bluuebunny/crossref_metadata_2025 huggingface dataset:
Autocrine/paracrine regulation of renal Na+-phosphate cotransport by dopamine
Authors: Glahn R. P. , Onsgard M. J. , Tyce G. M. , Chinnow S. L. , Knox F. G. , Dousa T. P.
Published: December 2017

Pages F618–F622: R. P. Glahn, M. J. Onsgard, G. M. Tyce, S. L. Chinnow, F. G. Knox, and T. P. Dousa. “Autocrine/paracrine regulation of renal Na+-phosphate cotransport by dopamine.” As a result of a systematic computational error, all values for transport rates presented in Tables 1 and 2 are 36-fold greater than the actual measured values. Because the same calculation error was applied to all conditions (including controls), the interpretations, significance, and conclusions of this study are still valid. However, for the convenience of the readership, the corrected Tables 1 and 2 are printed here. Additionally, the value in the legend of Fig. 1 for basal Na+-dependent uptake rate of 32Pi should be 2.09 ± 0.19 nmol·5 min-1mg protein-1
Postnatal maturation of potassium transport in rabbit cortical collecting duct
Authors: Satlin L. M.
Published: December 2017

Clearance studies in newborns demonstrate low rates of urinary excretion of potassium, suggesting that the neonatal kidney contributes to the conservation of potassium necessary for growth. Because the cortical collecting duct (CCD) is a primary site for potassium secretion in the adult, we sought to examine the transport capacity of this segment for potassium during postnatal maturation. CCDs isolated from rabbits of various ages (5-6 animals/age group) were microperfused in vitro with solutions simulating plasma. The concentrations of potassium in samples of collected fluid, measured by helium glow photometry, were used to calculate net transport. At a flow rate of approximately 1.6 nl.min-1 x mm-1 net potassium secretion was absent at birth, first became evident at 4 wk of age (-11.08 +/- 2.39 pmol.min-1 x mm-1), and increased sharply thereafter to reach mature rates (-23.08 +/- 3.47 pmol.min-1 x mm-1; P < 0.05) by 6 wk of age. To determine whether low distal tubular flow rates limit net potassium secretion in the neonate, we perfused CCDs at two or more flow rates in the 0.5–5 nl.min-1 x mm-1 range. In CCDs taken from animals > or = 6 wk of age, potassium secretion showed a significant linear correlation with flow rate (y = -10.0x - 7.45; r = 0.87; n = 12).(ABSTRACT TRUNCATED AT 250 WORDS)
Angiotensin II suppression is a major factor permitting excretion of an acute sodium load in humans
Authors: Singer D. R. , Markandu N. D. , Morton J. J. , Miller M. A. , Sagnella G. A. , MacGregor G. A.
Published: December 2017

We examined the role of circulating angiotensin II (ANG II) in the excretion of an acute Na+ load in eight healthy subjects given 2 liters 0.9% saline in a placebo-controlled crossover study. On the control day, plasma ANG II decreased to 40-60% of basal values and 35 +/- 9 (SE) mmol of Na+ was excreted in the 5 h after the start of saline infusion. When ANG II was infused to maintain plasma ANG II levels at around basal values (6.6 +/- 1.6 pmol/l), only 7 +/- 8 mmol of Na+ was excreted in the same period (P < 0.05). In a previous similar study in which the fall in aldosterone was prevented by infusion of aldosterone, 16 +/- 16 mmol of Na+ was excreted vs. 36 +/- 16 mmol on the control day in comparable 5-h periods. Suppression of ANG II is one of the major factors permitting the acute increase in Na+ excretion after an intravenous Na+ load. ANG II has direct actions on Na+ excretion in addition to its effects on aldosterone.
Molecular adaptations of GLUT1 and GLUT2 in renal proximal tubules of diabetic rats
Authors: Dominguez J. H. , Camp K. , Maianu L. , Feister H. , Garvey W. T.
Published: December 2017

The renal reabsorption of glucose is mediated by two major classes of transporters. Initially, luminal glucose is concentrated in tubules by Na(+)-glucose cotransporters (Na(+)-GLUT). Afterwards, glucose reaches the blood space through facilitative glucose transporters, low-Michaelis constant (Km) GLUT1 and high-Km GLUT2. Hence, the transtubular flux of glucose could be impaired in hyperglycemia because the outwardly directed glucose gradient, from tubule to blood, is potentially lowered. However, in diabetic rats, transtubular glucose flux is not reduced but increased. In this work the molecular mechanism underlying this adaptation was examined. We tested the hypothesis that upregulation of renal tubular high-Km GLUT2 gene may compensate for the decrease in the tubule to blood glucose gradient. In rat tubules, GLUT1 protein and mRNA steady-state levels were reduced, and GLUT2 protein and mRNA levels were increased in rats after 2, 3, and 4 wk of uncontrolled streptozotocin-induced diabetes. These molecular adaptations were associated with augmented facilitative glucose flux. In summary, changes in GLUT1 and GLUT2 gene expression are important to the preservation of renal glucose reabsorption in hyperglycemia.
Why renal cysts grow
Authors: Gardner K. D. , Glew R. H. , Evan A. P. , McAteer J. A. , Bernstein J.
Published: December 2017

The growth of renal cysts leads to morbidity, contributes to mortality, and is a lesson in applied physiology. The fact that chemical, electrical, osmotic, and hydrostatic driving forces determine cyst volumes is inferred from observations that cyst fluids vary in their concentrations of osmotically active substances. Most have concentrations of sodium-based salts that are lower and nonsodium-based salts that are higher than those present in normal plasma. Nonsodium solutes include osmolytes that normally are present in higher concentrations inside of cells, including potassium, amino acids, and so-called idiogenic osmoles. The basic process of cyst growth therefore involves 1) the osmotic equilibration of water across cysts walls that have variable permeability characteristics and 2) the replacement of sodium salts with other osmotically active solutes in cyst fluids. Cyst volume is governed by the amounts and kinds of osmolytes that enter and become entrapped in lumina. Proliferation and necrosis of mural cells are events that are fundamental to the growth of renal cysts.
Differential regulation of angiotensin II and losartan binding sites in glomeruli and mesangial cells
Authors: Chansel D. , Bizet T. , Vandermeersch S. , Pham P. , Levy B. , Ardaillou R.
Published: December 2017

The aim of the present report was to examine the effect of several agents on angiotensin II (ANG II) and losartan receptors using 125I-[Sar1,Ala8]ANG II and [3H]losartan as radiolabeled ligand, respectively. ANG II receptors were downregulated in glomeruli from rats infused with ANG II during 3 wk or rats receiving losartan orally during 1 wk. The number of sites (Bmax) was reduced, but the dissociation constant (Kd) value was unchanged. Losartan receptors were downregulated in glomeruli from rats receiving losartan, but remained unchanged in glomeruli from rats infused with ANG II. Since in vivo administration of losartan results in increase of plasma ANG II and formation of metabolites, in vitro studies using human mesangial cells were performed to better analyze the present findings. Treatment of mesangial cells during 4 days by ANG II, losartan, or its metabolite, EXP-3174, also produced downregulation of 125I-[Sar1,Ala8]ANG II binding sites with a decreased Bmax and unchanged Kd value. Only treatment of mesangial cells by ANG II or EXP-3174 produced downregulation of [3H]losartan binding sites. In contrast, exposure of these cells to losartan resulted in upregulation of [3H]losartan binding sites. Under all conditions, only Bmax was modified. Whereas internalization of [3H]losartan in mesangial cells was negligible under all experimental conditions, there was an increase of the percentage of internalized 125I-[Sar1,Ala8]ANG II after exposure of the cells to ANG II or AT1 antagonists. No change was observed in mesangial cell AT1 receptor mRNA levels. This study demonstrates that 1) AT1 mRNA is expressed in human mesangial cells; 2) the characteristics of 125I-[Sar1,Ala8]ANG II and [3H]losartan binding sites in rat glomeruli and human mesangial cells are different, with Kd and Bmax values greater in both preparations when [3H]losartan was utilized; 3) both types of binding sites obey different regulations, and the effects of losartan in vivo are due in part to the associated increase in plasma ANG II levels and the transformation of the drug into its metabolite, EXP-3174; 4) downregulation of AT1 receptors does not depend on changes in mRNA expression but is associated with increased relative internalization.
Effect of insulin on water and urea transport in the inner medullary collecting duct
Authors: Magaldi A. J. , Cesar K. R. , Yano Y.
Published: December 2017

The effect of insulin on water and urea transport was examined in normal isolated rat inner medullary collecting duct (IMCD). Hydraulic conductivity (Lp, x 10(-6) cm.atm-1.s-1), diffusional water permeability (Pdw, x 10(-5) cm/s) and [14C]urea permeability (x 10(-5) cm/s) were studied at 37 degrees C and pH 7.4. Insulin (6 x 10(-8) M; 200 microU/ml) added to the bath fluid enhanced Lp from 0.40 +/- 0.10 to 1.21 +/- 1.40 (P < 0.01) and Pdw from 42.40 +/- 3.40 to 58.50 +/- 5.00 (P < 0.02) and also stimulated Lp in a dose-dependent manner. In the presence of antidiuretic hormone (ADH)-stimulated Pdw (10 microU/ml), insulin increased Pdw even more. Prostaglandin E2 (10(-5) M) added to the bath reversibly increased insulin-induced Lp. Forskolin (10(-4) M) blocked the action of insulin. Colchicine (10(-4) M) and V1-receptor antagonist (10(-4) M) inhibited the development but not the maintenance of insulin-stimulated Pdw. Vanadate (2.5 x 10(-6) M) enhanced Pdw. Polymyxin B (10(-5) M) inhibited the insulin-stimulated Pdw, whereas in a glucose-free medium insulin did not enhance Pdw. Urea transport was not affected by insulin. These data suggest that insulin may enhance water transport, probably by stimulating glucose transporters, which would serve as a water channel. We cannot rule out the possibility that insulin may be eliciting existing ADH-like mechanisms of water transport, beyond the microtubule step, to establish water transport.
Expression of the D2 subfamily of dopamine receptor genes in kidney
Authors: Gao D. Q. , Canessa L. M. , Mouradian M. M. , Jose P. A.
Published: December 2017

The D2, D3, and D4 dopamine receptors cloned from brain correspond to the classically described "D2" receptors. Although radioligand binding and biochemical and functional studies have demonstrated the presence of D2-like receptors in the kidney, the expression of D2, D3, or D4 receptor genes has not been conclusively demonstrated in the kidney. Since Northern blot analysis may have precluded demonstration of dopamine receptor mRNAs because of their relative low abundance, the expression of the D2long and D3 receptor genes was studied by reverse transcription-polymerase chain reaction (RT-PCR). We were able to amplify PCR products of the predicted size using mRNA from glomeruli, proximal tubules, outer medulla, inner medulla, and renal microvessels from normotensive Wistar-Kyoto rats (WKY). Specificities of the amplified products were confirmed by restriction analysis and by sequencing the D2long product and Southern blotting the D3 product. Because some studies have suggested that D2-like receptor actions may be different between WKY and spontaneously hypertensive rats (SHR), similar studies were performed in this rat strain. In the SHR, as in WKY, PCR products of the predicted size were amplified, and restriction enzyme digestion patterns were as predicted from the cDNA sequence. The PCR-generated cDNA from the glomeruli of SHR was subcloned and sequenced and was revealed to be identical to the D2long receptor cDNA from WKY. We conclude that the D2long and D3 receptor genes are expressed in specific regions of the kidney including the glomeruli. No differences in the sequence of the D2long receptor cDNA in part of the 3rd cytoplasmic loop were noted between WKY and SHR. These studies do not rule out the possibility that mutations in other segments of the receptor exist in the SHR.
Second messengers involved in the control of renin secretion in cultured human nephroblastoma cells
Authors: Devlin A. M. , Leckie B. J.
Published: December 2017

We aimed to study the intracellular signaling mechanisms involved in the stimulation and suppression of renin secretion from human nephroblastoma cells. Isoproterenol and forskolin increased intracellular adenosine 3',5'-cyclic monophosphate (cAMP) concentration and stimulated renin secretion as did the addition of dibutyryl-cAMP. Atrial natriuretic peptide (ANP) suppressed basal renin secretion and increased the concentration of extracellular guanosine 3',5'-cyclic monophosphate (cGMP). When ANP was added in the presence of isoproterenol or forskolin, the increase in cGMP was reduced. ANP attenuated the cAMP response to isoproterenol but not forskolin. Nephroblastoma cell membranes contained the guanosine-binding proteins Gs and Gi2, and the isoforms were similar to those in vascular smooth muscle. A functional role for Gi was indicated because the ANP-induced suppression of basal renin secretion was blocked by pertussis toxin. We conclude that cAMP stimulates and cGMP suppresses basal renin secretion, but neither fully accounts for the suppression of stimulated renin secretion by ANP.
Estimation of renal interstitial adenosine and purine metabolites by microdialysis
Authors: Baranowski R. L. , Westenfelder C.
Published: December 2017

It has been proposed that adenosine, derived from ATP and released into the renal interstitium, mediates a reduction in renal function in ischemic acute renal failure. Because no direct measurements of interstitial adenosine are available, we evaluated an in vivo microdialysis technique to assess the levels of adenosine and its metabolites in the cortex of the normal rat kidney (n = 6). Microdialysis probe implantation did not alter cortical renal blood flow, glomerular filtration rate, or fractional sodium excretion. The interstitial concentration of adenosine was 199 +/- 53 nM, and relative concentrations of inosine, hypoxanthine, xanthine, and uric acid were 99 +/- 47, 182 +/- 29, and 183 +/- 70 nM and 1.8 +/- 0.4 microM, respectively. Infusion of ATP-MgCl2 (n = 5) resulted in a significant increase in the dialysate levels of adenosine (67 +/- 11 to 378 +/- 97 nM), inosine (230 +/- 102 to 803 +/- 219 nM), and uric acid (3.5 +/- 1.3 to 6.9 +/- 1.7 microM). In conclusion, this study demonstrates that the microdialysis technique is suited to monitor metabolically important substances in the renal interstitium.
Effects of growth hormone on rat renal epidermal growth factor expression
Authors: Rogers S. A. , Rasmussen J. , Miller S. B. , Hammerman M. R.
Published: December 2017

The kidney is a site of synthesis for several polypeptide growth factors including epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I). Interactions between growth hormone (GH) and growth factors have been described that regulate renal growth factor expression. For example, GH and EGF each enhances the expression of IGF-I in kidney. To further define interrelationships in this renal GH-growth factor axis, we characterized the effect of GH on renal EGF expression in hypophysectomized, pituitary-intact (normal) rats, and hypersomatotropic rats. Levels of extractable immunoreactive mature EGF, levels of a 142-kDa EGF-precursor present in renal membrane fractions, and levels of EGF mRNA were significantly reduced in kidneys from hypophysectomized rats compared with levels in normal rats. Each was increased significantly after the administration of GH to hypophysectomized rats. In contrast, induction of hypersomatotropism in normal rats by injection of GH for 17 days did not affect levels of extractable mature EGF or EGF mRNA measured in kidneys. We conclude that GH enhances the renal synthesis of EGF in hypopituitary, but not in hypersomatotropic states.
Luminal acidification in K-replete OMCDi: contributions of H-K-ATPase and bafilomycin-A1-sensitive H-ATPase
Authors: Armitage F. E. , Wingo C. S.
Published: December 2017

We have previously demonstrated that basolateral addition of the gastric H-K-adenosinetriphosphatase (H-K-ATPase) inhibitor Sch-28080 (10 microM) profoundly reduced net total CO2 flux (JtCO2) in the inner stripe of the outer medullary collecting duct (OMCDi) of K-replete rabbits. In the present studies, we first addressed whether the inhibitory effect of Sch-28080 is dependent on the side of the membrane to which it is added. Second, we reassessed the relative magnitude of contribution of H-K-ATPase. Third, we formally tested whether a bafilomycin-A1 (BAF)-sensitive H-ATPase also contributes to luminal acidification in the OMCDi under K-replete dietary conditions. We found that luminal addition of the structurally and functionally dissimilar gastric H-K-ATPase inhibitor A80915A (10 microM) profoundly reduced JtCO2 while transepithelial voltage (VT) was unchanged. This degree of inhibition was statistically indistinguishable from our previous results when Sch-28080 was applied basolaterally. Inhibition of JtCO2 by the less membrane-permeable N-methyl cation of Sch-28080, H224/25, was significant when applied luminally but was not significant when applied basolaterally. VT was not significantly affected by either the luminal or basolateral addition of H224/25. To evaluate the possible contribution of an H-ATPase, the effect of both 5.0 nM and 10.0 nM luminal BAF on JtCO2 and VT was examined. At 5.0 nM, BAF significantly inhibited JtCO2). However, this observation was significantly less (P < 0.05) than the inhibition observed with 10 microM A80915A. No additional inhibition was observed by increasing the concentration of BAF to 10.0 nM.(ABSTRACT TRUNCATED AT 250 WORDS)
Cellular mechanisms of glucose-induced myo-inositol transport upregulation in rat mesangial cells
Authors: Chatzilias A. A. , Whiteside C. I.
Published: December 2017

Uptake of myo-inositol (MI) is necessary to maintain normal cellular phosphoinositide signaling and function. MI transport is up-regulated in the cells of diabetic rat glomeruli compared with normal rat glomeruli [C. I. Whiteside, J. C. Thompson, and J. Ohayon. Am. J. Physiol. 260 (Renal Fluid Electrolyte Physiol. 29): F138-F144, 1991]. To identify mechanisms associated with upregulation of MI transport, rat mesangial cells were cultured in high (25.6 mM) vs. normal (5.6 mM) glucose. Specific Na(+)-dependent [3H]MI uptake (> 97%), using L-[14C]glucose as the nonspecific marker, was linear for 120 min in high and normal glucose. In high glucose, compared with normal glucose, there was no change in Michaelis-Menten constant values [29.1 +/- 0.6 vs. 30.3 +/- 0.7 microM (SE)], whereas maximum velocity (Vmax) was increased (2.024 +/- 52 vs. 1.132 +/- 115 fmol.mg protein-1.min-1, P < 0.001). Mannitol (20.0 mM), used as an osmotic control, had no effect on the upregulation of MI transport. Maximum upregulation of MI transport measured by Vmax (control taken as 100%) was observed after 8 h of exposure to high glucose (222 +/- 6% above control, P < 0.0001) or galactose (20.0 mM) (194 +/- 6%, P < 0.0001) and was sustained for up to 48 h. The protein synthesis inhibitors cycloheximide (20 micrograms/ml) or actinomycin D (5 micrograms/ml), the F-actin depolymerizing agent cytochalasin D (2 micrograms/ml), and the aldose reductase inhibitor Tolrestat (0.3 mM) independently prevented glucose- or galactose-induced upregulation of MI transport.(ABSTRACT TRUNCATED AT 250 WORDS)
Maturation of proximal tubule Na+/H+ antiporter activity in sheep during transition from fetus to newborn
Authors: Guillery E. N. , Karniski L. P. , Mathews M. S. , Robillard J. E.
Published: December 2017

We have studied maturational changes in the kinetics of the proximal tubule Na+/H+ antiporter. Microvillus membrane vesicles were prepared from renal cortex of fetal and newborn lambs. Amiloride-sensitive uptake of 22Na+ by these vesicles was measured and Woolf-Augustinsson-Hofstee plots were used to determine the Michaelis constant (Km) and rate of maximal uptake (Vmax). Initial studies of fetal lambs at 130-132 days gestation (n = 5; term is 145 days) and 3- to 4-day-old lambs (n = 5) revealed no maturational change in Km (7.27 +/- 1.25 for fetuses and 9.01 +/- 1.03 mM for lambs); however, there was a 242% increase in Vmax (from 1.28 +/- 0.33 in the fetuses to 4.37 +/- 0.85 nmol.s-1.mg protein-1 in the lambs, P = 0.005). Further definition of the developmental change in Na+/H+ antiporter Vmax was obtained when 144-day-gestation fetuses (n = 5) were compared with 24-h-old sibling lambs (n = 5) that had been delivered by cesarean section at 144 days gestation. Again, no significant difference was seen in Na+/H+ antiporter Km (14.9 +/- 6.5 for fetuses and 12.5 +/- 3.4 mM for lambs); however, a significant increase in Na+/H+ antiporter Vmax occurred (from 1.41 +/- 0.51 in the fetuses to 3.32 +/- 0.37 nmol.s-1.mg protein-1 in the lambs, P < 0.01). This study shows that there is a maturational increase in renal cortical Na+/H+ antiporter Vmax during the transition from fetal to newborn life. This increase parallels the increase in renal tubular Na+ reabsorption that occurs at this time.
Vasopressin activates a chloride conductance in cultured cortical collecting duct cells
Authors: Nagy E. , Naray-Fejes-Toth A. , Fejes-Toth G.
Published: December 2017

In rabbit cortical collecting duct (CCD) cells, arginine vasopressin (AVP) causes a transient increase followed by a sustained depression of transepithelial potential difference (PDte). Mechanisms underlying the decrease in PDte are not well understood. In this study, we used primary cultures of rabbit CCD cells to study effects of AVP. Basolateral addition of AVP caused a dose-dependent increase in transepithelial conductance (Gte) and a corresponding decrease in PDte. A significant effect was observed at 1 pM AVP, and half-maximal response occurred at 30 pM AVP; 1 nM AVP increased Gte and decreased PDte. Replacement of apical Na+ with N-methyl-D-glucamine did not prevent the effect of AVP on Gte, suggesting that it is not mediated by an increase in apical Na+ conductance. Similarly, apical Ba2+ (1 mM) or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 0.1 mM) failed to prevent the effect of AVP. On the other hand, 5-nitro-2(3-phenylpropylamino)benzoic acid (0.1 mM) caused partial inhibition, whereas substitution of apical Cl- with gluconate or cyclamate almost completely prevented the AVP-induced increase in Gte. In unidirectional ion-flux studies, 1 nM AVP caused only a modest increase in apical-to-basolateral (A-->BL) flux of 22Na and had no effect on transepithelial flux of 86Rb in either direction. On the other hand, AVP caused a pronounced increase in A-->BL flux and BL-->A flux of 36Cl, resulting in an increased net Cl- absorption. The effect of AVP on Gte could be mimicked by 8-bromo-adenosine 3',5'-cyclic monophosphate (8-bromo-cAMP) and isoproterenol, and effects of AVP and isoproterenol were not additive.(ABSTRACT TRUNCATED AT 250 WORDS)
Physiological and structural responses to chronic experimental renal allograft injury
Authors: Junaid A. , Kren S. M. , Rosenberg M. E. , Nath K. A. , Hostetter T. H.
Published: December 2017

Chronic rejection necessitates a return to dialysis or retransplantation for a significant number of patients with renal allografts. Although alloresponses between donor organ and recipient importantly determine this process, the detailed immunologic processes and organ physiology of chronic rejection are unclear; in consequence its mechanism and therapy are uncertain. A model of chronic rejection in the rat was used to examine several facets of this process. Fisher-to-Lewis (F-L), allogeneic, and Lewis-to-Lewis (L-L), syngeneic, renal transplants were performed in nephrectomized recipients. All rats were treated with cyclosporin A (5 mg.kg-1.day-1) for 10 days from the time of grafting. At 6 wk, allogeneically grafted animals had a higher protein excretion rate (F-L, 47 +/- 30 mg/day; L-L, 17 +/- 6 mg/day; P <0.05) and an increase in glomerular capillary pressure (F-L, 69 +/- 5 mmHg; L-L, 58 +/- 8 mmHg; P <0.05) and fractional cortical interstitial volume (F-L, 29.8 +/- 4.3%; L-L, 19.5 +/- 4.0%; P < 0.01). This model of chronic rejection is characterized by glomerular capillary hypertension, proteinuria, and cortical interstitial expansion. Because these findings are also present in other models of chronic renal injury, mechanisms in addition to alloresponses may operate in chronic rejection.
Morphometric analysis of effects of angiotensin II on glomerular structure in rats
Authors: Pagtalunan M. E. , Rasch R. , Rennke H. G. , Meyer T. W.
Published: December 2017

Micropuncture and morphometric studies related the effects of angiotensin II (ANG II) on glomerular function and structure. Compared with control animals, rats receiving an intrarenal infusion of ANG II at 40 ng.kg-1.min-1 exhibited a marked reduction in the glomerular ultrafiltration coefficient (Kf) (0.84 +/- 0.13 vs. 1.43 +/- 0.05 microliters.s-1.mmHg-1, ANG II vs. control), which caused a decrease in glomerular filtration rate (GFR) (1.04 +/- 0.11 vs. 1.27 +/- 0.11 ml/min) despite an increase in glomerular transcapillary hydraulic pressure difference (46 +/- 1 vs. 40 +/- 1 mmHg). Morphometric studies showed that these hemodynamic changes were not associated with any reduction in glomerular volume (1.27 +/- 0.05 vs. 1.31 +/- 0.07 x 10(6) microns3, ANG II vs. control), glomerular capillary volume (4.25 +/- 0.36 vs. 4.41 +/- 0.33 x 10(5) microns3), or glomerular peripheral capillary surface area (2.24 +/- 0.11 vs. 2.29 +/- 0.30 x 10(5) microns2). Higher-power electron micrographs showed that ANG II also did not alter mean foot process width (478 +/- 14 vs. 491 +/- 18 nm, ANG II vs. control), reduce the total filtration slit length overlying the peripheral capillary wall (7.0 +/- 0.6 vs. 6.6 +/- 0.5 x 10(5) microns), or reduce the average width of individual filtration slits (45 +/- 2 vs. 43 +/- 2 nm). ANG II infusion thus caused a 40% reduction in the value of Kf without causing detectable changes in epithelial cell or filtration slit structure.
Sodium entry mechanisms in distal convoluted tubule cells
Authors: Gesek F. A. , Friedman P. A.
Published: December 2017

Sodium transport across apical membranes of distal convoluted tubules is thought to be mediated by Na-Cl cotransport and conductive Na entry. Immortalized mouse distal convoluted tubule cells were used to characterize Na entry pathways. Chlorothiazide, an inhibitor of Na-Cl cotransport, and amiloride, which blocks epithelial Na channels, reduced ouabain-suppressible oxygen consumption by 40 and 35%, respectively. In simple buffer solutions, free of bicarbonate, phosphate, or formate, chlorothiazide inhibited Na uptake by 44% and Cl uptake by 48%. Michaelis constants of 21 mM for Na and 14 mM for chloride were calculated. Amiloride inhibited Na uptake by 49% and had no effect on Cl uptake. The calculated Hill coefficient of 1.07 and the equivalence of chlorothiazide-sensitive Na and Cl uptake are consistent with the presence of Na-Cl cotransport. Na-Cl cotransport and amiloride-sensitive Na influx account for 85% of Na entry in distal convoluted tubule cells in the absence of phosphate and formate. The selective Na/H exchange inhibitor ethylisopropyl amiloride had no effect on Na uptake; however, it abolished formate-stimulated Na uptake. The anion exchange blocker 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) inhibited Na uptake. These findings are consistent with parallel Na/H and Cl/formate exchange. Na uptake was inhibited 8% by the selective Na/Ca exchange inhibitor, dimethylbenzamil. An additional 7% of Na entry was phosphate dependent and was abolished by phosphonoformic acid, a competitive inhibitor of Na-Pi cotransport. In summary, the majority of Na entry into distal convoluted tubule cells occurs through Na-Cl cotransport and an amiloride-sensitive pathway (75% in presence of phosphate and formate). An additional 13% may enter by Na/H exchange, with the remainder mediated by Na/Ca exchange and Na-Pi cotransport.
Bicarbonate transport along the loop of Henle effects of adrenal steroids
Authors: Unwin R. , Capasso G. , Giebisch G.
Published: December 2017

The role of adrenal steroids in the regulation of bicarbonate absorption in the loop of Henle was studied by in vivo microperfusion. Bicarbonate transport (JHCO3) was measured by microcalorimetry and fluid reabsorption by [14C]inulin, 7-10 days after surgery, in 1) sham-operated control rats, 2) adrenalectomized (Adx) rats, 3) Adx rats receiving dexamethasone (1.2 micrograms.100 g body wt-1.24 h-1) and a low dose of aldosterone (0.5 micrograms.100 g body wt-1.24 h-1), 4) Adx rats receiving dexamethasone, 5) Adx rats receiving a low dose of aldosterone, and 6) Adx rats receiving a high dose of aldosterone (1.0 micrograms.100 g body wt-1.24 h-1). JHCO3 along the loop of Henle was decreased by 40% in Adx rats. JHCO3 was increased by dexamethasone alone and by dexamethasone plus a low dose of aldosterone to rates observed in fully supplemented Adx rats. Aldosterone given alone at a low physiological dose had no effect, but, when administered at a high dose, returned JHCO3 to normal.
Cloning of a rabbit renal Na-Pi cotransporter, which is regulated by dietary phosphate
Authors: Verri T. , Markovich D. , Perego C. , Norbis F. , Stange G. , Sorribas V. , Biber J. , Murer H.
Published: December 2017

Previously, we isolated a cDNA (NaPi-1) related to a rabbit renal proximal tubular Na-Pi cotransporter (A. Werner, M.L. Moore, N. Mantei, J. Biber, G. Semenza, and H. Murer. Proc. Natl. Acad. Sci. USA 88:9608-9612, 1991.). In this study, we isolated an additional (rabbit renal) cDNA (NaPi-6), which induces Na-dependent Pi uptake in Xenopus laevis oocytes. Substrate specificity and kinetic properties corresponded to those known for rabbit renal brush-border membrane (BBM) Na-Pi cotransport. NaPi-6 was cloned by homology using NaPi-2 cDNA, a rat renal BBM Na-Pi cotransporter (S. Magagnin, A. Werner, D. Markovich, V. Sorribas, G. Stange, J. Biber, and H. Murer. Proc. Natl. Acad. Sci. USA 90: 5979-5983, 1993). NaPi-6 encodes a protein of 642 amino acids, exhibiting at least eight transmembrane domains. NaPi-6 mRNA and protein in kidneys of rabbits fed a low-Pi diet (LPD; 0.11% Pi) for 1 wk were increased by 1.5- and 4-fold, respectively, compared with those of rabbits fed a high-Pi diet (HPD; 1.20% Pi). This effect was correlated with an increase in Na-Pi cotransport of BBM vesicles isolated from animals adapted to LPD (2.5-fold with respect to HPD). In contrast, NaPi-1 mRNA and protein were not altered in response to LPD. Thus rabbit proximal tubular BBMs contain two different Na-Pi cotransport systems: NaPi-1 (type I) and NaPi-6 (type II). Only the type II transport system seems to be under regulatory control in response to low-Pi dietary intake.
Modulation of intracellular Ca2+ by glucose in MDCK cells: role of endoplasmic reticulum Ca(2+)-ATPase
Authors: Lien Y. H. , Wang X. , Gillies R. J. , Martinez-Zaguilan R.
Published: December 2017

Intracellular free calcium ([Ca2+]i) has multiple functional roles in renal epithelia, including mediating ligand- and volume-activated K+ and Cl- channels, modulating the permeability of apical membrane to Na+, and regulating tubuloglomerular feedback. We investigated glucose effects on intracellular pH (pHi) and [Ca2+]i in Madin-Darby canine kidney (MDCK) cells using fluorescent probes, SNARF-1 and fura 2, respectively. The addition of glucose decreased both pHi and [Ca2+]i in a dose-dependent fashion. Thapsigargin (TG) and cyclopiazonic acid (CPA), well-known endoplasmic reticulum (ER) Ca(2+)-adenosinetriphosphatase (Ca(2+)-ATPase) inhibitors, abolished the glucose-induced [Ca2+]i decrease. Without glucose, 1 microM TG induced a sustained elevation in [Ca2+]i, which increased further with glucose addition, whereas 15 microM CPA induced a transient increase in [Ca2+]i that was not affected by further addition of glucose. The sustained elevation in [Ca2+]i induced by TG was dependent on extracellular Ca2+. TG-induced [Ca2+]i increase was modulated by glucose, i.e., at higher glucose concentrations, TG induced a larger and more rapid rise in [Ca2+]i. We conclude that glucose has dual effects on [Ca2+]i regulation. Glucose alone reduces [Ca2+]i by activating ER-type Ca(2+)-ATPase, since this phenomenon is TG and CPA sensitive. In the presence of TG, glucose increases [Ca2+]i probably by increasing Ca2+ entry. Our data suggest a model in which TG activates capacitative Ca2+ entry by depletion of the ER Ca2+ pool. Glucose increases TG-induced [Ca2+]i elevation by further enhancing capacitative Ca2+ entry.
Activation of protein kinase C stimulates cAMP phosphodiesterase in rat renal collecting tubule
Authors: Tetsuka T. , Kusano E. , Takeda S. , Homma S. , Yoshida I. , Ando Y. , Asano Y.
Published: December 2017

The present study was undertaken to evaluate whether protein kinase C (PKC) affects adenosine 3',5'-cyclic monophosphate phosphodiesterase (cAMP-PDIE) activity in microdissected rat renal medullary collecting tubules (MCT). Phorbol 12-myristate 13-acetate (PMA, 10(-8) M), an activator of PKC, significantly stimulated cAMP-PDIE activity in intact MCT (29.5 +/- 1.5 to 38.3 +/- 3.7 fmol.min-1.mm-1, P < 0.05) but not in the proximal straight tubule [4.7 +/- 0.8 vs. 4.8 +/- 0.8, not significant (NS)] or the medullary ascending limb of Henle's loop (20.5 +/- 2.1 vs. 22.4 +/- 2.8, NS). PMA-stimulated cAMP-PDIE activity was reversed by PKC inhibitors, staurosporine (10(-8) M) and calphostin C (10(-8) M), but not by the cyclooxygenase inhibitor, indomethacin (5 x 10(-6) M). 1,2-Dioctanoyl-sn-glycerol (50 micrograms/ml), a synthetic analogue of diacylglycerol that stimulates PKC, also increased cAMP-PDIE activity in broken-cell preparations from MCT. This stimulation was also suppressed by staurosporine (10(-8) M) and calphostin C (10(-8) M). The stimulatory effect of PMA on cAMP-PDIE activity was lost with rolipram (10(-4) M), a type IV PDIE inhibitor, whereas it was preserved with N-(6-amino-hexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) (10(-4) M), a calmodulin inhibitor, or vinpocetin (10(-4) M), a direct inhibitor of type I PDIE. From these results, we suggest that activation of PKC specifically stimulates rolipram-sensitive cAMP-PDIE, but not the calmodulin-sensitive isozyme, in rat MCT.
Coordinate regulation of rat renal parathyroid hormone receptor mRNA and Na-Pi cotransporter mRNA and protein
Authors: Kilav R. , Silver J. , Biber J. , Murer H. , Naveh-Many T.
Published: December 2017

Parathyroid hormone (PTH) acts on the kidney by binding to the PTH receptor, leading to a decrease in the active renal reabsorption of phosphate by the Na-Pi cotransporter, which is also independently activated by hypophosphatemia. We have studied the effects of hypo- and hyperparathyroidism and hypophosphatemia on PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. Both surgical parathyroidectomy and hypophosphatemia, which itself leads to hypoparathyroidism, led to an upregulation of the PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. Parathyroidectomized rats fed a low-Pi diet had an increase in PTH receptor and Na-Pi cotransporter mRNAs. Diet-induced hyperparathyroidism had no effect on PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. The effect of hypoparathyroidism and hypophosphatemia to increase both PTH receptor mRNA and Na-Pi cotransporter mRNA and protein shows that there is a tight coordinate regulation of these factors, which are both involved in PTH action.
The parietal sheet of Bowman's capsule of rat renal glomerulus: a target of endothelin and PAF
Authors: Marchetti J. , Meneton P. , Lebrun F. , Bloch-Faure M. , Rajerison R. M.
Published: December 2017

On the basis of intracellular calcium concentration ([Ca2+]i) measurements, we have previously reported that the parietal sheet of Bowman's capsule was sensitive to cholinergic agonists. The aim of the present work was to investigate whether this structure could be also a target of endothelin and platelet-activating factor (PAF), since we observed [Ca2+]i increases in response to both agonists in the glomerulus, but which were very different from that induced by carbachol. For this purpose, we measured [Ca2+]i on single microdissected parietal sheets, using a fura 2 microfluorescence technique and compared the effects of maximal concentrations of the three agonists (10(-7), 10(-8), and 10(-4) M for endothelin, PAF, and carbachol, respectively) under various experimental conditions. We observed that, like in the glomerulus, endothelin and PAF induced, in the parietal sheet, [Ca2+]i responses that differed in many respects from those found with carbachol. Thus, in the presence of 2 mM external calcium, 1) endothelin and PAF responses spontaneously declined to basal level, whereas a stationary plateau was observed after a sharp peak of [Ca2+]i with carbachol; 2) the magnitude of [Ca2+]i peak was smaller with endothelin and PAF than with carbachol; and 3) endothelin and PAF, but not carbachol, induced a homologous dose-dependent desensitization. Moreover, in the absence of external calcium, endothelin and PAF responses were smaller than carbachol response, although all three responses apparently resulted from release of calcium ions from the same internal pool. In additional experiments, we observed that, like carbachol, endothelin and PAF contracted the parietal sheet, which is only composed of myoepithelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)
ROMK inwardly rectifying ATP-sensitive K+ channel. II. Cloning and distribution of alternative forms
Authors: Boim M. A. , Ho K. , Shuck M. E. , Bienkowski M. J. , Block J. H. , Slightom J. L. , Yang Y. , Brenner B. M. , Hebert S. C.
Published: December 2017

The rat ROMK gene encodes inwardly rectifying, ATP-regulated K+ channels [K. Ho, C. G. Nichols, W. J. Lederer, J. Lytton, P. M. Vassilev, M. V. Kanazirska, and S. C. Hebert. Nature Lond. 362: 31–38, 1993; H. Zhou, S. S. Tate, and L. G. Palmer. Am. J. Physiol. 266 (Cell Physiol. 35): C809-C824, 1994], and mRNA encoding these channels is widely expressed in distal cortical and outer medullary nephron segments [see companion study; W.-S. Lee and S. C. Hebert. Am. J. Physiol. 268 (Renal Fluid Electrolyte Physiol. 37): F1124-F1131, 1995]. Using approaches based on homology to ROMK1, we have identified two additional ROMK isoforms, ROMK2b and ROMK3. Analysis of the nucleotide sequences of the ROMK isoforms indicates that molecular diversity of ROMK transcripts is due to alternative splicing at both the 5'-coding and 3'-noncoding regions. The splicing at the 5' end of ROMK gives rise to channel proteins with variable-length NH2 termini containing different initial amino acid sequences. Functional expression of these isoforms in Xenopus oocytes showed that they form functional Ba(2+)-sensitive K+ channels. The nephron distribution of mRNAs encoding alternatively spliced isoforms of ROMK (ROMK1-ROMK3) was investigated by reverse transcription-polymerase chain reaction (RT-PCR) of nephron segments dissected from rat kidney. Nondegenerate PCR primer pairs were designed to span at least one intron and to amplify specific alternatively spliced forms of ROMK.(ABSTRACT TRUNCATED AT 250 WORDS)
Angiotensin II regulates nephrogenesis and renal vascular development
Authors: Tufro-McReddie A. , Romano L. M. , Harris J. M. , Ferder L. , Gomez R. A.
Published: December 2017

To test the hypothesis that angiotensin II (ANG II) is necessary for normal embryonic and postnatal kidney development, the effect of angiotensin receptor blockade or angiotensin converting enzyme inhibition on nephrovascular development was studied in newborn Sprague-Dawley rats and in Rana catesbeiana tadpoles undergoing prometamorphosis. Blockade of ANG II type 1 receptor (AT1) in newborn rats induced an arrest in nephrovascular maturation and renal growth, resulting in altered kidney architecture, characterized by fewer, thicker, and shorter afferent arterioles, reduced glomerular size and number, and tubular dilatation. Inhibition of ANG II generation in tadpoles induced even more marked developmental renal abnormalities. Blockade of ANG II type 2 receptor (AT2) in newborn rats did not alter renal growth or morphology. Results indicate that ANG II regulates nephrovascular development, a role that is conserved across species.
Polarity and transport properties of rabbit kidney proximal tubule cells on collagen IV-coated porous membranes
Authors: Genestie I. , Morin J. P. , Vannier B. , Lorenzon G.
Published: December 2017

A high degree of functional polarity has been obtained in primary cultures of rabbit kidney proximal tubule cells grown on collagen IV-coated porous membranes. Tight confluency was attained 6 days after seeding and maintained for at least 6 more days, as shown by analysis of paracellular inulin diffusion. From day 6 onward, L-lactate, ammonia, and D-glucose concentration gradient and a pH difference of approximately 1 unit developed between the two nutrient medium compartments. Confluent monolayers expressed organic ion transport properties higher than those formerly reported for other cell models. Transcellular transport of 20 microM tetraethylammonium was directed from basal to apical compartment and was specifically inhibited by mepiperphenidol (1 mM). Unidirectional transport of 2.4 microM p-aminohippurate also occurred from basal to apical compartment, was saturable, and specifically inhibited by probenecid (1 mM). These results suggest that rabbit kidney proximal tubule cells, cultured under the experimental conditions described here, may be a useful model for the in vitro study of highly polarized renal transport processes.
Characterization of Na(+)-phosphate cotransporters in renal cortical endosomes
Authors: Loghman-Adham M. , Motock G. T. , Wilson P. , Levi M.
Published: December 2017

To determine the role of membrane recycling in proximal tubular P(i) transport, we studied the transport functions of simultaneously prepared rat renal cortical endosomal vesicles (EV) and brush-border membrane vesicles (BBMV). Initial P(i) uptake was Na+ gradient-dependent in both vesicles. Kinetic studies showed a lower apparent maximal rate (Vmax) for P(i) uptake in EV compared with BBMV (446 +/- 69 vs. 1493 +/- 117 pmol.mg protein-1 x 15 s-1, respectively; n = 4, P < 0.05), with no difference in the apparent Michaelis constant (Km). Endosomal Na(+)-P(i) cotransport was inhibited by phosphate analogues, phosphonoformic acid and arsenate, but not by the anion-exchange inhibitor, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Na(+)-P(i) cotransport was electroneutral in both vesicles, with a stoichiometry of 2 for BBMV and 1 for EV. The nonpermeant sulfhydryl reagent, 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), produced a marked inhibition of Na(+)-P(i) cotransport in EV but not in BBMV, suggesting accessible sulfhydryl groups, an "inside-out" orientation in EV, and "right-side-out" orientation in BBMV. The EV and the BBMV differed significantly in their phospholipid composition and lipid fluidity. The Na(+)-P(i) cotransporter protein (NaPi-2) abundance, determined by Western blots, was five times lower in EV than in BBMV (0.25 +/- 0.05 vs. 1.36 +/- 0.20 arbitrary units). Renal cortical endosomes contain Na(+)-P(i) cotransporters, albeit at a lower density, suggesting involvement of membrane recycling in the regulation of proximal tubular P(i) transport.
Magnitude of TGF-initiated nephron-nephron interactions is increased in SHR
Authors: Chen Y. M. , Yip K. P. , Marsh D. J. , Holstein-Rathlou N. H.
Published: December 2017

We compared the tubuloglomerular feedback (TGF)-initiated nephron-nephron interaction in spontaneously hypertensive rats (SHR) and normotensive Sprague-Dawley (SD) rats. Interaction strength was assessed by measuring stop-flow pressure (delta SFP) responses in pairs of nephrons, where only one nephron of the pair was microperfused. The vascular connection was determined from casts of the nephrons and vessels; length of arteriolar separation between the two glomeruli was measured on the casts. When microperfusion rate was increased from 5 to 50 nl/min, delta SFP in perfused nephrons was 10.6 +/- 0.6 and 10.2 +/- 0.7 mmHg [not significant (NS)] in SD and SHR, respectively. In the matched unperfused nephrons from the same cortical radial artery, delta SFP was 1.3 +/- 0.2 and 2.9 +/- 0.7 mmHg (P < 0.05) in SD and SHR. When the perfused and unperfused nephron originated from different cortical radial arteries, delta SFP in the unperfused nephrons was -0.1 +/- 0.2 and 0.0 +/- 0.3 mmHg (NS) in SD and SHR, respectively. In both strains, interaction strength varied inversely with glomerular separation. When the dependence on distance was taken into account, interaction strength was about threefold greater in SHR than in SD. We conclude that the nephron-nephron interaction is significantly greater in SHR. The dependence of interaction strength on distance separating the glomeruli suggests that the interaction is propagated along the preglomerular vasculature.
Effects of chronic Cl depletion alkalosis on proximal tubule transport and renal production of ammonium
Authors: DuBose T. D. , Good D. W.
Published: December 2017

The role of renal ammonium excretion in the maintenance of chronic metabolic alkalosis is poorly defined, particularly under conditions in which the alkalosis is associated with secondary potassium depletion. Therefore, free-flow micropuncture experiments were performed to examine the effects of chronic chloride depletion metabolic alkalosis (CDAlk) on renal ammonium production, urinary ammonium excretion, and proximal convoluted tubule (PCT) ammonium transport in the rat in vivo. CDAlk was generated by peritoneal dialysis against NaHCO3 and maintained for 6–7 days by dietary Cl- restriction. Pair-fed controls were dialyzed against NaCl. Rats with CDAlk had elevated plasma HCO3- concentration, hypokalemia, and hypochloremia. HCO3- excretion was negligible in both control and CDAlk rats. Glomerular filtration rate and urine pH did not differ. CDAlk reduced urinary ammonium excretion by 35% but had no significant effect on whole kidney ammonium production. Net secretion of ammonium by the PCT was decreased by 70% and absolute delivery of ammonium out of the PCT was decreased by 55% in the CDAlk rats. The decrease in PCT ammonium secretion was the combined result of a decrease in net ammonium secretion along the early PCT and an increase in net ammonium absorption along the late PCT.(ABSTRACT TRUNCATED AT 250 WORDS)