bluuebunny/crossref_metadata_2025 huggingface dataset:
Whole-cell currents in rat cortical collecting tubule: low-Na diet increases amiloride-sensitive conductance
Authors: Frindt G. , Sackin H. , Palmer L. G.
Published: December 2017

Individual principal cells within the rat cortical collecting tubule were studied under voltage-clamp conditions using the whole-cell variation of the patch-clamp technique. Isolated tubules were split to expose the apical membrane surface and bathed in NaCl medium at 23 degrees C. When carboxyfluorescein was included in the patch pipette, the dye diffused rapidly into the cell being clamped but did not spread to neighboring cells, indicating a lack of cell-to-cell coupling. Average cell capacitance under whole-cell clamp conditions with KCl in the pipette was 18 +/- 2 pF (n = 10 cells) in rats maintained on a normal diet, consistent with that expected from morphometric measurements of cell surface area. The capacitance increased to 36 +/- 7 pF (n = 8 cells) for rats kept on a low-Na diet, indicating that cell membrane area was increased under these conditions. The amiloride-sensitive whole-cell conductance (GNa), assumed to equal the conductance through apical Na channels, was determined as the slope of the current-voltage relation near zero holding potential. GNa was 6.0 +/- 1.7 nS/cell (n = 12) for rats maintained on a low-Na diet compared with 0.06 +/- 0.08 nS/cell (n = 13) for rats kept on a normal diet. The amiloride-insensitive whole-cell conductance averaged 9.1 +/- 2.0 nS/cell, with no significant difference between low-Na and normal groups. Sodium channel density (N) was estimated from GNa, the mean open probability of the channel, and the single-channel conductance. N equals 3,000 channels/cells in rats on a low-Na diet compared with N less than 100 channels/cell for rats on a normal diet.
Mechanism of aluminum-induced calcium efflux from cultured neonatal mouse calvariae
Authors: Sprague S. M. , Bushinsky D. A.
Published: December 2017

Aluminum has been shown to increase unidirectional 45Ca efflux from prelabeled bones in vitro; whether aluminum affects net calcium efflux and, if so, by what mechanism has not been studied. To examine the effects of aluminum on net calcium flux from bone we cultured live and dead neonatal mouse calvariae with and without graded concentrations of aluminum (10(-8) to 10(-5) M). Aluminum induced a dose-dependent net calcium efflux from live bone after 24 h, but not 3 h, which was similar in magnitude to that produced by 10(-8) M parathyroid hormone. The normal calcium influx into dead bone was not altered by aluminum. Release of beta-glucuronidase, a lysosomal enzyme released by osteoclasts, increased after a 24-h incubation in aluminum-containing medium and was correlated with net calcium efflux. Calcitonin, an inhibitor of osteoclastic bone mineral dissolution, abolished the increase in beta-glucuronidase release and nullified the aluminum-induced net calcium efflux. Thus aluminum induces cell-mediated net calcium efflux from bone and increases beta-glucuronidase release. Calcitonin inhibits the increase in both calcium efflux and beta-glucuronidase release, suggesting that aluminum stimulates osteoclasts to release bone mineral.
Effects of a kinin antagonist on renal function in rats
Authors: Nakagawa M. , Stewart J. M. , Vavrek R. J. , Nasjletti A.
Published: December 2017

We contrasted the effects of D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-DPhe-Thi-Arg-TFA (kinin receptor antagonist), of aprotinin (kallikrein inhibitor), and of combined treatment with captopril (kininase II inhibitor) and phosphoramidon (neutral endopeptidase 24.11 inhibitor) on renal function of rats with and without 14-day deoxycorticosterone pretreatment (DOC, 25 mg.kg-1.wk-1 sc). Neither the kinin antagonist nor aprotinin affected renal function in rats with and without DOC pretreatment. Combined treatment with captopril and phosphoramidon caused in rats with and without DOC pretreatment augmentation (P less than 0.05) of kinin excretion (50-64%), glomerular filtration rate (12-11%), and sodium excretion (46-48%). In DOC-pretreated rats undergoing infusion of captopril and phosphoramidon, the superimposed administration of either the kinin antagonist or aprotinin caused the lowering of renal plasma flow, glomerular filtration rate, and sodium excretion. These effects of the kinin antagonist and aprotinin in rats infused with kininase inhibitors may be the consequence of blockade, respectively, of the renal actions and synthesis of kinins that, when in excess, elicit renal vasodilation and increase glomerular filtration rate and sodium excretion. Collectively, these observations suggest regulatory influence of kinins during conditions featuring increased renal kinin levels.
Role of xanthine oxidase in ischemia/reperfusion injury
Authors: Linas S. L. , Whittenburg D. , Repine J. E.
Published: December 2017

Oxygen metabolites formed during reperfusion of ischemic kidneys prevent recovery of renal function after short periods of renal ischemia. Xanthine oxidase has been proposed as a source of toxic oxygen metabolites during reperfusion of ischemic kidneys. To determine whether the enzyme is converted from the non-oxygen metabolite-producing dehydrogenase (type D) to the oxygen metabolite-producing oxidase (type O), we measured type D and type O (total, reversible, and irreversible) xanthine oxidase in renal cortical homogenates after 30 min of ischemia in vivo and 60 min of reperfusion by the isolated perfused kidney technique. Total enzyme activity (type D plus type O) was not altered by ischemia or reperfusion. Compared with nonischemic conditions, ischemia increased total type O (53 +/- 5 vs. 21 +/- 3%, P less than 0.01) and reversible type O (15.4 +/- 1.5 vs. 2.1 +/- 1.4 U/g) xanthine oxidase activities. Reperfusion further increased total type O (82 +/- 3%) and reversible type O (27.7 +/- 3.3 U/g, both P less than 0.01 vs. nonischemic perfusions) xanthine oxidase activities. To determine the physiological role of xanthine oxidase in renal ischemia, we depleted rats of xanthine oxidase by feeding tungsten. After 4 wk of tungsten, renal xanthine oxidase levels were reduced by greater than 90% and renal function was markedly improved during reperfusion.(ABSTRACT TRUNCATED AT 250 WORDS)
Glomerular effects of angiotensin II require intrarenal factors
Authors: Wiegmann T. B. , MacDougall M. L. , Savin V. J.
Published: December 2017

Glomerular ultrafiltration coefficient (Kf) of glomeruli isolated from kidneys of normovolemic rats decreases following infusion of angiotensin II (ANG II). Kf from isolated glomeruli after ANG II infusion in vivo and from isolated perfused kidneys following infusion of ANG II in vitro was measured to determine whether the decrease required the presence of systemic factors. Filtration was induced in vitro and the maximum rate of change in glomerular volume was used to calculate Kf. Glomerular capillary hydraulic conductivity (Lp) was calculated from Lp = Kf/A where the basement membrane area A was calculated as 3 X pi X D2. ANG II infusion in vivo in rats diminished Lp from 3.19 +/- 0.19 to 1.96 +/- 0.13 and to 1.82 +/- 0.11 microliters.min-1.mmHg-1.cm-2, respectively. ANG II infusion into isolated kidneys caused a similar decrease in Lp (3.55 +/- 0.11 to 2.37 +/- 0.07). ANG II infusion either in vivo or during isolated kidney perfusion decreases Kf and Lp. ANG II effects do not require the presence of extrarenal factors but depend on perfusion in situ since incubation of isolated glomeruli with ANG II did not alter Kf.
Chronic hyperinsulinemia and blood pressure regulation
Authors: Hall J. E. , Coleman T. G. , Mizelle H. L. , Smith M. J.
Published: December 2017

The aims of this study were to determine whether chronic hyperinsulinemia, comparable to that found in obese hypertensives, elevates mean arterial pressure (MAP) or potentiates the hypertensive effects of angiotensin II (ANG II). Studies were conducted in conscious dogs with kidney mass reduced by 70% in order to increase their susceptibility to hypertensive stimuli. Insulin infusion (0.5 or 1.0 mU.kg-1.min-1 iv) for 7 days with plasma glucose held constant raised plasma insulin more than fivefold but did not increase MAP in four dogs on 138 meq/day Na intake. In seven dogs maintained on a high Na intake (319 meq/day), insulin infusion (1.0 mU.kg-1.min-1) for 28 days raised fasting insulin from 9.8 +/- 1.5 to 56-78 microU/ml but did not increase MAP, which averaged 106 +/- 2 mmHg during control and 102 +/- 2 mmHg during 28 days of insulin infusion. Insulin caused transient sodium and potassium retention followed by renal "escape" that was associated with increased glomerular filtration rate (12-27%). Plasma renin activity and plasma aldosterone were not altered by insulin. In five dogs infused with ANG II (2.0 ng.kg-1.min-1) to cause mild hypertension, insulin infusion (1.0 mU.kg-1.min-1) for 6-28 days did not increase MAP further. Thus chronic hyperinsulinemia did not elevate MAP, even when kidney mass was reduced, and did not potentiate the hypertensive effects of ANG II. These findings suggest that additional factors besides hyperinsulinemia per se are responsible for obesity-associated hypertension.
Basolateral transport of lactate in dog thick ascending limbs
Authors: Vinay P. , Senecal J. , Noel J. , Tejedor A. , Berteloot A. , Gougoux A.
Published: December 2017

Basolateral membrane vesicles (BLMV) isolated from both red outer medulla or from thick ascending limb segments isolated from the dog kidney were used to examine the process of lactate transport in this nephron segment. The BLMV preparation was enriched in Na(+)-K(+)-adenosinetriphosphatase (Na(+)-K(+)-ATPase) that represented 96% of the total ATPase activity of this preparation and the vesicles were largely under the right side-out orientation. On application of a OH- or HCO3- gradient (inside greater than outside), a secondary active lactate accumulation was observed, with characteristic transient overshoot. This phenomenon was shown to occur irrespective of the presence or absence of Na+, K+, or Cl-. The pH, but not the bicarbonate-driven, overshoot was abolished by nigericin (in presence of K+). Studies with valinomycin and K+ demonstrated that the generation of a membrane potential was not responsible for the acceleration of lactate transport, even if the amplitude of lactate accumulation was reduced in the presence of a bicarbonate gradient and valinomycin. A significant trans-stimulation of [14C]lactate transport by cold lactate was observed (under voltage-clamp condition). The transport was 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid insensitive but sensitive to furosemide (IC50 = 0.1 mM) and alpha-hydroxycyanocinnamate (IC50 = 1 mM). The kinetic parameters of the transporter revealed a single carrier with an apparent Michaelis constant of 1.7 mM and an apparent Vmax of 9.7 nmol.mg protein-1.30 s-1. The transporter was shown to be distinct from that of proximal tubule brush-border membrane or mitochondria (pyruvate). Thus thick ascending limbs possess a carrier-mediated lactate transport that can be used for lactate uptake (aerobic condition) or for lactate release (anaerobic glycolysis) according to metabolic processes imposed by the local oxygenation condition.
Comparative effects of diuretics and atrial peptide in chronic caval dogs
Authors: Levy M.
Published: December 2017

Anesthetized chronic caval dogs with ascites were given atrial natriuretic peptide (ANF) at 75 ng.kg-1.min-1 iv and were characterized as to natriuretic response [n = 21; urinary excretion rate of Na (UNaV): 28-362 mu eq/min] or lack of response (n = 14; UNaV: 0-10 mu eq/min). Sixteen normal dogs showed an increment in UNaV between 38 and 288 mu eq/min following the same dose of ANF. Both caval "responders" and "nonresponders" were equivalent with regard to pre- and post-ANF hemodynamics and renal function. When various diuretic agents (glucagon, acetazolamide, furosemide, thiazide, amiloride) were administered, equivalent natriuretic responses were obtained between both groups of caval dogs and the controls. An acute saline load (7% body wt), however, produced a greater natriuresis in controls than in caval dogs, but in this latter group the response was similar in responders and nonresponders. When six caval dogs each were initially pretreated with either amiloride or ANF, and then treated with a second agent, an additive natriuretic effect was obtained. These results suggest that tubular resistance to diuretics in caval dogs is unique to ANF, and this peptide appears to act at an additional Na transport site separate from amiloride-inhibitable channels.
Gluco- and mineralocorticoids control adenylate cyclase in specific nephron segments
Authors: Doucet A. , Barlet-Bas C. , Siaume-Perez S. , Khadouri C. , Marsy S.
Published: December 2017

Adrenal insufficiency is associated with an impairment of kidney diluting and concentrating ability, defects that may result from alterations of vasopressin-induced adenosine 3',5'-cyclic monophosphate (cAMP) production. The purpose of this study were 1) to localize the sites of decreased vasopressin-stimulated adenylate cyclase (AC) activity along the nephron of adrenalectomized rats; 2) to determine whether the response of AC to other hormones is altered by adrenalectomy; 3) to evaluate whether changes in AC are due to the deficiency in mineralocorticoids and/or glucocorticoids; and 4) to characterize the mechanism of action of corticosteroids on the AC system. Results indicate that adrenalectomy reduced AC stimulation by vasopressin, glucagon, and calcitonin in the thick ascending limb, whereas only the response to vasopressin decreased in the collecting tubule. Glucocorticoid administration curtailed adrenalectomy-induced alterations of AC in the thick ascending limb, whereas that in the collecting tubule was prevented by mineralocorticoids. Adrenalectomy did not alter forskolin-stimulated AC, whereas it decreased responses to aluminum fluoride and cholera toxin. Finally, alterations of fluoride- and cholera toxin-stimulated AC were prevented by glucocorticoid and mineralocorticoid repletion in the thick ascending limb and collecting tubule, respectively.
Total CO2 transport in rat cortical collecting duct in chloride-depletion alkalosis
Authors: Gifford J. D. , Sharkins K. , Work J. , Luke R. G. , Galla J. H.
Published: December 2017

Previous studies in chloride-depletion metabolic alkalosis (CDA) generated by intraperitoneal dialysis have suggested major alterations in chloride and bicarbonate transport beyond the distal convoluted tubule. To investigate the possible role of the cortical collecting duct (CCD) in the pathophysiology of CDA, isolated CCD segments were perfused in vitro from either control (CON) rats dialyzed against Ringer-bicarbonate or those made alkalotic by peritoneal dialysis with 0.15 M NaHCO3. Tubules from CDA animals secreted CO2 for greater than or equal to 3 h after dissection (-22.4 +/- 7.2 pmol.mm-1.min-1) compared with CON tubules that absorbed CO2 (18.3 +/- 4.2 pmol.mm-1.min-1). Replacement of luminal chloride with gluconate in the perfusate abolished net total CO2 (tCO2) secretion in tubules from CDA animals (from -21.5 +/- 4.5 to -2.7 +/- 2.3 pmol.mm-1.min-1) but did not alter net tCO2 absorption in tubules from CON animals. In contrast, removal of bath chloride increased net tCO2 secretion (-12.1 +/- 2.9 to -26.1 +/- 3.6 pmol.mm-1.min-1) in CDA tubules, whereas net tCO2 flux was altered from absorption to secretion in CON tubules (15.5 +/- 4.0 to -13.6 +/- 9.2 pmol.mm-1.min-1). These results demonstrate that 1) CDA generated in vivo within 45 min results in stable net tCO2 secretion in vitro up to 240 min in the CCD; 2) luminal chloride is necessary for tCO2 secretion; 3) the shift of net tCO2 flux from absorption to secretion in CON tubules in vitro was not sustained in contrast to CDA tubules.(ABSTRACT TRUNCATED AT 250 WORDS)
Intracellular pH regulates Na(+)-independent Cl(-)-base exchange in JTC-12 (proximal tubule) cells
Authors: Fineman I. , Hart D. , Nord E. P.
Published: December 2017

The role of an anion exchange pathway in the regulation of intracellular pH (pHi) under alkaline load and steady-state conditions and the modulation of this transporter by pHi was investigated in confluent monolayers of cloned JTC-12 cells, derived from monkey kidney proximal tubule. Regulation of pHi was fluorometrically monitored using the pH-sensitive probe, 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Monolayers in which pHi was rapidly elevated by removal of HCO3(-)-CO2 from the bathing medium demonstrated an absolute requirement for Cl- to recover toward base-line pHi. The recovery process proceeded in the absence of Na+, was inhibited 80% by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, and was unaffected by 10(-4) M amiloride. When extracellular pH (pHo) was serially lowered from 7.4 to 6.7, recovery from an alkaline load induced by removal of HCO3(-)-CO2 from the medium occurred only when pHi was elevated above approximately 7.25. Below pHi approximately 7.25 no recovery toward initial pHi was observed. When pHi was elevated above approximately 7.25 with pHo maintained at 6.7, the recovery process ceased at pHi approximately 7.25 despite favorably oriented Cl- and OH- chemical gradients. Consistent with these observations, removal of Cl- from the medium of cells buffered with 25 mM HCO3(-)-5% CO2 at pHo 7.4 (in the absence of Na+) resulted in reversible elevation of pHi, whereas in a solution buffered to pHo 6.7 with 5 mM HCO3(-)-5% CO2, removal of Cl- failed to elevate pHi. Under steady-state conditions in the presence of 25 mM HCO3(-)-5% CO2 at pHo 7.4, pHi was 7.40 +/- 0.02 and reversibly decreased to 7.23 +/- 0.01 on removal of Na+ (in the presence of amiloride) from the bathing medium, indicating that the Cl(-)-base exchanger is operative under basal conditions and functions as a base extruder. In summary the JTC-12 cell possesses a Na(+)-independent Cl(-)-base exchange mechanism that is operative under alkaline load and steady-state conditions. pHi but not pHo modulates the activity of this transport pathway, and below pHi approximately 7.25 the exchanger is quiescent.
Influence of humoral and volume factors on altered osmoregulation of normal human pregnancy
Authors: Davison J. M. , Shiells E. A. , Philips P. R. , Lindheimer M. D.
Published: December 2017

These studies were designed to characterize mechanisms leading to decreased plasma osmolality (Posmol) and osmotic thresholds (T) for arginine vasopressin (AVP) release (TAVP) and thirst (Tthirst) in pregnancy. First, the influence of the pregnancy hormone, human chorionic gonadotrophin (hCG), was tested in six nonpregnant women who received hypertonic saline during the luteal phase of the menstrual cycle on two occasions (randomly allocated), once after 15,000 IU of hCG when Posmol, TAVP, and Tthirst decreased 6, 5, and 5 mosmol/kgH2O, respectively (P less than 0.01). In contrast, hCG pretreatment of males (n = 6) had no significant effect. Next, the role of decreased effective vascular volume (underfilling) was evaluated in seven women undergoing hypertonic saline infusion in the presence and absence of head-out water immersion (randomly allocated) during early and late pregnancy and postpartum. Posmol, TAVP, and Tthirst were not influenced by immersion and remained 10 mosmol/kgH2O lower in pregnancy (P less than 0.01). Central redistribution of intravascular volume consistently lowered hematocrit and rate of rise of PAVP per unit increment in Posmol (P less than 0.01). Although these data failed to support the hypothesis that the osmoregulatory change in human pregnancy is attributable to decrements in effective central volume (underfill), they do suggest that hCG may play a role.
Sulfated bile acids inhibit Na(+)-H+ antiport in human kidney brush-border membrane vesicles
Authors: Sellinger M. , Haag K. , Burckhardt G. , Gerok W. , Knauf H.
Published: December 2017

Patients with obstructive jaundice are at a high risk for acute renal failure after surgery. Direct toxic membrane effects of bile acids or bilirubin have been discussed as possible causes. Therefore, we investigated the influence of bile acids and conjugated bilirubin on Na(+)-H+ antiport and ion permeabilities in brush-border membrane vesicles isolated from the human kidney. Brush-border membrane vesicles were prepared by Mg2+ precipitation. These were highly purified as estimated from the 14-fold enrichment in the specific activity of alanine aminopeptidase. The pH-sensitive dye acridine orange was used to study the properties of proton uptake under different conditions. The brush-border membrane vesicles from human kidney cortex exhibited Na+ and K+ conductances, which were small compared with H+ conductance. Furthermore, these membranes possess an Na(+)-H+ antiporter that is sensitive to amiloride. Various bile acids (30 microM) had no significant effect on Na(+)-H+ antiport. However, the addition of sulfated bile acids resulted in a significant inhibition (greater than 50%) of the Na(+)-H+ antiporter. A nonspecific effect of sulfated bile acids on the vesicles was excluded by the use of ionophores to determine vesicle integrity and to estimate the various ion conductances. Therefore specific inhibition of the human renal Na(+)-H+ antiporter by sulfated bile acids occurs. This could result in an impaired cellular pH regulation and might play a role in postoperative acute renal failure in patients with obstructive jaundice.
Role of diacylglycerol in adrenergic-stimulated 86Rb uptake by proximal tubules
Authors: Baines A. D. , Drangova R. , Ho P.
Published: December 2017

We used rat proximal tubule fragments purified by Percoll centrifugation to examine the role of diacylglycerol (DAG) in noradrenergic-stimulated Na+ reabsorption. Tubular DAG concentration and ouabain-inhibitable 86Rb uptake increased within 30 s after adding norepinephrine (NE) and remained elevated for at least 5 min. NE (1 microM) increased DAG content 17% and ouabain-inhibitable 86Rb uptake 23%. Cirazoline-stimulated 86Rb uptake was not inhibited by BaCl, quinidine, or bumetanide (1-10 microM) or by the omission of HCO3- or Cl- from the medium, but it was completely inhibited by ouabain and furosemide. Oleoyl-acetyl glycerol, L-alpha-1,2-dioctanoylglycerol, and L-alpha-1,2-dioleoylglycerol (DOG) increased total 86Rb uptake 8-11%. 12-O-tetradecanoylphorbol-13-acetate (TPA) (5 nM) increased uptake by only 4%. Staurosporine at 5 nM inhibited DOG stimulation completely, whereas 50 nM staurosporine was required to inhibit NE stimulation completely. Sphingosine inhibited DOG stimulation by 66% but did not inhibit NE stimulation. Amiloride (1 mM) completely blocked DOG stimulation. Monensin increased 86Rb uptake 31% and completely blocked the DOG effect but reduced the NE effect by only 26% (P = 0.08). In tubules from salt-loaded rats, NE did not increase DAG concentration, but NE-stimulated 86Rb uptake was reduced by only 23% (P = 0.15). Thus DAG released by NE may stimulate Na+ entry through Na(+)-H+ exchange. NE predominantly stimulates Na(+)-K(+)-adenosinetriphosphatase (ATPase) by activating a protein kinase that is insensitive to DAG and TPA and is inhibited by staurosporine but not by sphingosine. NE may also stimulate K+ efflux through a BaCl-insensitive K+ channel that is inhibited by millimolar furosemide.(ABSTRACT TRUNCATED AT 250 WORDS)
Cyclooxygenase-dependent mediators of renal hemodynamic function in female rats
Authors: Munger K. A. , Blantz R. C.
Published: December 2017

Previous studies have revealed a sex-dependent difference in response to cyclooxygenase inhibition in anesthetized male and female rats. Female rats have shown an unexpected vasodilation in response to prostaglandin (PG) inhibition. The present studies were designed to further investigate the sex-dependent role of the PG system in the control of normal renal hemodynamics in female Munich-Wistar rats. Renal hemodynamic studies were performed on anesthetized female rats before and during acute cyclooxygenase inhibition using a variety of protocols. One group underwent subacute unilateral renal denervation. A separate group was chronically catheterized and plasma catecholamines were measured during the awake state and then after anesthesia under the euvolemic protocol. Another group was administered the angiotensin II blocker, saralasin, before and during cyclooxygenase inhibition. In a final group, flow to the distal nephron was interrupted via placement of a wax block into the late proximal tubule to determine the role of distal nephron flow and tubuloglomerular feedback in the glomerular response to cyclooxygenase inhibition. It was determined that neither the wax block nor saralasin administration attenuated the vasodilatory response observed in normal female rats due to cyclooxygenase inhibition; however, subacute unilateral renal denervation completely blocked the vasodilatory response to PG inhibition in these female Munich-Wistar rats. Plasma catecholamines were found to be similar whether awake or under anesthesia. These studies indicate the importance of the adrenergic system in modulating PG production in the acutely anesthetized intact female rat.
Control of renal function during intrarenal infusion of endothelin
Authors: Stacy D. L. , Scott J. W. , Granger J. P.
Published: December 2017

It has been demonstrated that bolus injections of a vasoconstrictor derived from endothelial cells, endothelin 1 (ET-1), constricts isolated arteries and increases blood pressure in animals when infused intravenously. The purpose of this study was to examine the direct effects of intrarenal infusions of ET-1 on renal function at doses that do not alter systemic arterial pressure. The effects of ET-1 on renal hemodynamics and electrolyte excretion were examined during 40 min of intrarenal infusions of ET-1 at rates of 1.15 and 5 ng.kg-1.min-1. Infusion of ET-1 (1.15 ng.kg-1.min-1) resulted in a transient increase in renal blood flow (RBF) followed by a progressive vasoconstriction, which reduced RBF by 23%. ET-1 decreased glomerular filtration rate (GFR) and had no significant effect on filtration fraction. Intrarenal infusion of ET-1 (1.15 ng.kg-1.min-1) had no effect on fractional excretion of sodium (FENa) or potassium. Infusion of ET-1 at a higher dose (5 ng.kg-1.min-1) produced further reductions in RBF, GFR, and FENa. These data indicate that ET-1 is a potent renal vasoconstrictor that could play a role in controlling renal hemodynamics.
Glomerular endothelial cells respond to calcium-mobilizing agonists with release of EDRF
Authors: Marsden P. A. , Brock T. A. , Ballermann B. J.
Published: December 2017

To determine whether glomerular endothelial cells (GEN) may play a role in the local control of glomerular function by releasing endothelium-derived relaxing factor (EDRF), the effect of several agonists on GEN cytosolic calcium concentration ([Ca2+]i) and GEN EDRF release was determined. Bradykinin, ATP, thrombin, and platelet-activating factor (PAF) all increased [Ca2+]i in GEN in a concentration-dependent manner, whereas serotonin, acetylcholine, phenylephrine, and endothelin-1 were without effect. Coincubation of glomerular mesangial cells (GMC) with GEN augmented mesangial cell guanosine 3',5'-cyclic monophosphate (cGMP) content five- to sixfold, Bradykinin elicited a further concentration-dependent increase in GMC cGMP content in the presence but not absence of GEN. The GEN-dependent bradykinin-stimulated GMC cGMP accumulation was abolished by hemoglobin and methylene blue, blunted by gossypol, and augmented by superoxide dismutase. Other agonists capable of augmenting GEN [Ca2+]i also stimulated GMC cGMP accumulation in the presence but not in the absence of GEN. Thus cultured GEN release a factor that stimulates cGMP accumulation in adjacent mesangial cells which has the pharmacological characteristics of EDRF.
Oxygen consumption and oxidant stress in surviving nephrons
Authors: Nath K. A. , Croatt A. J. , Hostetter T. H.
Published: December 2017

We investigated the effects of reduction of renal mass on rates of oxygen consumption, sodium transport, and indexes of oxidant stress in surviving nephrons. Rates of oxygen consumption in surviving nephrons were elevated by more than twofold compared with nephrons in intact kidneys in rats on standard protein intakes. Absolute rates of sodium reabsorption (TNa) in the surviving nephrons were increased with a lower ratio of TNa to oxygen consumption. To determine oxidant stress, we measured malondialdehyde (MDA) in the kidney and urine and the glutathione redox ratio in kidney tissue. MDA per nephron was increased in the subtotally nephrectomized model and was accompanied by increased absolute and fractional urinary excretion of MDA but not by an increase in kidney MDA per milligram protein. The glutathione redox ratios were similar. Since increased dietary protein intake worsens renal injury, we studied the effects of dietary protein manipulation (30 vs. 6%) on oxygen consumption, MDA levels, and the glutathione redox ratio. The kidneys of subtotally nephrectomized animals maintained on 30% protein diets exhibited increased rates of oxygen consumption. Increased dietary protein intake led to increased MDA per nephron, increased urinary excretion of MDA, and increased MDA per milligram protein in subtotally nephrectomized animals, and markedly increased the glutathione redox ratio. We conclude that, despite increased oxygen consumption, surviving nephrons compared with intact nephrons in rats on standard protein intake demonstrate no evidence of oxidant stress. Increased urinary clearance of MDA may provide a mechanism that prevents the buildup of lipid peroxidation. Subjecting the remnant nephron to increased protein increases oxygen consumption and imposes oxidant stress.(ABSTRACT TRUNCATED AT 250 WORDS)
Detection of specific mRNAs in single nephron segments by use of the polymerase chain reaction
Authors: Moriyama T. , Murphy H. R. , Martin B. M. , Garcia-Perez A.
Published: December 2017

We have developed a procedure to detect specific mRNAs in single renal nephron segments. This approach combines microdissection, reverse transcription (RT) of the target mRNA, and amplification of the resulting cDNA using the polymerase chain reaction (PCR). After microdissection, the sample is placed in a tube where it is permeabilized and where all reactions are performed directly without the need for isolation of the RNA. Our model target was the mRNA for aldose reductase. This enzyme catalyzes the conversion of glucose to sorbitol. Its expression is modulated by changes in extracellular osmolality in the renal medulla. RT-PCR of inner medullary collecting duct (1 mm) and glomeruli (6-10) yielded a product of the predicted length (670 base pairs) defined by the PCR primers. Its identity was confirmed by a specific oligonucleotide probe that differed from the primers. RT-PCR of proximal tubules (1 mm) resulted in no aldose reductase-specific amplification product. RT-PCR is generally applicable for measuring specific gene expression in single nephron segments or small numbers of cultured cells. Utility, limitations, and refinements of this approach are discussed.
Mechanism of organic cation transport in rabbit renal basolateral membrane vesicles
Authors: Sokol P. P. , McKinney T. D.
Published: December 2017

The transports of [3H]tetraethylammonium (TEA), [3H]procainamide (PCA), and N1-[3H]methylnicotinamide (NMN) were studied in rabbit renal basolateral membrane vesicles (BLMVs) by use of a rapid filtration assay. All three compounds exhibited a similar uptake profile into the BLMVs and reached equilibrium by 1 h. In the presence of valinomycin, a K+ ionophore and K+ gradients (in to out), an inside-negative potential difference (PD) was generated that stimulated the uptake of TEA, PCA, and NMN by 1.9-, 1.9-, and 2.1-fold, respectively. The effect of PD could be blocked by the organic cation transport inhibitor mepiperphenidol. An inside-negative PD was also generated by a pH gradient (inside acidic). An overshoot of TEA uptake was produced, which was blocked by a valinomycin voltage clamp. Counterflow studies revealed that 1 mM TEA was capable of trans-stimulating 50 microM [3H]TEA uptake and producing a peak overshoot of nearly three times the equilibrium value, which was not abolished in the presence of a valinomycin voltage clamp or a gramicidin pH clamp. When an inside-negative PD was imposed on 1 mM TEA-loaded BLMVs, the uptake of [3H]TEA was 33% less. In contrast, neither NMN nor PCA produced a trans-stimulation of [3H]NMN or [3H]PCA transport, respectively. In addition, the effect of several organic cations on the TEA-TEA exchange mechanism was studied. Mepiperphenidol, PCA, choline, cimetidine, and NMN all demonstrated cis inhibition (82, 81, 58, 51, and 20%, respectively). Arginine, a basic amino acid, and probenecid, an organic anion transport inhibitor, had no effect. Choline was capable of trans-stimulating TEA uptake.(ABSTRACT TRUNCATED AT 250 WORDS)
Developmental changes of Ca2+, PO4, and calcitriol metabolism in spontaneously hypertensive rats
Authors: Bourgouin P. , Lucas P. , Roullet C. , Pointillart A. , Thomasset M. , Brami M. , Comte L. , Lacour B. , Garabedian M. , McCarron D. A. , et al.
Published: December 2017

We have measured Ca and P balance, serum calcitriol, and vitamin D-dependent intestinal calcium-binding protein (CaBP9k) in the spontaneously hypertensive rat (SHR) and its normotensive control, the Wistar-Kyoto rat (WKY) at 4–5 and again at 13–14 wk of age. In rats on a 1% Ca diet, P balance was significantly more positive in the 5-wk-old SHR than WKY (P less than 0.01); Ca balance tended to be greater in the 5-wk-old SHR. In contrast, in the 14-wk-old SHR, P and Ca balance were less positive than in the WKY (P was 5.8 +/- 1.3 vs. 13 +/- 1.7 mg/day, P less than 0.01, and Ca was 53 +/- 4.6 vs. 67 +/- 2.7 mg/day, P less than 0.05). On the 1% Ca diet, plasma calcitriol levels of the 5-wk-old SHR were higher than those of the WKY (58 +/- 3.2 vs. 40 +/- 2.1 pg/ml, P less than 0.002), whereas at 12 wk there was no difference. On Ca-deficient (0.1%) diets, plasma calcitriol was increased in 5-wk-old and 12-wk-old SHR and WKY, compared with the 1% Ca diet (P less than 0.001). Calcitriol metabolic clearance rate was the same in the 13-wk-old SHR and WKY on either Ca diet. Intestinal CaBP9k content of the 5-wk-old (38 +/- 2.6 vs. 40 +/- 3.9 micrograms/mg) and the 12-wk-old (11 +/- 1.6 vs. 14 +/- 1.4 micrograms/mg) SHR and WKY were similar on the 1% Ca diet.(ABSTRACT TRUNCATED AT 250 WORDS)
Physiology and biochemistry of vitamin D-dependent calcium binding proteins
Authors: Gross M. , Kumar R.
Published: December 2017

The vitamin D-dependent calcium binding proteins (calbindins) are members of the troponin-C superfamily of proteins that occur in a number of calcium-transporting tissues such as the intestine, the distal tubule of the kidney, and the placenta. They are also present in other tissues such as the brain, peripheral nervous system, pancreas, parathyroid gland, and bone. In some tissues, such as the adult brain, the proteins occur in the absence of the vitamin. The proteins bind calcium in "EF" hand structures and are "calcium-sensitive" in that they undergo a conformational change on binding calcium. They appear to enhance transcellular calcium transport and are frequently present in tissues that contain the plasma membrane calcium pump.
Renal microvascular effects of endothelin
Authors: Edwards R. M. , Trizna W. , Ohlstein E. H.
Published: December 2017

The effects of endothelin 1, 2, and 3 (ET-1, -2, -3) on lumen diameter of individual afferent and efferent arterioles dissected from rabbit kidney were examined. ET-1 produced concentration-dependent and long-lasting decreases in lumen diameter in both arterioles. The 50% maximum response (EC50) values were 1.4 +/- 0.41 and 0.9 +/- 0.65 nM for afferent and efferent arterioles, respectively. In afferent arterioles, ET-2 produced decreases in lumen diameter (EC50 = 3.3 +/- 1.75 nM) that were indistinguishable from ET-1. However, ET-3 was considerably less potent (EC50 = 21.9 +/- 6.0 nM, P less than 0.05) than ET-1 or ET-2. Similar results were obtained in the efferent arteriole in which the EC50 for ET-2 (0.25 +/- 0.1 nM) was similar to ET-1, but ET-3 was significantly less potent (EC50 = 2.6 +/- 0.4 nM, P less than 0.05). Nicardipine (0.01-1 microM) produced concentration-dependent shifts in the ET-1 concentration-response curve in afferent arterioles. Verapamil (1 microM) also caused a significant shift in the ET-1 response curve. The contractile response to ET-1 was significantly more sensitive to nicardipine than was the response to norepinephrine. In contrast, the response of efferent arterioles to ET-1 and norepinephrine was unaffected by nicardipine or verapamil. The results demonstrate that ETs are potent vasoconstrictors of both the pre- and postglomerular microvasculature and may play a role in the regulation of renal hemodynamics.(ABSTRACT TRUNCATED AT 250 WORDS)
In vivo microperfusion of inner medullary collecting duct in rats: effect of amiloride and ANF
Authors: Sonnenberg H. , Honrath U. , Wilson D. R.
Published: December 2017

A method is described that allows perfusion of the inner medullary collecting duct (IMCD) of the rat kidney in situ and in vivo. Fine polyethylene catheters connected to a microperfusion pump were inserted into collecting ducts via the openings at the exposed papilla tip. Perfusate contained 22Na as well as [3H]inulin. During perfusion at 30 nl/min, urine was simultaneously collected. A decrease in the Na-to-inulin concentration ratio in the urinary sample, compared with the perfusate, was taken as indicating unidirectional efflux of Na from the perfused duct system. The effects of luminal amiloride (2 X 10(-4) M) or atrial natriuretic factor (ANF, 10(-8) M) were studied. Compared with control perfusions, both agonists reduced Na efflux from the IMCD to approximately 50%, indicating luminal sites of action. Combination of amiloride and ANF at their respective concentrations had no further effect. The lack of statistically significant additivity suggests, but does not prove, that ANF, administered from the luminal side, is able to block amiloride-sensitive Na channels in the apical membrane of IMCD cells.
Atrial peptide and cGMP effects on NaCl transport in inner medullary collecting duct
Authors: Rocha A. S. , Kudo L. H.
Published: December 2017

We examined the action of atrial natriuretic factor (ANF) on Na+ and Cl- transport in in vitro microperfused inner medullary collecting ducts (IMCD) isolated from rat kidneys. First we studied the isotopic fluxes at low perfusion rates (7 nl/min). The results showed that ANF added to bath decreased lumen-to-bath flux (Jl----b) of Na+ and increased Na+ bath-to-lumen flux (Jb----l). This was substantiated by a direct demonstration that ANF reduces net Na+ and Cl- absorption. The effect of ANF on Jl----b and Jb----l of Na+ was also observed at high perfusion rates (25 nl/min). The inhibitory effect of ANF was observed even when Na+ Jl----b was stimulated by vasopressin (VP). ANF (6 x 10(-11) M) added to bath increased Cl- Jb----l and generated a negative lumen potential difference (PD). These two effects were inhibited by furosemide and by the replacement of Na+ by choline and Cl- by SO4(2-) in the bath fluid. These observations are compatible with the existence of a Na(+)-Cl(-)-K+ cotransport mechanism stimulated by ANF. Moreover, the effects of guanosine 3',5'-cyclic monophosphate (cGMP) added to the bath on PD, Jl----b, and Jb----l of Na+ were similar to those observed with ANF. Thus, physiological concentrations of ANF inhibit directly Na+ and Cl- absorption in IMCD by two mechanisms, 1) by increasing cotransport Na(+)-Cl(-)-K+ secretion and 2) by inhibiting NaCl absorption both in the absence and in the presence of VP. These effects on NaCl transport appear to be mediated by cGMP.
Endothelin in experimental congestive heart failure in the anesthetized dog
Authors: Cavero P. G. , Miller W. L. , Heublein D. M. , Margulies K. B. , Burnett J. C.
Published: December 2017

Studies were performed in anesthetized dogs to determine plasma endothelin (ET) concentrations in the presence and absence of experimental congestive heart failure (CHF) produced by rapid ventricular pacing for 8 days. These studies were also designed to determine the effect of exogenous low-dose ET upon integrated cardiorenal and endocrine function in the presence and absence of CHF. In these studies, plasma ET was significantly elevated in CHF (3.25 +/- 0.39 pg/ml) compared with normal (1.03 +/- 0.21 pg/ml) or sham-operated (1.08 +/- 0.27 pg/ml) groups. Compared with the control group, which was characterized by a significant cardiorenal vasoconstrictor response to low-dose ET, a significant attenuation of the vasoconstrictor and antinatriuretic actions of ET was observed in the CHF group. Despite these differential responses, exogenous ET suppressed plasma renin activity (PRA) and activated aldosterone in both control and CHF groups. Thus these studies demonstrate for the first time that experimental CHF is characterized by elevated plasma ET in association with an attenuated cardiorenal response to exogenous ET. In contrast, low-dose ET inhibited PRC and activated aldosterone in the presence and absence of experimental CHF.
Epidermal growth factor accelerates renal repair in mercuric chloride nephrotoxicity
Authors: Coimbra T. M. , Cieslinski D. A. , Humes H. D.
Published: December 2017

Repair and recovery of ischemic or nephrotoxic acute renal failure (ARF) are dependent upon renal tubule cell regeneration. Because epidermal growth factor (EGF) is a potent growth promoter to renal tubule cells, experiments were undertaken to assess the effects of exogenous administration of EGF during the recovery phase of HgCl2-induced ARF. Rats were administered HgCl2 (5 mg/kg sc), and [3H]thymidine incorporation into renal tissue and blood urea nitrogen (BUN) and serum creatinine concentrations were measured at various times after toxin administration. EGF (20 microgram) was administered subcutaneously 2 or 4 h after HgCl2 injection. Exogenous EGF resulted in greater levels of renal [3H]thymidine incorporation into renal proximal tubule cells compared with those observed in nontreated animals at several time points in the first 48 h after toxic injury. Morphometric analysis of histoautoradiograph sections of renal tissue demonstrated that greater than 96% of labeled cells were tubular in all examined sections. This EGF-related acceleration in DNA synthesis was associated with significantly lower peak BUN and serum creatinine levels, averaging 213 +/- 23 and 6.54 +/- 0.72 (SE) mg/dl, respectively, at 3 days in EGF-treated nephrotoxic rats compared with peak levels of 359 +/- 40 and 9.92 +/- 1.67 mg/dl (P less than 0.001, n = 7-16) at 5 days in non-EGF-treated nephrotoxic rats. EGF treatment also was associated with a return to near normal BUN and serum creatinine levels approximately 4 days earlier than that observed in non-EGF-treated animals. These findings demonstrate that exogenous EGF accelerates the repair process of the kidney after a severe toxic insult.
Acidification mechanisms on the basolateral membrane of renal vesicles from the developing nephron
Authors: Baum M.
Published: December 2017

The present study examined acidification mechanisms on the basolateral membrane of the early renal vesicle, an undifferentiated ball of cells that will develop into parts of the glomerulus, proximal tubule, loop of Henle, and a portion of the distal convoluted tubule. Renal vesicles were dissected from newborn rabbit kidneys and bathed in vitro. To examine the basolateral membrane acidification mechanisms, intracellular pH (pHi) was measured by use of the pH-sensitive dye (2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. Evidence for Cl(-)-base exchange included the fact that removal of bath Cl-resulted in cell alkalinization (7.35 +/- 0.03 to 7.48 +/- 0.05; P less than 0.01). Cell alkalinization induced by Cl- removal was also observed in presence of a voltage clamp without bath Na+ (7.18 +/- 0.02 to 7.39 +/- 0.04; P less than 0.01) and was inhibited by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). pHi recovery after acute alkalinization resulting from CO2 removal was 0.20 +/- 0.03 pH units/min in the presence of Cl-, 0.07 +/- 0.01 in experiments with 0.2 mM DIDS, and 0.07 +/- 0.01 in absence of bath Cl-. The early renal vesicle also has a basolateral Na(+)-H+ antiporter. Removal of bath Na+ resulted in cell acidification (7.36 +/- 0.09 to 7.18 +/- 0.06; P less than 0.01), which was inhibited by 2 mM amiloride. Cell pH recovery after acute acidification (NH4Cl prepulse technique) was entirely dependent on bath Na+ and inhibited by amiloride. Thus the renal vesicle has basolateral membrane Na(+)-H+ and Cl(-)-base exchangers that can defend against cell acidification and alkalinization, respectively.
Thiazide-sensitive NaCl absorption in rat cortical collecting duct
Authors: Terada Y. , Knepper M. A.
Published: December 2017

The mechanism of transepithelial NaCl transport was investigated in isolated perfused cortical collecting ducts from the kidneys of deoxycorticosterone-treated rats. In the presence of vasopressin, hydrochlorothiazide (0.1 mM) markedly reduced the net rate of Na absorption, Cl absorption, and active fluid absorption but did not significantly change the transepithelial voltage. Similarly, in the absence of vasopressin, hydrochlorothiazide decreased the rate of sodium absorption by 50% without affecting transepithelial voltage. Amiloride (30 microM) completely eliminated the lumen-negative voltage but decreased net sodium absorption only by approximately 50%. In the presence of amiloride, chloride absorption occurred against an electrochemical gradient for chloride, indicating that there was active chloride absorption. Bumetanide (0.1 mM) did not affect chloride absorption or spontaneous fluid absorption in the presence of vasopressin. The combination of amiloride and hydrochlorothiazide inhibited net sodium absorption by a greater extent than did either agent alone. These results demonstrate the presence of the following two parallel sodium transport pathways in cortical collecting ducts from mineralocorticoid-replete rats: 1) an electrogenic pathway blocked by amiloride, which presumably involves an apical sodium channel, and 2) a thiazide-inhibitable electroneutral pathway, which presumably utilizes apical Na-Cl cotransport and mediates secondary active transport of chloride.
Effects of nifedipine and enalapril on glomerular injury in rats with deoxycorticosterone-salt hypertension
Authors: Dworkin L. D. , Levin R. I. , Benstein J. A. , Parker M. , Ullian M. E. , Kim Y. , Feiner H. D.
Published: December 2017

Male Munich-Wistar rats underwent right nephrectomy and were given weekly injections of deoxycorticosterone acetate (DOCA) and 1% saline (salt) to drink. Two studies were performed. In the first, rats given enalapril (ENP) were compared with controls. In the second, rats ingested either standard chow or chow to which the calcium-entry blocker nifedipine (NIF) had been added. Six to eight weeks after nephrectomy, both control DOCA-salt rats and those given ENP had severe hypertension and significant proteinuria. Rats given NIF excreted less protein, and glomerular lesions were not observed in this group. The effects of NIF on several parameters that have been associated with glomerular injury were examined. Micropuncture studies revealed that glomerular capillary pressure was increased in DOCA-salt rats and was not reduced by NIF. Platelet aggregation was also similar in NIF-treated and control rats. Morphometric studies revealed a tendency toward lower glomerular volume of NIF-treated rats; however, kidney weight and glomerular capillary radius were unaffected by therapy. Thus NIF, but not ENP, prevents DOCA-salt rats from developing hypertension and glomerular injury. This effect does not depend on reduction in glomerular pressure or inhibition of platelet aggregation.