bluuebunny/crossref_metadata_2025 huggingface dataset:
Renal brush-border chloride transport mechanisms characterized using a fluorescent indicator
Authors: Chen P. Y. , Illsley N. P. , Verkman A. S.
Published: December 2017

Cl transport mechanisms in brush-border membrane vesicles (BBMV) isolated from rabbit renal cortex were characterized using the Cl-sensitive fluorescent indicator 6-methoxy-N-[3-sulfopropyl]quinolinium (SPQ). In control experiments using 36Cl, SPQ did not itself alter rates for Cl transport. Cl fluxes (JCl) in nanomoles per second per milligram vesicle protein in response to gradients of Cl and other ions were calculated from the SPQ fluorescence time course, fluorescence vs. [Cl] calibration, and BBMV glucose space (microliter/mg protein). For a 50 mM inwardly directed Cl gradient in BBMV (voltage and pH clamped) JCl was 0.26 +/- 0.01 nmol.s-1.mg-1 (mean +/- SD, 4 vesicle preparations). JCl was not altered by a 50 mM inwardly directed Na gradient (0.25 +/- 0.01 nmol.s-1.mg-1) but increased to 0.52 +/- 0.04 nmol.s-1.mg-1 when driven by a 1.5 pH unit inwardly directed proton gradient. The pH driven increase in JCl was blocked by H2DIDS (100 microM, JCl = 0.38 +/- 0.02 nmol.s-1.mg-1). In the absence of a gradient, JCl increased by 64 +/- 4% in the presence of 1 mM formate on both sides of the membrane; the increase was blocked by H2DIDS. JCl increased to 0.38 +/- 0.02 nmol.s-1.mg-1 for a 60-mV K valinomycin diffusion potential (internal positive); the increase was inhibited by the Cl channel blocker diphenylamine-2-carboxylate (DPC, 100 microM, JCl = 0.26 +/- 0.03 nmol.s-1.mg-1). Br and I transport was also blocked by DPC.(ABSTRACT TRUNCATED AT 250 WORDS)
Modeling the proximal tubule: complications of the paracellular pathway
Authors: Weinstein A. M.
Published: December 2017

When the proximal tubule epithelium is represented as cellular and lateral intercellular (LIS) compartments, the presence of a paracellular pathway can render the overall phenomenologic equations quite an indirect representation of intraepithelial transport processes. 1) Active sodium transport into the LIS may create a hypertonic region that drives water movement from lumen to peritubular blood, i.e., a term for active water transport may appear in the overall transport equations. The correlate of this uphill water flux is a solute polarization effect, such that the measured epithelial water permeability is less than that of the cell membranes. 2) Basolateral uptake of potassium by the cell may lower the LIS concentration and promote diffusive entry of K across the tight junction. Even without cellular uptake of K from the lumen, the epithelial transport equations may contain a term for active K reabsorption. The solute polarization correlate is a low epithelial reflection coefficient that does not represent a convective flux of K through a specific channel. 3) When there is convective flux of Na and Cl through the tight junction but none through the cell, then a fluid circuit around junction and cell may be present, even when net epithelial volume flux is absent. In this case, part of the net epithelial Cl flux must be represented in the overall transport equations as electroneutral Na-Cl cotransport.
Decreased transport in renal basolateral membrane vesicles from hypertaurinuric mice
Authors: Mandla S. , Scriver C. R. , Tenenhouse H. S.
Published: December 2017

Basolateral membrane vesicles were prepared from mouse kidney by use of a Percoll density gradient method. The preparation was enriched ninefold in Na+-K+-ATPase with minimal contamination by other cellular membranes. The basolateral membranes were a mixture of sealed inside-out and right-side-out vesicles (30%) and leaky vesicles or sheets (70%). Taurine uptake into basolateral membrane vesicles was osmotically sensitive, sodium dependent, temperature sensitive, inhibited by beta-alanine, and saturable (apparent Km, 360 microM; Vmax, 25.4 pmol.mg protein-1.15 s-1), indicating transport by a carrier-mediated process. The function of this transporter was examined in an inbred mouse strain, C57BL/6J, which has selective hypertaurinuria, presumably a result of decreased basolateral membrane permeability to taurine [Rozen et al., Am. J. Physiol. 244 (Renal Fluid Electrolyte Physiol. 13): F150-F155, 1983]. The sodium-dependent component of taurine uptake was significantly lower in C57BL/6J vesicles relative to control (C3H/HeJ strain): 2.9 +/- 0.7 vs. 9.4 +/- 0.3 (SE) pmol.mg protein-1.15 s-1, respectively; P less than 0.001. The interstrain difference in uptake was specific for taurine and could not be ascribed to differences in vesicle purification, integrity, orientation, or size. These findings indicate that the renal basolateral membrane is the site of a transport defect, which explains decreased net taurine reabsorption in vivo in the C57BL/6J strain, and corroborate earlier observations in the renal cortical slice preparation.
Hormone effects on NaCl permeability of rat inner medullary collecting duct
Authors: Sands J. M. , Nonoguchi H. , Knepper M. A.
Published: December 2017

It has been proposed that regulation of NaCl excretion occurs in part by hormonal effects on NaCl permeability in the inner medullary collecting duct (IMCD). We carried out experiments in isolated perfused terminal IMCDs to determine whether atrial natriuretic factor (ANF), vasopressin, or deoxycorticosterone (DOC) affects NaCl permeability. Apparent Cl- or Na+ permeabilities (PCl and PNa) were determined by measuring ion fluxes resulting from imposed electrochemical gradients. Transepithelial resistance (RT) was calculated from voltage deflections at the perfusion and collection ends of the tubule, which resulted from perfusion end current injection (cable analysis). ANF [rat ANF-(1–28), 100 nM in the peritubular bath] significantly decreased PCl from 2.20 to 1.84 x 10(-5) cm/s and did not alter PNa (1.11 to 1.18 x 10(-5) cm/s). ANF also decreased PCl in IMCDs from DOC-treated rats (1.14 to 0.98 x 10(-5) cm/s). Vasopressin (10 nM in the peritubular bath) did not affect PCl. RT averaged 39.3 omega.cm2 in IMCDs from control rats and was significantly increased to 62.3 omega.cm2 in tubules from DOC-treated rats. Neither ANF nor vasopressin significantly affected RT in either group. We conclude the following: 1) the results do not support the hypothesis that ANF causes natriuresis by increasing the NaCl permeability of the terminal IMCD. Instead, ANF significantly decreases the chloride permeability. 2) Vasopressin does not affect NaCl permeability. 3) Mineralocorticoid-induced antinatriuresis may be due in part to reduced NaCl permeability in the terminal IMCD.
Influence of kinins and angiotensin II on the regulation of papillary blood flow
Authors: Roman R. J. , Kaldunski M. L. , Scicli A. G. , Carretero O. A.
Published: December 2017

The influence of kinins and angiotensin II on the regulation of renal cortical and papillary blood flow and sodium and water excretion was examined in rats. Superficial cortical and papillary blood flows were measured using a laser-Doppler flowmeter. Papillary blood flow increased 50% after enalaprilat (60 micrograms/kg) and phosphoramidon (5.5 micrograms.kg-1.min-1) were given along with 0.3 M sodium bicarbonate solution to inhibit degradation of kinins and enhance urinary kallikrein activity. Infusion of a kinin antagonist, D-Arg-Hyp-Thi-D-Phe-bradykinin (5 micrograms/min), returned papillary blood flow to control levels. Urine flow and sodium excretion increased after the administration of the kininase inhibitors and sodium bicarbonate, while glomerular filtration rate (GFR) and outer cortical blood flow were unaltered. The kinin antagonist did not alter sodium and water excretion in rats receiving the kininase inhibitors and bicarbonate. Administration of the kinin antagonist alone lowered papillary blood flow by 20%, without affecting outer cortical blood flow or GFR. Urine flow decreased and urine osmolality increased after the rats received the kinin antagonist, but sodium excretion remained unaltered. To assess the role of angiotensin II in the control of papillary blood flow, kinin receptors were blocked by infusion of an antagonist, and the effects of enalaprilat and saralasin were studied. Papillary blood flow increased after blockade of the angiotensin II system in rats receiving the kinin antagonist. These results indicate that the kallikrein-kinin and renin-angiotensin systems participate in the regulation of papillary blood flow.
ANF secretion and renal responses to volume expansion with equilibrated blood
Authors: Paul R. V. , Ferguson T. , Navar L. G.
Published: December 2017

To evaluate the role of atrial natriuretic factor (ANF) in the renal response to acute blood volume expansion without hemodilution, a reservoir syringe filled with donor rat blood was connected to the femoral artery and vein of anesthetized Sprague-Dawley rats to allow rapid equilibration of the reservoir with the intravascular blood. Volume expansion with blood from the reservoir in two steps (of 1 and 1.5% body wt, separated by 1 h, n = 5 rats) produced a mean peak increase in plasma immunoreactive ANF from 99 +/- 21 to 1,310 +/- 230 pg/ml (P less than 0.001); plasma ANF levels throughout these experiments correlated significantly with simultaneously measured urine flow (r = 0.74, P less than 0.005) and sodium excretion (r = 0.65, P less than 0.005). Another group (n = 7) underwent the same two-step procedure; after the second volume expansion, high-dose atriopeptin III infusion (0.4 microgram.kg-1.min-1 did not further increase fractional excretion of sodium (3.17 +/- 0.27 to 2.50 + 0.39%, P = NS). In another group (n = 9 rats), the same dose of atriopeptin III was started before any blood volume expansion. After the resulting hypotension was corrected by restoration of blood volume, an additional 1.5% body weight blood volume expansion did not further augment sodium excretion. We conclude that the diuresis and natriuresis, which occur in response to volume expansion without hemodilution, rise and fall in parallel with immunoreactive ANF in the plasma, and that ANF and acute blood volume expansion act on the kidney through a similar, saturable mechanism.
Cystine and lysine reabsorption in the isolated perfused rat kidney
Authors: Roby K. A. , Segal S.
Published: December 2017

Renal tubular reabsorption of cystine and lysine were studied in the isolated perfused rat kidney to bridge the gap between in vivo clearance studies, and in vitro transport studies of tubule fragments, cells, and brush-border membranes. Lysine was reabsorped by a saturable transport system shared by the dibasics. Cystine was also reabsorbed by a saturable transport system, which was shared in part by the dibasics (maximum inhibition 30%). The lysine threshold (Fmin) was 0.9 mumol.min-1.g-1, with a tubular maximum (TM) of 2.4 mumol.min-1.g-1. The cystine Fmin was 0.06 mumol.min-1.g-1; the TM could not be estimated because it was above the limit of cystine solubility. There was no evidence of cystine ,secretion.- The gamma-glutamyltransferase inhibitor, AT-125, decreased cystine excretion, but only in the presence of glutathione, glycine, glutamate, and the diabasic amino acids. This suggests that cystine from glutathione degradation at the brush border may contribute to urinary cystine (an explanation of the phenomenon of cystine secretion), but only under certain conditions.
Organic osmolytes in inner medulla of Brattleboro rat: effects of ADH and dehydration
Authors: Blumenfeld J. D. , Hebert S. C. , Heilig C. W. , Balschi J. A. , Stromski M. E. , Gullans S. R.
Published: December 2017

Inner medullary methylamine [glycerophosphorylcholine (GPC) and glycine betaine (betaine)] and polyol [sorbitol and myo-inositol (inositol)] osmolytes were measured in water-restricted and antidiuretic hormone (ADH)-infused Brattleboro (DI) rats. Compared with DI rats allowed water ad libitum, rats dehydrated for 3 days had higher urinary osmolality (Uosmol) (812 vs. 239 mosmol/kgH2O) and plasma osmolality (Posmol) (333 vs. 296 mosmol/kgH2O). Dehydration reduced betaine content (36 vs. 66 nmol/mg protein) but had no significant effect on GPC, sorbitol, or inositol. In separate protocols, DI rats, allowed water ad libitum, were infused for either 3 or 12 days with either ADH in saline (+ADH) or saline alone (-ADH). Compared with -ADH controls, 3- or 12-day ADH-infused rats were antidiuretic (Uosmol, 1,000–1,300 mosmol/kgH2O) but not dehydrated (Posmol, 297-300 mosmol/kgH2O). Three days of ADH infusion caused an increase in GPC (340%), betaine (80%), and sorbitol (248%) but not in inositol. After 12 days of ADH, further increases were observed in GPC (730%) and sorbitol (870%); inositol was also elevated (170%), whereas betaine was unchanged. Consequently, the total osmolyte content was significantly higher in +ADH than in -ADH [449 vs. 256 (3 days) and 778 vs. 199 (12 day) nmol/mg protein], whereas total osmolyte levels in dehydrated and control rats were similar (222 vs. 219 nmol/mg protein).(ABSTRACT TRUNCATED AT 250 WORDS)
Na+-independent sugar transport by cultured renal (LLC-PK1) epithelial cells
Authors: Mullin J. M. , McGinn M. T. , Snock K. V. , Kofeldt L. M.
Published: December 2017

The LLC-PK1 cell line has been well characterized concerning its proximal tubule-like Na+-dependent active sugar transporter in the apical membrane. In this study, we investigated the uptake of the glucose analogue, 2-deoxy-D-glucose (2DOG), a paradigm substrate for the facilitated diffusion, Na+-independent sugar transporter in the renal basolateral membrane. The uptake of 0.1 mM 2-[14C]DOG by confluent LLC-PK1 cell sheets at 25 degrees C is linear at least to 10 min, at which time greater than 90% of intracellular radioactivity is 2DOG phosphate. The uptake of this analogue by LLC-PK1 cells is Na+ independent, and the transporter appears to be localized to the basolateral cell membrane. Phlorizin is a much less effective inhibitor than its aglycon, phloretin. Cytochalasin B is also an effective inhibitor, but it causes morphological changes in the cells at concentrations required to inhibit transport. Specificity studies indicate that this transport system requires a hexose with a free hydroxyl at C-1, and that the hydroxyls at C-3 and C-4 be preferably in the equatorial position. Glucose starvation causes an increased rate of 2DOG uptake. Subconfluent (cycling) cultures of LLC-PK1 cells have a threefold greater rate of 2DOG uptake than that seen in confluent (noncycling) LLC-PK1 cells.
Glomerular dysfunction induced by polymorphonuclear leukocyte-derived reactive oxygen species
Authors: Yoshioka T. , Ichikawa I.
Published: December 2017

The role of reactive oxygen species (ROS) derived from polymorphonuclear leukocytes (PMN) on glomerular microcirculation was evaluated in Munich-Wistar rats. To induce local activation of PMN, phorbol myristate acetate (PMA, 20 micrograms) was infused unilaterally into the renal artery in six rats and the micropuncture study was performed 18 h later. In these animals, there was a significant (P less than 0.005) glomerular PMN accumulation in the PMA-injected kidney (on average 2.2 +/- 0.2 per 3-microns glomerular section) compared with the contralateral kidneys of the same rats or the vehicle-treated kidneys from a separate group of seven rats (0.2 +/- 0.1 and 0.2 +/- 0.1 per 3-microns glomerular section, respectively). In the PMA-treated kidneys, whole kidney glomerular filtration rate (GFR) was significantly reduced compared with that of the contralateral kidney of the same rats or vehicle-treated kidneys of a separate group of rats (0.61 +/- 0.04, 1.18 +/- 0.05, and 1.02 +/- 0.04 ml/min, respectively, P less than 0.05). This PMA-induced reduction in GFR was largely prevented by pretreatment of rats with a scavenger of hydrogen peroxide, catalase (1,000 kU; n = 7), or by depletion of PMN with an intravenous nitrogen mustard administration (1.7 mg/kg; n = 7). The reduction of GFR in the PMA-treated kidneys therefore appeared to be mediated by the ROS derived from activated PMN. The micropuncture measurement of glomerular microcirculatory dynamics in PMA-treated kidneys without above pretreatment revealed that there was a profound fall in single-nephron GFR and ultrafiltration coefficient and an increase in renal arteriolar resistances (greater in efferent than afferent arteriole).(ABSTRACT TRUNCATED AT 250 WORDS)
Immune complex activation of rat glomerular mesangial cells: dependence on the Fc region of antibody
Authors: Knauss T. C. , Mene P. , Ricanati S. A. , Kester M. , Dubyak G. R. , Emancipator S. N. , Sedor J. R.
Published: December 2017

Glomerulonephritis is frequently associated with immunoglobulin deposition in the mesangium. We had previously shown that contractile, rat mesangial cells in culture synthesize superoxide anion after binding immune complexes (IC) in a manner dependent on the Fc region of immunoglobulin G (IgG). We now studied the effects of soluble IC on mesangial cell cytosolic free calcium ([Ca2+]i) and phosphatidylinositol turnover as putative mechanisms of transmembrane signaling as well as prostaglandin biosynthesis and contraction. IC (500 micrograms specific antibody) raised [Ca2+]i in mesangial cells loaded with fura-2 from resting levels of 100.4 +/- 8.0 to a peak of 282.3 +/- 31.5 nM in a dose-dependent manner. Removal of extracellular Ca2+ by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid only slightly reduced peak, IC-stimulated [Ca2+]i to 236 +/- 18 nM but prevented the sustained phase of the response, indicating that IC both mobilized Ca2+ from intracellular stores and increased the influx of Ca2+ across the plasma membrane. IC did not increase water-soluble inositol phosphates, measured by anion-exchange chromatography of trichloroacetic acid-extracted cells but markedly stimulated PGE2 and thromboxane B2 synthesis in a dose- and time-dependent manner. Finally, IC (250 micrograms specific antibody) induced 45.8 +/- 10.1% of the cells to contract with an average decrease in cross-sectional surface area of 20.0 +/- 1.8% of basal as assessed by image-analysis microscopy. IC formed with F(ab')2 fragments of antibody and antigen or mixtures of antigen and nonimmune whole molecule antibody did not alter [Ca2+]i, induce prostaglandin synthesis, or stimulate mesangial cell contraction.(ABSTRACT TRUNCATED AT 250 WORDS)
The growth hormone-insulin-like growth factor axis in kidney
Authors: Hammerman M. R.
Published: December 2017

Growth hormone (GH) exerts a variety of metabolic and anabolic effects on skeletal and soft tissues including kidney. Some of these actions are mediated directly, whereas others result from GH-dependent synthesis and release of polypeptide growth factors designated insulin-like growth factors (IGFs). Receptors for GH are present in proximal tubule and GH directly stimulates gluconeogenesis at this site. IGF receptors are found in glomerulus and proximal tubule. Mechanisms for signal transduction by GH and IGFs have been characterized using proximal tubular basolateral membranes. IGFs regulate metabolic and transport processes in cultured glomerular mesangial cells and in isolated proximal tubular cells. IGF I is synthesized in cultured mesangial cells and is produced in a GH-dependent manner in cortical and medullary collecting duct. Evidence has accumulated that IGF I of renal origin functions as a paracrine growth factor in the settings of GH-induced hypertrophy and compensatory hypertrophy of the kidney, and in the setting of proximal tubular regeneration following ischemic injury. IGFs are embryonal mitogens and IGF II may act as a transforming agent for Wilms' tumor. Further characterization of the GH-IGF axis in kidney will provide additional insights into the roles of these peptides as regulators of renal function, growth, and development.
Increased cAMP in proximal tubules is acute effect of nicotinamide analogues
Authors: Campbell P. I. , Abraham M. I. , Kempson S. A.
Published: December 2017

The possible role of adenosine 3',5'-cyclic monophosphate (cAMP) in the mechanism of the acute inhibitory effects of nicotinamide and analogues on brush-border membrane (BBM) phosphate transport was investigated. Compared with basal values, cAMP content of rat renal proximal tubule suspensions was elevated two- to fivefold when incubated at 37 degrees C for 1 h with nicotinamide, 5-methylnicotinamide, or picolinamide at 1–3 mM and in the presence of a phosphodiesterase inhibitor. Thymidine had no effect on cAMP content. There was significant and specific inhibition of BBM transport of phosphate when proximal tubules were incubated with either nicotinamide or picolinamide at concentrations that increased tubule cAMP content. Thymidine had no effect on BBM transport of phosphate. These findings were independent of the dietary Pi intake of the rats. The absence of any effect of thymidine on phosphate transport strongly suggests that inhibition of poly(adenosine diphosphate ribose) polymerase does not play a role in nicotinamide action on phosphate transport. The change in phosphate transport induced by nicotinamide occurred with no change in NAD content. These findings indicate that an increase in cAMP, rather than NAD, is the important change that may mediate the acute inhibition of Na(+)-dependent phosphate transport by nicotinamide.
Isolation of mutant renal (LLC-PK1) epithelia defective in basolateral, Na(+)-independent glucose transport
Authors: Mullin J. M. , Snock K. V. , McGinn M. T. , Kofeldt L. M.
Published: December 2017

To obtain mutant renal epithelia defective in the uptake of 2-deoxyglucose (Na(+)-independent glucose transport), LLC-PK1 renal epithelial cells were subjected to a combination of DNA alkylation, tritium-suicide with 2-[3H]deoxyglucose, and replica plating. One of the mutant sublines obtained, LLC-PK1M-7A, possesses only 40% of the 2-deoxyglucose uptake rate of the parent line, LLC-PK1M, and this defect is stable over at least 30 population doublings. Initial rate of 3-O-methylglucose transport into these mutant cells is only 20% of the parental rate, and efflux of 3-O-methylglucose from the mutant cells is correspondingly low. Glucose metabolism in the mutant cells does not appear to be altered, nor is free glucose accumulated in the cells against a concentration gradient. The uptake rate of L-leucine is the same in both mutant and parent, whereas the (Na(+)-dependent) uptake of alpha-methyl-D-glucoside and methylaminoisobutyric acid is greater in the mutant than the parent. The population doubling times of LLC-PK1M and LLC-PK1M-7A cultures are similar. LLC-PK1M-7A cell morphology is similar to LLC-PK1M when cultures are subconfluent, but on reaching confluence, LLC-PK1M-7A appear larger than LLC-PK1M cells. Their measured cell volume is then 150% of the volume of the parent cells. Future studies of the LLC-PK1M-7A mutant, and acquisition of additional mutant sublines, should elucidate the roles of transepithelial glucose transport in proximal tubular cell physiology.
Arteriovenous acid-base disparity in circulatory failure: studies on mechanism
Authors: Adrogue H. J. , Rashad M. N. , Gorin A. B. , Yacoub J. , Madias N. E.
Published: December 2017

The normal relationship between arterial and venous acid-base composition is altered in hemodynamic compromise. Because the mechanism of this phenomenon remains conjectural, we have studied the acid-base profile and the end-tidal PCO2 of dogs with normal or depressed hemodynamic status in association with either normal ventilation or respiratory arrest. Reductions in cardiac output widened the arteriovenous difference in PCO2 and pH, largely due to arterial hypocapnia but also to venous hypercapnia, and decreased end-tidal PCO2. The arteriovenous gradients for PCO2 and pH of -5.1 +/- 0.4 mmHg and 0.02 +/- 0.01, respectively, during normal hemodynamics widened progressively with graded circulatory compromise reaching values of -30 +/- 5 mmHg for PCO2 (P less than 0.01) and 0.35 +/- 0.05 for pH (P less than 0.01) during cardiac arrest. Development of this disparity, however, required the presence of substantial pulmonary ventilation, since respiratory arrest obliterated the arteriovenous gradients. We propose that arterial hypocapnia, which occurs in association with reduced CO2 excretion, is secondary to an increased ventilation-to-perfusion ratio that reflects a disproportionate decrement in cardiac output. Venous hypercapnia, on the other hand, results from a greater than normal addition of CO2 per unit of blood traversing the capillaries of the hypoperfused peripheral tissues and a diminished CO2 excretion because of pulmonary hypoperfusion. Titration of bicarbonate stores by ongoing production of organic acids might also contribute to venous hypercapnia.
Combined intrarenal blockade of the renin-angiotensin system in the conscious dog
Authors: Siragy H. M. , Howell N. L. , Peach M. J. , Carey R. M.
Published: December 2017

We produced maximal or near-maximal acute intrarenal blockade of the renin-angiotensin system (RAS) by combining inhibitors. Intrarenal infusion of the renin inhibitor, ACRIP, the converting enzyme inhibitor, teprotide, and saralasin were administered individually or combined in random order. The inhibitors were infused for 20 min in doses that did not produce systemic effects in uninephrectomized conscious dogs in sodium balance at 10 meq/day. Significant increases in urine flow rate (UV; F = 97, P less than 0.0001), urinary sodium excretion (UNaV; F = 220, P less than 0.0001), glomerular filtration rate (GFR; F = 64, P less than 0.0001), and renal plasma flow (RPF; F = 108, P less than 0.0001) were observed with each blocker, whether alone or in combination except that ACRIP alone did not alter GFR or RPF. The increase in renal function was related to the number of blockers (3 greater than 2 greater than 1). With the three blockers combined UV increased approximately sixfold (from 0.5 +/- 0.06 to 2.9 +/- 0.03 ml/min), UNaV approximately 10-fold (from 3 +/- 0.4 to 34 +/- 2.8 mueq/min), GFR from 31 +/- 2 to 49 +/- 2 ml/min, RPF from 59 +/- 1 to 120 +/- 4 ml/min, and fractional excretion of sodium from 0.06 +/- 0.01 to 0.5 +/- 0.4% (all P less than 0.001). These changes did not occur where the inhibitors were infused systemically and the changes during intrarenal blocker administration were blocked completely with co-administration of angiotensin II intrarenally. The intrarenal RAS is a potent physiological regulator of renal function.(ABSTRACT TRUNCATED AT 250 WORDS)
Role of the kidney in primary hypertension: a renal transplantation study in rats
Authors: Rettig R. , Folberth C. , Stauss H. , Kopf D. , Waldherr R. , Unger T.
Published: December 2017

We have previously shown that transplantation of kidneys from genetically hypertensive to normotensive rats result in hypertension in renal graft recipients. To investigate whether this posttransplantation hypertension may have been the result of damage to the renal graft by high perfusion pressure before transplantation, we normalized blood pressure throughout life in spontaneously hypertensive rat (SHR) kidney donors by continuous antihypertensive treatment with the angiotensin-converting enzyme inhibitor ramipril (1 mg.kg-1.day-1 in drinking fluid). When kidneys from these rats were transplanted at age 20 wk to age-matched bilaterally nephrectomized F1 hybrids bred from SHR and Wistar-Kyoto (WKY) parents, posttransplantation hypertension still developed. In contrast, blood pressure did not change significantly in recipients of kidneys from ramipril-treated WKY rats. In the initial phase, recipients of SHR kidneys had a lower body weight and higher plasma urea concentrations than recipients of WKY kidneys. However, in the chronic phase, there were no significant differences between the two groups with respect to daily water intake, plasma urea concentration, glomerular filtration rate, renal blood flow, and weight of transplanted kidneys; no histological differences were observed between renal grafts from WKY and SHR donors, except for structural vascular hypertrophy in the latter group. We conclude that posttransplantation hypertension in recipients of SHR kidney grafts also develops, when the grafts have not been subjected to high renal perfusion pressure before transplantation. Our data support the hypothesis that SHR kidneys carry a primary defect, which can induce hypertension in renal graft recipients.
Effect of dopamine on the tubuloglomerular feedback mechanism
Authors: Schnermann J. , Todd K. M. , Briggs J. P.
Published: December 2017

Experiments were performed in anesthetized rats to examine whether infusion of dopamine is associated with a reduction in the tubuloglomerular feedback (TGF) response of stop-flow pressure (PSF) and early proximal flow rate (VEP) to increases of loop of Henle flow. The purpose of these studies was to test further the validity of the proposal that renal vasodilation is a nonspecific cause for diminished TGF responsiveness. When femoral arterial pressure was kept constant with a suprarenal aortic clamp, intravenous infusion of dopamine at rates of 4, 15, 35, and 75 micrograms.kg-1.min-1 induced a 10.9, 23.4, 31.3, and 30.1% decrease in renal vascular resistance. Maximum PSF and VEP responses were significantly reduced at all dose levels of dopamine, whereas V1/2, the flow rate required to produce the half-maximum response, was not altered. TGF blunting occurred within less than 10 min after starting the dopamine infusion. Peritubular infusion of dopamine reduced maximum PSF responses from 8.8 +/- 0.7 to 4.6 +/- 0.53 mmHg at 10(-4) M (P less than 0.01) and from 6.0 +/- 1.19 to 3.6 +/- 0.55 mmHg at 10(-3) M (P less than 0.05). The results are consistent with the notion that renal vasodilatation may modify TGF responses by preventing the full vasoconstrictor response to changes in luminal NaCl concentration.
Light-chain binding sites on renal brush-border membranes
Authors: Batuman V. , Dreisbach A. W. , Cyran J.
Published: December 2017

Immunoglobulin light chains are low-molecular-weight proteins filtered at the renal glomerulus and catabolized within the proximal tubular epithelium. Excessive production and urinary excretion of light chains are associated with renal dysfunction. They also interfere with proximal renal tubule epithelial functions in vitro. We studied the binding of 125I-labeled kappa- and lambda-light chains, obtained from the urine of multiple myeloma patients, to rat and human renal proximal tubular brush-border membranes. Light-chain binding to brush borders was also demonstrated immunologically by flow cytometry. Computer analysis of binding data was consistent with presence of a single class of low-affinity, high-capacity, non-cooperative binding sites with relative selectivity for light chains on both rat and human kidney brush-border membranes. The dissociation constants of light chains ranged from 1.6 X 10(-5) to 1.2 X 10(-4) M, and maximum binding capacity ranged from 4.7 +/- 1.3 X 10(-8) to 8.0 +/- 0.9 X 10(-8) (SD) mol/mg protein at 25 degrees C. Kappa- and lambda-light chains competed with each other for binding with comparable affinity constants. Competition by albumin and beta-lactoglobulin, however, was much weaker, suggesting relative site selectivity for light chains. These binding sites probably function as endocytotic receptors for light chains and possibly other low-molecular-weight proteins.
Parathyroid hormone action on phosphate transport is inhibited by high osmolality
Authors: Kempson S. A. , Helmle C. , Abraham M. I. , Murer H.
Published: December 2017

Parathyroid hormone (PTH) produces rapid inhibition of Na(+)-phosphate cotransport, characterized by a decreased maximal rate of transport, and the inhibition is independent of de novo protein synthesis. The present study determined whether the action of PTH on Na(+)-phosphate cotransport is mediated, at least in part, by rapid endocytic internalization of Na(+)-phosphate cotransporters present in the plasma membrane. Horseradish peroxidase, a fluid-phase marker, was used to demonstrate the presence of endocytosis in opossum kidney (OK) epithelial cells in monolayer culture. An increase in medium osmolality to 500 mosmol/kgH2O, by addition of sucrose, produced 80% inhibition of endocytosis within 1 h. The inhibition was reversed on returning the cells to normal medium. Incubation of OK cell monolayers with PTH (10(-8) M) for 3 h at normal osmolality (281 mosmol/kgH2O) inhibited Na(+)-phosphate cotransport (4 min uptakes) by 56-67%. In hyperosmolar medium (513 mosmol/kgH2O), when endocytosis was inhibited, PTH inhibited Na(+)-phosphate cotransport by only 25-39%, a change that was significantly different from the inhibition in normal medium. Hyperosomolality had no effect on PTH inhibition of Na(+)-H+ exchange or on PTH stimulation of intracellular adenosine 3',5'-cyclic monophosphate. We conclude that the full inhibitory action of PTH on Na(+)-phosphate cotransport may require an intact endocytic mechanism.
Early selective effects of gentamicin on renal brush-border membrane Na-Pi cotransport and Na-H exchange
Authors: Levi M. , Cronin R. E.
Published: December 2017

Gentamicin nephrotoxicity is associated with impairments in proximal tubular function. This study determined whether gentamicin administration to the rat, before a reduction in glomerular filtration rate (GFR), causes early and selective alterations in renal cortical brush-border membrane (BBM) enzyme and transport activity, lipid composition, and fluidity. Three days of gentamicin administration caused significant decreases in the Vmax of alkaline phosphatase, the Vmax of sodium gradient-dependent phosphate transport (Na-Pi cotransport), and the Vmax of pH gradient-dependent sodium transport (Na-H exchange). Gentamicin did not affect BBM-bound maltase or leucine aminopeptidase activities and sodium gradient-dependent glucose or proline transport activities. Gentamicin also caused a significant decrease in BBM sphingomyelin, significant increases in BBM phosphatidylcholine and phosphatidylinositol, a significant decrease in the phospholipid fatty acid saturation index, and a significant increase in BBM fluidity, i.e., decrease in the fluorescence anisotropy of diphenylhexatriene. These BBM functional and compositional effects of gentamicin were independent of endogenous parathyroid hormone activity. We conclude that gentamicin causes early and specific alterations in BBM enzyme and transport activity and also lipid composition, which may play an important role in the progression of renal cell injury.
Ca2+ and pH regulation of K+ channels in membrane vesicles of rabbit proximal tubule
Authors: Jacobsen C. , Mollerup S. , Sheikh M. I.
Published: December 2017

The effect of Ca2+ and pH on the renal epithelial K+ channel was investigated by measuring the Ba2(+)-sensitive 86Rb+ fluxes in membrane vesicles from pars convoluta of rabbit proximal tubule. It was found that the presence of nanomolar concentrations of Ca2+ in the internal compartment (cytoplasmic) of the vesicles ([Ca2+]i) substantially lowered the channel-mediated flux. Ba2(+)-sensitive 86Rb+ uptake was completely blocked by 10 microM [Ca2+]i. This inhibitory effect of Ca2+ was strongly dependent on pH. Thus 0.1 microM [Ca2+]i produced a maximal inhibition of 86Rb+ uptake at pH greater than 7.4 but had no effect at pH less than 7.0. The tracer fluxes measured in the absence of Ca2+ were pH independent over this range. The data are compatible with the model that Ca2+ blocks K+ channels by binding to a site composed of one or several deprotonated groups. The protonation of any one of these groups prevents Ca2+ from binding to this site but does not by itself block transport.
Superiority of salt restriction over diuretics in reducing renal hypertrophy and injury in uninephrectomized SHR
Authors: Benstein J. A. , Feiner H. D. , Parker M. , Dworkin L. D.
Published: December 2017

Spontaneously hypertensive rats (SHR) were uninephrectomized (UNX) at 6 wk of age and given either standard chow (CON), low-sodium chow (LSC), or standard chow and hydrochlorothiazide (HCTZ) added to the drinking water. Severe hypertension developed in all three groups. Forty-two weeks after UNX, proteinuria and glomerular sclerosis were significantly lower in LSC than in CON or HCTZ. The protective effect of salt restriction did not depend upon alterations in plasma renin concentration or glomerular hemodynamics. Micropuncture revealed that glomerular pressure was high in all three groups. Renal hypertrophy assessed by kidney weight, kidney-to-body weight ratio, glomerular volume, and glomerular capillary radius were reduced by salt restriction. These findings suggest that, in the setting of glomerular hypertension, hypertrophy promotes sclerosis. Salt restriction inhibits compensatory kidney growth and protects the kidney.
Characterization of the macula densa stimulus for renin secretion
Authors: Lorenz J. N. , Weihprecht H. , Schnermann J. , Skott O. , Briggs J. P.
Published: December 2017

These studies utilize the isolated perfused rabbit juxtaglomerular apparatus (JGA) to study the macula densa signal for renin secretion in the absence of the confounding influences of intravascular pressure and renal nerve activity. In the first experimental series, JGAs were perfused alternately with high- and low-NaCl solutions to determine the reversibility of the renin response to changes in NaCl concentration. Compared with high-NaCl controls, perfusion with a low-NaCl solution resulted in a fivefold increase in renin secretion rate (RSR) [2.1-10.0 nano-Goldblatt hog units (nGU)/min], and this response was largely reversible. When the solutions were presented in the reverse order, a similar inhibition by high NaCl was observed. In the second series, JGAs were perfused with high-, medium-, and low-NaCl solutions to determine the sensitive range of the renin response to NaCl concentration changes. The full renin response (3.2-16.6 nGU/min), similar in magnitude to that seen in series 1, was found to occur between 80 and 24 mM for Na+ and 61 and 7 mM for Cl-. In the third series, the NaCl concentration and flow rate of the perfusate were altered independently to separate the effects of flow rate, NaCl delivery, and NaCl concentration on RSR. Although a decrease in perfusate flow rate slightly increased RSR (3.4-8.1 nGU/min), a comparable decrease in NaCl concentration resulted in a much higher RSR (26.3 nGU/min). We conclude that in this preparation 1) RSR responds equally to both increases and decreases in macula densa NaCl concentration, and these changes are rapid and largely reversible, 2) the full renin response occurs within the concentration range normally occurring at the macula densa, i.e., below 80 mM Na+ and 61 mM Cl-, and 3) RSR responds with a larger change to alterations in NaCl concentration than in NaCl delivery or fluid flow rate.
Morphological and physiological responses to aldosterone: time course and sodium dependence
Authors: Wade J. B. , Stanton B. A. , Field M. J. , Kashgarian M. , Giebisch G.
Published: December 2017

The time course and Na+ dependence of morphological responses to a physiological elevation of aldosterone were examined in the renal cortical collecting duct. Male Sprague-Dawley rats were adrenalectomized and given a basal replacement infusion of aldosterone and dexamethasone by osmotic minipump. The animals were given additional aldosterone by either intravenous infusion or by a second implanted osmotic minipump for either 5 h or 1, 3, 7, or 14 days. Animals were fed either a Na(+)-replete or a low-Na+ diet. After these treatments we conducted a morphometric analysis on kidneys processed for electron microscopy by standard techniques. Increased aldosterone induced detectable proliferation of the basolateral membrane of principal cells after 24 h. Basolateral membrane continued to increase rapidly through 3 days of exposure and thereafter at a modest rate. Cell area and the surface density of the basolateral membrane were also increased by aldosterone treatment. The morphological response to aldosterone was markedly inhibited by maintaining animals on a low-Na+ diet. Acute exposure to elevated aldosterone levels (5 h) increased K+ secretion by principal cells without alterations in basolateral membrane area. Chronic (greater than 24 h) exposure to aldosterone induces morphological adaptations that are accompanied by further elevations in K+ secretion.
High proton flux through membranes containing antidiuretic hormone water channels
Authors: Harris H. W. , Kikeri D. , Janoshazi A. , Solomon A. K. , Zeidel M. L.
Published: December 2017

Antidiuretic hormone (ADH) stimulation of toad urinary bladder granular cells causes simultaneous increases in transepithelial water and H+ permeabilities (PF and PH+, respectively), suggesting that ADH-elicited water channels inserted into granular cell apical membranes might be permeable to both water and H+. We have previously used self-quenching fluorophores entrapped within endocytic vesicles selectively retrieved from water-permeable apical membranes to measure vesicle PF. The membranes of these vesicles possess an extremely high PF such that our measurements provide only minimum estimates of vesicle PF and have limited our ability to quantitate the properties of ADH water channels. We therefore quantitated vesicle PH+ using similar rapid mixing techniques. Vesicle PH+ was 5.1 +/- 0.5 x 10(-3) cm/s. Activation energy of this process was 3.6 +/- 0.6 kcal/mol, indicative of H+ flux through an aqueous channel. The mercurial reagent, para-chloromercuribenzenesulfonate (PCMBS), which inhibits ADH-stimulated transepithelial PF in intact bladders by 50-60%, inhibited vesicle PH+ by 55%. N-Ethylmaleimide and phloretin, which do not alter ADH-stimulated PF, did not affect vesicle PH+. We conclude that membranes containing ADH water channels possess substantial PH+ that likely reflects proton flux through water channels. The apparent high PH+ of the ADH water channel may have important implications for intracellular trafficking of these water channels in ADH-responsive epithelial cells.
Endogenous natriuretic peptides: effect on collecting duct function in rat kidney
Authors: Schulz-Knappe P. , Honrath U. , Forssmann W. G. , Sonnenberg H.
Published: December 2017

In vivo microcatheterization of the medullary collecting duct of rat kidney was used to compare the functional effects of intravenous infusion of rat atrial natriuretic factor, human urodilatin (a peptide produced in the kidney), or porcine brain natriuretic peptide. All three peptides resulted in striking and quantitatively similar inhibition of NaCl and water reabsorption in the duct. In addition, all three induced significant K+ secretion in this part of the nephron. These results could not be explained by functional alterations in upstream tubular segments. We conclude that the three peptides are equivalent in their transport effects on the inner medullary collecting duct system, when administered at the same (presumably supramaximal) dose.
Effects of intracellular calcium on proximal bicarbonate absorption
Authors: Liu F. Y. , Cogan M. G.
Published: December 2017

Using in vivo microperfusion in the proximal convoluted tubule (PCT) of the Munich-Wistar rat, we assessed the impact of raising intracellular calcium concentration on bicarbonate transport and its regulation by angiotensin II. Luminal perfusion with the calcium ionophore A23187 caused a dose-dependent increase in S1 PCT bicarbonate absorption, with a maximal change of 60 peq.mm-1.min-1. Subsequent addition of phorbol ester (PMA) after A23187 further increased transport but to a level no higher than previously shown with PMA alone, and A23187 was nonstimulatory when added after PMA. These results suggest that intracellular calcium affects the same pathway as protein kinase C in stimulating proximal acidification. Pretreatment with A23187 (+/- PMA) attenuated by approximately one-third the bicarbonate absorptive response in the S1 PCT usually observed after angiotensin II administration. A23187 had similar actions, but of smaller magnitude, in the S2 PCT. In conclusion, a rise in intracellular calcium increases bicarbonate absorption in the S1 and S2 PCT in vivo.
Tubuloglomerular feedback kinetics in spontaneously hypertensive and Wistar-Kyoto rats
Authors: Daniels F. H. , Arendshorst W. J.
Published: December 2017

The steady-state behavior of the tubuloglomerular feedback system has been studied in detail, but little is known about its dynamics. However, kinetic data can provide insight regarding the contribution of feedback to autoregulatory responses. Accordingly, experiments were conducted in anesthetized, euvolemic, spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats to characterize the time course of changes in proximal tubular stop-flow pressure after step changes in the rate of orthograde perfusion of Henle's loop. We studied the responses both to increase in perfusion rate, which produced preglomerular vasoconstriction, and decreases in perfusion rate, which produced preglomerular vasodilation. In both strains, the pattern of induced stop-flow pressure transients consisted of a pure delay followed by a monoexponential decay to a new steady state. In SHR rats, delay times were shorter than in WKY rats, but response time constants were not significantly different in the two strains. However, response time constants for dilation were longer than for constriction in both strains. The delay times and relatively large response time constants observed indicate that tubuloglomerular feedback cannot mediate rapid autoregulatory responses to fluctuations in renal perfusion pressure. The response time of tubuloglomerular feedback is probably limited by both the time lag associated with fluid transit through the loop of Henle and a relatively slow rate-limiting step in the signal transduction process at the macula densa.
Stimulation of renin secretion by ethacrynic acid is independent of Na(+)-K(+)-2Cl- cotransport
Authors: Park C. S. , Doh P. S. , Carraway R. E. , Chung G. G. , Fray J. C. , Miller T. B.
Published: December 2017

This study investigated the cellular mechanism of stimulation of renin secretion by the loop diuretic ethacrynic acid (EA) in rabbit renal cortical slices. The diuretic rapidly stimulated renin secretion reversibly and in a concentration-dependent manner. The stimulation was independent of the presence of Na+, Cl-, Ca2+, or other loop diuretics (furosemide and bumetanide) in the incubation media, suggesting that the stimulation in vitro was not dependent on the inhibitory effect of the diuretic on Na(+)-K(+)-2Cl-cotransport. The findings do not support the macula densa hypothesis. The stimulation by the diuretic was prevented and reversed by thiols such as cysteine and dithiothreitol, which also prevented and reversed the stimulation of renin secretion by the nondiuretic sulfhydryl reagent P-chloromercuriphenyl-sulfonate (PCMPS). These results suggest that EA stimulates renin secretion in vitro via reversible chemical reactions with specific membrane sulfhydryl groups that may have no functional role in the Na(+)-K(+)-2Cl- cotransport.