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M R Hammerman

Publications and source records attributed to M R Hammerman.

124 records · Page 7Linked to original sources
PubMed1983

Effect of parathyroid hormone on Na+-dependent phosphate transport and cAMP-dependent 32P phosphorylation in brush border vesicles from isolated perfused canine kidneys.

Concentrative uptake of 32Pi induced by the dissipation of a Na+ gradient (overshoot) was demonstrated in brush border membrane vesicles obtained from isolated perfused canine kidneys. Na+-dependent 32Pi transport was decreased in brush border vesicles from isolated kidneys perfused with parathyroid hormone (PTH) for 2 h compared to uptake measured in vesicles from kidneys perfused without PTH. Cyclic AMP-dependent 32P phosphorylation of a 62,000 Mr protein band was demonstrable on autoradiograms of sodium dodecyl sulfate-polyacrylamide gels of membrane suspensions from kidneys perfused +/- PTH. Evidence that perfusion with PTH resulted in cAMP-dependent phosphorylation in isolated kidneys from parathyroidectomized dogs (decreased cAMP-dependent 32P phosphorylation of the 62,000-Mr band in brush border vesicles) was obtained after 2-h perfusion with PTH. Decreased 32P phosphorylation was not observed if membranes were allowed to dephosphorylate prior to 32P phosphorylation in vitro. We conclude that brush border vesicles from isolated perfused canine kidneys can be used to study the action of PTH on Na+-Pi cotransport in brush border membranes and on cAMP-dependent phosphorylation of the membrane. It is strongly suggested that PTH effects changes in Na+-dependent 32Pi transport in isolated brush border vesicles and changes in 32P phosphorylation of vesicles via a direct action on the renal cortical cell rather than as a consequence of extrarenal actions of the hormone.

Animals↗
PubMed1983

Metabolic acidosis and parathyroidectomy increase Na+-H+ exchange in brush border vesicles.

Na+-H+ exchange across the brush border membrane of the renal proximal tubular cell is a mechanism for Na+ reabsorption and H+ secretion. An electroneutral Na+-H+ exchange activity has been identified in isolated renal brush border membrane vesicles from rat and dog kidney, and increased Na+-H+ exchange has been measured in brush border membrane vesicles from remnant kidneys of dogs with chronic renal failure. To ascertain whether changes in H+ secretion by the kidney observed in chronic metabolic acidosis and in states of altered parathyroid function might result from altered Na+-H+ exchange across the renal cortical cellular brush border membrane, we measured Na+-H+ exchange in brush border membrane vesicles from kidneys of dogs with chronic metabolic acidosis and from kidneys of thyroparathyroidectomized dogs. Increased amiloride-sensitive Na+-H+ exchange was demonstrated in brush border membrane vesicles from kidneys of both groups of dogs, suggesting that adaptations in H+ excretion in chronic metabolic acidosis and hypoparathyroidism might be explained by increased activity of a renal brush border membrane Na+-H+ exchanger.

Acidosis↗
PubMed1983

Pi transport, phosphorylation, and dephosphorylation in renal membranes from HYP/Y mice.

To ascertain whether cAMP-dependent phosphorylation could be demonstrated in brush border membrane vesicles (BBMV) isolated from kidneys of mice with X-linked hypophosphatemic rickets (HYP/Y) and normal littermates (+/Y) and, if so, to determine whether the absence of dephosphorylation might underlie differences in Na+-dependent 32Pi transport in BBMV, we measured 1) 32Pi transport, 2) cAMP-dependent phosphorylation, and 3) dephosphorylation in BBMV from +/Y and HYP/Y mice. Na+ gradient-dependent 32Pi transport was decreased in BBMV from HYP/Y mice as reflected in a decreased apparent Vmax. cAMP-dependent phosphorylation of a 62,000 Mr protein was demonstrated in sodium dodecyl sulfate polyacrylamide gels of BBMV from +/Y and HYP/Y mice and was associated with decreased Na+-dependent 32Pi transport. Dephosphorylation of the 62,000 Mr band was demonstrable in both types of membranes. Thus, both cAMP-dependent protein kinase and phosphoprotein phosphatase activities were demonstrable in BBMV isolated from +/Y and HYP/Y mice. These results are consistent with the renal tubular defect in the HYP/Y mouse reflecting an intrinsic abnormality of Pi transport in the brush border membrane independent from mediation of the phosphaturic effect of parathyroid hormone.

Animals↗
PubMed1982

Na+-dependent transport of glycine in renal brush border membrane vesicles. Evidence for a single specific transport system.

The uptake of glycine in rabbit renal brush border membrane vesicles was shown to consist of glycine transport into an intravesicular space. An Na+ electrochemical gradient (extravesicular greater than intravesicular) stimulated the initial rate of glycine uptake and effected a transient accumulation of intravesicular glycine above the steady-state value. This stimulation could not be induced by the imposition of a K+, Li+ or choline+ gradient and was enhanced as extravesicular Na+ was increased from 10 mM to 100 mM. Dissipation of the Na+ gradient by the ionophore gramicidin D resulted in diminished Na+-stimulated glycine uptake. Na+-stimulated uptake of glycine was electrogenic. Substrate-velocity analysis of Na+-dependent glycine uptake over the range of amino acid concentrations from 25 microM to 10 mM demonstrated a single saturable transport system with apparent Km = 996 microM and Vmax = 348 pmol glycine/mg protein per min. Inhibition observed when the Na+-dependent uptake of 25 microM glycine was inhibited by 5 mM extravesicular test amino acid segregated dibasic amino acids, which did not inhibit glycine uptake, from all other amino acid groups. The amino acids D-alanine, D-glutamic acid, and D-proline inhibited similarly to their L counterparts. Accelerative exchange of extravesicular [3H]glycine was demonstrated when brush border vesicles were preloaded with glycine, but not when they were preloaded with L-alanine, L-glutamic acid, or with L-proline. It is concluded that a single transport system exists at the level of the rabbit renal brush border membrane that functions to reabsorb glycine independently from other groups of amino acids.

Amino Acids↗
PubMed1982

Na+-independent L-arginine transport in rabbit renal brush border membrane vesicles.

Na+-independent L-arginine uptake was studied in rabbit renal brush border membrane vesicles. The finding that steady-state uptake of L-arginine decreased with increasing extravesicular osmolality and the demonstration of accelerative exchange diffusion after preincubation of vesicles with L-arginine, but not D-arginine, indicated that the uptake of L-arginine in brush border vesicles was reflective of carrier-mediated transport into an intravesicular space. Accelerative exchange diffusion of L-arginine was demonstrated in vesicles preincubated with L-lysine and L-ornithine, but not L-alanine or L-proline, suggesting the presence of a dibasic amino acid transporter in the renal brush border membrane. Partial saturation of initial rates of L-arginine transport was found with extravesicular [arginine] varied from 0.005 to 1.0 mM. L-Arginine uptake was inhibited by extravesicular dibasic amino acids unlike the Na+-independent uptake of L-alanine, L-glutamine, glycine or L-proline in the presence of extravesicular amino acids of similar structure. L-Arginine uptake was increased by the imposition of an H+ gradient (intravesicular pH less than extravesicular pH) and H+ gradient stimulated uptake was further increased by FCCP. These findings demonstrate membrane-potential-sensitive, Na+-independent transport of L-arginine in brush border membrane vesicles which differs from Na+-independent uptake of neutral and acidic amino acids. Na+-independent dibasic amino acid transport in membrane vesicles is likely reflective of Na+-independent transport of dibasic amino acids across the renal brush border membrane.

Amino Acids↗
PubMed1982

Cyclic AMP-dependent protein phosphorylation in canine renal brush-border membrane vesicles is associated with decreased phosphate transport.

It is known that the administration of parathyroid hormone to dogs results in phosphaturia and decreased phosphate transport in brush-border vesicles isolated from the kidneys of those dogs. Parathyroid hormone has been shown to activate adenylate cyclase at the basal-lateral membrane of the renal proximal tubular cell. It has been postulated that parathyroid hormone-induced phosphaturia is effected through phosphorylation of brush-border protein by membrane-bound cAMP-dependent protein kinase. An experimental system was designed such that phosphorylation of brush-border vesicles and Na+-stimulated solute transport could be studied in the same preparations. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane vesicles revealed cAMP-dependent phosphorylation of 2 protein bands (Mr = 96,000 and 62,000), which was enhanced by exposure of the inside of the membrane vesicles to ATP and cAMP. Cyclic AMP-dependent phosphorylation of brush-border vesicles was accompanied by inhibition of Na+-stimulated Pi but not D-glucose transport or 22Na+ uptake. When renal brush-border vesicles from parathyroidectomized and normal dogs were phosphorylated in vitro in the presence and absence of cAMP, both the cAMP-dependent phosphorylation and inhibition of Na+-stimulated Pi transport were greater in vesicles isolated from kidneys of parathyroidectomized dogs relative to control animals. We conclude that the cAMP-dependent phosphorylation of brush-border membrane-vesicle proteins is associated with specific inhibition of Na+-stimulated Pi transport. The phosphaturic action of parathyroid hormone (PTH) could be mediated through the cAMP-dependent phosphorylation of specific brush-border membrane proteins.

Animals↗
PubMed1982

Decreased luminal membrane transport of phosphate in chronic renal failure.

To examine the effects of the hyperparathyroidism that accompanies chronic renal failure (CRF) on Pi transport across the renal proximal tubular luminal membrane, brush border membrane vesicles (BBMV) were prepared from the viable portion of the kidney of dogs with CRF or normal renal function. Initial rates of Na+ gradient-dependent Pi transport were decreased in BBMV from dogs with CRF compared with normal dogs. Initial rates of 22Na+ uptake in BBMV from dogs with CRF, however, were increased, suggesting a more rapid dissipation of the Na+ gradient in these vesicles. Initial rates of Pi transport measured under Na+-equilibrated conditions were decreased in BBMV from dogs with CRF. Parathyroidectomy of dogs with CRF abolished this decrease in Pi uptake. We conclude that the adaptations in Pi excretion associated with CRF are accompanied by changes in Na+-dependent Pi transport in BBMV and, therefore, possibly across the luminal membrane of the renal tubule. Moreover, the activity of this transport system can be altered by parathyroidectomy of dogs prior to isolation of BBMV.

Animals↗
PubMed1982

Uptake of Pi in brush border vesicles after release of unilateral ureteral obstruction.

After release of complete unilateral ureteral obstruction, a decreased fractional excretion of phosphate (Pi) is observed in the postobstructed kidney compared with the nonobstructed (control) kidney. To determine whether this decrease in the urinary excretion of Pi is due to changes in Na+-dependent Pi transport across the renal brush border membranes of postobstructed and control kidneys, membrane vesicles were prepared from the brush borders of kidneys from dogs that had undergone complete unilateral ureteral obstruction. Alkaline phosphatase activity was decreased in membrane vesicles isolated from postobstructed kidneys. No differences were observed in Na+-dependent Pi transport or in Na+ uptake in membrane vesicles isolated from postobstructed as compared with control kidneys. The in vivo administration of parathyroid hormone decreased Na+-dependent Pi transport in membrane vesicles isolated from postobstructed and control kidneys despite the absence of a phosphaturic response. Our findings suggest that no intrinsic change in the transport characteristics of Pi across the luminal membrane of renal tubular cells occurs with unilateral ureteral obstruction. The findings are consistent with the suggestion that the low fractional excretion of Pi in the postobstructed kidney results from very low filtered loads of Pi on the postobstructed side.

Animals↗
PubMed1982

Increased Na+-H+ exchange in brush border vesicles from dogs with renal failure.

In the remnant kidney model of chronic renal failure, absolute reabsorption of Na+ in the proximal tubule of the remaining nephrons is increased over normal. Absolute proximal tubular reabsorption of bicarbonate and proximal tubular H+ excretion per nephron have also been shown to be increased over normal in his model of renal disease. Na+ uptake in membrane vesicles isolated from the brush border membrane of remnant kidneys of dogs with chronic renal failure is increased over uptake in membrane vesicles isolated from kidneys of normal dogs. In the present studies an amiloride-sensitive, electroneutral Na+-H+ exchanger was identified in canine renal brush border membrane vesicles. Na+ uptake in membrane vesicles in the presence of an initial H+ gradient (intravesicular pH less than extravesicular pH) was increased in membrane vesicles isolated from the remnant kidneys of dogs with chronic renal failure over that in membrane vesicles from kidneys of normal dogs. This increase was abolished by amiloride. It is possible that the alterations in Na+ and bicarbonate reabsorption and H+ excretion in the remnant kidney model of chronic renal failure can be explained on the basis of increased activity of the Na+-H+ exchanger in the renal brush border membrane.

Animals↗
PubMed1981

Parathyroid hormone inhibition of phosphate transport in renal brush border vesicles from phosphate-depleted dogs.

Dietary phosphate (Pi) restriction increases renal Pi reabsorption and induces resistance to the phosphaturic action of parathyroid hormone. Na+-gradient-stimulated Pi transport in membrane vesicles isolated from the renal brush border of experimental animals has been shown to parallel changes in renal Pi reabsorption induced by dietary Pi restriction and in vivo administration of parathyroid hormone. Dietary Pi restriction has been shown to markedly inhibit the phosphaturic response to parathyroid hormone in rats and dogs. Parathyroid hormone has been reported not to decrease the Na+-gradient-stimulated transport of Pi in brush border membrane vesicles isolated from dietary Pi restricted rats unless the rats were administered an acute Pi load prior to killing, however, thyroparathyroidectomy of rats fed a low Pi diet has been reported to increase Na+-gradient-stimulated Pi transport. Using the dietary Pi restricted dog, we demonstrated no significant decrease in renal reabsorption of Pi in response to parathyroid hormone administration. However, significant decreases in Pi transport in brush border membrane vesicles isolated from the kidneys of dietary Pi restricted dogs were observed in response to in vivo parathyroid hormone administration. These data demonstrate that the resistance to the phosphaturic action of parathyroid hormone observed in vivo does not include resistance to the inhibitory effect of parathyroid hormone on Pi transport in brush border membrane vesicles. Thus, the data suggest that parathyroid hormone continues to alter Pi transport characteristics of the brush border membrane in states of Pi depletion despite the resistance to parathyroid hormone seen in vivo.

Animals↗
PubMed1980

Regulation of canine renal vesicle Pi transport by growth hormone and parathyroid hormone.

Renal phosphate (Pi) reabsorption is increased by growth hormone (GH) and decreased by parathyroid hormone (PTH). Na+-stimulated Pi transport across the brush border membrane of the proximal tubule is the initial step in the process of Pi reabsorption. To determine whether changes in Pi reabsorption induced by GH or PTH are accompanied by changes in brush border membrane Na+-gradient-stimulated Pi transport, we examined the effect of in vivo GH and PTH administration and thyroparathyroidectomy on Pi transport by isolated brush border membrane vesicles prepared from canine kidney. In experiments in which the effect of PTH administration was examined, the same animal provided the control kidney (before PTH administration) and the experimental kidney (after PTH administration). The Na+-gradient Pi overshoot in vesicles isolated from normal, GH-treated and thyroparathyroidectomized dogs was increased after in vivo PTH administration. GH administration and thyroparathyroidectomy increased the height of the overshoot compared to normal. PTH administration decreased the apparent V value by 44% in vesicles from normal animals. The apparent V value was increased, compared to normal, by GH (34%) and thyroparathyroidectomy (57%). PTH administration decreased the apparent V in both the latter groups. GH administration to thyroparathyroidectomized dogs further increased the apparent V. Changes in the apparent V paralleled changes in Pi reabsorption in vivo induced by experimental manipulations. We conclude that changes in renal Pi reabsorption induced by GH were like those induced by PTH, accompanied by changes in the Na+-stimulated Pi transport system in the renal brush border membrane, and that the effect of PTH on vesicular Pi transport in GH-treated dogs did not differ from the effect on vesicles from normal animals.

Animals↗
PubMed1980

myo-Inositol transport in renal brush border vesicles and it inhibition by D-glucose.

We examined the mechanism of myo-inositol uptake by rabbit renal proximal tubule brush border membrane vesicles and characterized the relationship between the transports of myo-inositol and D-glucose. A 100 mM Na+ electrochemical gradient (extravesicular medium > intravesicular medium) stimulated the initial rate of myo-inositol uptake 20- to 60-fold. Other cation gradients were ineffective. The Na+ myo-inositol co-transport system was shown to be electrogenic. The Na+ electrochemical gradient-dependent uptake of myo-inositol saturated at about 1 mM myo-inositol, with an apparent Km of 94 micro M at an initial 100 mM Na+ gradient. D-Glucose was an inhibitor of the Na+ gradient-dependent uptake of myo-inositol. D-Glucose, but not L-glucose, elicited accelerative exchange diffusion of myo-inositol. myo-Inositol did not significantly inhibit the Na+ gradient-dependent transport of D-glucose. We suggest that D-glucose inhibits myo-inositol uptake by dissipating the membrane potential and sharing the myo-inositol carrier. The inhibition of myo-inositol transport across the brush border membrane by D-glucose explains how glycosuria could produce inosituria in patients with diabetes mellitus.

Animals↗
PubMed1977

Transport of amino acids in renal brush border membrane vesicles. Uptake of the neutral amino acid L-alanine.

The transport of L-alanine was studied using membrane vesicles derived from the brush borders of the rabbit renal proximal tubule. Preincubation of the renal membranes with L-alanine, but not D-alanine, accelerated exchange diffusion of L-alanine, i.e. stereospecific counter transport. The equilibrium uptake of L-alanine decreased with increasing medium osmolarity. Extrapolation to infinite medium osmolarity, i.e. zero intravesicular space, indicates no uptake. These findings demonstrate that the uptake of L-alanine represents transport into membrane vesicles and not surface binding to the membrane. The presence of a Na+ gradient between the external incubation medium and the intravesicular medium stimulated L-alanine uptake. Accumulation of the amino acid in the vesicles was maximal at 5 min and then decreased, indicating efflux. The final level of uptake in the presence of the Na+ gradient was identical with that obtained in the absence of the gradient, suggesting that equilibrium was established. At the peak of the "overshoot" the uptake of L-alanine was slmost twice the final equilibrium value. These results suggest that the imposition of a large extravesicular to intravesicular gradient of Na+ effects the transient movement of L-alanine into renal brush border membrane vesicles against its concentration gradient. Stimulation of L-alanine uptake was specific for Na+. When the intravesicular medium contained no Na+, the rate of uptake enhanced with increases in the concentration of Na+ in the external medium. Increasing the Na+ gradient lowered the apparent Km for L-alanine. In the absence of the Na+ gradient, the rates of uptake of L- and D-alanine were identical. In the presence of the Na+ gradient, the rate of D-alanine uptake was stimulated, but significantly less than that of L-alanine. The uptake of L-alanine, at a given concentration of amino acid reflected the sum of the contributions from Na+ gradient-dependent and -independent transport systems. The dependent system was saturated at about 2 mM L-alanine. The independent system exhibited minimal saturability and may itself represent the sum of passive diffusion and a "carrier"-mediated system. At physiological concentrations of L-alanine, the rate of the Na+ gradient-dependent uptake was 5-fold that in the absence of the gradient. Valinomycin enhanced the Na+ gradient-dependent uptake of L-alanine, provided a K+ gradient (vesicle greater than medium) was present. This finding indicates that the Na+ gradient-dependent transport of L-alanine into renal brush border membrane vesicles is an electrogenic process and suggests that the membrane potential is a determinant of L-alanine transport. In the presence of a Na+ gradient, the uptake of L-alanine was strongly inhibited by other neutral L-amino-acids. Imino acids and glycine also inhibited, but acidic and basic amino acids were without effect. In the absence of a Na+ gradient, little selective competition was found...

Alanine↗
PubMed1977

Transport of amino acids in renal brush border membrane vesicles. Uptake of L-proline.

The findings (a) stereospecific counter transport; (b) equilibrium uptake of L-proline decreased with increasing medium osmolarity; and (c) L-proline and Na+ were taken up into identical intravesicular spaces, indicate that uptake of L-proline by rabbit renal brush border membranes represents transport into membrane vesicles, not surface binding to the membrane. An Na+ gradient between extravesicular and intravesicular media stimulated the initial rate of L-proline uptake about 10 times. Accumulation of the imino acid was maximal at 2 min, then decreased until the equilibrium level was attained. At the peak of this "overshoot" uptake of L-proline was 3-fold greater than the final equilibrium value. These results suggest that the electrochemical Na+ gradient drives the transient movement of L-proline into the membrane vesicles against its concentration gradient. Stimulation of L-proline uptake was specific for Na+. Increasing the Na+ gradient lowered the apparent Km for L-proline. Efflux of L-proline from the membrane vesicles, like uptake, showed stimulatory effects when the Na+ gradient and L-proline were on the same side, and inhibitory effects when the Na+ gradient and the imino acid were on opposite sides of the membrane. Uptake of L-proline, at a given concentration, reflected the sum of contributions from Na+ gradient-dependent and -independent transport systems. The dependent system was saturated at 4 mM L-proline. The independent system did not saturate but may represent the sum of passive diffusion and a "carrier"-mediated system. At physiological concentrations the rate of the Na+ gradient-dependent uptake was 5 times that in the absence of the gradient. In K+-loaded vesicles, valinomycin, but not nigericin, enhanced the Na+ gradient-dependent uptake of L-proline. Gramicidin diminished uptake. These findings indicate that the Na+ gradient-dependent transport of L-proline is an electrogenic process and suggest that the membrane potential is a determinant of L-proline transport. The Na+ gradient-dependent rate of L-proline uptake was strongly inhibited by other imino acids, suggesting that L-imino acids have a common transport system. Glycine and neutral amino acids inhibited the rate of L-proline uptake moderately, acidic amino acid and D-glucose were very weak inhibitors, and basic amino acids were without effect. In the absence of Na+, the rate of L-proline uptake was independent of the presence of other amino acids. These findings indicate that the brush border membrane is a site of amino acid recognition during vectorial transepithelial transport. It is proposed that imino and neutral amino acids inhibit transport of L-proline by competitive interaction with the L-proline "carriers" in addition to competition for the electrochemical Na+ gradient or membrane potential. The findings are relevant to the understanding of genetic amino acid transport disorders, such as iminoglycinuria.

Amino Acids↗