Fractional Excretion Of Pi Research Articles

MO566THE BONE EXPRESSION OF WNT INHIBITORS IN THE EXPERIMENTAL MODEL OF EARLY CHRONIC KIDNEY DISEASE

Abstract Background and Aims Molecular mechanisms implicated in the initial stages of inorganic phosphate (Pi) imbalance in chronic kidney disease (CKD) remain poorly understood.The aim of the study was to evaluate whether canonical Wnt pathway inhibitors (iWnt) involved in early response to Pi retention in CKD. Methods Mild CKD was induced by 3/4 nephrectomy (NE) in spontaneously hypertensive rats (SHR) fed rat chow diet containing 0.6 % phosphate. Controls were sham operated SHR (SO). Duration of experimental exposure (NE or SO) was 2 and 6 months. Serum levels of creatinine (Cr), inorganic phosphate (Pi), fractional Pi excretion (FEPi), intact parathyroid hormone (PTH), fibroblast growth factor 23 (FGF23), alfa-Klotho (KL), sclerostin (SOST) and Dickkopf-1 (DKK1) were measured. The following morphological characteristics by light microscopy of bone metaphysis and kidney tissues: the area of renal interstitial fibrosis (RF) (Masson's trichrome), bone matrix volume (MV), the active osteoblasts to trabecular cells number ratio (aOB/cells), eroded surface to bone surface ratio (ES/BS) (hematoxylin & eosin), and bone SOST and DKK1 proteins expression (by IHC) were analyzed and calculated quantitatively. Statistical comparisons among groups were performed using Mann–Whitney U-test and Kruskal-Wallis H-test. Results Serum Cr, RF and indices of Pi exchange in the experimental model corresponded to early CKD (Table). Pi elevated in NE6 suggestive for its renal retention. KL level decreased (Table) in all experimental groups vs control. No differences were observed in serum levels FGF23 (p=0.62) and PTH (p=0.63). Serum SOST and DKK1 levels were significantly higher in NE6 group compared to SO6 (Table). The bone SOST and DKK1 expression increased in NE6 compared to SO6 (Figure). aOB/cells were lower in NE2, SO6 and NE6 vs SO2 (all p-values<=0.041). ES/BS increased in NE2 (vs SO2) while being lowest in NE6 and SO6 animals (Table). SOST and DKK1 metaphyseal expression increased in NE6 compared to SO2, SO6, NE2 (Figure). Osteocyte SOST expression increased in SO6 compared to SO2 and NE2 without differences in later groups. Osteoblast SOST expression was also higher in SO6 vs SO2 (Figure). Conclusion Increased serum levels of sclerostin and Dickkopf-1 and their bone expression are apparent in early stages of experimental CKD associating with hyperphosphatemia. Alterations of bone resorption and osteoblast depopulation occurred before the increase of serum Pi likely reflecting incipient stages of renal Pi retention. These molecular and cellular events seem to be independent of systemic FGF23 and PTH response.

The kinetics of inorganic phosphate excretion in the acidotic rabbit during intravenous phosphate loading: a pseudo-ruminant model

The rabbit is a much-used experimental animal in renal tubule physiology studies. Although a monogastric mammal, the rabbit is a known hindgut fermenter. That ruminant species excrete inorganic phosphate (Pi) mainly through the digestive system while non-ruminants eliminate surplus phosphate primarily through the renal system are acknowledged facts. To understand phosphate homeostasis in the acidotic rabbit, anaesthetized animals were infused with hydrochloric acid, after which they underwent intravenous phosphate loading. Biofluids were collected during the infusion process for analysis. Plasma Pi increased (7.9 ± 1.7 mmoles.Litre−1 (N = 5) vs 2.2 ± 0.4 mmoles.Litre−1 (N = 10) pre-infusion, (p < 0.001)), while urinary phosphate excretion was also enhanced (74.4 ± 15.3 from a control value of 4.7 ± 3 µmol.min−1 (N = 9), pre-infusion, p < 0.001)) over an 82.5 minute Pi loading period. However, the fractional excretion of Pi (FePi) only increased from 14.2 ± 5.4% to a maximum of 61.7 ± 19% (N = 5) over the infusion period. Furthermore, the renal tubular maximum reabsorption rate of phosphate to glomerular filtration rate (TmPi/GFR) computed to 3.5 mmol.L−1, while a reading of 23.2 µmol.min−1.Kg.0.75 was obtained for the transport maximum for Pi (TmPi). The high reabsorptivity of the rabbit nephrons coupled with possibly a high secretory capacity of the salivary glands for Pi, may constitute a unique physiological mechanism that ensures the rabbit hindgut receives adequate phosphate to regulate caecal pH in favour of the resident metabolically - active microbiota. The handling of Pi by the rabbit is in keeping with the description of this animal as a monogastric, pseudo-ruminant herbivore.

Bone and Mineral Metabolism and Fibroblast Growth Factor 23 Levels After Kidney Donation.

Introduction: Prospective studies to better understand the effects of a mild reduction in kidney function on bone metabolism in otherwise healthy individuals is warranted. Materials and Methods: We measured renal function and mineral metabolites longitudinally for 1 year (days (D) 1, 2, 14, 180, & 365 post-operatively) in 74 healthy individuals who underwent kidney live donation. Changes were assessed by repeated measures ANOVA. Results: eGFR (MDRD)decreased to 59% of it's baseline on D2 (109±21 to 65±15, p<0.01) and started to increase at D3 (71±14, p<0.01), to its maximum at D180 (74±16, p<0.01), wile FGF23 increased from 60.61 (25th-75th percentile 19-81 RU/mL) at baseline to 110.99±144.56 (p<0.01) on D1 and remained higher through the study (D365: 65.87-25th-75th percentile 28-89 RU/mL). PTH rose maximally on D1 (64.9 ± 30.3RU/mL) and remained higher than baseline throughout the study (D365:52.4 ± 22.1RU/mL). Total serum Calcium levels decreased from 9,2 mg/dL to a nadir of 7.9 mg/dL on D1 (p<0.01). Serum Phosphate levels reached their nadir on D2 (2.7 mg/dL; p<0.01). At D14 total calcium and phosphate levels had returned to baseline, but phosphate levels returned down on POD4 (3.47 mg/dL;p<0.05) and D5 (3,33 mg/dL; p<0.01). Urinary fractional excretion of Pi increased from base line (11.4±5.1%) up to 16.1±10.7% until D360. Conclusions: Mild to moderate reduction in eGFR induces physiological increases in FGF23 and PTH, and decreases in serum Ca and Pi. The changes in Pi, PTH and FGF23 remains modestly yet significantly different from baseline throughout the first year after nephrectomy. The clinical consequences of elevated FGF23 following kidney donation require further studies

Activation of a Non-cAMP/PKA Signaling Pathway Downstream of the PTH/PTHrP Receptor Is Essential for a Sustained Hypophosphatemic Response to PTH Infusion in Male Mice

PTH increases urinary Pi excretion by reducing expression of two renal cotransporters [NaPi-IIa (Npt2a) and NaPi-IIc (Npt2c)]. In contrast to acute transporter regulation that is cAMP/protein kinase A dependent, long-term effects require phospholipase C (PLC) signaling by the PTH/PTHrP receptor (PPR). To determine whether the latter pathway regulates Pi through Npt2a and/or Npt2c, wild-type mice (Wt) and animals expressing a mutant PPR incapable of PLC activation (DD) were tested in the absence of one (Npt2a(-/-) or Npt2c(-/-)) or both phosphate transporters (2a/2c-dko). PTH infusion for 8 days caused a rapid and persistent decrease in serum Pi in Wt mice, whereas serum Pi in DD mice fell only transiently for the first 2 days. Consistent with these findings, fractional Pi excretion index was increased initially in both animals, but this increase persisted only when the PPR Wt was present. The hypophosphatemic response to PTH infusion was impaired only slightly in PPR Wt/Npt2c(-/-) or DD/Npt2c(-/-) mice. Despite lower baselines, PTH infusion in PPR Wt/Npt2a(-/-) mice decreased serum Pi further, an effect that was attenuated in DD/Npt2a(-/-) mice. Continuous PTH had no effect on serum Pi in 2a/2c-dko mice. PTH administration increased serum 1,25 dihydroxyvitamin D3 levels in Wt and DD mice and increased levels above the elevated baseline with ablation of either but not of both transporters. Continuous PTH elevated serum fibroblast growth factor 23 and blood Ca(2+) equivalently in all groups of mice. Our data indicate that PLC signaling at the PPR contributes to the long-term effect of PTH on Pi homeostasis but not to the regulation of 1,25 dihydroxyvitamin D3, fibroblast growth factor 23, or blood Ca(2+).

Regulation of Intestinal Phosphate Transport II. Metabolic acidosis stimulates Na+-dependent phosphate absorption and expression of the Na+-Pi cotransporter NaPi-IIb in small intestine

During metabolic acidosis, P(i) serves as an important buffer to remove protons from the body. P(i) is released from bone together with carbonate buffering protons in blood. In addition, in the kidney, the fractional excretion of phosphate is increased allowing for the excretion of more acid equivalents in urine. The role of intestinal P(i) absorption in providing P(i) to buffer protons and compensating for loss from bone during metabolic acidosis has not been clarified yet. Inducing metabolic acidosis (NH(4)Cl in drinking water) for 2 or 7 days in mice increased urinary fractional P(i) excretion twofold, whereas serum P(i) levels were not altered. Na(+)-dependent P(i) transport in the small intestine, however, was stimulated from 1.89 +/- 3.22 to 40.72 +/- 11.98 pmol/mg protein (2 days of NH(4)Cl) in brush-border membrane vesicles prepared from total small intestine. Similarly, the protein abundance of the Na(+)-dependent phosphate cotransporter NaPi-IIb in the brush-border membrane was increased 5.3-fold, whereas mRNA levels remained stable. According to immunohistochemistry and real-time PCR NaPi-IIb expression was found to be mainly confined to the ileum in the small intestine, and this distribution was not altered during metabolic acidosis. These results suggest that the stimulation of intestinal P(i) absorption during metabolic acidosis may contribute to the buffering of acid equivalents by providing phosphate and may also help to prevent excessive liberation of phosphate from bone.

Alterations in Serum Parathyroid Hormone and Electrolyte Concentrations and Urinary Excretion of Electrolytes in Horses with Induced Endotoxemia

Hypocalcemia and hypomagnesemia are common in horses with sepsis and endotoxemia. We hypothesize that endotoxemia triggers a systemic inflammatory response that results in hypocalcemia and hypomagnesemia. The goal of this study was to determine the effect of endotoxin (lipopolysaccharide [LPS]) administration to healthy horses on serum parathyroid hormone (PTH), ionized calcium (Ca2+) and total calcium (tCa), ionized magnesium (Mg2+) and total magnesium (tMg), phosphate (Pi), potassium (K+), sodium (Na+), chloride (Cl-), and insulin concentrations, and on the urinary excretion of these electrolytes. Twelve mares were infused with Escherichia coli LPS (30 ng/kg/h i.v.) for 1 hour. Six mares were infused with saline (controls). In LPS-infused horses, heart rate increased significantly from (mean +/- SD) 40.0 +/- 1.3 to 70.0 +/- 9.0 beats/min, respiratory rate from 12.7 +/- 1.0 to 21.1 +/- 3.0 breaths/min, body temperature from 37.4 +/- 0.3 to 38.9 +/- 0.6 degrees C, and tumor necrosis factor-alpha concentrations from 6.6 +/- 3.5 to 507 +/- 260 pg/mL (P < .05). White blood cell count decreased significantly from 7570 +/- 600 to 1960 +/- 560 cells/ microL. Serum concentrations of Ca2+ decreased from 6.5 +/- 0.3 to 6.0 +/- 0.3 mg/dL, of Mg2+ from 0.53 +/- 0.06 to 0.43 +/- 0.04 mM, of tMg from 0.78 +/- 0.05 to 0.62 +/- 0.08 mM, of K+ from 4.3 +/- 0.4 to 3.0 +/- 0.5 mEq/L, and of Pi from 3.4 +/- 0.5 to 1.7 +/- 0.5 mg/dL (all P < .05). PTH increased significantly from 1.3 +/- 0.4 to 6.0 +/- 5.2 pM; however, in some horses (n=2), PTH did not increase despite hypocalcemia. Insulin increased significantly from 9.4 +/- 3.6 to 50.5 +/- 9.6 microIU/mL (n=3). Urinary fractional excretion of Ca2+ decreased significantly from 4.7 +/- 1.4 to 1.7 +/- 1.2%, of Mg2+ from 36.6 +/- 6.5 to 11.7 +/- 7.3%, and of K+ from 37.9 +/- 11.3 to 17.7 +/- 6.2%. Fractional excretion of Pi increased from 0.02 +/- 0.02 to 0.14 +/- 0.07% and of Na+ from 0.26 +/- 0.13% to 1.2 +/- 0.5%. No changes were found in serum tCa, Na+, and Cl- concentrations. In conclusion, endotoxemia in horses resulted in electrolyte abnormalities that included hypocalcemia, hypomagnesemia, hypokalemia, hypophosphatemia, and increased serum PTH and insulin concentrations.

Thyroid hormones stimulate Na+?Pi transport activity in rat renal brush-border membranes: Role of membrane lipid composition and fluidity

Antecedent studies have suggested that lipid composition and fluidity of cellular membranes of various organs are altered in response to thyroid hormone status. To date, the effects of thyroid hormone status on these parameters have not been examined in rat renal apical membrane in regard to sodium-dependent phosphate transport. In the present study, we determined the potential role of alterations in cortical brush-border membrane lipid composition and fluidity in modulation of Na+-Pi transport activity in response to thyroid hormone status. Thyroid hormone status influences the fractional excretion of Pi, which is associated with alteration in renal brush-border membrane phosphate transport. The increment in Na+-Pi transport in renal BBMV isolated from Hyper-T rats is manifested as an increase in the maximal velocity (Vmax) of Na+-Pi transport. Further, the cholesterol content was significantly increased in renal BBM of Hypo-T rats and decreased in Hyper-T rats as compared to the Eu-T rats. The molar ratio of cholesterol/phospholipids was also higher in renal BBM from hypo-T rats. Subsequently, fluorescence anisotropy of diphenyl hexatriene (rDPH) and microviscosity were significantly decreased in the renal BBM of the Hyper-T rats and increased in the Hypo-T rats as compared to Eu-T rats. The result of this study, therefore, suggest that alteration in renal BBM cholesterol, cholesterol/phospholipid molar ratio, and membrane fluidity play an important role in the modulation of renal BBM Na+-Pi transport in response to thyroid hormone status of animals.

Expression and regulation of the renal Na/phosphate cotransporter NaPi-IIa in a mouse model deficient for the PDZ protein PDZK1

Inorganic phosphate (P(i)) is reabsorbed in the renal proximal tubule mainly via the type-IIa sodium-phosphate cotransporter (NaPi-IIa). This protein is regulated tightly by different factors, among them dietary P(i) intake and parathyroid hormone (PTH). A number of PDZ-domain-containing proteins have been shown to interact with NaPi-IIa in vitro, such as Na(+)/H(+) exchanger-3 regulatory factor-1 (NHERF1) and PDZK1. PDZK1 is highly abundant in kidney and co-localizes with NaPi-IIa in the brush border membrane of proximal tubules. Recently, a knock-out mouse model for PDZK1 (Pdzk1(-/-)) has been generated, allowing the role of PDZK1 in the expression and regulation of the NaPi-IIa cotransporter to be examined in in vivo and in ex vivo preparations. The localization of NaPi-IIa and other proteins interacting with PDZK1 in vitro [Na(+)/H(+) exchanger (NHE3), chloride-formate exchanger (CFEX)/putative anion transporter-1 (PAT1), NHERF1] was not altered in Pdzk1(-/-) mice. The abundance of NaPi-IIa adapted to acute and chronic changes in dietary P(i) intake, but steady-state levels of NaPi-IIa were reduced in Pdzk1(-/-) under a P(i) rich diet. This was paralleled by a higher urinary fractional P(i) excretion. The abundance of the anion exchanger CFEX/PAT1 (SLC26A6) was also reduced. In contrast, NHERF1 abundance increased in the brush border membrane of Pdzk1(-/-) mice fed a high-P(i) diet. Acute regulation of NaPi-IIa by PTH in vivo and by PTH and activators of protein kinases A, C and G (PKA, PKC and PKG) in vitro (kidney slice preparation) was not altered in Pdzk1(-/-) mice. In conclusion, loss of PDZK1 did not result in major changes in proximal tubule function or NaPi-IIa regulation. However, under a P(i)-rich diet, loss of PDZK1 reduced NaPi-IIa abundance indicating that PDZK1 may play a role in the trafficking or stability of NaPi-IIa under these conditions.

Regulation of renal NaPi-2 expression and tubular phosphate reabsorption by growth hormone in the juvenile rat.

Growth hormone (GH) is an important factor in the developmental adaptation to enhance P(i) reabsorption; however, the nephron sites and mechanisms by which GH regulates renal P(i) uptake remain unclear and are the focus of the present study. Micropuncture experiments were performed after acute thyroparathyroidectomy in the presence and absence of parathyroid hormone (PTH) in adult (14- to 17-wk old), juvenile (4-wk old), and GH-suppressed juvenile male rats. While the phosphaturic effect of PTH was blunted in the juvenile rat compared with the adult, suppression of GH in the juvenile restored fractional P(i) excretion to adult levels. In the presence or absence of PTH, GH suppression in the juvenile rat caused a significant increase in the fractional P(i) delivery to the late proximal convoluted (PCT) and early distal tubule, so that delivery was not different from that in adults. These data were confirmed by P(i) uptake studies into brush-border membrane (BBM) vesicles. Immunofluorescence studies indicate increased BBM type IIa NaP(i) cotransporter (NaPi-2) expression in the juvenile compared with adult rat, and GH suppression reduced NaPi-2 expression to levels observed in the adult. GH replacement in the [N-acetyl-Tyr(1)-d-Arg(2)]-GRF-(1-29)-NH(2)-treated juveniles restored high NaPi-2 expression and P(i) uptake. Together, these novel results demonstrate that the presence of GH in the juvenile animal is crucial for the early developmental upregulation of BBM NaPi-2 and, most importantly, describe the enhanced P(i) reabsorption along the PCT and proximal straight nephron segments in the juvenile rat.

P-glycoprotein inhibitors stimulate renal phosphate reabsorption in rats

Dipyridamole (Dip) was previously shown to increase renal phosphate (Pi) reabsorption in humans. However, the mechanism(s) underlying this renal tubular effect is not fully elucidated. It is known that Dip inhibits the activity of the P-glycoprotein (Pgp) multidrug resistance protein 1 (MDR1) expressed on the apical membrane of renal proximal tubular cells where the Na-Pi cotransporter (NPT2) is also expressed. We hypothesized that Dip could increase renal Pi reabsorption by inhibiting Pgp activity. To test this hypothesis, the effects of Dip, verapamil (Ver), and cyclosporine A (CsA), three unrelated Pgp inhibitors, were studied on the renal Pi reabsorption in rats. All three drugs decreased the fractional excretion of Pi (FE(Pi)) in a dose-dependent manner within one hour after beginning the drug infusion, without altering the glomerular filtration rate or serum parathyroid hormone concentration. Sodium-dependent Pi uptake but not Na-glucose transport was increased in brush-border membrane vesicles (BBMVs) when comparing treated with untreated rats. Western blot analysis showed that NPT2 protein was increased in BBMVs from treated rats. Dip and Ver had no effect when applied directly to BBMVs prepared from untreated rats. Pretreatment of rats with colchicine prevented the effects of Dip on the FE(Pi) and NPT2 expression in brush-border membranes. Our results suggest that inhibition of Pgp in the proximal tubule increases Pi uptake and NPT2 translocation to the apical membrane.

Evidence for a PTH-independent humoral mechanism in post-transplant hypophosphatemia and phosphaturia

Patients undergoing successful kidney transplantation often manifest overt hypophosphatemia associated with exaggerated phosphaturia during the early post-transplant period (2 weeks to 3 months). The mechanism for this phenomenon has not been fully elucidated. We tested the hypothesis that a circulating serum factor [non-parathyroid hormone (non-PTH)], which operates during chronic renal failure (CRF) to maintain phosphate (Pi) homeostasis, can increase fractional excretion of Pi (FE(PO4)) in normal functioning kidney grafts during the early post-transplant period, thereby causing phosphaturia and hypophosphatemia. Five groups of patients were studied: control subjects (group 1, N = 16), "early" (2 weeks to 1 month) post-transplant patients (group 2, N = 22), "late" (9 to 12 months) post-transplant patients (group 3, N = 14), patients with advanced CRF (glomerular filtration rate = 30 to 40 mL/min; group 4, N = 8), and patients who suffered from end-stage renal failure and were treated by chronic hemodialysis (group 5, N = 14). Group 2 manifested significant hypophosphatemia and phosphaturia when compared with groups 1 and 3 (Pi = 0.9 +/- 0.003 mg/dL, FE(PO4) = 68+/- 5%, P < 0.0005 vs. groups 1 and 3). Sera were taken from each of the five subject groups and applied to the proximal tubular opossum kidney (OK) cells. The activity of Na/Pi-type 4 (that is, OK-specific type II transporter) was evaluated by measuring Na(+)-dependent (32)Pi flux. The expression of Na/Pi type II mRNA and the abundance of Na/Pi protein were determined by Northern and Western blot assays, respectively. When compared with sera from groups 1 and 3, 10% sera taken from groups 2, 4, and 5 (incubated overnight with OK cells) inhibited (32)Pi flux by 25 to 30% (P < 0.0003). Both Na/Pi mRNA and the expression of Na/Pi protein were markedly augmented under the same conditions (P < 0.05 groups 2, 4, and 5 vs. groups 1 and 3). Time-course analysis revealed that the up-regulation of Na/Pi protein by sera from groups 2, 4, and 5 was observed as early as four hours of incubation, whereas augmented abundance of Na/Pi mRNA was only seen after eight hours of incubation. The addition of PTH (1-34) to sera from groups 2, 4, and 5 abolished the augmented expression of NaPi protein. We labeled OK cell surface membrane proteins with N-hydroxysuccinimide bound to biotin (NHS-SS-biotin). Biotinylated transporters incubated with the different sera were precipitated by strepavidin and identified by Western blot analysis. Cells incubated in sera from group 2 showed increased membrane bound transporter when compared with control sera, whereas the intracellular pool of the transporter was comparable between the two groups. A non-PTH circulating serum factor (possibly phosphatonin) that increases FE(PO4) during CRF is also responsible for phosphaturia and hypophosphatemia in the early period following successful kidney transplantation. The putative factor inactivates Na/Pi activity along with inhibition of the transporter trafficking from the cell membrane into the cytosol.

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Npt2 gene disruption confers resistance to the inhibitory action of parathyroid hormone on renal sodium-phosphate cotransport.

PTH inhibition of renal sodium-phosphate (Na-Pi) cotransport is associated with the endocytic retrieval of the type II Na-Pi cotransporter, Npt2, from the renal brush border membrane into the late endosomal/lysosomal compartment. The aim of the present study was to determine whether mice homozygous for the disrupted Npt2 gene (Npt2-/-) exhibit decreased renal Pi reabsorption in response to PTH. We demonstrate that PTH has no effect on the serum Pi concentration, fractional excretion of Pi, or Na-dependent Pi transport in renal brush border membrane vesicles in Npt2-/- mice. In contrast, PTH elicits a fall in the serum Pi concentration, an increase in urinary Pi excretion, a decrease in brush border membrane Na-Pi cotransport, and a corresponding reduction in the relative abundance of Npt2 protein in wild-type mice (Npt2+/+). Both Npt2-/- and Npt2+/+ mice exhibit a significant rise in the urinary cAMP/creatinine ratio in response to PTH, indicating that generalized resistance to PTH cannot account for the absence of the PTH response in Npt2-/- mice. In addition, we demonstrate that Pi-depleted normal mice respond to PTH with a decrease in renal brush border membrane Na-Pi cotransport and Npt2 protein, indicating that Pi deficiency per se does not account for PTH resistance in Npt2-/- mice. Taken together, our data provide compelling evidence that Npt2 gene expression is crucial for PTH effects on renal Pi handling.

Gender differences in renal growth and function after uninephrectomy in adult rats

It is known that compensatory renal growth (CRG) following unilateral nephrectomy (UNX) increases both the size of the kidney and its functional capacity; however, few studies have investigated whether differences in CRG exist between the sexes. Our study examined the sex-related differences in remnant kidney growth and function two months following UNX. Adult male and female Wistar rats underwent either left UNX or sham operation and recovered for 8 to 10 weeks. Another group of female rats underwent ovariectomy (Ox), with vehicle, estrogen, or testosterone replacement: two-weeks postsurgery animals underwent UNX and recovered for 8 to 10 weeks. Metabolic studies, acute renal function studies [response to acute saline volume expansion (2 to 4% of body wt) or phosphate (Pi) infusions in thyroparathyroidectomized rats (to determine the transport maximum (TmPi)], and renal morphology were assessed at the end of the experimental period. Two months post-UNX, male remnant kidneys grew 114+/-7% of their excised kidney weight (KW), whereas female remnant kidneys grew only 57+/-4% (P<0.05). There was a significant increase in the glomerular volume of male remnant kidneys (126.2+/-13.4%, P<0.001) compared with control kidney volume, whereas there was no change in glomerular volume in female remnant kidneys (20.2+/-16.1%, P = NS). There was also glomerular and tubular damage in the male remnant kidneys, whereas female remnant kidneys were intact. Studies in Ox female rats supplemented with gonadal steroids determined that testosterone is the driving force for the enhanced remnant kidney growth and glomerular hypertrophy. Renal function studies determined that UNX males had significantly higher glomerular filtration rates (GFRs) than UNX females, although the GFR/single KW was not different between the sexes, indicating a proportional increase in GFR. Basal urinary sodium excretion and urine flow rates were significantly higher in anesthetized UNX rats than their sham-operated controls, and urinary sodium excretion and urine flow rates in UNX males were significantly higher than in UNX females. Both male and female UNX rats responded to volume expansion with an exaggerated initial sodium and urine excretion compared with their controls. Phosphate handling was not altered in UNX male rats; however, UNX female rats had increases in fractional Pi excretion that were associated with significant reductions in the maximum capacity for Pi reabsorption (2.10+/-0.07 vs. 3.43+/-0.24 micromol/ml GFR in female controls, P<0.0001). This difference was also observed in Ox rats treated with estrogen and testosterone (2.31+/-0.07 vs. 3.12+/-0.11 micromol/ml GFR, P<0.0007). These findings indicate that sexual dimorphism exists in remnant kidney growth and function two months following UNX. Indeed, morphological abnormalities and impairment in renal phosphate handling are affected by gonadal steroids by two-months post-UNX. The fact that renal Pi transport was reduced in female but not male UNX rats may also have important implications during periods of high metabolic demand for phosphate in the female.

Locally formed 5-hydroxytryptamine stimulates phosphate transport in cultured opossum kidney cells and in rat kidney.

Renal proximal tubular cells have been shown to express aromatic L-amino acid decarboxylase (L-AAAD), which converts L-dopa into dopamine and 5-hydroxytryptophan [(OH)Trp] into 5-hydroxytryptamine (5-HT; serotonin). Because 5-HT receptors have been demonstrated in proximal cells, we hypothesized that 5-HT may act as an autocrine/paracrine modulator of proximal transport. We evaluated this possibility in opossum kidney (OK) cells, a renal epithelial cell line with a proximal phenotype expressing 5-HT1B receptors, and in intact anaesthetized rats. 5-HT synthesis by OK cells increased with incubation time and (OH)Trp concentration, and was abolished by benserazide, an L-AAAD inhibitor. 5-HT reversed parathyroid hormone (PTH)-induced cAMP accumulation in a pertussis toxin-sensitive manner and reduced the PTH inhibition of P(i) uptake without affecting the NaP(i)-4 mRNA level. The effects of 5-HT on cAMP generation and Na-P(i) co-transport were reproduced by (OH)Trp, except in the presence of benserazide, and by L-propranolol and dihydroergotamine, two 5-HT1B receptor agonists. In rats, (OH)Trp and dihydroergotamine decreased fractional P(i) excretion. Benserazide abolished the effect of (OH)Trp but not that of dihydroergotamine. (i) locally generated 5-HT blunts the inhibitory effect of PTH on Na-P(i) co-transport in OK cells; (ii) endogenous 5-HT decreases P(i) excretion in rats; and (iii) 5-HT is a paracrine modulator involved in the physiological regulation of renal P(i) transport.

Maturation of rat renal phosphate transport: effect of triiodothyronine.

1. Prevention of weaning prevents the increase in the carrier affinity for Na(+)-cotransported phosphate (Pi) and the normal elevation of plasma 3,5,3'-triiodothyronine (T3) which occur between postnatal days 14 and 21. 2. This study examines the possible role of T3 in the control of the maturation process of Pi transport. Clearance experiments and brush-border membrane studies were performed on 14-day-old rats given T3. 3. The fractional excretion of Pi of T3-treated rats was 33% lower compared with controls (P < 0.01). After Pi perfusion, it remained at a lower level, and the amount of Pi reabsorbed per minute, corrected for the glomerular filtration rate (RPi/GFR), was higher. 4. The membrane vesicles from 14-day-old rats given T3 showed a 30% increase in carrier affinity for Na(+)-cotransported Pi. In addition to this maturational effect of T3, a 46% increase in Vmax was also observed. 5. The amount of immunoreactive Pi transporter, detected using anti-(NaPi-2) antiserum, was increased in T3-treated rats. 6. Glucose transport, another Na(+)-dependent transport system, was not altered by T3. 7. It is concluded that exogenous T3 given before the third postnatal week specifically induced precocious maturation of renal Pi transport in 14-day-old rats, suggesting that thyroid hormone is normally involved in this maturation.

Restoration of a phosphaturic response to parathyroid hormone in the immature rat.

Recent studies have shown that immature rats display a diminished sensitivity to the phosphaturic effects of parathyroid hormone (PTH), and that the responsiveness to PTH increases with age. The attenuated phosphaturia may reflect an inability of the neonate to respond to the hormone because of functional immaturity of the developing kidney. Alternatively, PTH may actually inhibit tubular phosphate reabsorption in the neonate but, due to other phosphate conservation mechanisms, no phosphaturia occurs. Our objective was to determine whether a phosphaturic response to PTH would be elicited in immature rats during infusion of moderate amounts of phosphate (Pi). Clearance experiments were performed on 26 acutely thyroparathyroidectomized immature Wistar rats (3-5 wk of age) fed a normal Pi diet (0.63%). In response to infusion of either Pi (1 mumol/min.100 g) (group I) or PTH (8.3 ng/min.100 g) (group II) alone, the fractional excretion of phosphate rose minimally (from 0.01 +/- 0.01% to 4.9 +/- 1.9% and from 0.12 +/- 0.12% to 2.9 +/- 1.4% for groups I and II, respectively). However, when Pi and PTH were combined either Pi first followed by PTH (group III) or PTH first followed by Pi (group IV), the fractional excretion of Pi rose dramatically (from 0.01 +/- 0.01 to 21.8 +/- 3.5% and from 0.04 +/- 0.04 to 27.7 +/- 3.3% for groups III and IV, respectively). A significant increase in urinary cAMP excretion occurred during infusion of PTH even when Pi excretion was minimal, but there was no further increase in urinary cAMP during the combined infusion of Pi and PTH.(ABSTRACT TRUNCATED AT 250 WORDS)

Endogenous ANP augments fractional excretion of Pi, Ca, and Na in rats with reduced renal mass.

Atrial natriuretic peptide (ANP) infusion increases fractional excretion of many solutes including sodium, chloride, bicarbonate, phosphate, calcium, and magnesium. Because fractional excretion of these solutes increases with advancing renal disease, and because plasma ANP levels are known to be elevated in chronic renal failure, we sought to determine whether ANP mediates increased solute excretion rates per nephron in rats following extensive renal ablation, a model of chronic renal failure. Because sodium restriction decreases plasma ANP levels in the setting of reduced renal mass, we also determined the effect of sodium restriction on sodium, phosphate, calcium, and magnesium excretion rates in rats with 5/6 nephrectomy (NX). We also assessed whether high endogenous ANP levels influence fractional sodium, phosphate, calcium, and magnesium excretion in rats with 5/6 NX, by inhibiting ANP action via infusion of a high-affinity ANP antiserum. Whole-kidney glomerular filtration rate in 5/6 NX rats averaged approximately one-third that of shams, and plasma ANP levels were significantly elevated in these rats above those of shams, but to a lesser extent in rats on low- vs. high-salt intakes. Fractional sodium, phosphate, and calcium, but not magnesium excretion rates were significantly greater in 5/6 NX rats on the higher sodium intake compared with those in 5/6 NX rats on the lower sodium intake. Moreover, in 5/6 NX rats on the higher sodium intake, ANP antiserum significantly reduced fractional sodium, phosphate, and calcium excretion, but was without effect on magnesium excretion. These data implicate endogenous ANP in promoting the adaptive increase in sodium, phosphate, calcium, but not magnesium excretion per nephron in chronic renal disease.

Adaptation to phosphorus depletion: effect of streptozotocin-induced diabetes mellitus.

The effect of streptozotocin-induced diabetes mellitus (DM) on the adaptive response to phosphorus depletion (PD) was investigated in order to examine if DM has any influence on the adaptation to PD in rats. PD for 7 days caused a marked reduction in serum phosphate (Pi) levels and increase in serum calcium (Ca) concentrations in control rats. In contrast, the increase in serum Ca concentration caused by PD was almost entirely eliminated in DM rats. Similarly, while bone Ca and P content was decreased by 7 days of PD in control rats, no significant changes in bone mineral contents were observed in DM rats during PD. There was a marked reduction in fractional excretion of Pi and an increase in fractional excretion of Ca during PD in both control and DM rats. Serum somatomedin A levels measured by radioreceptor assay were lower in DM rats compared to those in control rats, but PD caused no significant changes in either group of animals. These results demonstrate that the development of hypercalcaemia and reduction in bone mineral content in response to PD were inhibited while the renal tubular responses to PD were not affected in DM rats. It is suggested that the inhibition of the hypercalcaemic response to PD in DM rats is mainly due to an inhibition of the resorptive response of bone to PD, and that insulin either directly or indirectly may play a permissive role in the development of the resorptive response of bone to PD.

Effects of 1,25-dihydroxycholecalciferol on phosphate transport in vitamin D-deprived rats.

To clarify the role of vitamin D in renal phosphate transport, weanling rats were fed a vitamin D-deficient diet containing 1.8% calcium and 1.2% phosphorus. After 5-6 wk, the rats were normocalcemic, normophosphatemic, and had normal levels of PTH. Assays of vitamin D metabolites revealed undetectable plasma levels of 25(OH)D, and 1,25(OH)2D levels of 92 +/- 16 pg/ml in partially vitamin D-depleted (PVDD) rats and 169 +/- 58 pg/ml in normal rats. PVDD rats had increased phosphate excretion, both absolute and fractional, and a decrease in Na+ gradient-dependent Pi transport in proximal tubular brush border membrane vesicles (BBMV) prepared from their kidneys. Vitamin D repletion of PVDD rats with 1,25(OH)2D3, 15 pmol/100 g body wt, decreased fractional excretion of Pi from 22.6 +/- 1.9 to 13.5 +/- 1.3%; the latter values were similar to normal rats. Repletion with 1,25(OH)2D3 also increased Na+-dependent phosphate transport in BBMV from 322 +/- 24 pmol X mg protein-1 X 15 s-1 in BBMV from PVDD rats to 698 +/- 70 pmol X mg protein-1 X 15 s-1. Repletion with larger doses of 1,25(OH)2D3 produced hypercalcemia and hyperphosphatemia from intestinal absorption, an increase in phosphate excretion, and a blunted response of Pi transport to 1,25(OH)2D3. Prevention of hyperphosphatemia by dietary adjustments allowed full expression of the stimulatory effects of 1,25(OH)2D3 on Pi transport. These later data may partially explain the inhibitory effects reported in prior studies in which plasma Pi was not controlled and the larger doses of 1,25(OH)2D3 administered.(ABSTRACT TRUNCATED AT 250 WORDS)