bluuebunny/crossref_metadata_2025 huggingface dataset:
Promoter analysis of the gene encoding the β-subunit of the rat amiloride-sensitive epithelial sodium channel
Authors: Bremner Harry Robert , Freywald Tanya , O'Brodovich Hugh M. , Otulakowski Gail
Published: December 2017

The amiloride-sensitive epithelial Na+channel (ENaC), found in the apical membrane of Na+-absorptive epithelia, is made up of three differentially regulated subunits: α, β, and γ. We undertook a study of the 5′-end of the gene encoding the β-ENaC subunit in the rat. 5′-Rapid amplification of cDNA ends and RNase protection assays indicated multiple transcription start sites over a 50-bp region. Sequencing 1.3 kb of the 5′-flanking DNA revealed putative binding sites for PEA3, Sp1, activator protein (AP)-1 and Oct-1 but neither a TATA box nor consensus sites for steroid hormone receptor binding. Transient transfections of reporter constructs driven by β-ENaC 5′-flanking DNA in the representative epithelial cell lines Madin-Darby canine kidney, MLE-15, and Caco-2 revealed a negative element present between positions −424 and −311 that affected basal transcription rates. Gel shift assays showed protein-DNA binding activity of an AP-1 consensus site in this region; however, mutation of the AP-1 site did not abrogate the repressive activity of the region in transient transfections. Deletion of two clusters of Sp1 consensus binding sites between −1 and −51 bp and between −169 and −211 bp indicated that the proximal cluster was essential to basal promoter activity in transfected cell lines. In a comparison of these data with those in published studies on α- and γ-ENaC promoters, the β- and γ-subunit promoters appear to be more similar to each other than to the α-promoter.
Immunodissection: use of monoclonal antibodies to isolate specific types of renal cells
Authors: Smith W. L. , Garcia-Perez A.
Published: December 2017

Several laboratories have described antibodies that are directed against cell surface antigens unique to different renal cell types. It can reasonably be assumed that each different type of renal cell possesses one or more unique antigenic determinants. In principle, it should be possible to prepare monoclonal antibodies against these cell-specific antigens and to use the antibodies as immunoaffinity reagents to isolate populations of specific renal cell types. We employed this approach in one instance to isolate canine cortical collecting tubule (CCT) cells. Approximately 10(7) canine CCT cells can be obtained from 5 g of canine renal cortex. This compares with an estimate of 10(3) to 10(4) CCT cells that can be reasonably obtained by microdissection. The availability of relatively large numbers of cells makes it possible to study in greater detail the cellular physiology and biochemistry of tubule-specific processes such as solute and water transport and hormone action. Our experiences in attempting to isolate canine CCT cells and other renal epithelia have indicated that the ideal monoclonal antibody for immunodissection should 1) interact with only one type of renal cell, 2) be directed against a determinant present in relative abundance, 3) be noncytotoxic, and 4) be an immunoglobulin G. Described in this review are some conceptual and practical aspects of the methods that can be used to develop B lymphocyte-myeloma hybrids producing such ideal monoclonal antibodies and to isolate renal cells using cell-specific monoclonal antibodies.
Effects of dietary potassium depletion and mineralocorticoid excess on renal Cl-conservation in the dog
Authors: Hulter H. N. , Licht J. H. , Sebastian A.
Published: December 2017

Preexisting dietary K+ depletion (KD) in dogs exaggerates the renal acid excretory response to mineralocorticoid hormone (MCH) and attenuates the renal Cl- reabsorptive response without altering the Na+ reabsorptive response. The exaggerated acid excretory response has been postulated to be an electrophysiological consequence of a defect in renal Cl- reabsorption caused by KD. To investigate the specific effects of KD on renal Cl- transport in dogs, we assessed renal Cl- conservation during dietary Cl- restriction in KD adrenalectomized dogs maintained on physiological replacement doses of MCH. After a 16-day period of dietary K+ restriction and physiological MCH replacement, reduction of dietary NaCl from 5.0 to 0.25 mmol X kg-1 X 24 h-1 was attended by reduction in urinary Cl- excretion to values less than intake and to significantly lower values than in K+ -replete controls. In a subsequent experimental period of continued Cl- restriction and administration of DOC (15 mg/24 h, i.m.), urinary Cl- excretion decreased further in both groups to stable values, but the values were significantly greater in KD (2.7 +/- 0.4 vs. 1.1 +/- 0.1 meq/24 h, P less than 0.05) and the cumulative retention of urinary Cl- was significantly less (10.3 +/- 1.4 vs. 29.5 +/- 6.7 meq, P less than 0.05). These findings demonstrate that preexisting dietary KD accelerates chronic renal Cl- conservation in response to dietary Cl- restriction under conditions in which MCH supply is normal and fixed but that it impairs maximal renal Cl- -conserving ability in response to MCH excess.
Synthesis of prostaglandin E2 in different segments of isolated collecting tubules from adult and neonatal rabbits
Authors: Schlondorff D. , Satriano J. A. , Schwartz G. J.
Published: December 2017

Prostaglandin E2 (PGE2) inhibits the action of the antidiuretic hormone (ADH) in isolated collecting tubules. A negative feedback loop has been postulated whereby ADH stimulates PGE2 synthesis. Furthermore, lysyl-bradykinin (LBK) inhibits the antidiuretic effect of ADH, probably via PGE2. Enhanced PGE2 synthesis has also been implicated as contributing to the inability to maximally concentrate urine during the neonatal period. We investigated PGE2 synthesis in microdissected cortical (CCT), medullary (MCT), and branched cortical (BCT) collecting tubules from adult and in corticomedullary collecting tubules (CT) from newborn rabbits. Isolated BCT produced significantly less PGE2 (12 +/- 2 pg X mm-1 X 20 min-1) than CCT (65 +/- 9) or MCT (76 +/- 8) from kidneys of adult rabbits. CT from newborn rabbits produced only 19 +/- 3 pg/mm, significantly less than either CCT or MCT from adults. A large variability in basal PGE2 production and hormonal response was observed from tubule to tubule. Under either basal conditions or in the presence of 2 microM arachidonic acid, LBK enhanced PGE2 synthesis in CCT and MCT from adults. ADH enhanced PGE2 production in MCT under basal conditions and in CCT in the presence of arachidonic acid. Neither LBK nor ADH stimulated PGE2 synthesis in neonatal CT. A23187 consistently stimulated PGE2 synthesis in CCT and MCT from adults and, to a lesser extent, in CT from newborn rabbits. Our results support the hypothesis that ADH and LBK enhance PGE2 synthesis in the collecting tubule. This response is, however, subject to large variations from tubule to tubule and depends on the in vitro incubation conditions.
ATP and cAMP system in the in vitro response of microdissected cortical tubules to PTH
Authors: Kiebzak G. M. , Yusufi A. N. , Kusano E. , Braun-Werness J. , Dousa T. P.
Published: December 2017

Responsiveness of proximal convoluted tubule (PCT) and distal convoluted tubule (DCT) microdissected from mouse kidney to PTH, in terms of cAMP accumulation and stimulation of adenylate cyclase, was examined. In both PCT and DCT, the cell-free adenylate cyclase was stimulated at least 10-fold by the same dose (10 U/ml) of PTH, and activity of cAMP phosphodiesterase was about 80% higher in DCT than in PCT. In intact tubules, while the incubation with PTH increased cAMP content in DCT more than 10-fold, it failed to increase the cAMP levels in PCT. To explain discrepancies between cell-free and intact cell incubations, ATP content in microdissected tubules was determined with use of a microbioluminescence assay. ATP content in PCT (4.0 +/- 1.3 fmol/mm, n = 30) was dramatically lower than ATP content of DCT (376.8 +/- 54.3 fmol/mm, n = 25). Incubation with 1 microM rotenone reduced markedly (delta -98%) the ATP content in DCT. In DCT, with ATP depleted by 1 microM rotenone, PTH failed to increase the cAMP, although 1 microM rotenone did not inhibit the adenylate cyclase activity. When 0.1 mM of 1-methyl-3-isobutylxanthine (MIX) was added to the incubation medium, PTH caused a marked elevation in tubular cAMP in PCT and to even a greater degree in DCT. Present results show that various segments of microdissected tubules differ greatly in their ability to maintain adequate ATP levels for cAMP generation in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)
Physiological and morphological responses of the rat kidney to reduced dietary protein
Authors: Schmidt-Nielsen B. , Barrett J. M. , Graves B. , Crossley B.
Published: December 2017

Renal physiological and morphological adjustments to a reduced protein diet were studied in young Munich-Wistar rats. Two groups of animals were used for the correlative physiological-morphological studies: normal protein (NP, 24% dietary protein) rats and reduced protein (LP, 8% dietary protein) rats. Both groups were fed their respective diets for 4-5 wk and had free access to drinking water. Physiological measurements of GFR and urea clearance were made on five animals from each group. These data showed that the changes in renal function specifically and almost exclusively affected the handling of urea. There was no difference in GFR between the NP and LP rats. Urea clearance was substantially reduced in LP rats. Morphological analyses were made on perfusion-fixed kidneys of five animals from each group. Selected slices were examined and photographed by light and electron microscopy. These data showed no difference in size and number of elements within the vascular bundles but showed significantly smaller lumina of the thin limbs of the short-looped nephrons and a significant thinning of the wall of the thin descending limbs of the long-looped nephrons. These morphological changes may in part be responsible for the observed physiological adjustments to a reduced protein diet. An additional group of rats (6 NP and 5 LP, all dehydrated) were analyzed for distribution of solutes within the inner medulla. The data showed that the concentration of urea, but not that of Na+, was reduced at the papillary tip in LP rats.
Proton/hydroxyl permeability of proximal tubule brush border vesicles
Authors: Ives H. E.
Published: December 2017

The net H+/OH- permeability of rabbit renal proximal tubule brush border membrane vesicles was determined by measuring the rate of collapse of preformed pH gradients using acridine orange. The membranes were voltage clamped using valinomycin and [K+]in = [K+]out. Internal buffer capacity was determined by titration of lysed vesicles and by titration of measured Na+/H+ exchange rates with exogenously added buffers. Both methods revealed an intravesicular buffer capacity of 125-135 mM/pH unit at pH 6.0 and 20 degrees C. Using this buffer capacity, the net H+/OH- permeability was found to be 5 X 10(-3) cm/s in brush border vesicles prepared by Mg2+ aggregation. The rate of collapse of pH gradients in brush border vesicles prepared by sucrose density gradient centrifugation was virtually identical to the rate in vesicles prepared with Mg2+, indicating that the high H+/OH- permeability was not an artifact of Mg2+ preparation. Activation energy of the H+/OH- permeability pathway was 4.9 kcal/mol, whereas activation energy of the Na+/H+ antiporter was 11.4 kcal/mol. Since the rate of H+/OH- diffusion was not affected by amiloride, it is concluded that H+/OH- permeate through brush border membranes by a pathway separate from the Na+/H+ antiporter. This pathway is not inhibited by dicyclohexylcarbodiimide at concentrations up to 2 mM but is inhibited by 0.2-5 mM p-chloromercuribenzenesulfonate, suggesting the presence of a sulfhydryl group in the pathway.(ABSTRACT TRUNCATED AT 250 WORDS)
Acute and chronic actions of bradykinin on renal function and arterial pressure
Authors: Granger J. P. , Hall J. E.
Published: December 2017

The present study was designed to examine the acute and chronic effects of increased levels of circulating bradykinin (BK) on control of renal hemodynamics, electrolyte excretion, and arterial pressure. Intrarenal infusion of BK (50 ng X kg-1 X min-1) for 60 min in five anesthetized dogs with renal perfusion pressure maintained at a constant level of 108 +/- 1 mmHg had no significant effect on glomerular filtration rate (GFR), whereas it increased renal blood flow (RBF) from a control value of 230 +/- 14 to 282 +/- 18, 266 +/- 15, and 253 +/- 17 ml/min after 15, 30, and 60 min of infusion, respectively. Acute intrarenal infusion of BK also increased urine volume (UV) from 0.255 +/- 0.044 to 0.523 +/- 0.103 ml/min and urinary sodium excretion (UNaV) from 5.72 +/- 1.5 to 13.7 +/- 3.4 mueq/min. To determine whether the potent acute effects of BK on RBF, UV, and UNaV lead to a chronic reduction in arterial pressure, BK (50 ng X kg-1 X min-1) was infused intrarenally for 7 days in conscious dogs. Intrarenal infusion of BK for 7 days had no significant effect on GFR, UNaV, UV, or arterial pressure. However, BK elevated renal plasma flow and decreased renal vascular resistance throughout the 7 days of infusion. Chronic intrarenal BK infusion caused no significant changes in plasma renin activity or plasma aldosterone concentration. Results from these studies indicate that although increased levels of bradykinin in the renal circulation can have potent acute effects on RBF, UV, and UNaV, these effects on UV and UNaV are not sustained and therefore do not result in long-term changes in arterial pressure.
Vasoconstrictor-evoked prostaglandin synthesis in cultured human mesangial cells
Authors: Ardaillou N. , Hagege J. , Nivez M. P. , Ardaillou R. , Schlondorff D.
Published: December 2017

We examined the influence of angiotensin II (ANG II), arginine vasopressin (AVP), and platelet activating factor (PAF) on prostaglandin (PG) synthesis and cell contractility in human glomerular mesangial cells in culture. Addition of sodium butyrate to the culture medium for 40 h significantly increased synthesis of both 6-keto-PGF1 alpha and PGE2 in the presence of exogenous arachidonic acid and of PGE2 under basal conditions. To optimize conditions in all further experiments, cells cultured with butyrate were studied. Under basal conditions, cultured mesangial cells produced predominantly 6-keto-PGF1 alpha and much less PGE2. Addition of either ANG II, AVP, or PAF all resulted in a rapid (within minutes) two- to threefold stimulation of 6-keto-PGF1 alpha and PGE2. Threshold stimulations were obtained at 10 pM for ANG II, 1 nM for AVP, and 10-100 pM for PAF. Preincubation of the cells with [Sar1,Ala8]ANG II, an antagonist of ANG II, inhibited ANG II-enhanced PG production, and preincubation with 1-desamino-8-D-arginine vasopressin, an antidiuretic analogue, blunted AVP-enhanced PG production. Under phase-contrast microscopy, PAF, ANG II, and, to a lesser degree, AVP caused decrease in cell surface area of mesangial cells cultured without butyrate at concentrations similar to those stimulating PG synthesis. Only PAF contracted cells cultured with butyrate, indicating attenuation of the vasoactive effects of ANG II and AVP when synthesis of PG was increased. However, a lower dose of PAF was only active when PG synthesis was inhibited, suggesting the same feedback mechanism for the three agonists.
Effects of angiotensin II on plasma ADH, prostaglandin synthesis, and water excretion in normal humans
Authors: Usberti M. , Federico S. , Di Minno G. , Ungaro B. , Ardillo G. , Pecoraro C. , Cianciaruso B. , Cerbone A. M. , Cirillo F. , Pannain M. , et al.
Published: December 2017

To verify whether angiotensin II (ANG II) stimulates ADH release in humans and to evaluate whether endogenous prostaglandins influence the resulting renal effect of ADH, nonpressor and low pressor doses of ANG II were infused in nine normal volunteers under normal conditions (control study) and after prostaglandin synthesis inhibition with aspirin (ASA study). During ANG II infusion plasma ADH increased in both conditions. Plasma PGE2, urinary PGE2, and urinary 6-keto-PGF1 alpha increased only in the control study, whereas they were undetectable in the plasma and significantly reduced in the urine in the ASA study. ANG II caused a significant fall of glomerular filtration rate, renal plasma flow (with an increase in filtration fraction), fractional sodium excretion, and urine output in both studies. Despite the reduced urine output, urine osmolality decreased significantly in the control study, whereas it increased after aspirin administration. These results suggest that intravenous ANG II stimulates ADH release in humans but that the renal effects of the resulting increase in plasma ADH are different depending on the presence or absence of endogenous prostaglandins.
Regulation of water permeability in toad urinary bladder at two barriers
Authors: Kachadorian W. A. , Sariban-Sohraby S. , Spring K. R.
Published: December 2017

The effects of prostaglandin synthesis inhibition by naproxen were studied in toad bladder. Luminal membrane water permeability was evaluated both by the frequency of intramembranous particle aggregates in granular cell luminal membrane and by direct assessment of the rate of change of cell volume during perfusion of an anisosmotic solution. Total tissue water permeability was assessed by transbladder osmotic water flow. Inhibition of prostaglandin synthesis caused luminal membrane water permeability to increase much more than expected from tissue permeability measurements. The addition of a very low dose of antidiuretic hormone (ADH) (0.125 mU/ml) during prostaglandin synthesis inhibition increased luminal membrane water permeability to the same level as maximal stimulation with ADH, while tissue water permeability failed to increase proportionately. The results imply the presence of a regulatable barrier to water movement across toad bladder that is distal to the luminal membrane and subject to control by either prostaglandins or ADH.
Electrochemical profile of K+ and Na+ in the amphibian early distal tubule
Authors: Teulon J. , Froissart P. , Anagnostopoulos T.
Published: December 2017

Double-barreled microelectrodes selective to either potassium or sodium were used to determine the transepithelial potential difference (VTE) and the intraluminal activity of potassium (alpha LuK) or sodium (alpha LuNa) in the early distal tubule (EDT) of Triturus waltlii in vivo; luminal activities were compared with the corresponding plasma ion activities, alpha PtK and alpha PtNa. The transepithelial equilibrium potentials for potassium (EK(TE] and sodium (ENa(TE] were computed from the respective transmural chemical distributions: they were used to assess the transepithelial electrochemical potential differences [(V-EK)TE and (V-ENa)TE]. By dividing the raw data into three groups of 30% total tubular length (0-30, 31-60, 61-90%), the following results were obtained. 1) VTE increases from +15 to +20 mV (lumen positive) between the first and second portion of the EDT but remains constant thereafter. 2) The alpha LuK/alpha PtK ratio decreases steadily along the EDT from 1.92 to 1.66 and then to 1.32. 3) The values of alpha LuNa/alpha PtNa in the same three subdivisions are 0.79, 0.44, and 0.45. 4) The (V-EK)TE difference is largely positive along the whole EDT: +32, +33, and +27 mV. 5) The (V-ENa)TE difference declines from +9 mV (first portion) to values statistically not different from zero in the last two thirds of the EDT.(ABSTRACT TRUNCATED AT 250 WORDS)
Antidiuretic and PGE2 responses to AVP and dDAVP in subjects with central and nephrogenic diabetes insipidus
Authors: Moses A. M. , Scheinman S. J. , Schroeder E. T.
Published: December 2017

Vasopressin stimulates renal prostaglandin (PGE2) production at several loci and in turn PGE2 modulates the antidiuresis. We have found the time courses of increased urinary PGE2 in subjects with central diabetes insipidus (DI) parallel the antidiuretic responses to AVP and dDAVP. The antidiuretic response to 4 micrograms dDAVP in these subjects was far greater than the response to 5 U (12.5 micrograms) AVP, but the PGE2 response to the dDAVP was only marginally greater than that which followed the AVP. Therefore, dDAVP disproportionately stimulates antidiuresis in relation to PGE2 production, whereas the reverse holds for AVP. In subjects with nephrogenic DI 12.5 micrograms AVP caused no antidiuresis but stimulated PGE2 excretion as well as in subjects with central DI. There was an intermediate relationship between antidiuresis and PGE2 excretion in subjects with central DI given AVP and subjects with nephrogenic DI injected with dDAVP. In summary, 1) the normal PGE2 response to AVP in subjects with nephrogenic DI is consistent with other evidence that non-antidiuretic actions of vasopressin are not impaired in these subjects. 2) The limited capability of dDAVP to stimulate PGE2 may be a factor in the augmented antidiuretic response to dDAVP in subjects with central DI. 3) Antidiuretic and PGE2 responses to vasopressin can be dissociated, thus allowing further consideration of mechanisms by which each may be independently controlled and interrelated.
High potassium intake selectively increases urinary PGF2 alpha excretion in the rat
Authors: Nasjletti A. , Erman A. , Cagen L. M. , Brooks D. P. , Crofton J. T. , Share L. , Baer P. G.
Published: December 2017

This study was designed to investigate relationships between dietary potassium and the renal prostaglandin system in rats. The potassium content of the diet was 0.162 mmol/g during the control period and 0.004, 0.162, 1.351, or 2.702 mmol/g during the experimental period. Relative to control data in rats fed a 0.162 mmol/g potassium diet, the urinary excretion of 6-keto-PGF1 alpha was not affected by high potassium intake but increased (P less than 0.05) by 25% in rats fed a low potassium diet for 13 days and was associated with reduction of plasma potassium and with elevation of both plasma renin and net release of 6-keto-PGF1 alpha from renal inner medulla slices incubated in Krebs solution. The excretion of PGF2 alpha was not affected by low potassium intake but increased (P less than 0.05) by about twofold in rats fed a potassium-rich diet (1.351 and 2.702 mmol/g) for 13 days and was associated with elevation of plasma potassium concentration, renal prostaglandin 9-keto-reductase activity, and urinary excretion of kallikrein and vasopressin. The urinary excretion of PGE2 was not altered in rats fed either low or high potassium diets. Altogether, these results indicate selective influence of dietary potassium on the urinary excretion of prostaglandins in the rat.
A distributed model of peritoneal-plasma transport: analysis of experimental data in the rat
Authors: Flessner M. F. , Dedrick R. L. , Schultz J. S.
Published: December 2017

Transport of uncharged, water-soluble substances (ranging in molecular weight from 180 to 5,000) between the fluid in the peritoneal cavity and plasma was studied in anesthetized female Sprague-Dawley rats. In certain experiments the effect of fluid shifts on the transport was observed by manipulating the effective osmotic pressure or the hydrostatic pressure of the dialysis fluid. Parameters for the distributed model outlined in previous work were obtained from the experimental data for the substances tested. Capillary membrane transport was modeled by pore theory. A single pore radius of 40 A and a pore density of 600 cm-2 were satisfactory. Tissue diffusivities for these substances were found to correspond closely to those in the literature. Additional simulations were performed with a three-compartment model and the results were compared with those of the distributed model.
Adaptive changes in renal acidification in response to chronic respiratory acidosis
Authors: Tannen R. L. , Hamid B.
Published: December 2017

To examine whether chronic respiratory acidosis results in adaptive changes in renal acidification, rats were housed for 3 days in an environmental chamber with an ambient CO2 content of 10% and their kidneys were perfused in vitro according to two protocols. To assess hydrogen ion secretory capacity of the distal nephron, perfusions were carried out with a low bicarbonate concentration, in the absence of ammoniagenic substrate, and with saturating quantities of the buffer creatinine. Under these conditions, the titration of creatinine at a pH less than 6.0 (TA pH 6.0) reflects the H+ secretory capacity of a discrete functional segment of the distal nephron. Kidneys from rats with chronic respiratory acidosis exhibited a significantly lower urine pH and higher rate of TA pH 6.0 than controls perfused in this fashion, indicative of an adaptive increase in the distal nephron capacity for proton transport. This adaptation was comparable with that reported previously for rats exposed to chronic metabolic acidosis. Furthermore, evidence of adaptation persisted in the presence of amiloride (10(-5) M), suggesting that it reflects, at least in part, a sodium-independent mechanism of proton transport. Hydrogen ion secretion by the proximal nephron was assessed by performing standard bicarbonate titration curves with kidneys from rats with chronic respiratory acidosis, chronic metabolic acidosis, and controls using a perfusate equilibrated with 95% O2/5% CO2.(ABSTRACT TRUNCATED AT 250 WORDS)
Phosphate loading attenuates renal tubular dysfunction induced by maleic acid in the dog
Authors: Al-Bander H. , Etheredge S. B. , Paukert T. , Humphreys M. H. , Morris R. C.
Published: December 2017

The metabolic pathogenesis of the complex renal tubular dysfunction of type II renal tubular acidosis and Fanconi's syndrome (RTA II/FS) acutely induced by maleic acid could depend on the occurrence of a positive feedback loop in cells of the proximal renal tubule: impaired mitochondrial oxidation----increased glucose uptake----increased formation and concentration of phosphorylated glycolytic intermediates----limitation on availability of cellular inorganic phosphate----more severely impaired mitochondrial oxidative metabolism. To test this hypothesis we intravenously administered maleic acid both alone and after initiating intravenously administered neutral sodium phosphate, sodium sulfate, or sodium chloride to 10 unanesthetized trained female dogs undergoing water diuresis. We made the following observations: 1) Administration of maleic acid alone predictably induced dose-dependent increments in urine flow (V) and in renal clearance of HCO3-, Na+, K+, and alpha-aminonitrogen and a pronounced increase in the renal clearance and excretion of citrate. 2) Prior phosphate loading, which increased the plasma concentration of phosphate from 2.5 +/- 0.20 to 11.3 +/- 2 mg/dl: a) attenuated the increment in renal clearance of HCO3- by one-half even though the filtered load of bicarbonate was higher by 37%, owing to the higher values of both GFR and plasma bicarbonate concentration that obtained with phosphate loading; b) prevented the increment in renal clearance and excretion of alpha-aminonitrogen; c) significantly attenuated the increments in V and renal clearance of K+; but d) did not affect the increment in renal clearance and excretion of citrate.(ABSTRACT TRUNCATED AT 250 WORDS)
Lactate production in isolated segments of the rat nephron
Authors: Bagnasco S. , Good D. , Balaban R. , Burg M.
Published: December 2017

Lactate production was measured directly in individual segments of the rat nephron. Tubules were dissected and then incubated in vitro with glucose as the only metabolic substrate. Each segment was incubated with and without antimycin A, an inhibitor of oxidative metabolism. Proximal tubules produced no lactate with or without antimycin A. The distal segments all produced lactate. The rate of lactate production without antimycin A ranged from 0.4 to 0.9 pmol X min-1 X mm-1 in all distal segments except one, the inner medullary collecting duct, which produced lactate at the significantly higher rate of 2.8 pmol X min-1 X mm-1. Antimycin A increased lactate production significantly in all of the distal segments. The increase was largest in medullary thick ascending limbs (1,400%) and cortical (798%) and outer medullary collecting ducts (357%). Increments were smaller in cortical thick ascending limbs (98%) and distal convoluted tubules (98%) and least in the inner medullary collecting ducts (28%). We conclude that lactate production occurs only in distal segments of the nephron and that under anoxic conditions significant amounts of ATP are produced by anaerobic glycolysis in these segments.
Na-K-ATPase in isolated rabbit tubules after unilateral nephrectomy and Na+ loading
Authors: Scherzer P. , Wald H. , Czaczkes J. W.
Published: December 2017

Na-K-ATPase activity was studied in tubule segments from the cortex and medulla of rabbit kidneys under normal conditions, after unilateral nephrectomy, and after chronic salt loading. After unilateral nephrectomy kidney weight increased by 37% and Na-K-ATPase activity rose significantly in all nephron segments by 36-200% (P less than 0.01). Oral salt loading for 2 wk with 0.5% NaCl caused an increase in GFR and in absolute sodium excretion as well as reabsorption; plasma aldosterone decreased by 44% (P less than 0.005). In the proximal segments (PCT, CPST, OMPST, and TDL) there were no marked changes in Na-K-ATPase activity, whereas along the whole length of the ascending limb of Henle's loop (iMTAL, MTAL, and CTAL) there was a significant rise in the enzymatic activity of 30-200% (P less than 0.02). In the distal segments (DCT, CCD, and OMCD) there was a marked decrease of 50-60% (P less than 0.005) in Na-K-ATPase activity after the salt loading. We conclude that unilateral nephrectomy caused a general increase in Na-K-ATPase activity along the whole length of the nephron, and salt loading caused a selective increase in enzyme activity along the ascending limb of Henle's loop and decrease in the distal segments.
NH4Cl inhibition of transport in the rabbit cortical collecting tubule
Authors: Hamm L. L. , Gillespie C. , Klahr S.
Published: December 2017

Ammonium has previously been found to inhibit transport in a number of tissues. The present experiments were designed to evaluate the effect of ammonium chloride on transepithelial voltage (VTE) and cation transport in the isolated rabbit cortical collecting tubule perfused in vitro. Peritubular NH4Cl (2-10 mM) inhibited VTE in these tubules independent of bath or lumen pH. Luminal NH4Cl had a similar effect. However, VTE did not change with bath NH4Cl in tubules treated with amiloride or ouabain. Furthermore, when bath PCO2 was lowered simultaneously with the addition of NH4Cl to the bath, little change in VTE occurred, raising the possibility that intracellular pH falls after addition of NH4Cl to the bath. Consistent with the voltage effects, 5 mM NH4Cl in the bathing solution inhibited net sodium reabsorption by 36% and potassium secretion by 44%. Unidirectional lumen-to-bath tracer fluxes were consistent with predominant inhibition of the sodium reabsorptive flux and the potassium secretory flux. These findings may have relevance to metabolic acidosis in vivo because ionic ammonium concentrations attain the levels used in this study.
Postprandial regulation of renal hemodynamics: role of pancreatic glucagon
Authors: Premen A. J. , Hall J. E. , Smith M. J.
Published: December 2017

The aim of this study was to ascertain the quantitative importance of glucagon in mediating protein-induced increases in renal blood flow (RBF) and glomerular filtration rate (GFR) during the postprandial state. Six chronically instrumented conscious dogs were each subjected to four protocols: normal protein control meal (C); high protein meat meal (M); somatostatin infusion and meat (S + M); and glucagon infusion. C produced small increases in RBF (12 +/- 2%) and GFR (14 +/- 2%) without changing arterial plasma glucagon. M produced marked increases in RBF (38 +/- 4%), GFR (41 +/- 5%), and glucagon (from 23.3 +/- 3.6 to 73.3 +/- 7.1 pg/ml) over a 3-h period. During S + M, RBF and GFR failed to increase while glucagon was suppressed by 36 +/- 8% over a 2-h period. When S was stopped, glucagon rose to 100 +/- 13.9 pg/ml over the next hour, yet RBF and GFR increased by only 14 +/- 4 and 10 +/- 3%, respectively. Glucagon infusion (3 ng X kg-1 X min-1, i.v.) markedly elevated plasma glucagon to 131.6 +/- 27.3 pg/ml, yet neither RBF nor GFR significantly changed. These data indicate that while a protein-rich meat meal does elevate arterial plasma glucagon, the rise is not great enough to elicit significant changes in renal hemodynamics. Thus, elevated plasma levels of glucagon cannot account for protein-mediated increases in RBF and GFR during the postprandial state.
Effect of PTH on blood pressure and response to vasoconstrictor agonists
Authors: Saglikes Y. , Massry S. G. , Iseki K. , Nadler J. L. , Campese V. M.
Published: December 2017

Parathyroid hormone (PTH) is known to be a vasodilator and to exert a hypotensive action. The present study examined the possible mechanisms involved, with special emphasis on the role of vasodilatory prostaglandins. The effects of the intact 1-84 PTH and that of its amino-terminal 1-34 fragment on mean arterial pressure (MAP) and on the vascular response to norepinephrine (NE) or angiotensin II (ANG II) were examined in rats before and after pretreatment with indomethacin. Bolus injections of 30 U of both 1-84 and 1-34 PTH produced a significant (P less than 0.01) decrease in MAP; however, the hypotensive response to 1-34 PTH (-28 +/- 4.6 mmHg) was more marked (P less than 0.01) than to 1-84 PTH (-9 +/- 1.8 mmHg). The infusion of 1-84 PTH (30 U/h) did not alter MAP, whereas the infusion of 1-34 PTH (30 U/h) led to a decrease in MAP from 124 +/- 2 to 103 +/- 4.0 mmHg (P less than 0.01) and to a rise in heart rate from 359 +/- 30 to 437 +/- 13 beats/min (P less than 0.02). Both 1-84 and 1-34 PTH antagonized the pressor effects produced by bolus injections of NE and ANG II. Pretreatment with indomethacin (5 mg/kg body wt) abolished the inhibitory effect of 1-84 and 1-34 PTH on the response of MAP to NE and ANG II and the hypotensive action of 1-34 PTH. Infusion of 1-84 and 1-34 PTH produced a significant rise in urinary excretion of 6-keto-PGF1 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)
Tubular capacity for phosphate reabsorption in superficial and deep nephrons
Authors: Haramati A.
Published: December 2017

The present studies were performed to determine the capacity for phosphate reabsorption in superficial and deep nephron proximal tubules in vivo. Micropuncture experiments were performed in 20 acutely thyroparathyroidectomized (TPTX) Munich-Wistar rats fed a normal phosphate diet (0.7%). Four groups were infused with differing amounts of phosphate (0,2,4, or 6 mumol/min) to increase the filtered phosphate load. The sites selected for micropuncture were the superficial early distal tubule and the deep nephron loop of Henle, which reflect fractional phosphate delivery (FDPi%) from superficial and deep nephron proximal tubules, respectively. In response to phosphate infusions, plasma phosphate increased from 3.03 +/- 0.09 to 7.01 +/- 0.58 mM, and fractional phosphate excretion rose from 2 +/- 1 to 58 +/- 5%. FDPi% increased from both superficial (14 +/- 1 to 58 +/- 2%) and deep nephron proximal tubules (4 +/- 1 to 27 +/- 5%) but always remained lower from deep nephrons, reflecting more avid reabsorption by deep nephron proximal tubules. The maximal rate of phosphate reabsorption (max RPi/SNGFR) in the superficial proximal tubule was significantly less than in the deep nephron proximal tubule (3.2 +/- 0.4 vs. 5.1 +/- 0.1 pmol/nl). In seven of the phosphate-infused rats, parathyroid hormone (PTH, 33 U/kg bolus; 1 U X kg-1 X min-1) was added to the infusion following the initial collections. In the presence of PTH, the RPi/SNGFR was significantly lower in deep than in superficial proximal tubules (0.4 +/- 0.5 vs. 1.6 +/- 0.4 pmol/nl). Thus, the maximum capacity for phosphate reabsorption was greater in deep than in superficial nephrons in TPTX rats. Furthermore, in the presence of phosphate infusions, PTH inhibited phosphate reabsorption to a greater extent in deep than in superficial proximal tubules.
Cation dependence of renal outer cortical brush border membrane L-glutamate transport
Authors: Fukuhara Y. , Turner R. J.
Published: December 2017

The cation dependence of L-glutamate transport in renal outer cortical brush border membrane vesicles was studied. Both cis sodium and trans potassium were required for optimal L-glutamate flux. Relative to simultaneous sodium (out greater than in) and potassium (in greater than out) gradient conditions, flux was reduced 50-fold by potassium removal and more than 100-fold by sodium removal. The effect of potassium removal in this preparation was markedly larger than that observed in other renal brush border membrane preparations. No other monovalent cation tested was effective in replacing sodium. However, Rb+ and Cs+ and to a lesser degree NH+4 were found to be effective potassium substitutes. Kinetic analysis of the Na/K-dependent component of L-glutamate flux indicated a single transport system of Km = 13 microM and Vmax = 1.3 nmol X min-1 X mg protein-1. Studies of the dependence of L-glutamate flux on potassium and sodium concentrations yielded Hill coefficients of 0.9 and 1.9, respectively, consistent with involvement of one potassium ion and two or more sodium ions in the L-glutamate transport event. Efflux studies indicated that sodium and potassium act at different steps in the transport cycle, sodium to facilitate the translocation of the glutamate-carrier complex, and potassium to facilitate the return of the unloaded carrier. A model for renal Na/K-dependent L-glutamate transport is suggested on the basis of these results.
Direct visualization of the isolated and perfused macula densa
Authors: Kirk K. L. , Bell P. D. , Barfuss D. W. , Ribadeneira M.
Published: December 2017

Direct examination of the structure and function of the macula densa is compromised by the relative inaccessibility and small size of this cell plaque. We report the isolation and perfusion of rabbit nephron segments with attached glomeruli and the direct visualization of the macula densa with differential interference-contrast microscopy. We used this technique to examine the structural sensitivity of the macula densa to perturbations in luminal osmolality and NaCl concentration. Reducing luminal osmolality from 290 to 70 mosmol/kg by removing NaCl resulted in a dilation of the lateral intercellular spaces that was both reversible and specific to the region of the macula densa. Associated with the dilation of the intercellular spaces was a small (congruent to 10%), but statistically significant, increase in the height of the macula densa cells. These structural events were related to the reduction in luminal osmolality, since isosmotic replacement of NaCl with mannitol resulted in no detectable structural changes. Thus, the macula densa may represent a small water-permeable plaque of cells within the remaining water-impermeable thick ascending limb of Henle's loop.
Responses of the normal rat kidney to sequential ischemic events
Authors: Zager R. A. , Jurkowitz M. S. , Merola A. J.
Published: December 2017

This study was undertaken to help define how one episode of renal ischemia, insufficient to cause acute renal failure, influences the susceptibility of the kidney to a second more severe ischemic event. Female Sprague-Dawley rats underwent either 15 min of bilateral renal artery occlusion (RAO) or sham RAO. They were subjected 30 min, 3.5 h, or 24 h later to 25 min of RAO. Renal function (GFR, BUN, creatinine), histology, and adenine nucleotide concentrations were compared before and after the 25-min ischemic event. Only the rats with a 30-min hiatus between the 15- and 25-min bouts of RAO had significantly worse renal failure than controls subjected to a single 25-min ischemic event. Three findings were noted only in the rats with increased susceptibility: tubular cell swelling and luminal membrane injury prior to 25 min of RAO and a relative failure of ATP formation immediately following 25 min of RAO. Susceptibility to 25 min of RAO did not correlate with preischemia ATP content. Conclusion: prior mild ischemic injury transiently lowers renal resistance to a second ischemic event. Normal resistance is rapidly restored once improvements in prior cell membrane injury, cell volume regulation, and cellular energetics occur. However, resistance to additional ischemia can be normal despite persisting depressions in renal ATP content.
Renal hydroxybutyrate and acetoacetate reabsorption and utilization in the rat
Authors: Barac-Nieto M.
Published: December 2017

Is the renal conservation of ketones effected through saturable transport systems? Does the renal reabsorption of ketoacids limit their utilization? The reabsorptive transport (T) and metabolic utilization (Q) of D-3-hydroxybutyrate (HB) and acetoacetate (AA) in the kidney were measured in anesthetized rats by use of clearance and arteriovenous extraction determinations. THB and TAA increased in proportion with increasing HB and AA filtered loads. Fractional THB decreases from 0.97 to 0.72 upon increases in the plasma HB. Fractional TAA also decreases from 0.89 to 0.73, at a constant filtered AA load, when the filtered HB load increased. Thus both saturable and nonsaturable components effect the reabsorption of HB and AA. Renal QHB increased from 0.4 to 4 mumol X g-1 X min-1 on increase in the plasma HB. THB always exceeded the simultaneously measured QHB. However, QHB was significantly correlated with THB, suggesting that QHB may be limited by delivery of HB to the renal cells through reabsorptive pathways. There was no net renal conversion of HB and AA. Both HB and AA were utilized by the kidney. Thus, utilized ketoacids must have entered pathways for complete oxidation or for synthesis (lipid?) in the kidney.
Rat renal peritubular transport and metabolism of plasma [35S]glutathione
Authors: Rankin B. B. , Wells W. , Curthoys N. P.
Published: December 2017

More than 80% of the plasma glutathione is extracted during a single pass through the kidney. The peritubular component of this extraction was characterized by in situ arterial infusion of [35S]glutathione and [3H]inulin. The peak of 35S-labeled material recovered in the renal venous effluent was delayed approximately 10 S compared with the peak of [3H]inulin. As a result, the initial fractions exhibited a decreased 35S/3H ratio, indicating that 35S-labeled material is transported out of the postglomerular peritubular capillaries. Later fractions exhibited a normalized 35S/3H ratio greater than 1, consistent with the subsequent addition of a 35S-labeled metabolite to the venous circulation. An identical profile was observed when perfusion experiments were repeated using gamma-[35S]glutamyl-S-methylcysteine and [3H]inulin. Renal venous plasma samples obtained from a rat perfused with [35S]glutathione were reduced with sodium borohydride, reacted with monobromobimane, and analyzed by high-pressure liquid chromatography. More than 70% of the recovered 35S-labeled material was identified as cysteine and 20% was recovered as unmetabolized glutathione. Pretreatment of rats with a single injection of AT-125 resulted in 96% inactivation of renal gamma-glutamyltranspeptidase. Under these conditions, the percent of glutathione converted to cysteine (35%) was significantly less than the observed level of renal extraction (61%). Two injections of AT-125 caused a complete inhibition of cysteine formation. However, the residual level of renal extraction (41%) was still significantly greater than the filtration fraction (26%). The peritubular transport of glutathione is stimulated by prior depletion of renal glutathione with buthionine-L-sulfoximine and is competitively inhibited by simultaneous infusion of gamma-glutamylcysteine.(ABSTRACT TRUNCATED AT 250 WORDS)
Blockade of renal effects of dopamine in the dog by the DA1 antagonist SCH 23390
Authors: Frederickson E. D. , Bradley T. , Goldberg L. I.
Published: December 2017

Dopamine (DA) acts on two receptor subtypes, DA1 and DA2. The purpose of this study was to determine which subtype is involved in the increments in renal blood flow (RBF) and electrolyte excretion produced by DA. Mongrel dogs were anesthetized with pentobarbital sodium. Phenoxybenzamine (10 mg X kg-1 ia) and propranolol (5 mg X kg-1 iv) were administered to exclude effects mediated by alpha- and beta-adrenoceptors. DA was infused into the renal artery before and after administration of either the selective DA1 antagonist SCH 23390 or the selective DA2 antagonist domperidone. With DA alone, RBF increased by 52 +/- 7%, Na+ excretion increased by 35 +/- 8%, and K+ excretion increased by 35 +/- 5%. Infusion of SCH 23390 (0.5 micrograms X kg-1 X min-1) completely blocked DA-induced increase in RBF and electrolyte excretion. Intravenous infusion of domperidone (1 microgram X kg-1 X min-1) did not attenuate the responses to DA. Neither SCH 23390 nor domperidone affected base-line RBF or electrolyte excretion, suggesting that in these experiments endogenous DA was not active. In conclusion, these data indicate that the effects of DA to increase RBF and electrolyte excretion are the result of action on DA1 receptors.
Phorbol esters inhibit gluconeogenesis in canine renal proximal tubular segments
Authors: Rogers S. , Gavin J. R. , Hammerman M. R.
Published: December 2017

Gluconeogenesis is a major metabolic function of the renal proximal tubular cell. To characterize the regulation of this process in proximal tubule, glucose production from gluconeogenic precursors was measured in proximal tubular segments prepared from dog kidney. Production of glucose was a linear function of time for up to 120 min of incubation at 37 degrees C under a variety of conditions. Lowering the pH of incubation media from 7.5 to 7.0 increased glucose synthesis. Production of glucose was inhibited by 3-mercaptopicolinate. Incubation of proximal tubular segments with insulin diminished synthesis of glucose. Incubation of segments with tumor-promoting phorbol esters, 12-O-tetradecanoylphorbol-13-acetate or phorbol 12,13-dibutyrate, resulted in decreased production of glucose. This effect was not observed following incubation with the inactive phorbol ester 4 alpha-phorbol. Changes in glucose synthesis could not be attributed to alterations in cell viability or in rates of glucose oxidation induced by experimental maneuvers. Our findings confirm the usefulness of proximal tubular segments for characterization of metabolic processes in this portion of the nephron. The experimental results are consistent with a role for protein kinase C in the control of gluconeogenesis in proximal tubule.