bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of glucocorticoids on renal cortical NHE-3 and NHE-1 mRNA
Authors: Baum M. , Moe O. W. , Gentry D. L. , Alpern R. J.
Published: December 2017

Glucocorticoids play an important role in modulating proximal tubule acidification. Chronic systemic administration of dexamethasone increases the rate of bicarbonate absorption in isolated perfused proximal convoluted tubules and Na+/H+ antiporter activity in renal brush-border membrane vesicles. The proximal tubule expresses mRNA corresponding to two known Na+/H+ antiporter isoforms: NHE-3, an amiloride-resistant apical membrane Na+/H+ antiporter; and NHE-1, an amiloride-sensitive Na+/H+ antiporter found on most mammalian cells. Administration of dexamethasone for 1 and 2 days (600 micrograms/kg twice daily and 2 h before animals were killed) increased NHE-3 mRNA abundance 1.3- and 2.5-fold, respectively, but had no effect on NHE-1 mRNA abundance. Aminoglutethimide-induced glucocorticoid deficiency had no effect on NHE-1 or NHE-3 mRNA abundance. Incubation of proximal tubules for 3 h with 10(-5) M dexamethasone increased proximal tubule Na+/H+ antiporter activity from 0.69 +/- 0.04 to 0.92 +/- 0.03 pH units/min (P < 0.01); however, there was no increase in NHE-3 or NHE-1 mRNA abundance. Similarly, there was no effect on NHE-3 or NHE-1 mRNA abundance in rabbit renal cortex 4 h after intravenous administration of 600 micrograms/kg dexamethasone. Thus chronic dexamethasone increases NHE-3 but not NHE-1 mRNA abundance. The acute increase in Na+/H+ antiporter activity induced by dexamethasone occurs by mechanisms independent of changes in NHE-1 and NHE-3 mRNA abundance.
Two types of K+ channel in thick ascending limb of rat kidney
Authors: Wang W. H.
Published: December 2017

We have used the patch-clamp technique to study the apical K+ channels in the thick ascending limb (TAL) of the rat kidney. Two types of K+ channels, a low-conductance and an intermediate-conductance K+ channel, were identified in both cell-attached and inside-out patches. We confirmed the previously reported intermediate-conductance K+ channel (72 pS), which is inhibited by millimolar cell ATP, acidic pH, Ba2+, and quinidine (4). We now report a second K+ channel in apical membrane of the TAL. The slope conductance of this low-conductance K+ channel is 30 pS, and its open probability is 0.80 in cell-attached patches. This channel is not voltage dependent, and application of 2 mM ATP in the bath inhibits channel activity in inside-out patches. In addition, 250 microM glyburide, an ATP-sensitive K+ channel inhibitor, blocks channel activity, whereas the same concentration of glyburide has no inhibitory effect on the 72-pS K+ channel. Channel activity of the 30-pS K+ channel decreases rapidly upon excision of patches (channel run down). Application of 0.1 mM ATP and the catalytic subunit of adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase A (PKA) restores channel activity. Furthermore, addition of 0.1 mM 8-(4-chlorophenylthio)-cAMP or 50-100 pM vasopressin in the cell-attached patches increases channel activity. In conclusion, two types of K+ channels are present in the apical membrane of TAL of rat kidney, and PKA plays an important role in modulation of the low-conductance K+ channel activity.
Presence and distribution of endothelin-1 gene expression in human kidney
Authors: Pupilli C. , Brunori M. , Misciglia N. , Selli C. , Ianni L. , Yanagisawa M. , Mannelli M. , Serio M.
Published: December 2017

To investigate the presence and the distribution of preproendothelin-1 (prepro-ET-1) mRNA in human kidney, eight human kidneys obtained at surgery from patients affected by localized renal tumors were studied. Northern blot analysis using a human prepro-ET-1 cDNA probe labeled with 32P showed the presence of a single band of approximately 2.3 kb that was present both in the renal cortex and medulla of all the kidneys studied. Densitometric analysis of hybridization signals demonstrated that prepro-ET-1 mRNA levels in the renal medulla were 2.2-fold higher than those in the renal cortex. The distribution of prepro-ET-1 mRNA in human kidney was investigated by in situ hybridization using a human prepro-ET-1 RNA probe labeled with 35S. The greatest density of prepro-ET-1 mRNA was observed in the renal medulla, where hybridization signal was demonstrated in vasa recta bundles and capillaries and in collecting ducts. By combining in situ hybridization with immunohistochemical detection of von Willebrand factor, we demonstrated that 93 +/- 2.5% of nontubular medullary cells containing prepro-ET-1 mRNA were endothelial cells. In the cortex, prepro-ET-1 mRNA was localized in the endothelial layer of arcuate and interlobular arteries and veins and in the endothelial cells of afferent arterioles. The results of the present study demonstrate that ET-1 gene expression is present in vascular and tubular structures of the human kidney. It is possible that ET-1 synthesized locally in the human kidney represents a local system affecting renal hemodynamics and functions through paracrine and/or autocrine actions on different renal structures.
Water and urea diffusive permeabilities in isolated proximal tubule cells
Authors: Echevarria M. , Gonzalez E. , Gutierrez A. , Whittembury G.
Published: December 2017

We measured the water (3H2O) and urea ([14C]urea) diffusive permeabilities (Pd) in intact proximal tubule cells (PTC). Isolated rabbit PTC were packed in polyethylene tubes to measure the diffusion coefficients for 3H2O and [14C]urea at 14 degrees C. Pd values for water and urea were estimated from the diffusion coefficients in intact cells, in extracellular and in intracellular media using a model of parallel and series diffusion in the packed PTC. Pd was 65 +/- 8 and 18 +/- 3 microns/s for water and urea, respectively. We examined the effect of different pharmacological agents on the Pd values for water and for urea. p-Chloromercuribenzenesulfonate at 2 mM markedly inhibited the Pd of water and urea. Phloretin (1 mM) inhibited the Pd for urea, whereas it increased Pd for water. KMnO4 (30 microM) markedly inhibited urea Pd but did not alter water Pd. The values for energy of activation of Pd for water and for urea were 2.9 +/- 2 and 7.3 +/- 4 kcal/mol, respectively. These results show that water and urea do not share the same pathway across the PTC membrane.
In situ hybridization localization of mRNA encoding inducible nitric oxide synthase in rat kidney
Authors: Ahn K. Y. , Mohaupt M. G. , Madsen K. M. , Kone B. C.
Published: December 2017

We used in situ hybridization with a digoxigenin-labeled cRNA for inducible nitric oxide synthase (iNOS) to characterize the intrarenal distribution of iNOS transcripts in normal and lipopolysaccharide (LPS)-treated rats. In normal rats, the S3 segment of the proximal tubule, the cortical and medullary thick ascending limb, the distal convoluted tubule, and the cortical and inner medullary collecting duct were intensely labeled, whereas the thin limbs of Henle, proximal convoluted tubule, outer medullary collecting duct, and medullary interstitial cells were weakly labeled. LPS-treated rats exhibited a similar labeling pattern, but with increased staining of mesangial cells, medullary interstitial cells, and papillary surface epithelium. The renal vasculature, including the afferent arteriole, was not labeled in either group. No cellular labeling was observed when the sections were hybridized with the sense iNOS probe. These results indicate that iNOS mRNA is tonically and differentially expressed along the normal rat nephron and that LPS induces iNOS gene expression in normally quiescent mesangial cells, medullary interstitial cells, and papillary surface epithelium.
Cellular mechanisms of acute and chronic adaptation of rat renal P(i) transporter to alterations in dietary P(i)
Authors: Levi M. , Lotscher M. , Sorribas V. , Custer M. , Arar M. , Kaissling B. , Murer H. , Biber J.
Published: December 2017

Recently, the cDNA for a Na-P(i) cotransport system of rat kidney cortex (NaPi-2) has been identified by expression cloning. Using polyclonal antibodies raised against this renal Na-P(i) cotransport system, and using the polymerase chain reaction after reverse transcription of mRNA in microdissected nephron segments, we recently demonstrated that NaPi-2-related mRNA and protein is expressed in the brush-border membranes (BBM) of the proximal tubules of rat kidney. The purpose of the present study was to study the cellular mechanisms involved in adaptation of rat renal Na-P(i) cotransporter to acute and chronic alterations in dietary P(i). Compared with rats fed chronically (7 days) a high-P(i) diet (1.2%), in rats fed chronically a low-P(i) (0.1%) diet the 3.4-fold increase in BBM Na-P(i) cotransport rate (chronic upregulation) was associated with a 2.2-fold increase in renal cortical NaPi-2 mRNA and a 4.9-fold increase in BBM NaPi-2 protein abundances. In contrast, compared with rats fed chronically (7 day) a high-P(i) diet, in rats fed acutely (2 h) a low-P(i) diet the 1.5-fold increase in Na-P(i) cotransport rate (acute upregulation) was associated with a 1.8-fold increase in NaPi-2 protein but no change in NaPi-2 mRNA abundance. Similarly, compared with rats fed chronically a low-P(i) diet, in rats fed acutely (2 h) a high-P(i) diet the 1.9-fold decrease in Na-P(i) cotransport rate (acute downregulation) was associated with a 3.8-fold decrease in NaPi-2 protein but no change in NaPi-2 mRNA abundance.(ABSTRACT TRUNCATED AT 250 WORDS)
Angiotensin II regulates H(+)-ATPase activity in rat cortical collecting duct
Authors: Tojo A. , Tisher C. C. , Madsen K. M.
Published: December 2017

Angiotensin II (ANG II) plays an important role in the regulation of solute transport in the kidney, and its effect on proximal tubule sodium and fluid transport has been studied extensively. Although there is evidence that ANG II receptors are present also in the distal nephron and collecting duct, little is known about the physiological role of ANG II in these segments of the renal tubule. Preliminary studies in our laboratory suggest that ANG II may have both structural and functional effects on intercalated cells in the cortical collecting duct (CCD). Therefore, the present study examines the effect of ANG II on H(+)-adenosinetriphosphatase (H(+)-ATPase) and H(+)-K(+)-ATPase activity in individual CCD segments microdissected from collagenase-treated rat kidneys. The H(+)-ATPase was measured as bafilomycin-sensitive ATPase activity, and H(+)-K(+)-ATPase was measured as Sch-28080-sensitive ATPase activity, by a fluorometric microassay. Preincubation of CCD segments with ANG II, 10(-10)-10(-5) M, caused a dose-dependent decrease in H(+)-ATPase activity with maximum inhibition at 10(-8) M of ANG II. The inhibitory effect of ANG II was abolished when tubules were incubated with ANG II in the presence of 10(-6) M losartan, indicating that the inhibition was mediated via specific AT1 receptors. The AT2-receptor antagonist, PD-123319, had no effect on the ANG II-mediated inhibition of H(+)-ATPase activity. Preincubation of CCD segments with 10(-10) or 10(-7) M ANG II had no effect on H(+)-K(+)-ATPase activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Role of CD11a and CD11b in ischemic acute renal failure in rats
Authors: Rabb H. , Mendiola C. C. , Dietz J. , Saba S. R. , Issekutz T. B. , Abanilla F. , Bonventre J. V. , Ramirez G.
Published: December 2017

Leukocytes, particularly neutrophils, have been implicated in ischemic-reperfusion organ injury (IRI). However, their role in kidney IRI is controversial. Leukocytes express the adhesion molecules CD11/CD18 on their surface, which mediate many functions that can lead to tissue damage. To determine the role of CD11a and CD11b in IRI in the kidney, uninephrectomized Sprague-Dawley rats were pretreated with monoclonal antibodies (MAbs) directed against CD11a and CD11b or control MAbs. The serum creatinine (SCr), complete blood count, and kidney histopathological damage scores (PDS) (scale: 0-4) were assessed prior to and 24 h after 60 min of ischemia. Mean SCr 24 h after ischemia was significantly decreased in the anti-CD11a- and -CD11b-treated group compared with the control MAb-treated group (2.5 +/- 0.3 mg/dl vs. 3.4 +/- 0.2 mg/dl, P <0.05). PDS were also reduced in the CD11a and CD11b group compared with controls (2.7 +/- 0.2 vs. 3.5 +/- 0.1, P < 0.001). These data show that the CD11/CD18 leukocyte adhesion pathway plays a role in mediating ischemic acute renal failure in rats.
Determinants of intrarenal oxygenation. I. Effects of diuretics
Authors: Brezis M. , Agmon Y. , Epstein F. H.
Published: December 2017

To study renal cortical and medullary oxygen tensions, we used sensitive Clark-type O2 microelectrodes, inserted by micromanipulators into the cortex and medulla of kidneys of anesthetized rats. As previously reported, under basal conditions, medullary PO2 was significantly lower than cortical PO2. Furosemide, which inhibits reabsorptive transport in the medullary thick ascending limb, increased medullary PO2 from 16 +/- 4 to 35 +/- 4 mmHg (P < 0.0005) without altering cortical PO2. This effect, reproduced by ethacrynic acid and bumetanide, was selective for loop diuretics and was directly due to decreased tubular O2 consumption, since medullary blood flow was remarkably reduced by furosemide (-28 +/- 6% from baseline, P < 0.0001, as measured by a laser-Doppler probe). By contrast, acetazolamide, which decreases proximal tubule metabolism, selectively increased cortical PO2. These data are, in general, consistent with tubular metabolism as a major determinant of intrarenal oxygenation and suggest, in particular, that medullary reabsorptive work is at least in part responsible for renal medullary hypoxia.
Modulation by Gly, Ca, and acidosis of injury-associated unesterified fatty acid accumulation in proximal tubule cells
Authors: Weinberg J. M. , Venkatachalam M. A. , Goldberg H. , Roeser N. F. , Davis J. A.
Published: December 2017

We have examined the dependence of unesterified fatty acid accumulation by intact, freshly isolated proximal tubules on Ca2+, pH, and the cytoprotective amino acid, glycine, during injury induced by hypoxia, antimycin, or antimycin plus ionomycin. In the absence of glycine, similarly high levels of fatty acid accumulation were seen during all three injury conditions irrespective of whether tubules were incubated in normal 1.25 mM Ca2+ medium or in medium where Ca2+ was buffered to 0.1 microM, a maneuver which prevented injury-associated increase of cytosolic-free Ca2+ as measured with fura 2. In the presence of glycine, which strongly suppressed development of lethal membrane damage for at least 60 min and did not have any apparent direct effects on fatty acid accumulation, both Ca(2+)-independent and Ca(2+)-dependent components of fatty acid accumulation were discernible. The Ca(2+)-independent component accounted for approximately 2/3 of fatty acid accumulation and did not vary as Ca2+ ranged from 10 nM to 1 microM. Unequivocal Ca(2+)-dependent accumulation occurred when Ca2+ exceeded 10 microM. Lowering pH to 6.9 had a moderate, generalized suppressive effect on fatty acid accumulation, including the major Ca(2+)-independent component, irrespective of the presence of glycine. These data emphasize the role of Ca(2+)-independent fatty acid accumulation during proximal tubule cell injury, clarify the modulatory actions of the potent, intrinsic cytoprotective factors, glycine and reduced pH, and provide insight into the relationship between fatty acid accumulation and lethal membrane damage.
Identification of a rat glomerular mesangial cell mitogenic 5-HT2A receptor
Authors: Nebigil C. G. , Garnovskaya M. N. , Spurney R. F. , Raymond J. R.
Published: December 2017

Previous studies have demonstrated the presence of mitogenic serotonin [i.e., 5-hydroxytryptamine (5-HT)] receptors on glomerular mesangial cells and have linked those receptors to a complicated array of intracellular and autocrine/paracrine signaling pathways [T. Knauss and H. E. Abboud. Am. J. Physiol. 251 (Renal Fluid Electrolyte Physiol. 20): F844-F850, 1986; and N. Takuwa, M. Ganz, Y. Takuwa, R. B. Sterzel, and H. Rasmussen. Am. J. Physiol. 257 (Renal Fluid Electrolyte Physiol. 26): F431-F439, 1989]. Those studies suggested that the mesangial subtype of 5-HT receptor is a member of the 5-HT2 receptor family, which consists of three known members, designated as subtypes A, B, and C. The purpose of the current study was to identify the subtype of 5-HT2 receptor present on mesangial cells. Northern blot showed detectable mRNA for a putative 5-HT2A receptor, but not for 5-HT1A or 5-HT2C receptors. Reverse transcription-polymerase chain reaction (RT-PCR) with degenerate oligonucleotides derived from the putative third and sixth transmembrane domains of cloned 5-HT2 receptors yielded a 580-nucleotide (nt) fragment. RT-PCR with primers highly specific for the 5-HT2A receptor and designed to amplify > 95% of its coding block yielded a product of 1,320 nt. Nested PCR reactions yielded products of the predicted sizes for the 5-HT2A receptor. Partial sequence information was obtained, and the sequence corresponded exactly (627/627 nt) to that published for the cloned rat brain 5-HT2A receptor. These studies identify the mesangial cell mitogenic 5-HT receptor as a 5-HT2A receptor subtype.
Periodic high extracellular glucose enhances production of collagens III and IV by mesangial cells
Authors: Takeuchi A. , Throckmorton D. C. , Brogden A. P. , Yoshizawa N. , Rasmussen H. , Kashgarian M.
Published: December 2017

We examined the effects of periodic changes in extracellular glucose concentration on matrix production and proliferation using three groups of cultured rat mesangial cells (MCs): 1) MCs in medium with continuous 5 mM glucose (CL), 2) MCs in medium alternating daily between 5 and 25 mM glucose (PH), and 3) MCs in medium with continuous 25 mM glucose (CH). MCs cultured in PH for 10 days produced 329 and 110% more type III collagen protein than MCs cultured in CL and CH, respectively. MCs cultured in PH induced 31 and 14% more type IV collagen than MCs cultured in CL and CH, respectively. Extracellular glucose concentration had no effect on the amount of type I collagen produced. MCs cultured in PH or CH for 5 days also expressed increased levels of type I, III, and IV collagen mRNA compared with MCs cultured in CL. MCs cultured in PH for 8-10 days also produced significantly more DNA than MCs in CL or CH. These data suggest that the temporal pattern of exposure to high extracellular glucose plays a role in regulating matrix formation and cellular proliferation by MCs. Furthermore, periodic elevations of extracellular glucose had a greater stimulatory effect on collagen production than a sustained elevation. These results suggest that decreasing the variability of blood glucose concentration may decrease the adverse effect of elevated glucose levels on MC matrix production and the progression of diabetic glomerulopathy.
Validation of miniature ultrasonic transit-time flow probes for measurement of renal blood flow in rats
Authors: Welch W. J. , Deng X. , Snellen H. , Wilcox C. S.
Published: December 2017

This study validates the accuracy of miniature ultrasonic transit-time flow probes for measuring renal blood flow (RBF) in the rat. Probes for 1-mm and 2-mm vessels were calibrated ex vivo using excised arteries at varying flow rates and hematocrit (Hct). Correlation between measured and true flow rates for the 2-mm probe were identical (r = 1.0) at both normal and subnormal Hct values. Correlation for the 1-mm probe was high (r = 0.994) at normal Hct, but varied at both high flow rates and subnormal Hct values. In vivo correlation of RBF measurements using the 1-mm probe with the clearance and extraction of p-aminohippuric acid showed a high correlation (r = 0.84; n = 72, P < 0.0001) over a wide range of flow rates (0.5-21 ml/min) and Hct (36-74%). Zero flow levels remained steady, averaging -0.2 +/- 0.2 ml/min during occlusion in the living animal and -0.1 +/- 0.3 ml/min after exsanguination. This study shows that the ultrasonic transit-time flowmeter (1-mm and 2-mm probes) is a reasonably accurate and reliable method with which to measure RBF in the anesthetized, acute-instrumented rat.
Effects of endothelins on renin secretion from isolated mouse renal juxtaglomerular cells
Authors: Ritthaler T. , Scholz H. , Ackermann M. , Riegger G. , Kurtz A. , Kramer B. K.
Published: December 2017

This study was performed to examine the effects of endothelin (ET)-1, ET-2, and ET-3 on renin secretion from cultured mouse renal juxtaglomerular (JG) cells. Although different ETs had no consistent effect on basal renin secretion, they equipotently inhibited adenosine 3',5'-cyclic monophosphate (cAMP)-stimulated renin release with a concentration of approximately 3 nM inhibiting 50% of maximal response. ETs did not significantly affect renin release stimulated by the nitric oxide donor sodium nitroprusside (100 microM) or that stimulated by low [2 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] or high (3 mM CaCl2) extracellular calcium. The inhibitory effect of ETs on cAMP-dependent renin secretion was abolished by lowering extracellular calcium concentration to the nanomolar range. However, the action of ETs was not changed by the ETA receptor antagonist BQ-123 (100 nM) and was mimicked by ETB receptor agonists IRL-1620 (1 microM), sarafotoxin S6b (1 microM), and [Ala1,3,11,15]ET-1 (1 microM). All ETs induced calcium oscillations in JG cells that were dependent on extracellular calcium and were associated with prominent calcium-activated chloride currents. These findings suggest that ETs inhibit rather selectively the cAMP-activated pathway of renin secretion through a calcium-sensitive process. The action of ETs on renal JG cells appears to be mediated via ETB receptors and is presumably related to activation of phospholipase C and subsequent events.
Expression of vascular endothelial growth factor and its receptors in human renal ontogenesis and in adult kidney
Authors: Simon M. , Grone H. J. , Johren O. , Kullmer J. , Plate K. H. , Risau W. , Fuchs E.
Published: December 2017

Vascular endothelial growth factor (VEGF) may modulate vascular permeability, chemotaxis for monocytes, and protease activity. In addition, VEGF may play a role in embryonic and tumor angiogenesis. In fetal mouse kidney, VEGF mRNA and protein expression have been demonstrated. This finding led to the hypothesis that VEGF might be involved in renal growth and development. To further elucidate the role of VEGF in human kidney, expression of VEGF and its receptors, the specific tyrosine kinase receptors, fit-1 and KDR, were studied. In fetal (6-24 gestational wk; mesonephros and metanephros) and adult kidney, VEGF mRNA and protein could be colocalized in glomerular epithelia and collecting duct cells by in situ hybridization and immunohistology. By reverse transcription-polymerase chain reaction, mRNA of three VEGF isoforms, VEGF121, VEGF165, and VEGF189, were found in fetal kidney and cortex, isolated glomeruli, and medulla of adult human kidney. KDR and flt-1 mRNA were coexpressed in endothelia of glomeruli and in peritubular capillaries in fetal and adult kidney. These data support the assumption that VEGF and its receptors may influence renal ontogenesis. We speculate that the constitutive expression of VEGF in adult kidney may be required for the function of VEGF receptor positive-fenestrated endothelia in glomeruli and postglomerular vessels. The expression of VEGF in collecting duct and of its receptors in medullary capillaries may in addition be relevant for maintaining medullary osmolality.
IL-1 beta increases laminin B2 chain mRNA levels and activates NF-kappa B in rat glomerular epithelial cells
Authors: Richardson C. A. , Gordon K. L. , Couser W. G. , Bomsztyk K.
Published: December 2017

The amount and distribution of laminin in the glomerular basement membrane (GBM) change in the course of many types of glomerular disease. Because interleukin-1 (IL-1) is thought to play a role in the pathogenesis of glomerulonephritis, it raises the possibility that this and other cytokines might regulate laminin gene expression. To determine whether laminin B2 chain mRNA levels change in response to cytokines, total mRNA from rat glomerular epithelial cells (GEC) grown in culture was analyzed by Northern blots. These studies showed an increase in laminin B2 chain mRNA levels in GEC treated with IL-1 beta. Nuclear factor kappa B (NF-kappa B) is a cytokine-responsive factor that regulates transcription of many genes from the cognate kappa B enhancer element. The mouse laminin B2 chain promoter contains several kappa B-like motifs, suggesting that NF-kappa B might be involved in IL-1-induced laminin B2 chain gene expression. Nuclear extracts from IL-1 beta-treated GEC showed increased binding to the immunoglobulin kappa B enhancer element and to a kappa B consensus sequence from the murine laminin B2 chain promoter in an electrophoretic mobility-shift assay (EMSA). The immunoglobulin kappa B and the laminin B2 chain kappa B-like motifs competed for the same DNA binding activity in nuclear extracts from IL-1 beta-treated GEC. Pretreatment of these nuclear extracts with antibodies to either the p50 or p65 subunits of NF-kappa B abrogated the DNA binding activity recognized by either of the two DNA motifs.(ABSTRACT TRUNCATED AT 250 WORDS)
Amiloride-sensitive Na+/H+ antiporter in basolateral membrane of hamster ascending thin limb of Henle's loop
Authors: Fujiwara I. , Kondo Y. , Igarashi Y. , Inoue C. N. , Takahashi N. , Tada K. , Abe K.
Published: December 2017

The mechanisms of intracellular pH (pHi) regulation were investigated in the in vitro microperfused hamster ascending thin limb (ATL) of Henle's loop with the fluorescent pH indicator, 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. pHi of ATL cells was 7.05 +/- 0.02 (n = 30) when microperfused with a CO2/HCO(3-)-buffered solution. In HEPES-buffered solution, pHi was 7.10 +/- 0.02 (n = 16), which was significantly higher than the value in CO2/HCO(3-)-buffered solution (P < 0.05, n = 16). In HEPES-buffered solution, elimination of Na+ and addition of 1 mM amiloride to basolateral solution decreased the pHi by 0.12 +/- 0.03 (n = 6) and 0.11 +/- 0.02 (n = 5) at 1 min, respectively. The same manipulations in the luminal solution had no effect on pHi. One millimolar of N-ethylmaleimide (NEM) added to either side of ATL caused no significant change in pHi. Elimination of K+ on either side of ATL did not alter pHi. After adding 20 mM NH4Cl to basolateral solution, pHi instantaneously increased from 7.17 +/- 0.01 to 7.51 +/- 0.03 (n = 3), and then returned to steady-state level of 7.21 +/- 0.05 (n = 15) in 3 min. Removal of NH4Cl from basolateral solution then caused a rapid fall in pHi to 6.31 +/- 0.05 (n = 15), followed by spontaneous recovery at a rate of 0.43 +/- 0.06 unit/min (n = 15).(ABSTRACT TRUNCATED AT 250 WORDS)
Regulation of renal cortical Na-HCO3 cotransporter. II. Role of G proteins
Authors: Ruiz O. S. , Qiu Y. Y. , Wang L. J. , Arruda J. A.
Published: December 2017

We examined the regulation of the renal cortical basolateral Na-HCO3 cotransporter by G proteins. Na-HCO3 cotransporter activity was measured in highly purified rabbit renal cortical basolateral membranes (BLMV) as the difference in 22Na uptake in presence of HCO3- and gluconate. HCO(3-)-dependent 22Na uptake was significantly inhibited by 10 microM guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S), a G protein activator. In contrast, addition of 50 microM guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), an inhibitor of G protein, prevented the inhibition of the Na-HCO3 cotransporter activity by GTP gamma S. AlF4-, another G protein activator, also inhibited the activity of the Na-HCO3 cotransporter. This inhibitory effect of G protein on the Na-HCO3 cotransporter activity was not prevented by dideoxyadenosine, an adenylate cyclase inhibitor, or by the protein kinase A inhibitor, suggesting a direct effect of G protein on the cotransporter. To identify the G proteins that mediate the regulation of the Na-HCO3 cotransporter, purified BLMV were ADP ribosylated in presence of cholera toxin or pertussis toxin. Autoradiograms of BLMV incubated with [32P]NAD showed that cholera and pertussis toxins caused ADP ribosylation of 42- and 41-kDa G proteins, respectively. To determine whether the ADP ribosylation by cholera or pertussis toxin was associated with alterations of the Na-HCO3 cotransporter activity, we measured HCO(3-)-dependent 22Na uptake in BLMV treated with 20 micrograms/ml cholera toxin or with 100 ng/ml pertussis toxin. Na-HCO3 cotransporter activity was significantly decreased by both cholera and pertussis toxins.(ABSTRACT TRUNCATED AT 250 WORDS)
Molecular characterization of renal calcium channel beta-subunit transcripts
Authors: Yu A. S. , Boim M. , Hebert S. C. , Castellano A. , Perez-Reyes E. , Lytton J.
Published: December 2017

An apical, hormone-regulated, calcium entry channel in the distal convoluted tubule and/or connecting tubule (DCT/CNT) is thought to play an important role in controlling renal calcium excretion. We previously identified a gene transcript encoding the pore-forming alpha 1-subunit of a calcium channel (alpha 1A, or CaCh4) which may be a candidate for such a molecule. The properties of voltage-dependent calcium channels are known to be modulated by their beta-subunits. To identify the accessory beta-subunit of DCT/CNT calcium channels, degenerate primers based on published beta-subunit sequences were used to amplify rat kidney cDNA by the polymerase chain reaction (PCR), and the products were subcloned and sequenced. Alternatively spliced transcripts of three beta-subunit genes (beta 2, beta 3, and beta 4) were identified. Northern blot analysis indicated that beta 4-subunit is preferentially expressed in kidney cortex. Transcripts of all three beta-subunit genes were detected by PCR in microdissected nephron segments, but only beta 4-subunit was found in DCT/CNT. As the beta 4- and alpha 1A-subunits colocalize to the DCT/CNT, we hypothesize that they may be constituent subunits of a renal calcium channel regulated by a hormone(s).
Apolipoprotein AI levels are increased in part as a consequence of reduced catabolism in nephrotic rats
Authors: Kaysen G. A. , Hoye E. , Jones H.
Published: December 2017

Apolipoprotein AI (apo AI) synthesis, measured as the turnover of 125I-labeled apo AI-labeled high-density lipoprotein (HDL), was increased significantly in rats with Heymann nephritis (HN) vs. control Sprague-Dawley (SD) rats. However, fractional apo AI catabolic rate was also significantly less in HN vs. SD. We used 125I-apo AI tyramine cellobiose HDL, a marker retained at the catabolic site, to establish where apo AI catabolism decreased in six HN rats, seven rats with adriamycin (Adria)-induced nephrosis, and six control SD. Total renal apo AI catabolism, plus urinary losses, were the same in all three groups, despite significant urinary apo AI in HN and Adria rats. Apo AI catabolism was reduced in skin in both nephrotic groups, accounting for approximately 44% of reduced in apo AI catabolism. Thus a significant fraction of apo AI is catabolized in skin of normal male rats. Reduced apo AI catabolism in skin contributes to increased plasma levels in nephrotic rats.
Drawbacks of the constant-infusion technique for measurement of renal function
Authors: Van Acker B. A. , Koomen G. C. , Arisz L.
Published: December 2017

We investigated the validity of the steady-state constant infusion method (CIM), in which quantitative urinary recovery and constant plasma concentrations of the solute infused are required. Successive 3-h clearances of inulin and p-aminohippuric acid (PAH) were determined for 27 h in 25 patients with renal disease. Results were compared with the standard method of bladder clearance (StM) and with a modified CIM (ModCIM). The 24-h urinary recovery was incomplete for both inulin and PAH. Mean 24-h ModCIM inulin clearance overestimated StM by 4.5 ml.min-1 x 1.73 m-2 (range 0–9, P < 0.001) independent of the extent of renal impairment and pointed to slow distribution and/or extrarenal clearance of inulin. For PAH, the difference between ModCIM and StM clearance was related to the average PAH clearance by ModCIM and StM (r = 0.78). Furthermore, neither plasma inulin nor PAH became completely constant, because of the circadian rhythm in renal function. In conclusion, the conditions of the steady-state CIM technique are not fulfilled, and the method is not suitable for accurate measurement of inulin and PAH clearance, especially when the clearance is low.
Role of chloride in the variable response of the kidney to cyclooxygenase inhibition
Authors: Yin K. , McGiff J. C. , Bell-Quilley C. P.
Published: December 2017

The role of prostanoids in renal function remains unclear, as inhibitors of cyclooxygenase (COX) have contrasting effects. We postulated that these inconsistencies were related to differential effects of the prevailing chloride concentration on COX-dependent mechanisms. In oncotically perfused rat kidneys, in the presence of either high (117 mM) or low (87 mM) chloride with sodium held constant, low chloride resulted in a higher glomerular filtration rate (GFR) than with high chloride, i.e., 1.2 +/- 0.2 and 0.5 +/- 0.1 ml/min, respectively, for the last clearance period. Water and electrolyte excretion and levels of immunoassayable prostaglandins were higher with low chloride. Indomethacin (10 microM) had opposite effects on renal function depending on the chloride levels, although prostaglandin release was inhibited similarly. For example, indomethacin substantially reduced the elevated urine flow and sodium excretion in the low-chloride group, which, by the last period, were reduced from 111 +/- 32 to 37 +/- 3 microliters/min and 8.3 +/- 2.9 to 3.1 +/- 0.6 mu eq/min, respectively, whereas the lower urine flow and sodium excretion in the high-chloride group increased from 32 +/- 8 to 109 +/- 15 microliters/min and 2.5 +/- 0.8 to 7.1 +/- 1.6 mu eq/min, respectively. In summary, inhibition of COX has differential effects depending on the prevailing chloride concentration, or conversely, high and low chloride have contrasting effects on renal function, which are reversed by COX inhibition. We suggest that prohypertensive and antihypertensive COX-dependent mechanisms are linked to chloride; the latter is an integral component in the development of salt-sensitive hypertension.
Hypotonicity increases basolateral taurine permeability in rabbit proximal convoluted tubule
Authors: Breton S. , Marsolais M. , Laprade R.
Published: December 2017

The permeabilities of the basolateral membrane of rabbit proximal convoluted tubule (PCT) to taurine (PTau) and glucose (PGlc) were estimated under control and hypotonic conditions using the initial rate of increase in cellular volume (CV) induced on isotonic replacement of 40 mM mannitol by one or the other of these substrates. Under control conditions, addition of taurine led to an increase in CV at an initial rate of 7.1 +/- 1.7%/min, leading to a cell swelling of 30.2 +/- 4.8% after 5 min (n = 6). Addition of glucose led to an increase in CV at an initial rate of 30.0 +/- 3.8%/min, leading to a cell swelling of 25.7 +/- 3.1% after 5 min (n = 7). After a period of recovery of 5 min in the absence of taurine or glucose, a 40 mosmol/kg hypotonic shock induced a cell swelling of 14.2 +/- 1.3 and 16.1 +/- 5.2%, respectively, followed by an almost complete volume regulatory decrease after 5 min. At that time, addition of taurine under continuous hypotonicity induced an increase in CV at an initial rate 2.57 +/- 0.17 times larger than that observed under the isotonic condition (P < 0.005), while addition of glucose induced an initial increase in CV identical to that observed under the isotonic condition. The increases in CV observed on addition of taurine were completely abolished in the absence of sodium under both isotonic and hypotonic conditions. The permeability to K+ was also estimated, in the absence of sodium, using the initial rate of increase in CV induced on isotonic replacement of 40 mM N-methyl-D-glucamine by K+.(ABSTRACT TRUNCATED AT 250 WORDS)
Thyroid hormones regulate development of energy metabolism enzymes in rat proximal convoluted tubule
Authors: Wijkhuisen A. , Djouadi F. , Vilar J. , Merlet-Benichou C. , Bastin J.
Published: December 2017

Ketone bodies represent preferred energy substrates in the adult rat proximal tubule. They are abundant in the plasma of suckling rats and might represent an important oxidative substrate for the immature proximal tubule. The postnatal development of two enzymes involved in ketone body oxidation pathway, 3-ketoacid-CoA transferase and acetoacetyl-CoA thiolase, and of citrate synthase and carnitine acetyltransferase was studied in microdissected rat proximal convoluted tubule (PCT) at 1, 8, 16, and 21 days after birth. The enzyme levels in PCT of juxtamedullary and subcapsular nephrons were compared at 8, 16, and 21 days. A role of thyroid hormones in regulating the development of these enzymes was investigated by studying 8- and 21-day-old pups made hypothyroid by propylthiouracyl (PTU) treatment, as well as 21-day hyperthyroid rats. PTU treatment had no effect on enzyme activities on day 8. In contrast, the activity of all mitochondrial enzymes, except acetoacetyl-CoA thiolase, was significantly decreased in 21-day-old hypothyroid pups. In hypothyroid animals, the normal development of 3-ketoacid-CoA transferase, citrate synthase, and carnitine acetyltransferase could be restored after treatment by triiodothyronine (T3). In addition, one single injection of T3 to 8-day-old control pups induced a precocious rise in the activity of 3-ketoacid-CoA transferase, citrate synthase, and carnitine acetyltransferase in juxtamedullary PCT and in the activity of citrate synthase and carnitine acetyltransferase in subcapsular PCT. Altogether, these results point out the importance of the postnatal physiological rise in T3 in triggering the development of some mitochondrial oxidative enzymes in the PCT.
Regulation of transcription by the rat EGF gene promoter in normal and ischemic murine kidney cells
Authors: Price P. M. , Megyesi J. , Saggi S. , Safirstein R. L.
Published: December 2017

Epidermal growth factor (EGF) is a small polypeptide belonging to a class of molecules that can mediate cell growth, differentiation, and acute phase responses. EGF mRNA is transcribed primarily in cells of the salivary gland and the kidney. We have found that the tissue and cellular specificities of EGF gene expression are controlled by a promoter region located upstream from the start of mRNA transcription. In a variety of experimentally induced forms of acute renal failure, the mRNA and protein levels for kidney EGF fall markedly and remain low for a prolonged period. This decrease was determined by nuclear runoff transcription to be a consequence of diminished transcription of the EGF gene rather than increased instability of the mRNA. Using transgenic mice, we found the effect of renal ischemia on EGF mRNA transcription to be a result of a disease-mediated interruption of the function of this upstream promoter region, presumably from alterations in the activity of one or more cellular trans-acting factors.
Convective uphill transport of NaCl from ascending thin limb of loop of Henle
Authors: Stephenson J. L. , Jen J. F. , Wang H. , Tewarson R. P.
Published: December 2017

In this paper we describe a mathematical model of the renal inner medulla based on a previously proposed model [A.S. Wexler, R.E. Kalaba, and D.J. Marsh. Am. J. Physiol. 260 (Renal Fluid Electrolyte Physiol. 29): F368–F383, 1991] in which in the inner medullary ascending thin limb of Henle's loop (ATL) and collecting duct (CD) exchange with a local capillary node with the reabsorbed water and solutes flowing radially toward a central vascular bundle. Our model differs in that ascending and descending vasa recta and surrounding interstitial space are replaced by a central core. Our analysis of the coupled ATL-CD system shows that it is theoretically capable of transporting NaCl out of the ATL into the central vascular space (approximated by the central core) against a concentration gradient, which in the absence of radial diffusion can be arbitrarily large. By numerical solution of the model with the radial diffusion coefficient (D(r)) for NaCl of 0, we find that the ATL can be more than 100 mosmol/l hypotonic with respect to the core. We also find that with restricted diffusion the osmolality of the CD at the papilla is significantly greater than that of the loop of Henle. As D(r) approaches the diffusion coefficient of NaCl in free solution, the osmolality of the loop increases and that of the CD decreases. Thus, overall, contrary to intuitive expectations, the radial separation and uphill transport of NaCl do not give any significant increase in loop concentration, which depends primarily on the quantity of urea reabsorbed from the CD.
Clearance of endogenous lithium in humans: altered dietary salt intake and comparison with exogenous lithium clearance
Authors: Folkerd E. , Singer D. R. , Cappuccio F. P. , Markandu N. D. , Sampson B. , MacGregor G. A.
Published: December 2017

We compared endogenous with exogenous lithium clearance (CLi) and studied the effects of dietary salt intake on endogenous CLi in healthy volunteers. Lithium was detectable within a narrow fourfold range in serum and in urine in all 25 subjects studied [serum (n = 25), mean 0.27 +/- 0.02 mumol/l, range 0.13-0.55 mumol/l; urine (n = 20), range 1.49–7.32, mean 4.09 +/- 0.36 mumol/24 h]. Mean clearance and fractional excretion of endogenous lithium were lower (15.2 +/- 2.0 ml/min and 16.4 +/- 2.1%, respectively) compared with results obtained using the exogenous CLi technique (25.5 +/- 1.7 ml/min and 27.9 +/- 2.1%; P < 0.01 and P < 0.05, respectively; n = 17). In a separate group of six normal subjects, absolute (8.7 +/- 2.9 vs. 20.7 +/- 3.8 ml/min) and fractional excretion of lithium (8.3 +/- 2.9 vs. 18.0 +/- 5.1%) were significantly lower on 5 days of low (31 +/- 10 mmol/day) vs. high sodium intake (357 +/- 78 mmol/day; P < 0.05). Use of endogenous CLi precludes the need for lithium tablets. This could be a particular advantage in population studies and permits serial measurement of CLi on different days. Our results show that it is important to take dietary sodium intake into account in studies of endogenous CLi. Lower values for endogenous compared with exogenous CLi could reflect differences in renal handling depending on the plasma lithium concentration. This clearly requires further study.
Chloride transport in the rat S1 proximal tubule
Authors: Wong K. R. , Berry C. A. , Cogan M. G.
Published: December 2017

In vivo microperfusion was used to elucidate the modes and regulation of the powerful chloride transport system resident in the rat early (S1) proximal convoluted tubule (PCT). From a complete, glomerular ultrafiltrate-like perfusate, omission of organic solutes reduced chloride absorption by 93 peq.mm-1.min-1 (302 +/- 10 to 209 +/- 24, P < 0.001). From a high-chloride perfusate (a relatively pure NaCl solution devoid of bicarbonate and organic solutes), luminal addition of the active transport inhibitor cyanide reduced chloride absorption by 153 peq.mm-1.min-1 (632 +/- 17 to 479 +/- 9, P < 0.001). Active transport was also estimated directly as 121 +/- 4 peq.mm-1.min-1 using a solution in which sodium isethionate isosmotically replaced bicarbonate and organic solutes, preventing development of a chloride gradient. Intravenous angiotensin II caused a stimulation of chloride absorption from a high-chloride perfusate by 55 peq.mm-1.min-1 (632 +/- 17 to 687 +/- 14, P < 0.05), which was partially cyanide-sensitive (510 +/- 6 peq.mm-1.min-1). In conclusion, the components of the normal S1 PCT chloride reabsorption (approximately 300 peq.mm-1.min-1) from the glomerular ultrafiltrate consist of the following: active transport (40–50%), which can be regulated by angiotensin II; sodium-coupled organic solute transport (30%); and passive, chloride concentration gradient-driven transport (20–25%).
Neutral phosphate administration generates and maintains renal metabolic alkalosis and hyperparathyroidism
Authors: Krapf R. , Glatz M. , Hulter H. N.
Published: December 2017

We examined the effects of chronic intravenous neutral phosphate administration on systemic acid-base equilibrium and parathyroid function in six normal, NaCl-replete male human subjects under metabolic balance conditions. The subjects received 4.35 mmol of neutral sodium phosphate.kg body wt-1.day-1 intravenously and continuously for 7 days and the same amount of sodium as NaCl during control and recovery. Blood pH increased from 7.388 to 7.411 (P < 0.001) and plasma bicarbonate from 23.5 to 26.0 mmol/l (P < 0.001). Urinary pH increased from 6.58 to 6.79 (P < 0.001). Net acid excretion increased from 59 to 100 mmol/24 h (P < 0.001). Plasma ionized calcium concentration decreased and plasma phosphate concentration increased transiently. Serum intact parathyroid hormone increased from 24 to 62 pg/ml (P < 0.001). Chronic phosphate administration also resulted in a significant increase in renal phosphate clearance (35 to 229 ml/min) and decrease in the fractional excretion of calcium (1.8 to 0.9%). Thus chronic intravenous phosphate administration generates and maintains renal metabolic alkalosis in salt-replete humans and induces hyperparathyroidism. The severity of metabolic alkalosis is mitigated by an apparent increase in effective endogenous acid production as evidenced by the significant increase in steady-state net acid excretion.
Disturbances in renal microcirculation induced by myoglobin and hemorrhagic hypotension in anesthetized rat
Authors: Vetterlein F. , Hoffmann F. , Pedina J. , Neckel M. , Schmidt G.
Published: December 2017

The question was studied of whether myoglobin (Mb), when released into the general circulation during hemorrhagic hypotension (HH), causes disturbances of renal blood flow. In anesthetized rats 250 mg/kg Mb was intravenously infused within 1 h; HH at 50 mmHg with subsequent retransfusion was induced for 30 min. By allowing two dyes to circulate for 1 and 3 min, respectively, and detecting their localization histologically after rapid freezing of the organ, intrarenal distribution of capillary blood flow was studied. In contrast to the results obtained with Mb or HH alone, when Mb was infused during HH, the development of large areas within cortex and medulla lacking any capillary perfusion was observed. In > 70% of the tissue, a distance > 60 microns to the next dye-labeled capillary was found (in controls 0%). At this time total renal flow had decreased from 5.3 to 0.20 ml/min (HH without Mb: 5.1 to 1.1 ml/min). It is concluded that the observed changes in renal blood flow contribute to the known direct nephrotoxic potential of Mb.