Stimulation by thyroid hormone of phosphate transport in primary cultured renal cells

The regulation by thyroid hormone of phosphate transport in primary cultured chick renal cells was examined. The more physiologically active L-analogs of triiodothyronine and thyroxine, but not the D-analogs of the hormones, stimulated the Na+-dependent phosphate uptake system. Na+-independent phosphate uptake and Na+-dependent uptakes of alpha-methylglucoside and L-proline were unaffected. The increase in Na+-dependent phosphate uptake was concentration dependent, exhibited an induction period, and was blocked by inhibitors of RNA and protein synthesis. The stimulation of phosphate uptake by triiodothyronine was due to an increased Vmax rather than to an altered affinity for phosphate. These findings demonstrate that thyroid hormone acts directly on renal cells to modulate phosphate transport and suggest that the renal cell system may serve as a model to examine the mechanism by which thyroid hormone controls gene expression and regulates plasma membrane transport function.     

Inhibition by glucocorticoids of phosphate transport in primary cultured renal cells

The regulation by glucocorticoids of phosphate transport in primary cultured chick renal cells was examined. Dexamethasone inhibited the Na+-dependent phosphate uptake system. Na+-independent phosphate uptake and Na+-dependent uptakes of alpha-methylglucoside and L-proline were unaffected. The mineralocorticoid aldosterone did not alter phosphate uptake. The inhibition of Na+-dependent phosphate uptake by dexamethasone was concentration-dependent, exhibited an induction period, was blocked by inhibitors of RNA and protein synthesis, and was rapidly reversed when the steroid was removed. Following reversal, the cells could respond a second time to the glucocorticoid. However, this time the response was rapid, could be evoked at least for 24 h after glucocorticoid withdrawal, and might be prevented by actinomycin D and cycloheximide. These findings demonstrate that glucocorticoids act on renal cells to modulate phosphate transport and suggest that the renal cell system provides an attractive model to examine the mechanism by which glucocorticoids control gene expression and regulate plasma membrane transport function.     

Divalent metal is required for both phosphate transport and phosphate binding to phosphorin, a proteolipid isolated from brush-border membrane vesicles

The Na+-dependent phosphate transport system in the brush border of rabbit kidney exhibits a positive requirement for a divalent metal ion. Treatment of the brush-border membrane vesicles (BBMV) with a divalent metal chelator in combination with the divalent metal ionophore A23187 dramatically and selectively decreased the Na+-dependent uptake of phosphate; Na+-independent uptake of phosphate was not affected. The combination of chelator plus A23187 also inhibited uptake of phosphate in the presence of Na+ but in the absence of a gradient for sodium across the BBMV. This indicates that the inhibitor is not a result of an alteration in the Na+ gradient by chelator plus ionophore. The inhibited Na+ gradient-dependent transport of phosphate was restored by removing the chelator and adding Mn2+ to the BBMV. The phosphate-binding proteolipid (phosphorin) isolated from rabbit kidney BBMV binds inorganic phosphate with high affinity and specificity. Binding of phosphate to phosphorin is also inhibited by divalent metal chelators and can be restored by addition of a divalent metal. We conclude that a divalent metal ion is required both for the Na+-dependent phosphate transport in BBMV and for the binding of phosphate to the proteolipid phosphorin. These findings are consistent with our suggestion that phosphorin is a component of the Na+-dependent phosphate transport system in renal brush-border membranes.     

Phorbol esters inhibit phosphate uptake in opossum kidney cells: a model of proximal renal tubular cells

The effects of phorbol esters and diacylglycerol on phosphate uptake in opossum kidney (OK) cells were investigated to assess the possible role of Ca2+-activated, phospholipid dependent protein kinase (protein kinase C) on renal phosphate handling. OK cells are widely used as a model of proximal renal tubular cells and are reported to possess a Na+-dependent phosphate transport system. Phorbol-12,13-dibutyrate (PDBu) inhibited phosphate uptake. This inhibitory effect was synergistically enhanced with A23187. 4 beta-phorbol 12,13-didecanoate inhibited phosphate uptake, while 4 alpha-phorbol 12,13-didecanoate did not. 1-oleoyl-2-acetyl-glycerol (OAG), a synthetic diacylglycerol, also exhibited an inhibitory effect on phosphate uptake. These data suggest the possible involvement of protein kinase C in proximal renal tubular phosphate transport.     

Effect of pH on the kinetics of Na+-dependent phosphate transport in rat renal brush-border membranes

The kinetics of Na+-dependent phosphate uptake in rat renal brush-border membrane vesicles were studied under zero-trans conditions at 37 degrees C and the effect of pH on the kinetic parameters was determined. When the pH was lowered it turned out to be increasingly difficult to estimate initial rates of phosphate uptake due to an increase in aspecific binding of phosphate to the brush border membrane. When EDTA or beta-glycerophosphate was added to the uptake medium this aspecific binding was markedly reduced. At pH 6.8, initial rates of phosphate uptake were measured between 0.01 and 3.0 mM phosphate in the presence of 100 mM Na+. Kinetic analysis resulted in a non-linear Eadie-Hofstee plot, compatible with two modes of transport: one major low-affinity system (Km approximately equal to 1.3 mM), high-capacity system (Vmax approximately equal to 1.1 nmol/s per mg protein) and one minor high-affinity (Km approximately equal to 0.03 mM), low-capacity system (Vmax approximately equal to 0.04 nmol/s per mg protein). Na+-dependent phosphate uptake studied far from initial rate conditions i.e. at 15 s, frequently observed in the literature, led to a dramatic decrease in the Vmax of the low-affinity system. When both the extra- and intravesicular pH were increased from 6.2 to 8.5, the Km value of the low-affinity system increased, but when divalent phosphate is considered to be the sole substrate for the low-affinity system then the Km value is no longer pH dependent. In contrast, the Km value of the high-affinity system was not influenced by pH but the Vmax decreased dramatically when the pH is lowered from 8.5 to 6.2. These results suggest that the low-affinity, high-capacity system transports divalent divalent phosphate only while the high-affinity, low-capacity system may transport univalent as well as divalent phosphate. Raising medium sodium concentration from 100 to 250 mM increased Na+-dependent phosphate uptake significantly but the pH dependence of the phosphate transport was not influenced. This observation makes it rather unlikely that pH changes only affect the Na+ site of the Na+-dependent phosphate transport system.     

Alpha-2-adrenergic modulation of the parathyroid hormone-inhibition of phosphate uptake in cultured renal (OK) cells

Parathyroid hormone enhances the formation of cAMP and decreases the Na+-dependent uptake of phosphate in cultured renal cells derived from the American opossum (OK cells). Epinephrine, acting as an alpha 2-adrenergic agonist, inhibits the PTH-induced synthesis of cAMP by a pertussis toxin-sensitive mechanism and blunts the inhibition of phosphate transport by PTH. Na+-dependent alpha-methylglucoside and Na+ uptakes by the cells are unaffected by PTH and epinephrine. These findings suggest that alpha 2-adrenergic agonists may selectively modulate PTH-sensitive phosphate transport in the renal proximal tubule.     

pH gradient as an additional driving force in the renal re-absorption of phosphate.

The effects of the Na+ gradient and pH on phosphate uptake were studied in brush-border membrane vesicles isolated from rat kidney cortex. The initial rates of Na(+)-dependent phosphate uptake were measured at pH 6.5, 7.5 and 8.5 in the presence of sodium gluconate. At a constant total phosphate concentration, the transport values at pH 7.5 and 8.5 were similar, but at pH 6.5 the influx was 31% of that at pH 7.5. However, when the concentration of bivalent phosphate was kept constant at all three pH values, the effect of pH was less pronounced; at pH 6.5, phosphate influx was 73% of that measured at pH 7.5. The Na(+)-dependent phosphate uptake was also influenced by a transmembrane pH difference; an outwardly directed H+ gradient stimulated the uptake by 48%, whereas an inwardly directed H+ gradient inhibited the uptake by 15%. Phosphate on the trans (intravesicular) side stimulated the Na(+)-gradient-dependent phosphate transport by 59%, 93% and 49%, and the Na(+)-gradient-independent phosphate transport by 240%, 280% and 244%, at pH 6.5, 7.5 and 8.5 respectively. However, in both cases, at pH 6.5 the maximal stimulation was seen only when the concentration of bivalent trans phosphate was the same as at pH 7.5. In the absence of a Na+ gradient, but in the presence of Na+, an outwardly directed H+ gradient provided the driving force for the transient hyperaccumulation of phosphate. The rate of uptake was dependent on the magnitude of the H+ gradient. These results indicate that: (1) the bivalent form of phosphate is the form of phosphate recognized by the carrier on both sides of the membrane; (2) protons are both activators and allosteric modulators of the phosphate carrier; (3) the combined action of both the Na+ (out/in) and H+ (in/out) gradients on the phosphate carrier contribute to regulate efficiently the re-absorption of phosphate.     

Phosphate transport by rat intestinal basolateral-membrane vesicles.

The characteristics of phosphate transport across intestinal basolateral membranes of the rat were determined by using enriched preparations in which uphill Na+-dependent D-glucose transport could not be demonstrated, but ATP-dependent Ca2+ transport was present. Phosphate transport was saturable, Na+-dependent and exhibited Michaelis-Menten kinetics. Vmax. was 51.1 +/- 4.2 pmol/10 s per mg of protein and Km was 14 +/- 3.9 microM. The transport process was electroneutral. Tracer-exchange experiments and counter-transport studies confirmed the presence of a Na+-Pi carrier at the basolateral membrane. The presence of inside-positive membrane potential did not enhance phosphate uptake, indicating that the Na+ effect is secondary to the presence of the Na+-Pi carrier rather than an induction of positive membrane potential. The stoichiometry of this carrier at pH 7.4 was 2 Na+:1 phosphate, as shown by direct studies utilizing the static-head method. These studies are the first to determine the presence of a phosphate carrier at the basolateral membrane.     

Glycine transport by cultured human fibroblasts

The transport of glycine was studied in cultured human fibroblasts. The amino acid entered the cell by Na+-dependent and Na+-independent mechanisms. Na+-independent glycine (0.1 mM) transport was less than 10% of total uptake and occurred by a mechanism formally indistinguishable from diffusion. Two distinct routes contributed to Na+-dependent glycine transport. The first route was identified with system A because it was inhibited by MeAIB and underwent adaptive regulation. The second route was identified with system ASC as it was inhibited by L-alanine, but not by MeAIB. Kinetic analysis revealed that the two systems operated glycine transport with the same Km of 1.6 mM, a value unusually high for system ASC.     

Interaction of cations with phosphate uptake by Saccharomyces cerevisiae. Effects of surface potential.

The effect of bivalent cations on phosphate uptake by Saccharomyces cerevisiae was investigated. Phosphate uptake via the Na+-dependent transport system at pH 7.2 is stimulated by bivalent cations. The apparent affinity of phosphate for the transport mechanism is increased, but the apparent affinity for Na+ is decreased. Uptake of phosphate via the Na+-independent transport system is accompanied by a net proton influx of 2H+ and an efflux of 1 K+ for each phosphate ion taken up. At pH 4.5 phosphate uptake via the Na+-independent system is stimulated by bivalent cations, whereas at pH 7.2 uptake is inhibited. The effect of bivalent cations on phosphate uptake can be ascribed to a decrease in the surface potential.     

Phorbol myristate acetate activates protein kinase C, stimulates the phosphorylation of endogenous proteins and inhibits phosphate transport in mouse renal tubules

Calcium-activated, phospholipid-dependent protein kinase (protein kinase C) has been implicated in the regulation of transport processes in a variety of tissues and cell lines. To establish whether protein kinase C participates in the regulation of renal phosphate transport, we examined the effect of phorbol myristate acetate (PMA), a potent activator of protein kinase C, on phosphate uptake in fresh preparations of mouse renal tubules, and we correlated the changes in transport activity with protein kinase C activation and phosphorylation of endogenous proteins. PMA inhibited Na+-dependent phosphate transport, elicited a rapid translocation of protein kinase C from the cytosolic to the particulate fraction and stimulated the phosphorylation of endogenous substrates in the cytosolic and brush border membrane fractions. Effects of PMA were maximal after a 10 min incubation of the tubules with the activator. 4 alpha-Phorbol, an inert analogue of PMA, did not elicit any of these effects. The present results demonstrate a temporal correlation between inhibition of Na+-dependent phosphate transport, translocation and activation of protein kinase C, and phosphorylation of endogenous proteins in mouse renal tubules. These data suggest that protein kinase C may play a regulatory role in phosphate transport in mammalian kidney.     

Characterization of a Pi transport system in cartilage matrix vesicles. Potential role in the calcification process.

The mechanisms by which calcium (Ca2+) and inorganic phosphate (Pi) accumulate into matrix vesicles (MV) have not been elucidated. In the present study the characteristics of Pi uptake into MV isolated from mildly rachitic chicken growth plate cartilage have been investigated. The results indicate that Pi accumulates into MV mainly via a Na(+)-dependent Pi transport system. In the absence of NaCl in the extravesicular medium, Pi uptake was a nonsaturable process. In the presence of 150 mM NaCl, the initial rate of Pi uptake was 4.38 +/- 1.02-fold higher than with 150 mM choline chloride (mean +/- S.E., n = 8, p less than 0.005). Other cations showed partial activity to drive Pi into MV as compared to Na+:Li+ (64.4%) greater than K+ (39.8%) greater than choline (39.0%) greater than tetramethylammonium (30.0%) greater than N-methylglucamine (26.3%). Na(+)-dependent Pi transport activity displayed saturability towards increasing extra-vesicular concentrations of Na+ and Pi. The apparent Km for Pi was 0.68 +/- 0.16 mM. The Na+ concentration producing half-maximum Pi transport activity was 106.2 +/- 11.0 mM. Kinetic analysis suggests that Na+ interacts with the Pi carrier with a stoichiometry of more than one Na+ ion with one Pi molecule. In MV isolated from normal chicken growth plate cartilage, this Na(+)-dependent Pi transport system was barely expressed. In contrast to the effect on Pi uptake by MV, the activity of alkaline phosphatase was not changed when NaCl was substituted for choline chloride in the assay medium. In addition to this observation which suggests that this enzyme is not related to the Pi transport activity described in this study, levamisole, which inhibited alkaline phosphatase activity did not affect the Na(+)-dependent uptake of Pi. Both arsenate and phosphonoformic acid, two inhibitors of the epithelial Na(+)-dependent Pi transport systems, were active inhibitors of the Na(+)-dependent Pi uptake by MV with a higher potency for phosphonoformic acid. Associated with the expression of a facilitated Na(+)-coupled Pi transport in MV, in vitro calcification assessed by 45Ca2+ uptake also showed a marked dependence on extravesicular sodium. This relationship was markedly attenuated in MV isolated from normal chicken growth plate cartilage expressing a weak Na(+)-facilitated Pi transport activity. In conclusion, a saturable Na(+)-dependent Pi carrier has been characterized which facilitates Pi transport in MV. Its potential role for Ca-Pi accumulation into MV and subsequent development of vesicular calcification followed by mineralization of the osteogenic matrix is proposed and remains to be further investigated.     

Expression of renal transport systems for inorganic phosphate and sulfate in Xenopus laevis oocytes.

As a first step within an experimental strategy (expression cloning) leading to the structural identification of the two brush-border membrane transport systems for phosphate and sulfate, we have studied the expression of Na(+)-dependent uptake of phosphate and sulfate in Xenopus laevis oocytes injected with rabbit kidney cortex poly(A)+ RNA (mRNA). Na(+)-dependent uptake of phosphate and sulfate was stimulated in a dose- and time-dependent manner up to 20-fold as compared to water-injected controls. After fractionation of the mRNA on a sucrose gradient (or by preparative gel electrophoresis), two neighboring fractions were identified to stimulate Na(+)-dependent phosphate uptake (average size: 3.4 kilobases) and Na(+)-dependent sulfate uptake (average size: 3.7 kilobases). The two transport systems can be discriminated by their inhibition by thiosulfate, which reduced sulfate uptake, but not phosphate uptake. Kinetic characterization of the expressed Na(+)-dependent transport activities results in properties similar to those described for transport activity in renal brush-border membrane vesicles.     

Identification of proximal tubular transport functions in the established kidney cell line, OK

OK cells, derived from an American opossum kidney, were analyzed for proximal tubular transport functions. In monolayers, L-glutamate, L-proline, L-alanine, and alpha-methyl-glucopyranoside (alpha-methyl D-glucoside) were accumulated through Na+-dependent and Na+-independent transport pathways. D-Glucose and inorganic sulfate were accumulated equally well in the presence or absence of Na+. Influx of inorganic phosphate was only observed in the presence of Na+. Na+/alpha-methyl D-glucoside uptake was preferentially inhibited by phlorizin and D-glucose uptake by cytochalasin B. An amiloride-sensitive Na+-transport was also identified. In isolated apical vesicles (enriched 8-fold in gamma-glutamyltransferase), L-glutamate, L-proline, L-alanine, alpha-methyl D-glucoside and inorganic phosphate transport were stimulated by an inwardly directed Na+-gradient as compared to an inwardly directed K+-gradient. L-Glutamate transport required additionally intravesicular K+. D-Glucose transport was similar in the presence of a Na+- and a K+-gradient. Na+/alpha-methyl D-glucoside uptake was inhibited by phlorizin whereas cytochalasin B had no effect on Na+/D-glucose transport. An amiloride-sensitive Na+/H+ exchange mechanism was also found in the apical vesicle preparation. It is concluded that the apical membrane of OK cells contains Na+-coupled transport systems for amino acids, hexoses, protons and inorganic phosphate. D-Glucose appears a poor substrate for the Na+/hexose transport system.     

Increases in transepithelial vectorial Na+ transport facilitates Na+-dependent L-DOPA transport in renal OK cells

The present study evaluated the hypothesis of whether increases in vectorial Na+ transport translate into facilitation of Na+-dependent L-DOPA uptake in cultured renal epithelial tubular cells. Increases in vectorial Na+ transport were obtained in opossum kidney (OK) cells engineered to overexpress Na+-K+-ATPase after transfection of wild type OK cells with the rodent Na+-K+-ATPase alpha1 subunit. The most impressive differences between wild type and transfected OK cells are that the latter overexpressed Na+-K+-ATPase accompanied by an increased activity of the transporter. Non-linear analysis of the saturation curve for l-DOPA uptake revealed a Vmax value (in nmol mg protein/6 min) of 62 and 80 in wild type and transfected cells, respectively. The uptake of a non-saturating concentration (0.25 microM) of [14C]-L-DOPA in OK-WT cells was not affected by Na+ removal, whereas in OK-alpha1 cells accumulation of [14C]-L-DOPA was clearly dependent on the presence of extracellular Na+. When Na+ was replaced by choline, the inhibitory profile of neutral l-amino acids, but not of basic and acidic amino acids, upon [14C]-L-DOPA uptake in both cell types, was significantly greater than that observed in the presence of extracellular Na+. It is concluded that enhanced ability of OK cells overexpressing Na+-K+-ATPase to translocate Na+ from the apical to the basal cell side correlates positively with their ability to accumulate L-DOPA, which is in agreement with the role of Na+ in taking up the precursor of renal dopamine.     

Characterization of the myo-inositol transport system in Trypanosoma cruzi.

myo-inositol is a growth factor for mammalian cells as well as for the pathogenic protozoa Trypanosoma cruzi. Most of the cell surface molecules in this organism rely on myo-inositol as the biosynthetic precursor for phosphoinositides and glycosylated phosphatidylinositols. The aim of this work was to investigate the process of myo-inositol translocation across the parasite cell membrane. myo-Inositol uptake was concentration-dependent in the concentration range 0.1-10 microM with maximal transport obtained at 8 microM. Using sodium-free buffers, where Na+ was replaced by choline or K+, myo-inositol uptake was inhibited by 50%. Furosemide, an inhibitor of the ouabain-insensitive Na+-ATPase, inhibited the Na+-dependent and Na+-independent myo-inositol uptake by 68 and 33%, respectively. In contrast, ouabain, an (Na++/K+) ATPase inhibitor, did not affect transport. Part of the myo-inositol uptake is mediated by active transport as it was inhibited when energy metabolism inhibitors such as carbonyl cyanide p-(trifluoromethoxy)-phenylhydrazone (34%), 2,4-dinitrophenol (50%), KCN (71%) and NaN3 (69%) were added to the medium, or the temperature of the medium was lowered to 4 degrees C. The addition of glucose (5-50 mM) or mannose (10 mM) did not change the myo-inositol uptake, whereas the addition of 10 mM nonlabeled myo-inositol totally inhibited this transport, indicating that the transporter is specific for myo-inositol. Phloretin (0.3 mM) and phoridzin (5 mM), but not cytochalasin B, were efficient inhibitors of myo-inositol uptake. A portion of the accumulated myo-inositol is converted to inositol phosphates and phosphoinositides. These data show that myo-inositol transport in T. cruzi epimastigotes is mediated by at least two specific transporters - one Na+-dependent and the other Na+-independent.     

Characteristics of D-alanine transport by luminal membrane vesicles from pars convoluta and pars recta of rabbit proximal tubule

Uptake of D-alanine against a concentration gradient has been shown to occur with isolated luminal-membrane vesicles from pars convoluta or pars recta of rabbit proximal tubule. Renal D-alanine transport systems, displaying the following characteristics, were shown: (1) In vesicles from pars convoluta, the uptake of D-alanine was mediated by both Na+-dependent and Na+-independent transport processes. It was found that an inwardly directed H+-gradient could drive the transport of D-alanine into the vesicles both in the presence and absence of Na+. Thus, in addition to Na+, the transport of D-alanine is influenced by the H+-gradient. (2) In vesicles from pars recta, the transient accumulation of D-alanine was strictly dependent on Na+, since no 'overshoot' was ever observed in the absence of Na+. Although the Na+-dependent uptake of D-alanine was stimulated at acid pH, H+ did not substitute for Na+, as it apparently does in pars convoluta, but instead potentiated the Na+ effect. (3) Addition of L-alanine to vesicle preparations, both from pars convoluta and from pars recta, specifically inhibited renal uptake of D-alanine. A comparison between the transport characteristics of D- and L-alanine indicated that these two isomers of alanine probably share common transport systems located along the proximal tubule of rabbit kidney.     

Studies on the thermotropic behavior of aqueous phosphatidylethanolamines

Transport of phosphate has been studied in subconfluent monolayers of LLC-PK1 cells. It was found that this transport system shows similar characteristics to those observed in the kidney. Uptake of phosphate is mediated by a Na+-dependent, substrate-saturable process with an apparent Km value for phosphate of 96 +/- 15 mumol/l. Kinetic analysis of the effect of Na+ indicated that at (pH 7.4) two sodium ions are cotransported with one HOP4(2-) ion (Hill coefficient 1.5) with an apparent Km value for sodium of 56 mmol/l. Pi uptake is inhibited by metabolic inhibitors (ouabain and FCCP). In the pH range of 6.6 of 7.4 Pi uptake rate does not change significantly, indicating that both the monovalent and the divalent form of phosphate are accepted by the transport system. It is suggested that phosphate is transported by LLC-PK1 cells together with sodium (2 Na+:1 HPO4(2-) in an electroneutral manner down a favourable sodium gradient.     

A phosphorylated phloretin derivative. Synthesis and effect on intestinal Na(+)-dependent phosphate absorption

2'-Phosphophloretin (2'-PP), a phosphorylated derivative of the plant chalcone, was synthesized. The effect of 2'-PP, on Na(+)-dependent phosphate uptake into intestinal brush-border membrane vesicles (BBMV) isolated from rabbit and rat duodenum and jejunum was examined. 2'-PP decreased Na(+)-dependent phosphate uptake into rabbit BBMV with an IC(50) of 55 nM and into rat BBMV with an IC(50) of 58 nM. 2'-PP did not affect Na(+)-dependent glucose, Na(+)-dependent sulfate, or Na(+)-dependent alanine uptake by rabbit intestinal BBMVs. 2'-PP inhibition of rabbit intestinal BBMV Na(+)-dependent phosphate uptake was sensitive to external phosphate concentration, suggesting that 2'-PP inhibition of Na(+)-dependent phosphate uptake was competitive with respect to phosphate. Binding of [(3)H]2'-PP to rabbit intestinal BBMV was examined. Binding of [(3)H]2'-PP was Na(+)-dependent with a K(0.5) for Na(+)(Na(+) concentration for 50% 2'-PP binding) of 30 mM. The apparent K(s) for Na(+)-dependent [(3)H]2'-PP binding to rabbit BBMVs was 58 nM in agreement with the IC(50) for 2'-PP inhibition of Na(+)-dependent phosphate uptake. These results indicate that 2'-PP bound to rabbit or rat intestinal BBMV Na(+)-phosphate cotransporter and inhibited Na(+)-dependent phosphate uptake. In rats treated with 2'-PP by daily gavage, the effect of 2'-PP on serum phosphate, serum glucose, and serum calcium was examined. In a concentration-dependent manner, 2'-PP reduced serum phosphate by 45% 1 wk after starting treatment. 2'-PP did not alter serum calcium or serum glucose. The apparent IC(50) for 2'-PP in vivo was 3 microM.     

Effect of parathyrin on the transport properties of isolated renal brush-border vesicles.

The transport properties of brush-border membrane vesicles isolated by a calcium-precipitation method from the renal cortex of normal and parathyrin (parathyroid hormone)-treated rats were studied by a rapid-filtration technique. Parathyrin elicited a dose-dependent decrease in the Na+-dependent phosphate uptake by the brush-border membrane vesicles, but the uptake of D-glucose, Na+ and mannitol was not affected. A maximum inhibition of 30% was observed after the application of 30 U.S.P. units intramuscularly 1 h before the animals were killed. Intravenous infusion of dibutyryl cyclic AMP (0.5-1.5 MG) also decreased the phosphate uptake by the brush-border vesicles. Both dibutyryl cyclic AMP and parathyrin were ineffective when added in vitro to brush-border membrane vesicles isolated from normal rats. These data suggest that parathyrin exerts its action on the phosphate reabsorption in the renal proximal tubule by affecting the Na+/phosphate co-transport system in the brush-border membrane. The effects of parathyrin on Na+ and glucose transport, however, seem to be due to alterations to the driving forces for transport and not to the brush-border transport systems.