bluuebunny/crossref_metadata_2025 huggingface dataset:
Natriuretic response to renal interstitial hydrostatic pressure during angiotensin II blockade
Authors: Haas J. A. , Lockhart J. C. , Larson T. S. , Henrikson T. , Knox F. G.
Published: December 2017

Increases in renal interstitial hydrostatic pressure (RIHP) increase urinary sodium excretion (UNaV). Experimentally increasing RIHP by direct renal interstitial volume expansion (DRIVE) has been shown to decrease proximal tubule sodium reabsorption. The purpose of the present study was to investigate whether the renin-angiotensin system modulates the natriuretic response to DRIVE. Unilateral nephrectomy and implantation of two polyethylene matrices were performed 3 wk before the acute experiment. Fractional sodium excretion (FENa), RIHP, and glomerular filtration rate (GFR) were measured before and after DRIVE in control rats (n = 9) and in rats receiving the angiotensin II (ANG II) receptor antagonist, losartan potassium (10 mg/kg i.v.; n = 10). DRIVE was achieved by infusing 100 microliters of 2.5% albumin solution directly into the renal interstitium. GFR remained unchanged by DRIVE in both groups. In control animals, DRIVE significantly increased both RIHP (delta 3.8 +/- 0.5 mmHg) and FENa (delta 0.92 +/- 0.19%). In the losartan-treated group, RIHP (delta 2.8 +/- 0.4 mmHg) and FENa (delta 1.93 +/- 0.41%) also significantly increased. The natriuretic response to DRIVE was significantly enhanced during ANG II receptor blockade compared with control animals (delta UNaV/delta RIHP = 2.01 +/- 0.67 vs. 0.44 +/- 0.17 mu eq.min-1 x mmHg-1, respectively; P < 0.05). These results suggest that the blockade of angiotensin enhances the natriuretic response to increased RIHP during DRIVE.
Augmentation of intrarenal angiotensin II levels by chronic angiotensin II infusion
Authors: Von Thun A. M. , Vari R. C. , el-Dahr S. S. , Navar L. G.
Published: December 2017

The objective of this study was to investigate the singular role of elevated angiotensin II (ANG II) levels in the development of two-kidney, one-clip (2K1C) Goldblatt hypertension in the rat and specifically in the altered intrarenal ANG II levels that occur in the nonclipped kidney. As a substitute for the clipped kidney, chronic delivery of ANG II (40 ng/min) via an osmotic minipump implanted subcutaneously was used to mimic plasma ANG II levels observed in 2K1C rats during the developmental phase of hypertension. Arterial pressure increased gradually over a period of 14 days, and a pressure profile similar in magnitude and temporal pattern to that of the 2K1C rats was observed. Systemic ANG II was elevated to similar levels in the 2K1C (60 +/- 13 fmol/ml) and ANG II-infused rats (72 +/- 15 fmol/ml) compared with intact two-kidney control animals (31 +/- 6 fmol/ml; P < 0.05) or uninephrectomized rats (13 +/- 1 fmol/ml; P < 0.05). Although renin content was markedly suppressed (80%), intrarenal ANG II content of the contralateral kidneys of the 2K1C groups (86 +/- 12 fmol/g) and the ANG II-infused group (150 +/- 17 fmol/g) was greater than that of the two-kidney control (53 +/- 7 fmol/g; P < 0.05) and uninephrectomized control animals (42 +/- 5 fmol/g; P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)
Effect of an endothelin-receptor antagonist on ischemic acute renal failure
Authors: Chan L. , Chittinandana A. , Shapiro J. I. , Shanley P. F. , Schrier R. W.
Published: December 2017

In the isolated perfused rat kidney, endothelin (ET) added to the perfusate at concentrations ranging from 50 to 500 pmol/l resulted in a dose-dependent reduction in renal perfusate flow (RPF) and inulin clearance (CIn). The decrease in RPF (17 +/- 3 vs. 34 +/- 3 ml.min-1 x g-1; P < 0.01 compared with control) and CIn (89 +/- 13 vs. 317 +/- 19 microliters.min-1 x g-1; P < 0.01 compared with control) by ET (500 pmol/l) was prevented by the ET antagonist BQ-123 (10 microM), with full recovery of RPF [36 +/- 2 vs. 34 +/- 3 ml.min-1 x g-1; not significant (NS) compared with control] and CIn (299 +/- 51 vs. 317 +/- 19 microliters.min-1 x g-1; NS compared with control). In the absence of ET, perfusion of the kidney with a similar concentration of BQ-123 (10 microM) did not induce any changes in RPF (36 +/- 5 vs. 34 +/- 3 ml.min-1 x g-1; NS compared with control) or CIn (320 +/- 14 vs. 317 +/- 19 microliters.min-1 x g-1; NS compared with control). After 60 min of arterial clamping, BQ-123 (10 microM) given before the onset of ischemia and during reflow improved CIn (88 +/- 4 vs. 19 +/- 3 microliters.min-1 x g-1; n = 6, P < 0.01) and net tubular sodium reabsorption (TNa) compared with no treatment. On the other hand, the same dose (10 microM) of BQ-123 given only during the reperfusion period was not effective in preventing the decreases in either CIn or TNa.(ABSTRACT TRUNCATED AT 250 WORDS)
Modulation of ANG II receptor and its mRNA in normal rat by low-protein feeding
Authors: Benabe J. E. , Wang S. , Wilcox J. N. , Martinez-Maldonado M.
Published: December 2017

Pages F660–F669: J. E. Benabe, S. Wang, J. N. Wilcox, and M. Martinez-Maldonado. “Modulation of ANG II receptor and its mRNA in normal rat by low-protein feeding.” Page F662, Table 1: for the third group (NP), the value for renal vascular resistance is incorrect and should be 9 ± 1 dyn·s·cm-5; also for the NP group, the plasma renin activity is incorrectly expressed as a negative value and should be 10.1 ± 1.3 ng·ml-1h–1.
Effect of dietary sodium chloride on insulin receptor number and mRNA levels in rat kidney
Authors: Sechi L. A. , Griffin C. A. , Schambelan M.
Published: December 2017

Insulin has been postulated to play a role in the pathophysiology of essential hypertension via an antinatriuretic action. To further explain the sodium insulin interaction in the kidney, we studied effects of different salt intakes on insulin receptor binding and mRNA levels in tissues from rats maintained on 0.07%, 0.3%, or 7.5% NaCl for 14 days. Scatchard analysis of competition data from in situ autoradiography studies resulted in curvilinear profiles, indicating the presence of either two classes of receptors or a single class of receptors with a negative cooperative hormone-receptor interaction. When data were analyzed using the two-site model, binding capacity of the high-affinity receptor site was significantly less in high-salt-fed rats. No significant differences between dietary groups were observed in apparent dissociation constants of the two receptor sites or in maximal binding capacity of the low-affinity, high-capacity site. Analysis of insulin binding to glomeruli, cortex, outer medulla, and inner medulla indicated that the high-salt diet was associated with decreased receptor density in all regions studied. Insulin receptor mRNA, as quantified by Northern and slot blot analysis, was inversely related to salt intake in absence of a change in plasma glucose, insulin, and corticosterone levels. Both 7.2- and 9.4-kb transcripts were similarly affected by dietary sodium content. Plasma renin concentration and renal renin mRNA levels were decreased but blood pressure was not affected by high-salt diet.(ABSTRACT TRUNCATED AT 250 WORDS)
Regulation of prostaglandin endoperoxide synthase gene expression in rat mesangial cells by interleukin-1 beta
Authors: Rzymkiewicz D. , Leingang K. , Baird N. , Morrison A. R.
Published: December 2017

In primary cultures of rat mesangial cells from passage 3 to 6, interleukin-1 beta (IL-1) induced a time-dependent increase in prostaglandin E2 (PGE2) formation and release into the extracellular medium. This increase was associated with a dramatic upregulation of the steady-state levels of mRNA for the prostaglandin endoperoxide synthetase (PES)-2 gene transcript as demonstrated by Northern analysis. In contrast, there did not appear to be a significant increase in the mRNA levels for a 2.8-kb transcript for the PES-1 gene. At 18 h of exposure to IL-1, the steady-state level of message for PES-2 remained elevated at 50% of the 2-h time point. Culturing the cells in the presence of cycloheximide and IL-1 demonstrated a superinduction of the PES-2 message without any change in PES-1 message. The tumor-promoting phorbol ester, phorbol myristate acetate (PMA), was also associated with an upregulation of the message for the PES-2 gene and did not influence the levels of the message for the PES-1 gene as demonstrated by Northern analysis. Dexamethasone (Dex) inhibited to control levels the induction by PMA, but the induction of the message by IL-1 was only inhibited 30%. Despite 70% of the message being present by 2 h of induction, Dex was capable of totally inhibiting the inductive effect of IL-1 with respect to PGE2 biosynthesis. Immunocytochemical studies demonstrated a dramatic induction of PES-2 protein by IL-1, which was inhibited by Dex. The data suggest that Dex inhibits the translation of the PES-2 protein.
Mechanism of endothelin activation of phospholipase A2 in rat renal medullary interstitial cells
Authors: Barnett R. L. , Ruffini L. , Hart D. , Mancuso P. , Nord E. P.
Published: December 2017

Previous studies from this laboratory have demonstrated that endothelin-1 (ET) stimulates phosphatidylinositol (PI) hydrolysis, activates dihydropyridine-insensitive Ca2+ channels, and promotes prostaglandin E2 (PGE2) accumulation in cultured rat renal medullary interstitial cells (RMIC). The mechanism whereby ET augments PGE2 production was explored in the current study. ET-evoked PGE2 accumulation proceeded independent of large increments in cytosolic free Ca2+ concentration ([Ca2+]i), derived from either extracellular or intracellular sources. Chelation of intracellular Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid eliminated ET-evoked PGE2 production, indicating that eicosanoid production was nonetheless a Ca(2+)-requiring process. Nanomolar concentrations of phorbol 12-myristate 13-acetate (PMA) alone did not stimulate PGE2 production, nor did PMA alter ET-stimulated PGE2 accumulation. Furthermore, downregulation of protein kinase C (PKC) by prolonged exposure of cells to PMA did not mitigate ET-mediated PGE2 production, demonstrating that PKC stimulation was not required for PGE2 production. ET stimulated PGE2 accumulation despite PI-specific phospholipase C (PI-PLC) inhibition by nanomolar concentrations of PMA, indicating that eicosanoid production was not a downstream event of PI hydrolysis. ET stimulated arachidonic acid metabolite release in parallel with a loss of label from membrane phospholipids. Phosphatidylethanolamine was the preferred substrate for ET-mediated activation of phospholipase A2 (PLA2). Immunocytochemicadl studies including immunostaining, immunoblotting, and immunoprecipitation confirmed the presence of cytosolic PLA2 (cPLA2) in RMIC. In summary, ET stimulation of PGE2 production in RMIC is mediated via agonist activation of cPLA2 independent of activation of PI-PLC, suggesting direct coupling to the ET receptor. Constitutive levels of [Ca2+]i rather than abrupt increments in [Ca2+]i are sufficient for activation of this receptor-effector system, with no obligatory requirement for PKC.
Interaction of rat kidney P-glycoprotein with a urinary component and various drugs including cyclosporin A
Authors: Charuk J. H. , Loo T. W. , Clarke D. M. , Reithmeier R. A.
Published: December 2017

The interaction of rat renal P-glycoprotein with various drugs and a hydrophobic component found in rat urine was studied to gain an understanding of both its transport function in kidney and its potential role in drug secretion and drug-induced nephrotoxicity. Rat kidney brush-border membranes (BBM) were photolabeled with [3H]azidopine, a calcium-channel blocker that covalently labeles P-glycoprotein. P-glycoprotein was immunoprecipitated with a rabbit polyclonal antibody against the human MDR1 protein (multidrug resistance gene class 1). The amount of [3H]azidopine incorporated into P-glycoprotein was quantitated following gel electrophoresis and fluorography. Photolabeling inhibition assays were conducted with a panel of drugs known to interact with P-glycoprotein in multidrug-resistant cells. Verapamil or quinidine [half-maximal inhibition constant (K0.5) = 1 microM], vinblastine (K0.5 = 3 microM), and doxorubicin or daunomycin (K0.5 = 10 microM) all blocked [3H]azidopine photolabeling of renal P-glycoprotein. Of the drugs tested, the immunosuppressant drug, cyclosporin A, interacted with kidney P-glycoprotein with the highest affinity (K0.5 = 50 nM). However, the cardiac glycoside, digoxin, failed to inhibit P-glycoprotein photolabeling. A hydrophobic rat urine extract prepared by reverse-phase chromatography also blocked photolabeling of renal P-glycoprotein. Our current hypothesis is that various drugs may inhibit urinary excretion of an endogenous substrate by virtue of their ability to bind with high affinity to P-glycoprotein. A hypothesis of drug-induced nephrotoxicity based on the interaction of various compounds like cyclosporin A with P-glycoprotein is presented.
Minealocorticoid receptors and 11 beta-steroid dehydrogenase activity in renal principal and intercalated cells
Authors: Naray-Fejes-Toth A. , Rusvai E. , Fejes-Toth G.
Published: December 2017

Aldosterone exerts complex effects on the cortical collecting duct (CCD): it increases Na+ and K+ transport, and it also influences H+ and HCO3 transport. Whether these latter effects represent direct action of aldosterone on intercalated cells (ICC) or are secondary to changes in the transport of other electrolytes is unclear. Because the presence of specific receptors is the prerequisite of a direct steroid action, and mineralocorticoid receptors (MR) have not yet been demonstrated in ICC, in this study we determined the density of MR directly in isolated principal cells (PC) and beta-ICC. Purified populations of these two cell types were obtained from rabbit renal cortex by immunodissection and fluorescence-activated cell sorting. We found that both PC and beta-ICC contained a significant number of MR, although receptor density was higher in PC than in beta-ICC (6,704 +/- 912 vs. 2,181 +/- 388 MR sites/cell; P < 0.001). 11 beta-Hydroxysteroid dehydrogenase (11 beta-OHSD), an enzyme that is present predominantly in mineralocorticoid target cells, exhibited a distribution similar to that of MR in the two cell types. 11 beta-OHSD activity, determined by measuring the rate of conversion of [3H]corticosterone to 11-dehydrocorticosterone, was 1.08 +/- 0.14 and 0.34 +/- 0.08 fmol.min-1 x 1,000 cells-1 (P < 0.001) in intact PC and beta-ICC, respectively. 11 beta-OHSD in both cell types utilized NAD as cofactor. These results suggest that beta-ICC are potential direct targets of aldosterone and that MR in both PC and beta-ICC are protected by 11 beta-OHSD.
Intracellular modulation of acid secretion in rat inner medullary collecting duct cells
Authors: Schwartz J. H. , Masino S. A. , Nichols R. D. , Alexander E. A.
Published: December 2017

In this study we defined some of the important elements in the acidification process of rat inner medullary collecting duct (IMCD) cells in culture. After cell acidification, i.e., cell pH (pHi) = 6.51 +/- 0.02, pHi increased 0.046 +/- 0.003 units/min. N-ethylmaleimide, N,N'-cyclohexylcarbodiimide, and bafilomycin reduced this rate by over 85%. In contrast, omeprazole and Sch-28080 had no effect. 1,2-Bis(2-aminophenoxy)ethane-N,N,N,N'-tetraacetic acid, which prevents a rise in cell Ca2+ concentration ([Ca2+]i) reduced the rate of pHi recovery to 0.013 +/- 0.002 units/min. Calmodulin inhibitors or disruption of cytoskeletal elements with cytochalasin B and colchicine also reduced pHi recovery significantly. In addition, these cells contain acidic vesicles and undergo pHi-regulated endocytosis and exocytosis, which are inhibited by disrupting the cytoskeleton. We conclude that, in our cultured line of rat IMCD cells, proton secretion is mediated by an H(+)-adenosinetriphosphatase. Changes in pHi produce alterations in acid secretion through a signal cascade that requires changes in [Ca2+]i, activation of calmodulin, an intact cytoskeleton, and alteration in the rate of exocytosis and endocytosis.
Renal cortical interstitium and fluid absorption by peritubular capillaries
Authors: Aukland K. , Bogusky R. T. , Renkin E. M.
Published: December 2017

Every minute, the cortical peritubular capillaries in a 1-g rat kidney take up more than 0.5 ml tubular reabsorbate. Studies of renal lymph and measurements of pressure in capillaries (Pc) and interstitium (Pi) indicate that normally the protein colloid osmotic pressure of peritubular capillary plasma (COPp) provides the necessary absorptive force, keeping Pi at 2-4 mmHg, i.e., 8-10 mmHg lower than Pc. At reduced COPp, continued delivery of fluid from the tubules automatically raises Pi to maintain capillary fluid uptake. The transient Pi response to sudden exposure of the kidney to subatmospheric pressure shows that such adjustment of forces may take place in only 5 s. Most remarkable, adjustment of forces may take place in only 5 s. Most remarkable, reabsorption continues during protein-free perfusion of the isolated rat kidney, apparently effected by a Pi exceeding Pc. A relative retardation of interstitial uptake of ferritin from plasma in this case suggests fluid reabsorption through both small and large pores in the capillary wall. Collapse of the capillaries is presumably prevented by tight tethering to the capillary wall, giving the narrow interstitium a very low compliance.
Metabolic acidosis stimulates carbonic anhydrase activity in rabbit proximal tubule and medullary collecting duct
Authors: Brion L. P. , Zavilowitz B. J. , Suarez C. , Schwartz G. J.
Published: December 2017

Both membrane-bound carbonic anhydrase (CA) (isozyme type IV) and cytosolic CA (type II) activities enhance urinary acidification. We have previously shown that chronic metabolic acidosis (CMA) accomplished by NH4Cl loading with food restriction induces soluble CA activity in rabbit renal cortical homogenates. The present study was designed to assess the effect of CMA on the activity of CA isozymes in cortical and outer medullary homogenates, as well as in major proton-secreting segments of the kidney. Segments were microdissected from proximal convoluted tubules (PCT) proximal straight tubules, cortical collecting ducts, and outer medullary collecting ducts (OMCD). Total CA activity was measured by a colorimetric endpoint method, and CA IV activity was assessed from the sodium dodecyl sulfate-resistant hydratase activity. In controls, CA IV activity accounted for 3% of total CA activity in tissue homogenates. CMA induced a threefold increase in CA IV activity in cortical homogenates, in the absence of renal or tubular hypertrophy. In the PCT, CMA induced a 78% increase in total CA activity, which comprised a 178% increase in CA IV activity, and a 58% increase in CA II activity. In the OMCD, CMA induced a 53% increase in total CA (probably CA II) activity. We conclude that CMA induces CA activity in the PCT (CA II and CA IV) and the OMCD (most likely CA II) of adult rabbit kidneys. The induction of CA activity accompanies the increase in urinary acidification observed in CMA.
Effect of luminal angiotensin II on proximal tubule fluid transport: role of apical phospholipase A2
Authors: Li L. , Wang Y. P. , Capparelli A. W. , Jo O. D. , Yanagawa N.
Published: December 2017

Recent micropuncture studies showed the existence of high concentrations of angiotensin II (ANG II) in proximal tubular fluid. In the present study, we have examined the effect of luminal ANG II, alone and in combination with peritubular ANG II, on fluid transport (JV) in the isolated perfused rabbit proximal convoluted tubule. In comparison with peritubular ANG II, luminal ANG II caused a similar but more potent biphasic effect on JV. At 10(-11) M, luminal ANG II maximally increased JV to 204 +/- 22% of the baseline compared with 142 +/- 10% by peritubular ANG II at 10(-10) M. At 10(-8) M, luminal ANG II suppressed JV to 9.7 +/- 16% of the baseline compared with 64 +/- 14% by peritubular ANG II. When luminal and peritubular ANG II were combined at concentrations that impose similar effect on JV, the effects of luminal and peritubular ANG II were not additive. However, when combined at concentrations that would otherwise impose opposing effects on JV, the stimulatory effect predominated. In support of the role of apical phospholipase A2 (PLA2) on the effect of luminal ANG II, ANG II stimulated PLA2 activity in isolated brush-border membrane vesicles, and addition of PLA2 inhibitor, mepacrine or dibucaine, to the luminal perfusate attenuated the effect of luminal ANG II on JV. In summary, these studies show a potent effect of luminal ANG II on proximal tubule JV involving activation of brush-border membrane PLA2. When combined, luminal and peritubular ANG II exert their effects in concert on proximal tubule JV.
ATP stimulates Ca2+ mobilization by a nucleotide receptor in glomerular endothelial cells
Authors: Briner V. A. , Kern F.
Published: December 2017

The present study investigates ATP effects on Ca2+ mobilization in bovine glomerular endothelial cells (GEC) and the receptors mediating ATP response. Extracellular ATP stimulated a rise in inositol 1,4,5-trisphosphate and cytosolic free Ca2+ concentration ([Ca2+]i) in a dose-dependent manner. Extracellular Ca2+ depletion did not prevent [Ca2+]i rise. ATP effects were not mediated by P1, P2x, and P2t purinoceptors, since the P1 receptor agonist adenosine and the P2x receptor agonist [alpha,beta-CH2]ATP had no effect on inositol 1-monophosphate (IP) formation and Ca2+ mobilization and ATP does not activate P2t receptors. The P2y receptor antagonist reactive blue (10(-3) M) had little inhibitory effect on ATP (10(-5) M)-stimulated IP formation (15.6 +/- 4.2%) and Ca2+ rise (7.0 +/- 3.0%). According to the classification of purinoceptors, ATP is less potent than 2-methylthioadenosine 5'-triphosphate (2-MeS-ATP) in stimulating P2y receptors. In GEC, however, the rank order of potency in stimulating IP and [Ca2+]i rise was ATP > 2-MeS-ATP > ADP. The pyrimidine nucleotide UTP (10(-3) M) induced maximal IP formation (653 +/- 37%) and Ca2+ mobilization (591 +/- 22 nM) similar to ATP (IP 647 +/- 27%; [Ca2+]i 583 +/- 15 nM). At submaximal (10(-5) M) but not at maximal (10(-3) M) doses ATP and UTP effects were additive. ATP and UTP induced specific cross-desensitization. It is concluded that the purinergic nucleotide ATP and pyrimidine nucleotide UTP mediate their effects by a common nucleotide receptor. This receptor differs from P2z and P2y1 receptors, since by definition UTP does not activate these receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
Mechanism of acidification along cortical distal tubule of the rat
Authors: Fernandez R. , Lopes M. J. , de Lira R. F. , Dantas W. F. , Cragoe Junior E. J. , Malnic G.
Published: December 2017

The cellular mechanism of luminal acidification (bicarbonate reabsorption) was studied in cortical distal tubules of rat kidney. The stopped-flow microperfusion technique was applied to early distal (ED) and late distal (LD) segments, perfused with bicarbonate Ringer solution to which specific inhibitors were added, to measure bicarbonate reabsorption [HCO3 flux (JHCO3)]. pH and transepithelial potential difference (Vt) were recorded by double-barreled H+ exchange resin/reference (1 M KCl) electrodes. Amiloride increased stationary pH and reduced Vt in both early and late segments. Hexamethylene-amiloride (HMA), a specific Na(+)-H+ exchange blocker, reduced JHCO3 in both segments (ED by 43.6 and LD by 40.3%) without affecting Vt. Benzamil, an Na(+)-channel blocker, reduced Vt by 75.9 in ED and 74.9% in LD but had no significant effect on acidification in both segments. The specific inhibitor of H(+)-ATPase, bafilomycin A1, inhibited LD JHCO3 at a concentration of 2 x 10(-7) M by 49%, but ED was inhibited by 24% only at 2 x 10(-6) M. Sch-28080, an inhibitor of gastric H(+)-K(+)-ATPase, reduced JHCO3 by 35% in LD of K(+)-depleted rats but not in control rats and had no effect on ED. These data indicate that, in ED, bicarbonate reabsorption is mediated mostly by Na(+)-H+ exchange. In LD, there is evidence for contribution of Na(+)-H+ exchange, vacuolar H(+)-ATPase, and H(+)-K(+)-ATPase (in K(+)-depleted rats) to bicarbonate reabsorption.
Expression of Na-K-ATPase alpha- and beta-subunit mRNA and protein isoforms in the rat nephron
Authors: Tumlin J. A. , Hoban C. A. , Medford R. M. , Sands J. M.
Published: December 2017

Na-K-ATPase is a heterodimeric complex composed of an alpha-catalytic and a glycosylated beta-subunit. Previous studies using in situ hybridization and Northern blot analysis to determine alpha- and beta-subunit mRNA isoform expression in the rat kidney have given conflicting results. This heterogeneity may be due to detection of alpha 2- or alpha 3-isoforms arising from nonrenal epithelial sources such as peripheral nerves or vascular smooth muscle. To address this possibility, we investigated alpha-subunit mRNA isoform expression in different nephron segments using tubule microdissection and reverse transcription-polymerase chain reaction amplification. Southern blot analysis of polymerase chain reaction products using isoform-specific primers and probes detected the expression of alpha 1- and alpha 3-mRNA isoforms in whole kidney, cortical collecting ducts (CCD), and proximal tubule S2 subsegments (S2). No evidence for alpha 2 was found in kidney or microdissected CCD or S2. To determine whether alpha 1- or alpha 3-mRNA in the CCD or S2 is translated into protein, Western blot analysis of total protein from microdissected S2, CCD, medullary thick ascending limb (MTAL), or outer medullary collecting duct outer stripe (OMCDos) was performed. Total protein was separated according to size by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, then probed using polyclonal antibodies specific for the alpha 1-, alpha 2-, alpha 3-, beta 1-, and beta 2-protein isoforms. Rat brain was used as a positive control and demonstrated that the antibodies could detect a single 97- and 35-kDa band for the alpha- and beta-isoforms, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Regulation of rat renal alpha 2 B-adrenergic receptors by potassium depletion
Authors: Muir J. C. , Huang L. , Harrison J. K. , Rosin D. L. , Okusa M. D.
Published: December 2017

Potassium depletion and alpha 2-adrenergic receptor (alpha 2-AR) agonists produce similar physiological effects on renal function. Both stimuli increase Na-H exchange in proximal tubule cells, inhibit water transport in collecting tubule cells, and alter blood pressure regulation. The purpose of this study was to determine whether potassium depletion and renal alpha 2-AR subtype expression were linked. Kidney membrane proteins and RNA were harvested from anesthetized rats fed a potassium-deficient diet for 4-20 days (LK 4 to LK 20). Using a selective alpha 2-AR antagonist, [3H]MK-912, we observed that potassium depletion led to a dramatic increase in maximum binding (270% of control) without a change in dissociation constant. Competitive binding studies in LK 14 kidney membranes employing chlorpromazine, prazosin, and oxymetazoline suggested that the increase in alpha 2-ARs in response to potassium depletion was due primarily to an increase in the B subtype of alpha 2-AR. Northern blot analysis demonstrated that renal alpha 2B-AR mRNA levels increased (190% of control) after 4 or 14 days on a potassium-deficient diet. In contrast, there was no difference in steady-state alpha 2A-receptor protein levels by Western blot analysis. We conclude that potassium depletion selectively increases the expression of the B subtype of alpha 2-AR with no detectable effect on alpha 2A-AR expression.
Erythropoietin enhances recovery from cisplatin-induced acute renal failure
Authors: Vaziri N. D. , Zhou X. J. , Liao S. Y.
Published: December 2017

Acute renal failure (ARF) is associated with erythropoietin (EPO) deficiency anemia. The present study was designed to determine whether the course of ARF can be altered by preventing EPO deficiency and the associated anemia. Sprague-Dawley rats were injected with a single dose of cisplatin (CP), 7 mg/kg intraperitoneally, and randomized into recombinant EPO-treated (EPO), placebo-treated (control), recombinant EPO-treated pair-fed (EPO-PF), and EPO-treated anemic (EPO-anemic) groups. They were then treated with daily injections of recombinant EPO, 100 U/kg, or placebo for 9 days. Animals in the EPO-anemic group received daily phlebotomies gauged to maintain hematocrits equal to those in the control group. Rats in the EPO-PF group were pair fed with the controls. The control and EPO-anemic groups showed a fall, whereas the EPO and EPO-PF groups showed a rise in hematocrit on day 9. Although blood volume on day 9 was significantly greater in the EPO group than in either the EPO-anemic group or the control group, it was comparable in the latter groups. An equally severe reduction in creatinine clearance (CCr) was found in all groups on day 4. However, measurements of CCr and inulin clearance on day 9 revealed a significantly greater functional recovery in the EPO, EPO-PF, and EPO-anemic groups than in the controls. The enhanced functional recovery with EPO administration was accompanied by an increased tubular regeneration and [3H]thymidine incorporation in the cortical tissue. No significant difference was found in either cortical tissue iron content or arterial blood pressure in the study groups.(ABSTRACT TRUNCATED AT 250 WORDS)
pHi determines rate of sodium transport in frog skin: results of a new method to determine pHi
Authors: Rick R.
Published: December 2017

The pH of the isolated frog skin epithelium was determined on a cellular and subcellular level based on the distribution of a weak organic acid, 4-bromobenzoic acid. The indicator is detectable by X-ray microanalysis due to the presence of an element label. The results show that the pH of principal cells, but not the Na concentration, is closely correlated with the rate of transepithelial Na transport. Acidification leads to an inhibition of Na transport, regardless of whether the change was spontaneous or experimentally induced. Under the conditions of this study, the pH of principal cells was not well regulated. At a bath pH of 7.0, large pH differences between the cell layers were detectable. In mitochondria-rich cells, the pH was a function of the intracellular Cl concentration but not the Na transport rate. The cytoplasmic pH consistently exceeded the nuclear pH. The nuclear-cytoplasmic pH differential in principal cells amounted to 0.3 pH units, which is equivalent to a nuclear potential of -17 mV. The results support the view that the intracellular pH (pHi) is an important regulator of transepithelial Na transport. Regulation is primarily achieved at the level of the apical Na channel, making the Na influx the rate-limiting step in Na reabsorption.
Systemic and renal effect of intravenous infusion of endothelin-1 in healthy human volunteers
Authors: Sorensen S. S. , Madsen J. K. , Pedersen E. B.
Published: December 2017

The effect of intravenous infusion of endothelin-1 (ET-1) at a rate of 1 pmol.min-1.kg-1 for 60 min (n = 9) or placebo (n = 9) was investigated in 18 healthy human volunteers with a mean age of 30 yr. In response to ET-1 infusion, concentration of ET-1 increased from 0.88 +/- 0.27 to 10.73 +/- 4.79 (SD) pmol/l. Diastolic blood pressure increased by 7.8% (P < 0.01) and heart rate decreased by 14.0% (P < 0.01), whereas systolic blood pressure did not change. Renal plasma flow decreased by 34.7%, glomerular filtration rate decreased by 16.1%, and renal vascular resistance increased by 66.0% (P < 0.01 all). Urinary sodium excretion decreased by 57.9% and urinary flow rate by 40.2% (P < 0.01 for both). As judged from the clearance of lithium, we found that ET-1 did not change absolute reabsorption of sodium and water in the proximal tubules, but in the distal tubules absolute reabsorption of both sodium and water decreased significantly. Plasma concentrations of angiotensin II, aldosterone, arginine vasopressin, and atrial natriuretic peptide did not change in response to ET-1 infusion. It is suggested that ET-1 at plasma concentrations found in certain pathophysiological conditions in humans may influence renal perfusion and renal sodium and water excretion.
Metabolic responses to nephrosis: effect of a low-protein diet
Authors: Choi E. J. , Bailey J. , May R. C. , Masud T. , Maroni B. J.
Published: December 2017

To determine whether dietary protein restriction (LPD) causes protein catabolism in adriamycin nephrosis, nephrotic and control rats were paired by weight and gavage fed an 8.5% protein diet for 3 days (protocol 1) or 12 days (protocol 2). Fasting whole body protein turnover was then measured using a constant infusion of L-[1-14C]leucine. After 3 days of LPD, proteinuria decreased slightly and body weight did not change in either group. In contrast, leucine oxidation and urinary urea nitrogen excretion in nephrotic rats decreased by 18% and 37%, respectively (P < or = 0.05). After 12 days of LPD, weight loss did not differ between groups. In contrast to protocol 1, proteinuria decreased by 45% in nephrotic rats fed LPD for 12 days, and leucine oxidation rats increased to the level of control rats. Rates of whole body protein synthesis (PS) and degradation (PD) did not differ between nephrotic and control rats receiving LPD for 3 or 12 days, but were significantly lower than rates measured in rats fed 22% protein. We conclude that 1) proteinuria stimulates protein conservation even when dietary protein intake is restricted; 2) the decrease in amino acid oxidation was dependent on moderate proteinuria, since prolonged LPD ameliorated nephrosis and leucine oxidation rates increased to control levels; and 3) since weight loss and rates of whole body PS and PD in nephrotic and control animals were indistinguishable, moderate proteinuria did not increase protein catabolism.
Immunolocalization of calcitriol receptor, 24-hydroxylase cytochrome P-450, and calbindin D28k in human kidney
Authors: Kumar R. , Schaefer J. , Grande J. P. , Roche P. C.
Published: December 2017

The precise localization of the calcitriol (1 alpha,25-dihydroxyvitamin D3) receptor (VDR) and the 25-hydroxyvitamin D3 [25(OH)D3] 24-hydroxylase cytochrome P-450 in the human kidney is unknown. Using newly developed polyclonal antibodies against the human VDR, we demonstrate that the receptor is present in cells of the distal tubule, the collecting duct, the proximal tubule, and in the parietal epithelial cells of the glomerulus. In the distal tubule and collecting duct not all cells contain epitopes for the receptor. The protein is not detected in glomerular capillaries, in the glomerular mesangium, in the interstitium, or in blood vessels. Specific polyclonal antibodies directed against the 25(OH)D3 24-hydroxylase cytochrome P-450 demonstrate epitopes for the cytochrome in cells of the proximal tubule, the distal tubule, glomerular parietal epithelial cells, and mesangial cells. The protein is absent from interstitial cells. Calbindin D28k is present exclusively in principal cells of the distal tubule and collecting duct. In the human kidney, the VDR is present in cells where vitamin D-inducible proteins are found; conversely it is absent from cells where vitamin D-dependent proteins are not present.
Synthesis and localization of C-type natriuretic peptide in mammalian kidney
Authors: Dean A. D. , Vehaskari V. M. , Greenwald J. E.
Published: December 2017

We have previously demonstrated the synthesis of atrial natriuretic factor (ANF) in the distal cortical nephron of the rat kidney. We now report the synthesis of C-type natriuretic peptide (CNP) in the rat, mouse, and human nephron. CNP mRNA was initially detected in the rat and mouse kidney, as well as in three transformed cell lines isolated from the proximal and distal nephron, using reverse transcription-polymerase chain reaction (RT-PCR). Confirmation of the kidney PCR product was performed by nucleotide sequence analysis and Southern hybridization. Northern hybridization of rat brain CNP cDNA to human kidney polyadenylated RNA detected a 1.4-kb gene transcript. Rat nephron segments were microdissected and subjected to RT-PCR to localize CNP mRNA. CNP mRNA was detected in the proximal convoluted tubule, cortical collecting duct, medullary thick limbs, and inner medullary collecting ducts. CNP, as detected by immunohistochemistry, was found to colocalize with CNP mRNA.
Feeding-induced increase in urinary dopamine excretion is independent of renal innervation and sodium intake
Authors: Muhlbauer B. , Osswald H.
Published: December 2017

Food intake increases urinary dopamine excretion. To investigate the role of renal nerve activity in this renal response to feeding, we studied urinary dopamine excretion after chronic bilateral renal denervation (DNX, n = 8) in male Sprague-Dawley rats. Controls were sham operated (CON, n = 6). In a paired crossover design, animals were studied both in fed and fasted state. Time course measurements of renal tissue norepinephrine (NE) content in a third group of DNX rats showed a decrease by 90% not earlier than 18 h post-DNX, remaining to be reduced to this level until day 11. Renal NE content in DNX animals was measured at the end of the study to confirm complete denervation. In 24-h urine, collected from fed and fasted conscious CON and DNX rats in metabolic cages, concentrations of dopamine and sodium were measured during two different sodium intake regimens, i.e., tap water and 1 g/dl NaCl as drinking water. No influence of renal innervation on urinary dopamine could be observed in fed and fasted animals. The marked changes in urinary volume and sodium output due to the different sodium regimens were not paralleled by urinary dopamine excretion. However, urinary dopamine was increased 2.1- to 2.4-fold (P < 0.01) because of feeding in all groups, independent of sodium balance and renal innervation. We conclude that in conscious rats both basal urinary dopamine excretion and its marked increase in response to feeding are independent of sodium balance and of renal nerve activity.
An Arg-Gly-Asp peptide stimulates Ca2+ efflux from osteoclast precursors through a novel mechanism
Authors: Yamakawa K. , Duncan R. , Hruska K. A.
Published: December 2017

We examined the effect of a peptide containing the Arg-Gly-Asp (RGD) sequence on 45Ca2+ efflux from osteoclast precursors. 45Ca(2+)-loaded osteoclast precursors were treated with GRGDSP (170 microM) for 10 min after 30 min of basal perfusion with a bicarbonate-containing buffer. GRGDSP significantly increased fractional efflux of Ca2+ from treated cells compared with vehicle-treated cells (P < 0.01) or cells treated with up to 200 micrograms/ml of a control peptide containing GRGESP. The effect of RGD was sustained for 15 min after the peptide was removed from the perfusate, but control levels of Ca2+ efflux returned by 1 h. The Ca2+ efflux effect of GRGDSP was most likely due to activation of the plasma membrane Ca(2+)-adenosinetriphosphatase (Ca(2+)-ATPase) pump, as indicated by its inhibition with vanadate and a calmodulin antagonist, N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide, and the absence of an effect of Na+/Ca2+ exchange inhibition. An inhibitor of cyclic nucleotide-dependent protein kinases, N-[2-(methylamino)ethyl]-5-isoquinoline-sulfonamide (0.1 mM), failed to inhibit GRGDSP-stimulated Ca2+ efflux. However, genistein and herbimycin A, inhibitors of protein-tyrosine kinases, blocked Ca2+ efflux stimulated by GRGDSP. The results indicate that RGD sequences of matrix proteins may stimulate Ca2+ efflux from osteoclasts through activation of protein-tyrosine kinases and suggest that GRGDSP-stimulated Ca2+ efflux is mediated via the plasma membrane Ca(2+)-ATPase.
Potential-induced changes in intracellular pH
Authors: Lyall V. , Biber T. U.
Published: December 2017

In a variety of cell types and tissues there is a strong dependence of intracellular pH (pHi) on membrane potential (Vm). Since cell Vm values can be altered by hormones, ion concentrations, and changes in membrane conductances, the potential-dependent changes in pHi may serve as an important mechanism by which cells can alter their pHi to an environmental stimulus. The H+ flux across the cell membranes is thought to take place via putative H+ channels that are blocked by low concentrations of divalent metal ions. However, in Na(+)-transporting epithelia, a major part of the H+ flux seems to be via the amiloride-sensitive apical Na+ channels, which are not sensitive to divalent metal ions. The H+ flux via the Na+ channels can be modulated by natriferic hormones and intracellular second messengers. The H(+)-conductive pathways may play an important role in signal transduction in some cells.
Impaired homologous upregulation of vitamin D receptor in rats with chronic renal failure
Authors: Koyama H. , Nishizawa Y. , Inaba M. , Hino M. , Prahl J. M. , DeLuca H. F. , Morii H.
Published: December 2017

We studied the homologous regulation of the vitamin D receptor (VDR) in the duodenum of rats with chronic renal failure. Mean basal nuclear 3H-labeled 1 alpha,25-dihydroxyvitamin D3 ([3H]1,25(OH)2D3) binding capacity was 48 and 43 fmol/mg protein for sham-operated and uremic rats with similar dissociation constants (Kd), respectively. These results coincided with those of immunoblot analysis, which found that VDR protein level of uremic rats was 87.6% that of sham-operated rats. In uremic rats, 1,25(OH)2D3, 2.0 micrograms/kg, failed to upregulate VDR protein levels until 24 h, in contrast to the significant increases produced in sham-operated rats at both 12 (1.55-fold) and 24 h (1.75-fold). Baseline level of VDR mRNA in uremic rats, determined by Northern blot analysis, was comparable to that in sham-operated rats. Treatment with 1,25(OH)2D3 slightly decreased VDR mRNA at 6-24 h in the sham-operated rats, in contrast to the increase seen at 6 h in uremic rats. These results suggest that the homologous upregulation of VDR was attenuated in rats with chronic renal failure because of an impairment at a translational and/or posttranslational step.
Renal function in hypertensive rats transgenic for mouse renin gene
Authors: Springate J. E. , Feld L. G. , Ganten D.
Published: December 2017

The recent development of a transgenic rat strain carrying the mouse ren-2 renin gene [TGR(mRen2)27] has provided a new model of hypertension characterized by suppressed plasma renin levels and marked hyperproreninemia. In this long-term study, we examined the kidney function of these animals. Transgenic rats had significantly (P < 0.01) higher blood pressures than control animals at 2, 4, and 8 mo of age. However, the severity of their hypertension diminished over time (225 +/- 8 mmHg at age 2 mo vs. 169 +/- 5 mmHg at age 8 mo, P < 0.001), indicating age-dependent transgene regulation. Whole kidney and single-nephron blood flows and glomerular filtration rates did not differ between control and TGR(mRen2)27 animals studied with micropuncture techniques at 4 and 8 mo of age. Preglomerular vasoconstriction was responsible for this normal autoregulatory response. Elevated preglomerular vascular resistance of transgenic rats prevented transmission of systemic hypertension to glomeruli at 4 but not 8 mo of age leading to increased glomerular capillary pressures in these older animals (53 +/- 1 vs. 48 +/- 1 mmHg, P < 0.05). Pathological albuminuria appeared as early as 2 mo of age but did not increase over the subsequent 6 mo of follow-up. The incidence of glomerulosclerosis, assessed at 4 and 8 mo of age, was greater in TGR(mRen2)27 than control animals (6.6 +/- 1.4% vs. 0.9 +/- 0.3%, P = 0.01) but did not differ between 4- and 8-mo-old transgenic rats. Glomerular ultrafiltration coefficients were significantly elevated (P < 0.05) in transgenic rats.(ABSTRACT TRUNCATED AT 250 WORDS)
Expression of Na-P(i) cotransport in rat kidney: localization by RT-PCR and immunohistochemistry
Authors: Custer M. , Lotscher M. , Biber J. , Murer H. , Kaissling B.
Published: December 2017

We have recently identified a rat kidney cortex Na-dependent transport system for phosphate (P(i)) by expression cloning (NaP(i)-2) (S. Magagnin, A. Werner, D. Markovich, V. Sorribas, G. Stange, J. Biber, and H. Murer. Proc. Natl. Acad. Sci. USA 90: 5979, 1993). In this study we have used reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry to establish the sites of expression of the NaP(i)-2-related mRNA and protein. RT-PCR was performed with single microdissected nephron segments. From these experiments we conclude that NaP(i)-2 mRNA is predominantly expressed in the proximal tubules of superficial and deep nephrons. No NaP(i)-2 mRNA was detected in the thick ascending limb of Henle's loop; however, faint NaP(i)-2 related PCR products were also observed in collecting ducts. Expression of the NaP(i)-2 protein was examined with the use of polyclonal antibodies raised against synthetic NaP(i)-2-derived peptides. Strong specific anti-NaP(i)-2 antiserum-mediated immunofluorescence was found in the convoluted part of proximal tubules and gradually decreased along the straight part. Immunofluorescence indicated that the NaP(i)-2 protein is present in the brush border of proximal tubular cells. In addition, NaP(i)-2-specific immunofluorescence was also observed in subapical vesicles. The described distribution of the NaP(i)-2 protein is in agreement with previously described nephron sites of P(i) reabsorption in the rat kidney and therefore suggests that the NaP(i)-2 transport system represents an Na-P(i) cotransporter involved in proximal tubular apical transport of phosphate.
Lithium reabsorption in perfused loops of Henle: effects of perfusion rate and bumetanide
Authors: Unwin R. J. , Walter S. J. , Shirley D. G.
Published: December 2017

The contribution of the loop segments to lithium reabsorption in sodium-replete, anesthetized Sprague-Dawley rats was examined by perfusing superficial loops of Henle between late proximal convolutions and early distal tubules. Preliminary experiments in which lithium was initially present only inside or outside the perfused loop confirmed substantial permeability of one or more of the loop segments to lithium. In subsequent experiments, in which lithium was infused intravenously and included in the perfusate so that the perfusate-to-plasma lithium concentration ratio was close to that found in late proximal tubules during lithium clearance studies, lithium reabsorption was inversely related to the perfusion rate: values for fractional lithium reabsorption (FRLi) at perfusion rates of 10, 20, and 30 nl/min were 58 +/- 3, 17 +/- 2, and 2 +/- 2%, respectively. Bumetanide (10(-6) M) markedly inhibited FRLi but also reduced water reabsorption, suggesting an effect on the pars recta at this dose; 10(-7) M bumetanide, which was without effect on water reabsorption, had only a small effect on FRLi at a perfusion rate of 10 nl/min but reduced FRLi by approximately 70% at 20 nl/min. We argue that the remarkable flow dependency of lithium reabsorption, together with its bumetanide sensitivity, provides evidence for significant voltage-driven lithium reabsorption in the thick ascending limb of Henle.
Distinct cellular localization of mRNAs for three subtypes of prostaglandin E receptor in kidney
Authors: Sugimoto Y. , Namba T. , Shigemoto R. , Negishi M. , Ichikawa A. , Narumiya S.
Published: December 2017

Distribution of the mRNAs for three subtypes of prostaglandin E (PGE) receptors in the mouse kidney was investigated by in situ hybridization. The mRNA for EP1 subtype, which is coupled to Ca2+ mobilization, was specifically localized to the collecting ducts from the cortex to the papilla. The mRNA for EP2 subtype, which is linked to stimulation of adenylate cyclase, was localized to the glomeruli. The mRNA for EP3 subtype, which is coupled to inhibition of adenylate cyclase, was located densely in the tubules in the outer medulla and in the distal tubules in the cortex. These results exhibit distinct cellular localization of three subtypes of PGE receptor in the kidney and suggest that PGE2 exerts multiple functions via these subtypes expressed in different segments of the nephron.