bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of an aldose reductase inhibitor on organic osmotic effectors in rat renal medulla
Authors: Yancey P. H. , Haner R. G. , Freudenberger T. H.
Published: December 2017

Renal medullary cells use sorbitol, betaine, and other organic compounds as osmotic effectors (osmolytes) to balance high extracellular NaCl. Excess sorbitol is also implicated in diabetes complications in several organs including kidneys. To study regulation of renal sorbitol, male Wistar rats were given an aldose reductase inhibitor, sorbinil, at 40 mg.kg-1.day-1 in food to block sorbitol formation from glucose. Inner medullas of kidneys were analyzed for osmolytes by high-performance liquid chromatography and atomic absorption. Animals on sorbinil had significantly reduced medullary sorbitol contents in a group on ad libitum water for 10 days (2.7 mmol/kg wet wt compared with 4.8 in controls) and in an antidiuretic group kept 7 days and an additional 3 days without water (3.8 mmol/kg wet wt compared with 7.2 in antidiuretic controls). In both groups, betaine contents were significantly elevated (9.2 mmol/kg wet wt compared with 5.5 in ad libitum water controls: 6.4 mmol/kg wet wt compared with 4.2 in antidiuretic controls). No other osmolytes differed. Total contents of nonurea organic osmolytes maintained a constant ratio to sodium contents; thus increased betaine concomitant with decreased sorbitol may have maintained constant cell volume. In contrast, in animals kept 21 days on sorbinil, there were significant decreases in urea and inositol contents. However, there were no significant differences in sorbitol or betaine compared with controls, suggesting a compensating increase in sorbitol production or in sorbinil removal.
Ion concentrations in the rat CCD: differences between cell types and effect of alkalosis
Authors: Gifford J. D. , Galla J. H. , Luke R. G. , Rick R.
Published: December 2017

We have previously shown that the isolated perfused cortical collecting duct (CCD) from chloride-depleted alkalotic (CDA) rats continues to secrete HCO3 for up to 3 h. To determine whether the sustained alteration in transport was associated with changes in intracellular ion concentrations, we performed energy-dispersive X-ray microanalysis in microdissected tubule bundles obtained from CDA rats and rats with normal acid-base status (CON). Before analysis, the bundles from both groups were incubated for 1 h in vitro in a modified Ringer solution (pH 7.4, 105 mM Cl). Principal (PC) and intercalated cells (IC) of the CCD from CON animals differed in the nuclear concentration of Na (17.0 vs. 24.7 mmol/l cell water), K (192.5 vs. 177.0 mmol/l cell water), and Cl (17.8 vs. 47.8 mmol/l cell water). Cells of the cortical thick ascending limb of Henle (CTAL) had the lowest Na and Cl values (11.5 and 14.8 mmol/l cell water, respectively). CDA resulted in no systematic Cl changes. In the IC the nuclear Na concentration was significantly increased (32.0 vs. 24.7 mmol/l cell water) and in all cells a small reduction in K concentration was detectable. These findings suggest that 1) the different transport functions of IC, PC, and CTAL are associated with differences in the intracellular ion composition, and 2) the sustained HCO3 secretion seen in CCD from CDA rats cannot be explained as the result of intracellular Cl depletion.
Extracellular matrix receptors in the kidney cortex
Authors: Simon E. E. , McDonald J. A.
Published: December 2017

Extracellular matrix (ECM) receptors anchor cells to substratum and impart positional information to cells. Within the group of ECM receptors known as integrins, alpha-subunits of these alpha beta heterodimers define ligand specificity, whereas beta-subunits define the subclass. We used immunofluorescence with anti-ECM receptor antibodies to examine distribution within human kidney cortex of all known alpha-subunits in the beta 1 subclass of integrins as well as a non-integrin 67-kDa elastin/lamin receptor. The alpha 1-subunit (alpha 1 beta 1 defines a collagen receptor) was present in mesangium and base of all tubule epithelial cells; alpha 2 (collagen) was present in mesangium and in distal but not proximal tubule cells; alpha 3 (collagen, laminin, fibronectin) was diffusely distributed within glomeruli but tubule staining was less intense; alpha 4 (fibronectin) was absent; alpha 5 (fibronectin) was present in blood vessels; and alpha 6 (laminin) was present along basolateral aspect of all tubule cells but absent in glomeruli. The elastin/laminin receptor was present in all tubule epithelial cells, but staining was heavier in distal tubules, especially intercalated cells. Thus striking heterogeneity in ECM receptor distribution was noted. For collagen receptors, differences in tubule staining were pronounced. Despite the presence of laminin within both glomeruli and tubules, laminin receptors also showed marked differences in staining between these structures. Both differences in ECM structure and intrinsic differences among different cells may underlie these differences in ECM receptor distribution.
Maturation of HCO3- transport in rabbit collecting duct
Authors: Mehrgut F. M. , Satlin L. M. , Schwartz G. J.
Published: December 2017

Newborn rabbits maintain a state of hypochloremic metabolic alkalosis with plasma HCO3- concentrations generally exceeding 27 mM. The large amounts of potential net base in mother's milk probably contribute to the generation of this alkalosis. We surmised that immature handling of H+ or HCO3- by the neonatal collecting duct helps maintain this alkalosis. Net HCO3- transport was measured in perfused collecting ducts taken from maturing rabbits, in solutions simulating adult rabbit plasma ultrafiltrate and then in presence of Cl(-)-free bathing solution. Cortical collecting ducts (CCD) from newborn and 4-wk-old rabbits failed to secrete HCO3- under baseline conditions and could not be stimulated to secrete HCO3- in Cl(-)-free bath. Neonatal segments perfused at very slow flow rates showed significant HCO3- absorption; inhibition of HCO3- secretion by removal of luminal Cl- revealed a substantial HCO3- absorptive flux. Segments from 6-wk-old and mature animals secreted net HCO3- and generally showed more than a fivefold increase in HCO3- secretion after Cl- removal from the bath. Outer medullary collecting ducts (OMCD) from newborn rabbits absorbed HCO3- at rates approaching that of mature segments. We conclude that relatively high rates of HCO3- absorption in OMCD and lack of HCO3- secretion in CCD may contribute to the metabolic alkalosis of the neonatal rabbit.
Inhibition of proximal tubule Na(+)-K(+)-ATPase activity requires simultaneous activation of DA1 and DA2 receptors
Authors: Bertorello A. , Aperia A.
Published: December 2017

This study examines the receptor mechanisms by which dopamine (DA) inhibits Na(+)-K(+)-adenosinetriphosphatase (ATPase) activity in single permeabilized proximal tubule (PCT). Na(+)-K(+)-ATPase activity was inhibited in the presence of both DA1- and DA2-specific agonists but not by either agonist alone. The inhibition induced by DA (10(-6) M) was attenuated in the presence of either of the two DA2-specific antagonists S-sulpiride and YM 09151 at 10(-5) M and in the presence of the DA1 antagonist SCH 23390 (10(-5) M). PCT adenosine 3',5'-cyclic monophosphate (cAMP) levels were significantly increased in the presence of DA and DA1 agonist, but DA2 agonist had no effect on cell cAMP levels. Na(+)-K(+)-ATPase activity was significantly inhibited in PCT incubated with DA2 agonist (10(-5) M) and dibutyryl (DB)-cAMP (10(-6) M) but not with DA2 agonist (10(-5) M) only. PCT Na(+)-K(+)-ATPase activity was also significantly inhibited in the presence of both DA2 agonist (10(-5) M) and forskolin (10(-6) M). Neither DBcAMP (10(-6) M) nor forskolin (10(-6) M) alone inhibited Na(+)-K(+)-ATPase activity. In tubules incubated with DA (10(-8) to 10(-9) M), the presence of DBcAMP (10(-6) M) enhanced the sensitivity by which DA inhibited Na(+)-K(+)-ATPase activity. We conclude that PCT Na(+)-K(+)-ATPase activity is inhibited by a synergistic action of the DA1 and DA2 receptors, with the DA1 receptor acting to increase cell cAMP levels.
Phosphate depletion increases cytosolic calcium of brain synaptosomes
Authors: Massry S. G. , Hajjar S. M. , Koureta P. , Fadda G. Z. , Smogorzewski M.
Published: December 2017

Phosphate depletion (PD) is associated with a rise in resting levels of [Ca2+]i in pancreatic islets. It is not known whether this derangement occurs in other cells, and the mechanisms by which PD affects [Ca2+]i have not been delineated. This study examined the effect of PD on [Ca2+]i of brain synaptosomes and evaluated potential mechanisms that may lead to rise in their [Ca2+]i. [Ca2+]i levels in synaptosomes of PD rats (460 +/- 18.3 nM) were higher (P less than 0.01) than those of pair-weighed (PW) rats (358 +/- 12.5 nM). Verapamil treatment of PD rats (PD-V) normalized [Ca2+]i in their synaptosomes (361 +/- 8.5 nM). In verapamil-treated PW rats (PW-V), synaptosomal [Ca2+]i (359 +/- 8.3 nM) was not affected. ATP content and Na(+)-K(+)-ATPase activity of synaptosomes were lower (P less than 0.01) in PD rats than in PW, PD-V, and PW-V rats. The values of these parameters from the latter three groups were not different. Km of synaptosomal Ca2(+)-ATPase was not affected by PD but Vmax of this enzyme (2.5 +/- 0.33 mumol Pi.mg protein-1.h-1) was lower (P less than 0.05) than in PW (5.4 +/- 0.66 mumol Pi.mg protein-1.h-1), PD-V, and PW-V rats. Our data indicate that PD raises [Ca2+]i in brain synaptosomes and suggest that PD increases calcium entry into synaptosomes. This would inhibit mitochondrial ATP production, with a consequent fall in ATP content of synaptosomes.(ABSTRACT TRUNCATED AT 250 WORDS)
Angiotensin II stimulates ammoniagenesis in canine renal proximal tubule segments
Authors: Chobanian M. C. , Julin C. M.
Published: December 2017

To determine whether angiotensin II (ANG II) affects ammoniagenesis in renal proximal tubule, ammonia production was measured in suspensions of canine renal proximal tubule segments (PCT) incubated with L-glutamine and varying concentrations of ANG II. Ammonia production from PCT was significantly increased by 15.5 +/- 1.1% in the presence of ANG II (10(-6) M) at 2 h. Similarly, glucose production significantly increased by 10.0 +/- 0.9%. Half-maximal stimulation occurred at approximately 10(-9) M ANG II. Stimulation of ammonia production by ANG II was blocked in the presence of the ANG II antagonist, [Sar1-Ile8]ANG II (10(-6) M). Enhancement of ammonia production in PCT by ANG II occurred in acidotic and neutral media but not in alkalotic medium. When extracellular [Na+] = intracellular [Na+] ANG II significantly increased ammonia production in PCT. Absence of extracellular Ca2+ or addition of trifluoperazine or N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) (Ca2(+)-calmodulin-dependent pathway inhibitors) blocked the action of ANG II to enhance ammonia production. We conclude that ANG II stimulates ammonia and glucose production in canine renal PCT via a receptor-mediated signal. The action of ANG II on ammoniagenesis may be mediated by a calcium-calmodulin-dependent pathway. Stimulation of ammoniagenesis in vitro under normal and acidotic conditions may reflect a role in vivo for ANG II in the regulation of renal acid-base metabolism.
Role of endogenous endothelin on renal functions in rats
Authors: Yamada K. , Yoshida S.
Published: December 2017

This study was conducted to determine the involvement of endogenous endothelin (ET), a novel potent vasoconstricting peptide, in systemic and renal hemodynamics and in the renin-angiotensin system by inhibiting ET action via infusion of a specific ET antiserum at a time of altered sodium balance. Infusion of 1:50 diluted ET antiserum, which completely inhibited renal vasoconstriction by the exogenously administered ET (0.25 to 1.0 nmol/kg), caused an increase in urinary sodium excretion and fractional excretion of sodium and a decrease in plasma renin concentration without significant changes in blood pressure, heart rate, glomerular filtration rate, renal plasma flow, and urine volume compared with the values with nonimmune serum in conscious rats fed a low-salt diet. A time control study showed no significant changes in all parameters. These results suggest that the state of low- compared to high-salt intake causes a relatively stronger activity of endogenous ET, and that the endogenous ET contributes to the adaptative modulations of sodium excretion via renal tubular action and renin release in association with the changed state of sodium balance.
Tubuloglomerular feedback dynamics and renal blood flow autoregulation in rats
Authors: Holstein-Rathlou N. H. , Wagner A. J. , Marsh D. J.
Published: December 2017

To decide whether tubuloglomerular feedback (TGF) can account for renal autoregulation, we tested predictions of a TGF simulation. Broad-band and single-frequency perturbations were applied to arterial pressure; arterial blood pressure, renal blood flow and proximal tubule pressure were measured. Data were analyzed by linear systems analysis. Broad-band forcings of arterial pressure were also applied to the model to compare experimental results with simulations. With arterial pressure as the input and tubular pressure, renal blood flow, or renal vascular resistance as outputs, the model correctly predicted gain and phase only in the low-frequency range. Experimental results revealed a second component of vascular control active at 100-150 mHz that was not predicted by the simulation. Forcings at single frequencies showed that the system behaves linearly except in the band of 33-50 mHz in which, in addition, there are autonomous oscillations in TGF. Higher amplitude forcings in this band were attenuated by autoregulatory mechanisms, but low-amplitude forcings entrained the autonomous oscillations and provoked amplified oscillations in blood flow, showing an effect of TGF on whole kidney blood flow. We conclude that two components can be detected in the dynamic regulation of renal blood flow, i.e., a slow component that represents TGF and a faster component that most likely represents an intrinsic vascular myogenic mechanism.
Single-nephron filtration rate and proximal reabsorption in aging rats
Authors: Corman B. , Roinel N.
Published: December 2017

Age-related changes in the function of individual nephrons were investigated by micropuncture experiments measuring single-nephron filtration rates (SNGFR) and proximal reabsorptions in 10-, 20-, and 30-mo-old rats. The animals were female WAG/Rij rats with low incidence of chronic progressive nephropathy, no loss of nephrons, and renal hypertrophy of both kidneys in the oldest animals. Mean SNGFR values per gram kidney weight were 41.4 +/- 1.1, 37.1 +/- 1.5, and 32.2 +/- 1.1 nl.min-1.g kidney wt-1 (n = 41) in the 10-, 20-, and 30-mo-old animals, respectively. This age-related decrease in filtration was no longer apparent when SNGFR values were expressed per nephron (means 24.3 +/- 0.7, 23.7 +/- 0.9, and 24.4 +/- 0.9 nl/min. Individual filtered loads of sodium, potassium, calcium, and magnesium and their absolute reabsorption by the proximal tubule were not different in the three age groups; however, absolute and fractional reabsorptions of phosphate decreased significantly in the 30-mo-old rats. These results indicate that, with the exception of phosphate, individual filtrations and proximal reabsorptions are well maintained in aging rats free of disease. This may be related to the observed renal hypertrophy.
Inhibitory effect of angiotensin II on renal tubular transport
Authors: Chatsudthipong V. , Chan Y. L.
Published: December 2017

This study was designed to examine the intracellular mechanism of inhibitory action of high concentration of angiotensin II (ANG II) on proximal tubular transport in rat kidneys by microperfusion methods. Perfusion of ANG II (10(-6) M) to peritubular capillaries caused a reduction of both fluid and HCO3- transport (Jv and JHCO3-, respectively) by 33 and 26%, respectively. These inhibitory effects were blocked by the ANG II-receptor antagonist [Sar1, Ile8]ANG II (10(-5) M). Similar degrees of inhibition on Jv and JHCO3- were observed when ionomycin (10(-7) and 10(-6) M), a Ca2+ ionophore, was added to capillary perfusate. Moreover, there was no additive effect when both ANG II and ionomycin were perfused together through capillaries, suggesting that both agents work via the same mechanism, presumably by increasing cytosolic Ca2+ concentration ([Ca2+]i). Inhibitory effects of ANG II on proximal tubular transport were still observed in a Ca2(+)-free perfusate containing ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid, indicating that these effects do not require influx of Ca2+ from extracellular medium. Furthermore, the observation that TMB-8, an agent that prevents intracellular Ca2+ mobilization, completely eliminated the effect of ANG II strongly suggests that intracellular Ca2+ rather than Ca2+ influx mediates effects of ANG II on proximal tubular transport. Direct measurement of [Ca2+]i by use of fura-2 in isolated proximal tubular cells showed slight but statistically significant increases in [Ca2+]i. Taken together, these observations support the idea that intracellular Ca2+ serves as a second messenger in the inhibitory effect of high concentrations of ANG II on Jv and JHCO3- in proximal tubule of kidney.
Transient ischemia or heat stress induces a cytoprotectant protein in rat kidney
Authors: Emami A. , Schwartz J. H. , Borkan S. C.
Published: December 2017

Sublethal heat exposure induces the production of heat stress protein (HSP) 72 kDa, a reported cytoprotectant, in several tissues including the rat kidney. However, the localization and time course of HSP 72 accumulation in the kidney in response to heat or other cell stresses such as transient ischemia have not been described. In anesthetized rats exposed to either 42 +/- 0.5 degrees C (heat stress) or 37 degrees C (sham) for 15 min, accumulation of HSP 72 in kidney homogenates, detected by immunoblot analysis using a specific monoclonal anti-HSP 72 antibody, peaked 4-6 h after heat stress and persisted for 10 days. HSP 72 appeared rapidly in renal papilla within 1 h and in medulla and cortex within 4 h after heat stress. No HSP 72 was detected in tissues from sham heat stress. HSP 72 was also detected within 3 h of at least 15 min of renal ischemia in situ. Accumulation was maximal after 60 min of ischemia and persisted for 5 days, whereas no HSP 72 was detected after 90 min of ischemia or in the contralateral nonischemic kidney at any time point. This study demonstrates that transient ischemia, like heat stress, results in the rapid cytosolic accumulation of HSP 72, a known cytoprotectant that may be important in mediating cell repair or increasing resistance to subsequent injury.
Cyclosporin inhibits phosphate transport and stimulates alkaline phosphatase activity in renal BBMV
Authors: Demeule M. , Beliveau R.
Published: December 2017

The activity of various enzymes and transport systems was studied in renal brush-border membrane vesicles (BBMV) isolated from rats injected daily with cyclosporin. Alkaline phosphatase (AP) was strongly stimulated: 55 and 113% increases were obtained in BBMV isolated from rats injected with 10 mg cyclosporin/kg for 5 and 10 days. The affinity of the enzyme remained unaltered, but maximal activity (Vmax) showed a strong increase of 2.4-fold between control and treated animals. In addition to the phosphatase activity, phosphate binding to AP also showed a dose-dependent stimulation by cyclosporin treatment: 44 and 70% increases in animals treated for 5 days with 5 and 10 mg cyclosporin/kg. However, the activity of aminopeptidase M was not affected by these treatments, and polyacrylamide gel electrophoresis of BBMV revealed no alterations in the profile of membrane proteins, suggesting the specificity of cyclosporin interaction with alkaline phosphatase. Na(+)-dependent amino acid and D-glucose transport systems remained unaffected by cyclosporin treatment. The Na(+)-independent transport system for lysine and the Na(+)-H+ antiporter activity were also unaltered. In contrast, the initial rate of phosphate uptake decreased by 28% after administration of cyclosporin (10 mg/kg) for 5 days: the Michaelis constant (Km) and Vmax decreased from 137 to 85 microM and from 1.49 to 1.07 pmol.micrograms-1.5 s-1, respectively. "In vitro" studies with membranes isolated from untreated rats were also undertaken by preincubating membranes with cyclosporin. Neither alkaline phosphatase nor the transport systems were affected under these conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
Aldosterone metabolism in combined isolated perfused rat liver and kidney
Authors: Egfjord M. , Daugaard H. , Olgaard K.
Published: December 2017

The metabolism of aldosterone (Aldo) at 4 nM was studied by radioimmunoassay and by high-performance liquid chromatography in isolated perfused liver (IPL), isolated perfused kidney (IPK), and combined isolated perfused liver and kidney (CIPLK) of male Wistar rats. Effect of D[4-(14)C]aldosterone (D [4-(14)C]Aldo) on the function of IPK of intact and adrenalectomized (ADX) rats was also studied. Aldo clearance in the liver was most important, 16.3 +/- 1.7 ml/min. In IPK, the total clearance of Aldo was 0.27 +/- 0.36 ml/min (39% of the glomerular filtration rate) (GFR). Fractional excretion (FE) of Aldo was 16 +/- 8%. Metabolic clearance of Aldo was (0.21 +/- 0.23 ml/min), 78% of total renal clearance. In CIPLK, the kidney inhibited hepatic clearance of Aldo by 23% when compared with IPL (P less than 0.05). Hepatic Aldo metabolites were predominantly eliminated by biliary excretion of polar metabolites. Several hepatic polar metabolites and tetrahydroaldosterone (THA) accumulated in perfusate and were excreted in the urine in a similar pattern. After hydrolysis of the polar metabolites, some coeluated with THA and dihydroaldosterone (DHA), whereas other metabolites remained more polar than Aldo. Without addition of Aldo, in IPK of ADX rats FENa was higher (P less than 0.01), and FEK was lower (P less than 0.01), resulting in three- to fourfold higher urinary Na-K ratio (P less than 0.01) when compared with IPK and CIPLK of intact rats. In IPK of ADX rats with Aldo in perfusate, only FEK was restored. Addition of Aldo to IPK of intact rats had no effect. However, only in CIPLK, addition of Aldo resulted in an increasing kaliuresis in three subsequent periods of 30 min (0.56-0.95, P less than 0.01). Thus the hepatic metabolites of Aldo could in part mediate the kaliuretic effect of Aldo.
ANP stimulates phospholipase C in cultured RIMCT cells: roles of protein kinases and G protein
Authors: Berl T. , Mansour J. , Teitelbaum I.
Published: December 2017

We examined the possibility that, in addition to stimulation of guanylate cyclase (GC), atrial natriuretic peptide (ANP) also activates phospholipase C (PLC) in cultured rat inner medullary collecting tubule (RIMCT) cells. ANP (10(-12)M) causes marked release of inositol trisphosphate (IP3) at a concentration that does not stimulate GC. Concentrations of ANP that stimulate GC (greater than or equal to 10(-10) M) result in attenuated IP3 release. Similarly, exogenous dibutyryl guanosine 3',5'-cyclic monophosphate (10(-6) M) markedly inhibits the response to 10(-10) M ANP. Inhibition of cyclic nucleotide-dependent protein kinase by H 8, but not inhibition of protein kinase C by H 7, restores the response to 10(-8) and 10(-6) M ANP. Therefore, activation of cyclic nucleotide-dependent protein kinase inhibits ANP-stimulated PLC activity. Activation of protein kinase C by phorbol 12-myristate-13-acetate (PMA) decreases ANP-stimulated IP3 production. Pretreatment with H 7, but not H 8, prevents inhibition by PMA. To explore a potential role for G proteins, we examined the effect of guanine nucleotide analogues on ANP-stimulated IP3 production in saponin-permeabilized cells. ANP-stimulated IP3 production is enhanced by GTP gamma S and is inhibited by GDP beta S. Similarly, preincubation with pertussis toxin prevents ANP-stimulated IP3 release. We conclude that ANP stimulates PLC in RIMCT cells via a pertussis toxin-sensitive G protein. Stimulation of PLC is inhibited on activation of either cyclic nucleotide or Ca2+-phospholipid dependent protein kinases.
Effects of purine derivatives on calcitriol metabolism in rats
Authors: Hsu C. H. , Patel S. R. , Young E. W. , Vanholder R.
Published: December 2017

The effects of theophylline and sodium urate on metabolic production (PR) and clearance rate (MCR) of calcitriol were determined by the constant isotope infusion method in normal rats. Calcitriol PR was significantly reduced after infusion for 20 h of either theophylline (1 mg/h, PR = 22.3 +/- 1.6 ng.kg-1.day-1, P less than 0.001, n = 5) or sodium urate (0.5 mg/h, PR = 18.6 +/- 1.2 ng.kg-1.day-1, P less than 0.001, n = 5) compared with control rats infused with saline (PR = 32.0 +/- 1.5 ng.kg-1.day-1, n = 5). Renal 1 alpha-hydroxylase activity of kidney homogenate was significantly inhibited in rats infused with theophylline or urate. Suppression of 1 alpha-hydroxylase activity was also observed when the kidney homogenate was preincubated for 3 h with various concentrations of xanthine (0.11-3.0 mg/dl). In addition, the MCR of calcitriol was decreased in rats infused with either theophylline (MCR = 21.0 +/- 0.88 microliter.min-1.100 g-1, P less than 0.005) or urate (MCR = 22.9 +/- 0.91 microliter.min-1.100 g-1, P less than 0.05) compared with saline-infused control rats (MCR = 25.2 +/- 0.41 microliter.min-1.100 g-1). Because calcitriol degradation is a receptor-mediated process that requires binding of the receptor-hormone complex to chromatin, we studied the binding affinity of labeled calcitriol receptor for DNA-cellulose in the presence of theophylline or urate. Both theophylline and urate inhibited receptor binding affinity for DNA-cellulose. We conclude that these purine derivatives suppress calcitriol synthesis and inhibit receptor binding affinity for DNA. The altered receptor binding affinity could explain the decreased MCR of calcitriol.
Sorbitol permease: an apical membrane transporter in cultured renal papillary epithelial cells
Authors: Garty H. , Furlong T. J. , Ellis D. E. , Spring K. R.
Published: December 2017

The efflux of sorbitol from the rabbit papillary epithelial cell line PAP-HT25 occurs through a specific transport pathway, which we denote the "sorbitol permease." The permease was studied by measuring cell volume changes that accompanied osmotic swelling and by determination of the sorbitol efflux from plasma membrane vesicles. The cell volume studies showed that sorbitol efflux in response to hypotonicity occurred only across the apical membrane of the cells and that loss of sorbitol was the primary mechanism for regulatory volume decrease (RVD) by these cells. Quinidine, a permeant inhibitor of the sorbitol permease, was shown to prevent RVD when added to either apical or basolateral bathing solution. Cell volume experiments also showed that the permease was present only on the apical membrane of cells that had been grown in isotonic medium and did not accumulate sorbitol. The permease could be demonstrated in membrane vesicles obtained from cells exposed to a hypotonic environment before being homogenized. Quinidine blocked the sorbitol efflux from vesicles indicating that it either directly inhibited the permease or a membrane-associated activation step.
Glomerular dysfunction in nephrotic humans with minimal changes or focal glomerulosclerosis
Authors: Guasch A. , Hashimoto H. , Sibley R. K. , Deen W. M. , Myers B. D.
Published: December 2017

Fractional clearances (theta) of uncharged dextrans (radii 28-60 A) were used to characterize glomerular dysfunction in 34 nephrotic humans with either minimal-change nephropathy (MCN) or focal, segmental glomerulosclerosis (FSGS). A theoretical analysis of theta of dextran with a heteroporous membrane model revealed a similar alteration, more marked in FSGS than MCN. The number of restrictive pores perforating the major membrane component was reduced in parallel with the prevailing glomerular filtration rate (GFR). Parallel shuntlike pores in the remaining membrane component were more prominent, pointing to impaired size selectivity. However, the theta of large (60 A) dextrans attributable to these shunts exceeded control in FSGS only, suggesting that coexistent impairment of charge selectivity contributed importantly to the proteinuria in MCN. Membrane properties returned toward normal when MCN remitted. Glomerular morphometry revealed the frequency of epithelial filtration slits to vary with the extent of membrane dysfunction. Despite offsetting hypertrophy of remnant glomeruli in FSGS, a loss of filtration surface due to sclerosis likely contributed to the more marked reductions in pore number and GFR observed in this disorder than in MCN.
Sublethal X-irradiation during acute puromycin nephrosis prevents late renal injury: role of macrophages
Authors: Diamond J. R. , Pesek-Diamond I.
Published: December 2017

Acute puromycin aminonucleoside (PA) nephrosis is associated with a surge in the glomerular macrophage number. Whole body X-irradiation (XI) was used to assess whether bone marrow depression, which depletes normal rat glomeruli of macrophages, is efficacious in ameliorating progressive glomerular disease. A solitary, sublethal dose of XI (600 rad) was administered 3 days after PA delivery to rats (PA/XI) that were followed for 18 wk. In contrast to a sham-irradiated nephrotic cohort (PA/Sham), the PA/XI rats had a complete prevention of the recurrent albuminuria and manifested a significant reduction in the percent of glomeruli exhibiting glomerulosclerosis (GS) lesions at 18 wk after PA. XI significantly reduced the glomerular and interstitial macrophage number as well as the circulating white blood and monocyte counts during peak albuminuria. This protection was independent of the magnitude of the acute albuminuria and elevations in the circulating lipid levels. These data show that whole body XI delivered during acute nephrosis is capable of preventing progressive glomerular disease in association with a reduction in the glomerular and interstitial macrophage number. Additionally, this study suggests that this immune effector cell is important in mediating the propagation of initial glomerular injury to GS.
Basolateral ionic permeabilities of macula densa cells
Authors: Lapointe J. Y. , Bell P. D. , Hurst A. M. , Cardinal J.
Published: December 2017

It has recently been shown that membrane ionic transport pathways of macula densa cells can be measured using conventional microelectrodes. To determine if conductances could be identified at the basolateral membrane of macula densa cells, cortical thick ascending limbs (CTAL) with attached glomeruli were continuously perfused with a 25 mM NaCl bicarbonate-free Ringer solution. Individual basolateral Na+, Cl-, NaCl, and K+ concentrations were altered by isosmotic replacement with N-methyl-D-glucamine and/or cyclamate. Reduction in basolateral [Na+] from 150 to 25 mM hyperpolarized basolateral membrane potential (Vbl) by 9.9 +/- 1.3 mV (n = 10; all data are corrected for changes in liquid junction potential at bath electrode). A decrease in bath [Cl-] from 150 to 25 mM depolarized Vbl by 20 +/- 2.4 mV (n = 13), whereas decreases in bath [NaCl] from 150 to 25 mM depolarized Vbl by 29 +/- 6.8 mV (n = 5). In the presence of 150 mM NaCl bathing solution, a stepwise increase in [K+] from 5 to 15 mM (by replacement of 10 mM NaCl with 10 mM KCl) depolarized Vbl by 3.3 +/- 1.1 mV (n = 8). After correction for individual transepithelial diffusion potentials, Cl conductance averaged 59 +/- 19% of the total basolateral conductance, whereas K+ (23 +/- 8%) and Na+ (17 +/- 10%) contributed significantly less to the overall basolateral conductance. These results indicate that membrane potential of macula densa cells may be very sensitive to alterations in intracellular Cl- activity and suggest that apical transport of NaCl through a furosemide-sensitive Na(+)-K(+)-2Cl- transporter may affect membrane potential in macula densa cells via a change in intracellular Cl- activity.
Muscarinic receptor activation inhibits AVP-induced water flow in rabbit cortical collecting ducts
Authors: Snyder H. M. , Fredin D. M. , Breyer M. D.
Published: December 2017

We examined the effect of carbachol, an acetylcholine analogue, on hydraulic conductivity (Lp) response to 10 microU/ml arginine vasopressin (AVP) in rabbit cortical collecting duct (CCD). In CCDs in which water flow had been established with AVP, subsequent addition of carbachol caused Lp (X10(-7) cm.atm-1.s-1) to fall from 251 +/- 32 to 146 +/- 19. Carbachol washout resulted in recovery of Lp to 217 +/- 38. In CCDs in which water flow had been established using 10(-4) M 8-chlorophenylthioadenosine 3',5'-cyclic monophosphate (8-CPT-cAMP), addition of carbachol had no effect. These posttreatment studies suggest that carbachol's effects on modulating established water flow occur at a "pre-cAMP" step. With carbachol added first, AVP-induced Lp was reduced from 233 +/- 24 (controls) to 105 +/- 19 (carbachol-pretreated). Pretreatment with 10(-6) M atropine, a muscarinic receptor antagonist, totally reversed the inhibitory effect of carbachol, consistent with a receptor-mediated effect of carbachol. Carbachol pretreatment also inhibited 8-CPT-cAMP-induced Lp, indicating that carbachol's effects also occur at a "post-cAMP" step. Pretreatment with 10(-7) M staurosporine, a protein kinase C (PKC) inhibitor, reversed inhibitory effect of carbachol on AVP-induced Lp (193 +/- 26), suggesting that carbachol's effects are mediated by PKC. Intracellular calcium concentration [( Ca2+]i) was measured in fura-2-loaded CCDs. Carbachol also increased [Ca2+]i from 229 +/- 120 to 389 +/- 160 nM.(ABSTRACT TRUNCATED AT 250 WORDS)
Platelet-activating factor alters glomerular barrier size selectivity for macromolecules in rats
Authors: Perico N. , Remuzzi A. , Dadan J. , Battaglia C. , Remuzzi G.
Published: December 2017

The effect of platelet-activating factor (PAF) on glomerular permeability to macromolecules was investigated in the isolated kidneys from normal male Sprague-Dawley rats perfused at constant pressure. Compared with basal values, infusion of PAF (10 nM final concentration) into the isolated kidneys induced a progressive and significant increase in protein excretion (6.7 +/- 2.7 vs. 40.7 +/- 10.4 micrograms/min, P less than 0.01), completely reversible 20 min after PAF infusion was discontinued (8.5 +/- 1.3 micrograms/min). In additional experiments, during PAF infusion the fractional clearance of small neutral dextrans (radius 24-48 A), defined as the ratio of the clearance of neutral dextrans to the clearance of creatinine, was comparable to preinfusion values, whereas fractional clearance of large dextrans (greater than 50 A) was significantly elevated (P less than 0.005) above preinfusion values. The specific PAF receptor antagonist L 652731 completely prevented the increased fractional clearance of large dextrans induced by PAF. Finally, lowering Ca2+ concentration in the perfusion medium from 2.5 to less than 0.1 mM markedly reduced proteinuria in isolated kidneys exposed to PAF (80.0 +/- 10.3, 42.8 +/- 3.1, and 22.0 +/- 7.6 micrograms/min, respectively, for 2.5, 1.25, and less than 0.1 mM). These results indicate that in isolated perfused kidneys 1) PAF-induced proteinuria is a functional phenomenon reversible on discontinuing PAF infusion, 2) PAF modifies glomerular size-selective properties by increasing transmural passage of large dextran molecules, and 3) PAF-induced change in glomerular permselective properties is dependent on Ca2+ concentration in the extracellular medium.
Indirect Na+ dependency of urate and p-aminohippurate transport in pig basolateral membrane vesicles
Authors: Werner D. , Roch-Ramel F.
Published: December 2017

Membrane vesicles were used to study the basolateral transport of urate and p-aminohippurate (PAH) in the proximal tubule of the pig kidney. Consistent with a cooperation between a Na(+)-2-oxoglutarate cotransporter and a 2-oxoglutarate-urate or a 2-oxoglutarate-PAH exchanger, urate and PAH uptakes were stimulated in presence of extravesicular 2-oxoglutarate when an inwardly directed Na+ gradient was applied. Both transports exhibited, however, different characteristics. The optimal 2-oxoglutarate concentration for stimulating uptakes was 10 microM for PAH and 150 microM for urate. Extravesicular chloride was required to observe a stimulation of PAH uptake but not of urate uptake. Transports of both PAH and urate exhibited different affinity sequences for various organic anions. Stimulated PAH uptake was inhibited by probenecid greater than cold PAH greater than urate = pyrazinoate greater than lactate, whereas stimulated urate uptake was inhibited by probenecid greater than cold urate greater than PAH and not by pyrazinoate or lactate. These results are consistent with independent transport processes for urate and PAH in pig basolateral membrane vesicles, both being indirectly driven by an inwardly directed Na+ gradient.
Effects of acute nitric oxide inhibition on rat glomerular microcirculation
Authors: Zatz R. , de Nucci G.
Published: December 2017

Endothelium-derived relaxing factor (EDRF), recently identified as nitric oxide (NO), has been shown to be released by glomerular endothelial cells and might influence the glomerular microcirculation. To examine this hypothesis, we studied in rats the renal effect of acute administration of NG-monomethyl-L-arginine (L-NMMA), a specific inhibitor of NO synthesis. Adult male Munich-Wistar rats were studied before and after intravenous administration of either pure saline or a bolus injection of L-NMMA (20 mg) followed by a continuous infusion of the inhibitor (0.4 mg/min). Although saline alone had no effect on systemic or glomerular hemodynamics, L-NMMA promoted marked systemic hypertension, glomerular arteriolar vasoconstriction, and glomerular hypoperfusion. Since efferent resistance was disproportionately increased, glomerular hydraulic pressure was also markedly elevated. The glomerular ultrafiltration coefficient (Kf) fell to 42% of control. Single-nephron glomerular filtration rate was unaffected. Striking polyuria was also observed. These findings suggest that EDRF exerts a basal relaxing effect on the glomerular microcirculation.
Release of arachidonic acid by complement C5b-9 complex in glomerular epithelial cells
Authors: Cybulsky A. V.
Published: December 2017

In experimental membranous nephropathy, C5b-9 induces noncytolytic glomerular epithelial cell (GEC) injury and proteinuria, which in some models is partially mediated by metabolites of arachidonic acid. In cultured GEC, sublytic C5b-9 increases cytosolic Ca2+ concentration ([Ca2+]i), activates phospholipase C (PLC), and releases arachidonic acid and eicosanoids. This study examined mechanisms of arachidonic acid production by C5b-9. In GEC labeled with [3H]arachidonate C5b-9 increased free [3H]arachidonic acid and 1,2-[3H]-arachidonoyl-diacylglycerol (DAG), an endogenous activator of protein kinase C (PKC). Elevated [Ca2+]i was not sufficient to account for increased free arachidonic acid. Moreover, in GEC that had been depleted of PKC by preincubation for 18 h with 2 microM phorbol myristate acetate, the C5b-9-induced arachidonate release was inhibited by greater than 75%. Reacylation of phospholipids was not decreased by C5b-9. Homogenates of GEC that had been stimulated with C5b-9 released more [14C]arachidonate from exogenously added 2-[14C]arachidonoyl-phosphatidyl-ethanolamine or 2-[14C]arachidonoyl-phosphatidylcholine than homogenates of unstimulated cells (assayed at a Ca2+ concentration of 2 mM). These experiments demonstrate directly that C5b-9 increased phospholipase A2 (PLA2) activity. PLA2 appeared to be stimulated as a result of PKC activation (probably secondary to increased DAG) in association with elevated [Ca2+]i. The C5b-9-induced activation of PLA2 may lead to release of eicosanoids, which may contribute toward impaired glomerular capillary wall permselectivity in experimental membranous nephropathy.
Transport of sodium chloride and water in rat ascending vasa recta
Authors: Pallone T. L.
Published: December 2017

Experiments were undertaken to test the hypothesis that transcapillary small solute (NaCl and urea) gradients drive water across ascending vasa recta (AVR). Axial gradients of NaCl and urea were eliminated with furosemide. AVR were perfused with buffer containing fluorescein isothiocyanate-labeled dextran and 22Na. Perfusion of AVR with isotonic buffer at 10 and 20 nl/min yielded collectate-to-perfusate 22Na ratios of 0.17 +/- 0.05 and 0.34 +/- 0.03, respectively, in AVR of 601 +/- 56 and 583 +/- 46 microns mean length, respectively. A 22Na permeability of 113.2 +/- 12.8 x 10(-5) cm/s was determined. AVR were perfused at 20 nl/min with buffer NaCl of 0 (hypotonic to papilla), 161 (isotonic), or 500 mM (hypertonic). Transcapillary volume flux was not significantly different in these groups (3.8 +/- 1.5, 4.6 +/- 1.5, and 2.1 +/- 1.4 nl.min-1.mm-1, respectively). AVR were perfused in the hydropenic kidney at 5 nl/min antegrade from tip to base and retrograde from base to tip, which was a maneuver designed to impose physiological transcapillary NaCl and urea gradients of opposite direction. Volume fluxes were -1.4 +/- 0.05 and -1.3 +/- 0.04 nl.min-1.mm-1 in these groups, respectively. These data demonstrate that the AVR are highly permeable to NaCl and that physiological small solute gradients do not influence water movement across the AVR wall.
Regulation of collecting duct water permeability independent of cAMP-mediated AVP response
Authors: Lankford S. P. , Chou C. L. , Terada Y. , Wall S. M. , Wade J. B. , Knepper M. A.
Published: December 2017

We have used the isolated perfused tubule technique, measurements of adenosine 3',5'-cyclic monophosphate (cAMP) content in single tubules, and freeze-fracture electron microscopy to study the basis of high vasopressin-independent (basal) osmotic water permeability (Pf) in the terminal inner medullary collecting duct (IMCD) of the rat. The results confirmed the observation that the basal Pf of the terminal IMCD is considerably higher than that of the initial IMCD. They also showed that the basal Pf of the terminal IMCD is regulated by in vivo factors related to water intake, such that a very high vasopressin-independent Pf can be induced in isolated tubules by prior in vivo thirsting. Tubules from thirsted rats did not display elevated urea permeabilities, nor did they exhibit measurable cAMP levels in the absence of exogenous vasopressin, indicating that the high basal Pf was not due to residual binding of vasopressin to its receptors. Freeze-fracture studies in thirsted rats demonstrated the presence of intramembrane particle (IMP) clusters in both initial and terminal IMCD, with more in the latter. Water loading of the rats suppressed the incidence of clusters almost entirely but did not fully suppress the basal Pf in the terminal IMCD, raising the possibility that a component of transepithelial water transport may occur independently of the vasopressin-regulated IMP clusters. On the basis of these results, we conclude that the vasopressin-independent Pf in the terminal IMCD can be stably elevated to very high levels in response to in vivo thirsting. This elevation appears to be due to a chronic conditioning effect mediated by unknown in vivo factors and is not due to the short-term cAMP-mediated regulatory effect of vasopressin.
Effect of protamine on ion conductance of ascending thin limb of Henle's loop from hamsters
Authors: Koyama S. , Yoshitomi K. , Imai M.
Published: December 2017

To evaluate the contribution of paracellular shunt pathway in ascending thin limb (ATL) of hamsters, we examined the effect of protamine, a selective blocker of paracellular conductance, on salt-diffusion voltage (dVT) and transmural resistance (RT) during in vitro microperfusion. Lumen-negative dVT generated on reduction of lumen NaCl concentration was increased further from -7.3 +/- 0.5 to -10.3 +/- 0.7 mV when 300 micrograms/ml protamine was added to the lumen, and calculated Na+/Cl- permeability ratio was decreased from 0.46 +/- 0.03 to 0.31 +/- 0.03. Although the effect of protamine persisted after removal of the agent from the lumen, addition of 30 U/ml heparin returned the dVT toward the control level. The effect of protamine was dose dependent from 30 to 300 micrograms/ml. Protamine also exerted its effect from the bath, and the effect was inhibited by heparin either from the lumen or from the bath. The inhibitory effect was almost the same when the orientation of imposed NaCl gradient was reversed. Inhibition of transcellular Cl- transport with 0.1 mM 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) in the bath caused lumen-positive dVT. This voltage was decreased significantly by protamine. Protamine markedly decreased the apparent transference number for Na+ but slightly increased the value for Cl-. Transmural cable analysis showed that 300 micrograms/ml protamine added to the lumen increased RT from 0.59 +/- 0.10 to 1.20 +/- 0.20 omega.cm2, with the effect being reversed by 30 U/ml heparin.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult and fetal human mesangial cells interact with specific laminin domains
Authors: Weeks B. S. , Kopp J. B. , Horikoshi S. , Cannon F. B. , Garrett M. , Kleinman H. K. , Klotman P. E.
Published: December 2017

Mesangial cells are centrally located pericytes in the renal glomerulus. They are surrounded by an extracellular matrix and directly contact the glomerular basement membrane in vivo. Because these interactions are critical for renal development and function, we have studied human mesangial cell interactions with laminin, a major adhesive component of basement membranes present in the extracellular matrix of the mesangium. Human fetal and adult mesangial cell attachment was stimulated by both laminin and the laminin-derived synthetic peptides YIGSR-NH2, CQAGTFALRGDNPQG-NH2, and CIKVAVS-NH2. Furthermore, mesangial cells spread on laminin as well as on both the RGD-containing and CIKVAVS peptides. When added in solution, all three peptides inhibited mesangial cell attachment to laminin, and the latter two peptides inhibited mesangial cell spreading on laminin. Laminin affinity column chromatography demonstrated several low-molecular-mass laminin-binding proteins ranging from between 35 and 42 kDa, which predominated in fetal mesangial cells, whereas a higher molecular mass laminin-binding protein of 65 kDa was predominant in adult mesangial cells. Western blot analysis with an anti-32-kDa laminin-binding protein antibody showed increased expression of both 31- and 42-kDa proteins in fetal mesangial cells when compared with the adult. The antisera to the 32-kDa laminin-binding protein also inhibited fetal mesangial spreading on the CIKVAVS peptide. Western blot analysis with an anti-67-kDa laminin-binding protein antibody revealed a 110-kDa protein in adult mesangial cells that was not present in fetal mesangial cells.(ABSTRACT TRUNCATED AT 250 WORDS)