Search PubMed⌕ Search

Biomedical subjects

H Murer

Publications and source records attributed to H Murer.

At least 361 records · Page 20Linked to original sources
PubMed1981

A high yield preparation for rat kidney brush border membranes. Different behaviour of lysosomal markers.

Rat kidney cortex slices were homogenized with a polytron in a isoosmotic medium containing 5 mmol/l EGTA. By two precipitations with MgCl2 (12 mmol/l) and differential centrifugation, brush border membranes were purified. The brush border marker enzymes alkaline phosphatase and aminopeptidase M were found to be enriched 17.0 +/- 5.3-fold and 16.7 +/- 3.7-fold, respectively. By this method, a high yield of brush border membranes was obtained (48.3 +/- 7.9% for alkaline phosphatase; 47.0 +/- 9.5% for aminopeptidase M). The acid phosphatase was enriched 5-fold, whereas other lysosomal enzymes (glucosaminidase, glucuronidase, cathepsin D) were enriched only 0.2-fold. Acid phosphatase activity could not be washed out, but could be separated from alkaline phosphatase and leucine aminopeptidase by means of free flow electrophoresis and sucrose density gradient centrifugation. Vesicles prepared by the presently described Mg/EGTA-method show better transport properties, compared to vesicles prepared by the calcium method of Evers et al. (Evers, C., Haase, W., Murer, H. and Kinne, R. (1978) Membrane Biochem. 1, 203-219), whereas by SDS-polyacrylamide gel electrophoresis, no differences in the protein patterns were observed.

Acid Phosphatase↗
PubMed1981

The influence of pH on phosphate transport into rat renal brush border membrane vesicles.

Sodium-dependent transport of inorganic phosphate into brush border membrane vesicles is strongly influenced by altering pH of the incubation medium (pHo). At constant total phosphate concentration an increase in pHo leads to an increase in the uptake of inorganic phosphate. Uptake of inorganic phosphate, however, is not affected by the intravesicular pH (pHi) or by transmembrane pH differences (pHo--pHi). If initial phosphate uptake is studied as a function of total phosphate concentration in the medium the half saturation concentration increases when pHo is raised from 6.3--6.9 but remains unaltered between pHo 6.9 and 7.8 Vmax increases about 3-fold between pHo 6.3 and 6.9 and by a factor of about 1.6 between pHo 6.9 and 7.4. The pHo- dependence of phosphate uptake is diminished by increasing sodium concentrations. Altering transmembrane electrical potential difference by potassium + valinomycin-induced diffusion potentials or by anion replacement fails to demonstrate electrogenicity of sodium-phosphate cotransport. Experiments using a potential-sensitive fluorescent dye, however, indicate a vesicle inside positive electrical potential difference when inorganic phosphate is added. The phosphate-induced alterations in the electrical potential difference are sodium-dependent and more pronounced at low pHo values. Together with earlier observations these results suggest that translocation of inorganic phosphate across the proximal tubular brush border membrane is mediated by cotransport of 2 sodium ions with one either monovalent or divalent phosphate molecule according to its availability in the tubular fluid. The pH sensitivity of this transport system is rather due to alterations in the transport system itself than to pH- dependent alterations in the ratio of monovalent to divalent phosphate.

Animals↗
PubMed1981

Transcellular transport of calcium and inorganic phosphate in the small intestinal epithelium.

Transport mechanisms involved in the small intestinal handling of inorganic phosphate and calcium have been studied by different in vitro methods during the last few years. In concordance with studies on intact epithelial preparations, studies with brush-border and basal-lateral membrane vesicles isolated from the small intestinal epithelial cell revealed that transcellular calcium and inorganic phosphate fluxes are coupled to transcellular sodium flux, i.e., secondary active via coupling to the primary active sodium flux. A sodium-coupled mechanism in the brush-border membrane leads to cellular accumulation of inorganic phosphate. A sodium-coupled mechanism leads to extrusion of calcium from the cell into the serosal interstitium. A primary active transport mediated by the Ca-ATPase and located in the basal-lateral membrane also exists for calcium. Regulation of transcellular phosphate and calcium flux proceeds via altered influx rates at the luminal cell pole.

Animals↗
PubMed1981

Sodium-coupled taurocholate transport in the proximal convolution of the rat kidney in vivo and in vitro.

Using the standing droplet technique in the renal proximal convolution and simultaneous microperfusion of the peritubular capillaries, the zero net flux transtubular concentration difference of taurocholate (DeltaC(TC-)) at 45 s was determined as a measure of active bile acid reabsorption in vivo. Starting with 0.1 mmol/liter taurocholate in both perfusates the control DeltaC(TC-) of 0.042 mmol/liter fell to 0.006 mmol/liter (P < 0.001) when the Na(+) concentration in the perfusates was reduced to zero. Removal of bicarbonate from the perfusates to alter pH had no influence on DeltaC(TC-). When glycocholate was added to the perfusates DeltaC(TC-) was decreased, while probenecid increased DeltaC(TC-). These observations were extended by studies performed with brush border membrane vesicles derived from renal cortex. The initial (20 s) uptake of 0.01 mmol/liter taurocholate in the presence of a Na(o) (+) > Na(i) (+) gradient was stimulated twofold compared with its uptake in the absence of a Na(+) gradient. Uptake of taurocholate was osmotically and temperature sensitive. Membranes preloaded with unlabeled glycocholate showed accelerated entry of labeled taurocholate (trans-stimulation) only in the presence of Na(+). Replacement of Na(+) in the media with K(+), Li(+), and choline(+) decreased initial taurocholate uptake by 49, 53, and 62%, respectively. Stimulation of taurocholate transport by cation gradient diffusion potentials was unlikely inasmuch as the addition of valinomycin under K(+) gradient conditions had no effect. A transmembrane pH gradient (pH(o) < pH(i)) did not influence initial uptake of taurocholate. Finally, in the presence of Na(+) taurocholate transport showed cis-inhibition with unlabeled bile acids and saturation kinetics with respect to increasing taurocholate concentrations. The micropuncture and vesicle data indicate that the net transport of taurocholate in the proximal tubule is the result of an electroneutral Na(+)-taurocholate cotransport across the brush border membrane.

Animals↗
PubMed1981

A simple isolation method for basal-lateral plasma membranes from rat kidney cortex.

Basal-lateral membranes were separated in a self-orienting Percoll (modified colloidal silica) gradient from a heavy microsomal membrane fraction by centrifugation at 48,000g for 0.5 h. The (Na+--K+)-ATPase activity as a marker enzyme for the basal-lateral plasma membrane was 20-fold enriched by this procedure. The adenylate-cyclase activity measured in the basal-lateral membrane fraction was stimulated 6-fold by parathyrin and only up to 1.5-fold by arginine-vasopressin, calcitonin, or isoproterenol. The yield of basal-lateral plasma membranes was 5 to 10 percent of the amount initially present in the homogenate. The method is also applicable to the pig kidney.

Adenylyl Cyclases↗
PubMed1981

Sulfate-sodium cotransport by brush-border membrane vesicles isolated from rat ileum.

Uptake of inorganic sulfate into brush-border membrane vesicles isolated by a calcium precipitation method from rat small intestine was investigated using a rapid filtration technique and 35Sulfur acid as tracer. Sulfate uptake by membrane vesicles was osmotically sensitive, suggesting transport into an intravesicular space rather than binding to or incorporation into the membrane. Transport of sulfate into brush-border vesicles isolated from rat ileum was only stimulated by sodium ions as compared with other monovalent cations. A typical "overshoot" phenomenon was observed in the presence of an inwardly directed NaCl gradient. Tracer sulfate exchange was faster in the presence of sodium than in the presence of potassium. Addition of the ionophores for monovalent cations, monactin, or gramicidin D, decreased the sodium gradient-driven sulfate uptake. Sulfate uptake showed a saturation phenomenon only in the presence of sodium. Transstimulaton of sodium-dependent sulfate transport was shown with MoO4(2-), but not with PO4(2-) and WO4(2-). Changing the electrical potential difference across the membrane vesicles by establishing different diffusion potentials (anion replacement; potassium gradient +/- valinomycin) did not alter sodium-dependent sulfate uptake. Stimulation of sulfate transport by sodium was greater in membrane vesicles from ileal segments than from duodenum or jejunum. It is concluded that isolated brush-border membranes of rat ileum contain an electroneutrol sodium-sulfate cotransport system.

Animals↗
PubMed1980

The use of isolated membrane vesicles to study epithelial transport processes.

Epithelia are multicompartment and multicomponent systems performing transcellular and paracellular transport in a very complex manner. One way to get a deeper understanding of the function of such a complex system is to dissect it into the single components and then, after having defined the components under well-controlled conditions, to try to describe the behavior of the whole system on the basis of the properties of the single components. This article deals with the analysis of isolated plasma membranes derived from the luminal and contraluminal face of epithelial cells, predominantly renal proximal tubular and small intestinal cells. It is aimed to give an overview of methods used to isolate and separate plasma membranes, to study their transport properties as membrane vesicles, and also to address the question of how information gained with the isolated membranes corresponds to observations made in the intact cell using other, notably electrophysiological, measurements. The review also critically evaluates the limitations of the approach and thereby tries to set the work on isolated membranes in the proper perspective within the field of transport physiology.

Animals↗
PubMed1980

The effects of potassium and membrane potential on sodium-dependent glutamic acid uptake.

The uptake of L-glutamic acid into brush-border membrane vesicles isolated from rat renal proximal tubules is NA+-dependent. In contrast to Na+-dependent uptake of D-glucose, pre-equilibration of the vesicles with K+ stimulates L-glutamic acid uptake. Imposition of a K+ gradient ([Ki+] > [Ko+]) further enhances Na+-dependent L-glutamic acid uptake, but leaves K+-dependent glucose transport unchanged. If K+ is present only at the outside of the vesicles, transport is inhibited. Intravesicular Rb+ and, to a lesser extent, Cs+ can replace intravesicular K+ to stimulate L-glutamic acid uptake. Changes in membrane potential incurred by the imposition of an H+-diffusion potential or anion replacement markedly affect Na+-dependent glutamic acid uptake only in the presence of K+. Experiments with a potential-sensitive cyanine dye also indicate that, in the presence of intravesicular K+ a charge movement is involved in Na+-dependent transport of L-glutamic acid. The data indicate that Na+-dependent L-glutamic acid transport can be additionally energized by a K+ gradient. Furthermore, intravesicular K+ render Na+-dependent L-glutamic acid transport sensitive to changes in the transmembrane electrical potential difference.

Animals↗
PubMed1980

Sodium ion/L-lactate co-transport in rabbit small-intestinal brush-border-membrane vesicles.

Uptake of L-lactate into rabbit jejunal brush-border-membrane vesicles prepared by a Ca2+-precipitation procedure was studied by a rapid filtration technique with L-[14C]-lactate as tracer. Transport of L-lactate into an intravesicular (osmotically reactive) space could be established. An inwardly directed NaCl gradient (outside 21 mM/inside 0mM) stimulated the uptake of L-lactate at 15 s 2-4-fold compared with that observed with an equal KCl gradient. A transient accumulation of L-lactate inside the vesicles (overshoot) was observed in the presence of an NaCl gradient. Gradients of LiCl, RbCl, CsCl or choline chloride were not able to replace NaCl in the stimulation of L-lactate uptake. L-Lactate uptake was saturable only in the presence of Na+. D-Lactate, DL-thiolactate (2-DL-mercaptopropionate), pyruvate and propionate inhibited the Na+-stimulated L-lactate uptake; D-lactate, thiolactate and pyruvate provoked trans-stimulation of L-lactate uptake. Artificially imposed diffusion potentials (inside negative) did not exert any effect on the Na+-dependent L-lactate uptake. The results are consistent with the existence of an electroneutral Na+/L-lactate co-transport system in the brush border of rabbit small intestine.

Animals↗
PubMed1980

A simple and fast method for the isolation of basolateral plasma membranes from rat small-intestinal epithelial cells.

A method was developed for the analytical and preparative isolation of basolateral plasma membranes from rat small intestine. They were separated on a self-orientating Percoll (modified colloidal silica) gradient starting with a heavy microsomal-membrane fraction and involving centrifugation at 48,000 g for 1 h. (Na+ + K+)-stimulated ATPase activity, used as a marker enzyme for the basolateral plasma membrane, is enriched 20-fold compared with that found in the homogenate of isolated intestinal epithelial cells.

Animals↗
PubMed1980

Polar distribution of sodium-dependent and sodium-independent transport system for L-lactate in the plasma membrane of rat enterocytes.

The uptake of L-lactate by rat small intestinal brush-border and basal-lateral plasma membrane vesicles has been studied. L-Lactate uptake by the isolated membrane vesicles is osmotically sensitive and represents predominantly transport into an intravesicular space and not binding to the membranes. The transport of L-lactate across the brush-border membrane is stimulated by sodium, whereas the transport across the basal-lateral plasma membrane is sodium-independent. In both types of membrane vesicles L-lactate is transported faster than D-lactate and L-lactate transport is inhibited by alpha-cyano-cinnamic acid. L-Lactate transport across basal-lateral membranes is inhibited by D-lactate and pyruvate and transstimulated by L-lactate and pyruvate. The polar distribution of transport system for L-lactate in the plasma membrane of rat enterocytes--a Na+/L-lactate cotransport system in the brush-border membrane and a facilitated diffusion system in the basal-lateral membrane--can explain the fact that in the intact epithelium L-lactate produced by cell metabolism is preferentially released on the serosal side and could enable the cell to perform vectorial, secondary active transport of L-lactate from the intestinal lumen to the serosal compartment.

Animals↗
PubMed1980

Lactate-sodium cotransport in rat renal brush border membranes.

Brush border membrane vesicles were obtained from rat kidney cortex through a calcium precipitation method and their transport properties for lactate were studied by a rapid-filtration technique. Transient concentrative uptake of L-lactate was observed in the presence of inwardly directed NaCl gradient, but not in the presence of a KCl, LiCl, RbCl, CsCl, or choline chloride gradient. The sodium-dependent L-lactate uptake was saturable and was inhibited by D-lactate. The activation curve with sodium was hyperbolic. Maneuvers that render the inside of the vesicle more negative stimulated sodium-dependent L-lactate uptake, suggesting an electrogenic transfer of L-lactate and sodium. An L-lactate gradient also accelerates the sodium movement across the brush border membrane. Studies on the pH dependency of L-lactate transport and on the effect of L-lactate on proton movements across the brush border membrane indicate that there is little contribution of nonionic diffusion and/or of lactate-H+ cotransport to the transfer of L-lactate across the renal brush border membrane. In summary, sodium-lactate cotransport is the major mechanism for L-lactate transfer across the renal brush border membrane.

Animals↗
PubMed1979

[Principles of epithelial transport in the kidney and intestines].

Epithelia of kidney and small intestine consist of one layer of cells which, at their luminal edge, are linked together by terminal bars. Solute transport proceeds either across the cells, which is true of all active transports, or it proceeds paracellularly through the basolateral spaces and terminal bars and is then passive. The driving force for the active transport of a substance is derived either directly from metabolism (primary active transport), or from the gradient of another solute, usually Na+, which in turn is created by primary active transport. In the latter case the transport is referred to as secondary active. The driving forces of passive transport are the electrochemical gradient of the respective substance and solvent drag. The proximal parts of the kidney as well as of the intestine are leaky so that a considerable part of net reabsorption proceeds passively. Their distal parts, however, where the transport is regulated, are tight so that large concentration differences can be created and maintained. Transcellular active transport is only possible if the cells are polar, i.e., the transport characteristics of the luminal cell membrane differ from those of the contraluminal cell membrane. By measuring the cellular electrical potential difference or by measuring transport into isolated plasma membrane vesicles from either cell side the driving forces for the two transport steps, the luminal and contraluminal, have been elucidated. Schemes for the transport steps in the proximal tubule and in the small intestine are given. They show the principal similarity of the transport of many substances in both epithelia.

Absorption↗
PubMed1979

Sulphate-ion/sodium-ion co-transport by brush-border membrane vesicles isolated from rat kidney cortex.

Uptake of SO(4) (2-) into brush-border membrane vesicles isolated from rat kindey cortex by a Ca(2+)-precipitation method was investigated by using a rapid-filtration technique. Uptake of SO(4) (2-) by the vesicles was osmotically sensitive and represented transport into an intra-vesicular space. Transport of SO(4) (2-) by brush-border membranes was stimulated in the presence of Na(+), compared with the presence of K(+) or other univalent cations. A typical ;overshoot' phenomenon was observed in the presence of an NaCl gradient (100mm-Na(+) outside/zero mm-Na(+) inside). Radioactive-SO(4) (2-) exchange was faster in the presence of Na(+) than in the presence of K(+). Addition of gramicidin-D, an ionophore for univalent cations, decreased the Na(+)-gradient-driven SO(4) (2-) uptake. SO(4) (2-) uptake was only saturable in the presence of Na(+). Counter-transport of Na(+)-dependent SO(4) (2-) transport was shown with MoO(4) (2-) and S(2)O(3) (2-), but not with PO(4) (2-). Changing the electrical potential difference across the vesicle membrane by establishing different diffusion potentials (anion replacement; K(+) gradient+/-valinomycin) was not able to alter Na(+)-dependent SO(4) (2-) uptake. The experiments indicate the presence of an electroneutral Na(+)/SO(4) (2-)-co-transport system in brush-border membrane vesicles isolated from rat kidney cortex.

Animals↗
PubMed1979

Effect of dietary phosphate intake on phosphate transport by isolated rat renal brush-border vesicles.

Renal brush-border membrane vesicles isolated from rats kept for 6-8 weeks on a low-phosphate diet (0.15% of dry matter) showed a markedly faster Na(+)-dependent phosphate uptake than did membrane vesicles isolated from animals kept on a high-phosphate diet (2% of dry matter). Phosphate-uptake rate by brush-border membrane vesicles isolated from animals on a low-phosphate diet remained significantly increased after acute parathyroidectomy. Dietary adaptation was also observed in animals that had been parathyroidectomized before exposure to the different diets. In animals on the low-phosphate diet parathyrin administration inhibited phosphate uptake by brush-border vesicles only if the animals were repleted with P(i) (5ml of 20mm-NaH(2)PO(4)) 1h before being killed. After acute phosphate loading and parathyrin administration the difference in the transport rate between the two dietary groups remained statistically significant. The results suggest that the adaptation of proximal-tubule phosphate transport to dietary intake of phosphate is reflected in the Na(+)/phosphate co-transport system located in the luminal membrane of the proximal-tubule cell. Since the dietary effects on phosphate transport by brush-border membranes are only partially reversed by acute changes in parathyrin concentration and are also observed in chronically parathyroidectomized animals, the adaptation of the Na(+)/phosphate co-transport system to dietary phosphate intake seems to involve an additional mechanism independent of parathyrin.

Alkaline Phosphatase↗