dipeptide, Pro-Hyp, on the apical side of the BBM vesicles. In parallel with the hydrolysis of the tripeptide, the dipeptide Pro-Hyp was identified in the BBM intravesicular space environment. We found that the transcellular transport of Pro-Hyp across the BBM was inhibited by the addition of a competitive substrate (Gly-Pro) for peptide transporter (PEPT1) and was pH-dependent. These results indicate that Gly-Pro-Hyp can be partially hydrolyzed by the brush-border membrane-bound aminopeptidase N to remove Gly, and that the resulting Pro-Hyp is, in part, transported into the small intestinal epithelial cells via the H+-coupled PEPT1. Gly-Pro-Hyp cannot cross the epithelial apical membrane in an intact form, and Pro-Hyp is highly resistant to hydrolysis by intestinal mucosal apical proteases.     

Effect of heat-stable and heat-labile enterotoxins of Escherichia coli on intestinal brush border membrane enzymes of mice

The activities of intestinal brush border membrane (BBM) enzymes alkaline phosphatase, maltase, lactase, sucrase, gamma-glutamyl transpeptidase and leucine aminopeptidase were determined in intestinal homogenates and purified BBMs from control, heat-stable and heat-labile enterotoxin treated mice. The activities of all the enzymes except lactase were decreased significantly (p less than 0.01) in homogenates while increased significantly (p less than 0.001) in BBMs of experimental groups as compared to controls. Calmodulin activities were increased significantly (p less than 0.01) as compared to control in heat-stable enterotoxin treated mice but remained unaltered in heat-labile enterotoxin treated mice. DNA contents of intestinal homogenates were decreased in experimental groups demonstrating the decrease in cell number in these groups. The altered BBM enzyme activities could not be attributed to changes in calmodulin activities. The increase in enzyme activities in BBMs may reflect a compensatory phenomenon in the remaining cells.     

Effect of antiserum to a 99 kDa polypeptide on the uptake of taurocholic acid by rat ileal brush border membrane vesicles

A 99 kDa polypeptide in rat ileal brush border membrane (BBM), regarded as a component of the active bile acid transport system on account of photoaffinity labeling, has been purified by affinity chromatography and preparative gel electrophoresis and utilized as an immunogen for raising polyclonal antibody. Immune serum, but not preimmune serum, specifically recognized a single band of 99 kDa protein on immunoblots of ileal and renal BBM. In contrast, no reactivity was observed with proteins in jejunal BBM. This polyclonal antibody, compared with preimmune serum and anticytosolic bile acid binding protein (14 kDa) serum, significantly inhibited the Na+ dependent uptake of [3H] taurocholate by BBM vesicles (p less than 0.01). [14C] D-glucose uptake by BBM vesicles was not influenced by the immune serum (p less than 0.01). Thus, these studies provide further support for the specific role of a 99 kDa protein in ileal BBM bile acid transport.     

Mechanism of regulation of rabbit intestinal villus cell brush border membrane Na/H exchange by nitric oxide

In the mammalian small intestine, coupled NaCl absorption occurs via the dual operation of Na/H and Cl/HCO(3) exchange on the villus cell brush border membrane (BBM). Although constitutive nitric oxide (cNO) has been demonstrated to alter gastrointestinal tract functions, how cNO may specifically alter these two transporters to regulate coupled NaCl absorption is unknown. In villus cells, inhibition of cNO synthase (cNOS) with l-N(G)-nitroarginine methylester (l-NAME) stimulated Na/H exchange whereas Cl/HCO(3) exchange was unaffected. In villus cell BBM vesicles (BBMV) prepared from rabbits treated with l-NAME, Na/H exchange was also stimulated. d-NAME, an inactive analog of l-NAME, and N(6)-(1-imonoethyl)-l-lysine dihydrochloride, a more selective inhibitor of inducible NO synthase, did not affect Na/H exchange. Kinetic studies demonstrated that the mechanism of stimulation is secondary to an increase in the maximal rate of uptake of Na, without an alteration in the affinity of the transporter for Na. Northern blot studies demonstrated an increase in the message for the BBM Na/H exchanger NHE3, and Western blot studies showed that the immunoreactive protein levels of NHE3 was increased when cNOS was inhibited. Thus these results indicate that cNO under nominal physiological states most likely maintains an inhibitory tone on small intestinal coupled NaCl absorption by specifically inhibiting BBM Na/H expression.     

Chronic cold stress-induced alterations in brush border membrane composition and enzyme activities in rat intestine

The effect of chronic cold stress on the composition and function of rat intestinal brush border membrane (BBM) was studied. Various lipid fractions from intestinal BBM viz. cholesterol (p < 0.01), phospholipids (p < 0.01), triglycerides (p < 0.05) and gangliosides (p < 0.05) were significantly reduced in cold stressed animals, as compared to controls. Analysis of membrane saccharide content revealed a significant increase in sialic acid (25%) and hexosamine (36%) contents and a reduction in fucose (19%) content in cold stressed rats. Determination of various enzyme activities in BBM showed significantly enhanced activities of alkaline phosphatase ( p < 0.01), lactase ( p < 0.001) and leucine aminopeptidase ( p < 0.001), whereas sucrase activity was reduced ( p < 0.05) under these conditions. The magnitude and site of these alterations across the crypt-villus axis varied from enzyme to enzyme. These findings suggest that chronic cold stress results in profound alterations in intestinal BBM. Altered structure and function of intestinal BBM may play a role in stress-induced derangements in gastrointestinal tract.     

Transport of divalent transition-metal ions is lost in small-intestinal tissue of b/b Belgrade rats

Belgrade rats exhibit microcytic, hypochromic anemia and systemic iron deficiency due to a glycine-to-arginine mutation at residue 185 in a metal ion transporter of a divalent metal transporter/divalent cation transporter/solute carrier 11 group A member 2 or 3 (DMT1/DCT1/SLC11A2), a member of the natural-resistance-associated macrophage protein (Nramp) family. By use of rabbit duodenal tissue, a calcein fluorescence assay has previously been developed to assess transport of divalent metal ions across the small-intestinal brush border membrane (BBM). The assay was readily applied here to rat BBM to learn if it detects DMT1 activity. The results demonstrate protein-mediated transport across the BBM of all tested ions: Mn(2+), Fe(2+), and Ni(2+). Transport into BBM vesicles (BBMV) from (b/b) Belgrade rats was below the detection limit. BBMV of +/b origin had substantial activity. The kinetic rate constant for Ni(2+) membrane transport for +/b BBMV was within the range for normal rabbit tissue. Vesicles from +/b basolateral membranes (BLM) showed similar activity to BBMV while b/b BLM vesicles (BLMV) lacked transport activity. Immunoblots using isoform-specific antibodies demonstrated that intestinal levels of b/b DMT1 were increased compared to +/b DMT1, reflecting iron deficiency. Immunoblots on BBMV indicated that lack of activity in b/b vesicles was not due to a failure of DMT1 to localize to the BBMV; an excess of specific isoforms was present compared to +/b BBMV or duodenal extracts. Immunoblots from BLMV also exhibited enrichment in DMT1 isoforms, despite their distinct origin. Immunofluorescent staining of thin sections of b/b and +/b proximal intestines confirmed that DMT1 localized similarly in mutant and control enterocytes and showed that DMT1 isoforms have distinct distributions within intestinal tissue.     

Role of N-acetylneuraminic acid in rat renal brush-border membrane vesicle aggregation by aminoglycosides

Interaction between aminoglycosides (AGs) and rat renal brush-border membrane (BBM) vesicles was investigated by the aggregation technique. The order of aggregation was gentamicin greater than dibekacin not equal to netilmicin greater than amikacin, and this order corresponds to the strength of the nephrotoxicity of the aminoglycosides in vivo rather than the number of amino groups in the aminoglycosides. BBM vesicles were aggregated through ionic interaction, as evident from the finding that aggregation ceased to occur at alkaline pH. By addition of N-acetylneuraminic acid (NANA) to the incubation medium, the vesicle aggregation induced by gentamicin was significantly inhibited. To affect the liberation of the NANA residue from BBM vesicles, the vesicles were treated with neuraminidase, resulting in an about 60% release with a significant decrease in the uptake of gentamicin into the vesicles. The decrease in the degree of vesicle aggregation was in proportion to the amount of NANA liberated. It follows from the findings that the NANA residue may in some way be responsible for the accumulation of aminoglycosides in renal proximal tubular cells.     

Role of scavenger receptors SR-BI and CD36 in selective sterol uptake in the small intestine

The serum lipoprotein high-density lipoprotein (HDL), which is a ligand of scavenger receptors such as scavenger receptor class B type I (SR-BI) and cluster determinant 36 (CD36), can act as a donor particle for intestinal lipid uptake into the brush border membrane (BBM). Both cholesterol and phospholipids are taken up by the plasma membrane of BBM vesicles (BBMV) and Caco-2 cells in a facilitated (protein-mediated) process. The protein-mediated transfer of cholesterol from reconstituted HDL to BBMV depends on the lipid composition of the HDL. In the presence of sphingomyelin, the transfer of cholesterol is slowed by a factor of about 3 probably due to complex formation between cholesterol and the sphingolipid. It is shown that the mechanism of lipid transfer from reconstituted HDL to either BBMV or Caco-2 cells as the acceptor is consistent with selective lipid uptake: the lipid donor docks at the membrane-resident scavenger receptors which mediate the transfer of lipids between donor and acceptor. Selective lipid uptake implies that lipid, but no apoprotein is transferred from the donor to the BBM, thus excluding endocytotic processes. The two BBM models used here clearly indicate that fusion of donor particles with the BBM can be ruled out as a major mechanism contributing to intestinal lipid uptake. Here we demonstrate that CD36, another member of the family of scavenger receptors, is present in rabbit and human BBM vesicles. This receptor mediates the uptake of free cholesterol, but not of esterified cholesterol, the uptake of which is mediated exclusively by SR-BI. More than one scavenger receptor appears to be involved in the uptake of free cholesterol with SR-BI contributing about 25% and CD36 about 35%. There is another yet unidentified protein accounting for the remaining 30 to 40%.     

Thiol redox and phosphate transport in renal brush-border membrane. Effect of nicotinamide

In the present study, the effect of thiol redox and its possible role in the inhibitory effect of nicotinamide on renal brush-border membrane (BBM) phosphate uptake was examined. Addition of thiol reducing agent, dithiothreitol (DTT, 5 mM), caused an increase, while addition of thiol oxidant, diamide (DM, 5 mM) caused a reversible decrease in sodium-dependent BBM phosphate uptake. Kinetic analyses revealed an increase in both Vmax and Km by DTT, and a decrease in Vmax by DM. These results suggest that thiol redox influences BBM phosphate uptake with sulfhydryl (SH) groups relate to its capacity and disulfide (SS) groups to its affinity for phosphate. Since changes in cytosolic NAD levels may affect BBM thiol redox through changes in redox states of NADP and glutathione systems, we have examined such possibility by studying the effect of nicotinamide (NM). Incubation of proximal tubules with NM (10 mM) induced an oxidative effect on redox states of cytosolic NAD, NADP systems as inferred from decreased cellular lactate/pyruvate, malate/pyruvate, respectively. Measurements of cytosolic glutathiones and BBM thiols also revealed that NM pretreatment shifted the cytosolic glutathione redox (GSH/GSSG) and BBM thiol redox (SH/SS) toward more oxidized state. On the other hand, incubation of proximal tubules with NM suppressed phosphate uptake by the subsequently isolated BBM vesicles. The lower phosphate uptake by NM-pretreated BBM vesicles was reversed by DTT and was resistant to the inhibitory effect of DM. These results thus suggest that BBM thiol oxidation may be involved in the inhibitory effect of NM on BBM phosphate uptake.     

How many Na+-dependent carriers for L-alanine and L-proline in the eel intestine? Studies with brush-border membrane vesicles

Using brush-border membrane (BBM) vesicles prepared from the intestine of the European eel, the specificity of L-alanine and L-proline Na+-dependent transport was investigated by measuring the uptake of isotopically labelled substrates. In the presence of Na+ ions, cross-inhibition between alanine and proline transports was observed; in addition alpha-(methylamino)isobutyric acid (MeAIB) inhibited proline but had no effect on alanine uptake. These results can be explained by the presence, in eel intestinal BBM vesicles, of at least two distinct agencies for Na+-dependent proline and alanine translocation. The first system is specific for alanine and short-chain neutral amino acids; the second system, specific for imino acids and the N-methylated analogues, is regulated by alanine concentration.     

Renal brush border membranes from mice with X-linked hypophosphatemia: protein composition, phosphate binding capacity, and protein kinase activity

Phosphate uptake by brush-border membrane (BBM) vesicles prepared from hypophosphatemic mice (Hyp) is reduced by half relative to BBM vesicles from normal mice. To investigate this abnormality, we studied the protein composition of BBM, their capacity to bind inorganic phosphate, and their protein kinase activity with and without the addition of exogenous cAMP, in normal and Hyp mice. Gradient polyacrylamide gel electrophoresis of BBM proteins showed 27 bands which were identical in normal and Hyp mice. Incubation of the membranes with ortho[32P]phosphate at 0 degrees C revealed a phosphate binding protein with an apparent molecular weight (Mr) of 79000, which has been previously identified in rats as the monomer of alkaline phosphatase. In normal mice, the Scatchard plot of phosphate binding was not linear, suggesting heterogeneity of the binding sites with two major components. At high substrate concentrations, the affinity (K) was 1.42 mM and maximal binding (Bmax) was 83 pmol/mg protein. At low substrate concentrations, these values were 0.07 mM and 10.9 pmol/mg, respectively. In Hyp mice BBM, only one binding system was found with K and Bmax values of 0.38 mM and 53.8 pmol/mg. Incubation of the membranes with 25 microM[gamma-32P]ATP resulted in the phosphorylation of 11 proteins. The major band (Mr: 79000) corresponded to the inorganic phosphate binding protein, i.e., to the alkaline-phosphatase monomer. The 11 proteins showed maximal phosphorylation at pH 10. The protein of 79000 Mr showed a second peak of phosphorylation at pH 7.5.(ABSTRACT TRUNCATED AT 250 WORDS)     

Thyroid hormones increase Na+–Pi co-transport activity in intestinal brush border membrane: Role of membrane lipid composition and fluidity

In the present study, we documented the promising role of thyroid hormones status in animals in modulation of Na⁺–P_i transport activity in intestinal brush border membrane vesicles (BBMV) which was accompanied with alterations in BBM lipid composition and fluidity. Augmentation of net P_i balance in hyperthyroid (Hyper-T) rats was fraternized with accretion of P_i transport across BBMV isolated from intestine of Hyper-T rats as compared to hypothyroid (Hypo-T) and euthyroid (Eu-T) rats while Na⁺–P_i transport across BBMV was decreased in Hypo-T rats relative to Eu-T rats. Increment in Na⁺–P_i transport in intestinal BBMV isolated from Hyper-T rats was manifested as an increase in the maximal velocity (V_max) of Na⁺–P_i transport system. Furthermore, BBMV lipid composition profile in intestinal BBM from Hyper-T was altered to that of Hypo-T rats and Eu-T rats. The molar ratio of cholesterol/phospholipids was higher in intestinal BBM from Hypo-T rats. Fluorescence anistropy of diphenyl hexatriene (rDPH) and microviscosity were significantly decreased in the intestinal BBM of Hyper-T rats and decreased in Hypo-T rats as compared to Eu-T rats which corroborated with the alteration in membrane fluidity in response to thyroid hormone status of animals. Therefore, thyroid hormone mediated change in membrane fluidity might play an important role in modulating Na⁺–P_i transport activity of intestinal BBM. (Mol Cell Biochem 278: 195–202, 2005)     

Effect of chronic inflammation on ileal short-chain fatty acid/bicarbonate exchange

Short-chain fatty acids (SCFA) have been demonstrated to at least partially ameliorate chronic intestinal inflammation. However, whether and how intestinal SCFA absorption may be altered during chronic intestinal inflammation is unknown. A rabbit model of chronic ileitis produced by coccidia was used to determine the effect of chronic inflammation on ileal SCFA/HCO(-)(3) exchange. SCFA/HCO(-)(3) exchange was present in the brush-border membrane (BBM) of villus but not crypt cells from normal rabbit ileum. An anion-exchange inhibitor, DIDS, significantly inhibited SCFA/HCO(-)(3) exchange. Extravesicular Cl(-) did not alter the uptake of SCFA, suggesting that SCFA/HCO(-)(3) exchange is a transport process distinct from Cl(-)/HCO(-)(3) exchange. In chronically inflamed ileum, SCFA/HCO(-)(3) exchange was also present only in BBM of villus cells. The exchanger was sensitive to DIDS and was unaffected by extravesicular Cl(-). However, SCFA/HCO(-)(3) exchange was significantly reduced in villus cell BBM vesicles (BBMV) from chronically inflamed ileum. Kinetic studies demonstrated that the maximal rate of uptake of SCFA, but not the affinity for SCFA, was reduced in chronically inflamed rabbit ileum. These data demonstrate that a distinct SCFA/HCO(-)(3) exchange is present on BBMV of villus but not crypt cells in normal rabbit ileum. SCFA/HCO(-)(3) exchange is inhibited in chronically inflamed rabbit ileum. The mechanism of inhibition is most likely secondary to a reduction in transporter numbers rather than altered affinity for SCFA.     

Effect of cisplatin on brush border membrane enzymes and anti-oxidant system of rat intestine

Cisplatin (CP) is a widely used antineoplastic agent which exhibits gastrointestinal toxicity. The present work was done to study the effect of administration of CP on brush border membrane (BBM) enzymes and anti-oxidant system of rat intestine. Male Wistar rats were given a single intraperitoneal dose of CP (6 mg/kg body weight) and then sacrificed 1, 3, 5 and 7 days after this treatment. Control animals were given saline only. The administration of CP led to significant decline in the specific activities of BBM enzymes both in the mucosal homogenates and isolated membrane vesicles. Kinetic studies showed that the V(max) of the enzymes was decreased in BBM vesicles from CP treated rats while the K(m) remained unchanged. The activities of catalase, Cu-Zn superoxide dismutase, glucose 6-phosphate dehydrogenase and glutathione reductase decreased while the activities of glutathione S-transferase and thioredoxin reductase increased in CP treated animals compared to the control group. Lipid peroxidation and total sulfhydryl groups were also altered upon CP treatment indicating the generation of oxidative stress. The maximum changes in all the parameters studied above were 3 days after administration of CP and then recovery took place on days 5 and 7. Thus, the administration of CP leads to significant alterations in the activities of BBM enzymes and the anti-oxidant status of rat intestine.     

ATP-driven copper transport across the intestinal brush border membrane

The divalent metal ion transporter DMT1 is localized in the brush border membrane (BBM) of the upper small intestine and has been shown to be able to transport Mn2+, Fe2+, Co2+, Ni2+, and Cu2+. Belgrade rats have a glycine-to-arginine (G185R) mutation in DMT1, which affects its function. We investigated copper transport with BBM vesicles of Belgrade rats loaded with calcein, which exhibits fluorescence quenching by various metal ions. Transport of copper was disrupted in unenergized BBM vesicle of b/b Belgrade rats, as had been described for iron transport, while +/b vesicles exhibited normal transport by DMT1. When either b/b or +/b vesicles were loaded with ATP and magnesium, similar high-affinity accumulation of copper was observed in both types of vesicles. Thus, brush border membranes possess an ATP-driven, high-affinity copper transport system which could serve as the primary route for copper uptake by the intestine.     

Characteristics of Na(+)-dependent intestinal nucleoside transport in the pig

Since the capacity of nucleic acid digestion and absorption appears to be comparatively high in the pig, we investigated the properties of transport of (3)H-labelled nucleosides across the porcine intestinal brush border membrane (BBM) using BBM vesicles isolated from the small intestine of slaughter pigs. In the presence of a transmembrane Na(+) gradient, uridine, thymidine and guanosine transiently accumulated in the vesicular lumen beyond the equilibrium (60 min) value suggesting the presence of Na(+)/nucleoside cotransporters in the BBM. The findings of inhibitory studies are consistent with the presence of two Na(+)-dependent nucleoside transporters with overlapping substrate specificity, one for pyrimidine nucleosides (N2) and one for purine nucleosides (N1). Guanosine appeared to be a specific substrate for N1, while this applies to thymidine for N2. Transport of thymidine and guanosine were also inhibited by 2 mmol/l D-glucose and alpha-methyl-D-glucoside. The maximal transport capacity (V(max)) for Na(+)-dependent thymidine and guanosine transport were much higher than reported for other monogastric species. Unlike in other species tested, there was no proximal-to-distal gradient, neither in nucleoside transport activity nor in the inhibition of nucleoside transport by monosaccharides in the porcine small intestine. The high intestinal nucleoside transport activity may contribute to the high digestive capacity for nucleic acids in the pig.