Potassium deprivation upregulates expression of renal basolateral Na+-HCO3 −cotransporter (NBC-1)

2000 ◽  
Vol 279 (3) ◽  
pp. F532-F543 ◽  
Author(s):  
Hassane Amlal ◽  
Khalid Habo ◽  
Manoocher Soleimani
Keyword(s):  
Proximal Tubule ◽  
Collecting Duct ◽  
Thick Ascending Limb ◽  
Mrna Levels ◽  
Time Intervals ◽  
Outer Medulla ◽  
Medullary Thick Ascending Limb ◽  
Medullary Collecting Duct ◽  
Potassium Deprivation ◽  
Expression And Activity

The purpose of the present experiments was to examine the effect of potassium deprivation on the expression of the renal basolateral Na+-HCO3 − cotransporter (NBC-1). Rats were placed on a K+-free diet for various time intervals and examined. NBC-1 mRNA levels increased by about threefold in the cortex ( P < 0.04) at 72 h of K+ deprivation and remained elevated at 21 days. NBC activity increased by ∼110% in proximal tubule suspensions, with the activity increasing from 0.091 in control to 0.205 pH/min in the K+-deprived group ( P < 0.005). The inner stripe of outer medulla and cells of medullary thick ascending limb of Henle (mTAL) showed induction of NBC-1 mRNA and activity in K+-deprived rats, with the activity in mTAL increasing from 0.010 in control to 0.133 pH/min in the K+-deprived group ( P < 0.004). K+ deprivation also increased NBC-1 mRNA levels in the renal papilla ( P < 0.02). We conclude that 1) K+ deprivation increases NBC-1 expression and activity in proximal tubule and 2) K+deprivation causes induction of NBC-1 expression and activity in mTAL tubule and inner medulla. We propose that NBC-1 likely mediates enhanced HCO3 − reabsorption in proximal tubule, mTAL, and inner medullary collecting duct in K+ deprivation and contributes to the maintenance of metabolic alkalosis in this condition.

scholarly journals Modulation of outer medullary NaCl transport and oxygenation by nitric oxide and superoxide

2011 ◽  
Vol 301 (5) ◽  
pp. F979-F996 ◽  
Author(s):  
Aurélie Edwards ◽  
Anita T. Layton
Keyword(s):  
Nitric Oxide ◽  
Active Transport ◽  
Collecting Duct ◽  
Thick Ascending Limb ◽  
Outer Medulla ◽  
Nacl Transport ◽  
Complex Interactions ◽  
Medullary Thick Ascending Limb ◽  
The Impact ◽  
Stimulation Of

We expanded our region-based model of water and solute exchanges in the rat outer medulla to incorporate the transport of nitric oxide (NO) and superoxide (O2−) and to examine the impact of NO-O2− interactions on medullary thick ascending limb (mTAL) NaCl reabsorption and oxygen (O2) consumption, under both physiological and pathological conditions. Our results suggest that NaCl transport and the concentrating capacity of the outer medulla are substantially modulated by basal levels of NO and O2−. Moreover, the effect of each solute on NaCl reabsorption cannot be considered in isolation, given the feedback loops resulting from three-way interactions between O2, NO, and O2−. Notwithstanding vasoactive effects, our model predicts that in the absence of O2−-mediated stimulation of NaCl active transport, the outer medullary concentrating capacity (evaluated as the collecting duct fluid osmolality at the outer-inner medullary junction) would be ∼40% lower. Conversely, without NO-induced inhibition of NaCl active transport, the outer medullary concentrating capacity would increase by ∼70%, but only if that anaerobic metabolism can provide up to half the maximal energy requirements of the outer medulla. The model suggests that in addition to scavenging NO, O2− modulates NO levels indirectly via its stimulation of mTAL metabolism, leading to reduction of O2 as a substrate for NO. When O2− levels are raised 10-fold, as in hypertensive animals, mTAL NaCl reabsorption is significantly enhanced, even as the inefficient use of O2 exacerbates hypoxia in the outer medulla. Conversely, an increase in tubular and vascular flows is predicted to substantially reduce mTAL NaCl reabsorption. In conclusion, our model suggests that the complex interactions between NO, O2−, and O2 significantly impact the O2 balance and NaCl reabsorption in the outer medulla.

scholarly journals Inhibition of calcium signaling in descending vasa recta endothelia by ANG II

2000 ◽  
Vol 278 (4) ◽  
pp. H1248-H1255 ◽  
Author(s):  
Thomas L. Pallone ◽  
Erik P. Silldorff ◽  
Zhong Zhang
Keyword(s):  
Collecting Duct ◽  
No Production ◽  
Vascular Bundles ◽  
Thick Ascending Limb ◽  
Ang Ii ◽  
Vasomotor Tone ◽  
Medullary Thick Ascending Limb ◽  
Vasa Recta ◽  
Medullary Collecting Duct ◽  
Peak Phase

The intracellular calcium ([Ca2+]i) response of outer medullary descending vasa recta (OMDVR) endothelia to ANG II was examined in fura 2-loaded vessels. Abluminal ANG II (10− 8 M) caused [Ca2+]i to fall in proportion to the resting [Ca2+]i ( r =0.82) of the endothelium. ANG II (10− 8 M) also inhibited both phases of the [Ca2+]i response generated by bradykinin (BK, 10− 7 M), 835 ± 201 versus 159 ± 30 nM (peak phase) and 169 ± 26 versus 103 ± 14 nM (plateau phase) (means ± SE). Luminal ANG II reduced BK (10− 7 M)-stimulated plateau [Ca2+]i from 180 ± 40 to 134 ± 22 nM without causing vasoconstriction. Abluminal ANG II added to the bath after luminal application further reduced [Ca2+]i to 113 ± 9 nM and constricted the vessels. After thapsigargin (TG) pretreatment, ANG II (10− 8 M) caused [Ca2+]i to fall from 352 ± 149 to 105 ± 37 nM. This effect occurred at a threshold ANG II concentration of 10− 10 M and was maximal at 10− 8 M. ANG II inhibited both the rate of Ca2+ entry into [Ca2+]i-depleted endothelia and the rate of Mn2+ entry into [Ca2+]i-replete endothelia. In contrast, ANG II raised [Ca2+]i in the medullary thick ascending limb and outer medullary collecting duct, increasing [Ca2+]i from baselines of 99 ± 33 and 53 ± 11 to peaks of 200 ± 47 and 65 ± 11 nM, respectively. We conclude that OMDVR endothelia are unlikely to be the source of ANG II-stimulated NO production in the medulla but that interbundle nephrons might release Ca2+-dependent vasodilators to modulate vasomotor tone in vascular bundles.

Regional and segmental localization of AE2 anion exchanger mRNA and protein in rat kidney

1995 ◽  
Vol 269 (4) ◽  
pp. F461-F468 ◽  
Author(s):  
F. C. Brosius ◽  
K. Nguyen ◽  
A. K. Stuart-Tilley ◽  
C. Haller ◽  
J. P. Briggs ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Chloride Concentration ◽  
Transepithelial Transport ◽  
Thick Ascending Limb ◽  
Cortical Collecting Duct ◽  
Base Exchange ◽  
Medullary Thick Ascending Limb ◽  
Nephron Segment ◽  
Medullary Collecting Duct

Chloride/base exchange activity has been detected in every mammalian nephron segment in which it has been sought. However, in contrast to the Cl-/HCO3- exchanger AE1 in type A intercalated cells, localization of AE2 within the kidney has not been reported. We therefore studied AE2 expression in rat kidney. AE2 mRNA was present in cortex, outer medulla, and inner medulla. Semiquantitative polymerase chain reaction of cDNA from microdissected tubules revealed AE2 cDNA levels as follows [copies of cDNA derived per mm tubule (+/- SE)]: proximal convoluted tubule, 688 +/- 161; proximal straight tubule, 652 +/- 189; medullary thick ascending limb, 1,378 +/- 226; cortical thick ascending limb, 741 +/- 24; cortical collecting duct, 909 +/- 71; and outer medullary collecting duct, 579 +/- 132. AE2 cDNA was also amplified in thin limbs and in inner medullary collecting duct. AE2 polypeptide was detected in all kidney regions. AE2 mRNA and protein were also detected in several renal cell lines. The data are compatible with the postulated roles of AE2 in maintenance of intracellular pH and chloride concentration and with its possible participation in transepithelial transport.

Immunolocalization of electroneutral Na-HCO3 − cotransporter in rat kidney

2000 ◽  
Vol 279 (5) ◽  
pp. F901-F909 ◽  
Author(s):  
Henrik Vorum ◽  
Tae-Hwan Kwon ◽  
Christiaan Fulton ◽  
Brian Simonsen ◽  
Inyeong Choi ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Electron Microscopic Level ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Electron Microscopic ◽  
Outer Medulla ◽  
Outer Stripe ◽  
Medullary Collecting Duct

An electroneutral Na-HCO3 − cotransporter (NBCN1) was recently cloned, and Northern blot analyses indicated its expression in rat kidney. In this study, we determined the cellular and subcellular localization of NBCN1 in the rat kidney at the light and electron microscopic level. A peptide-derived antibody was raised against the COOH-terminal amino acids of NBCN1. The affinity-purified antibody specifically recognized one band, ∼180 kDa, in rat kidney membranes. Peptide- N-glycosidase F deglycosylation reduced the band to ∼140 kDa. Immunoblotting of membrane fractions from different kidney regions demonstrated strong signals in the inner stripe of the outer medulla (ISOM), weaker signals in the outer stripe of the outer medulla and inner medulla, and no labeling in cortex. Immunocytochemistry demonstrated that NBCN1 immunolabeling was exclusively observed in the basolateral domains of thick ascending limb (TAL) cells in the outer medulla (strongest in ISOM) but not in the cortex. In addition, collecting duct intercalated cells in the ISOM and in the inner medulla also exhibited NBCN1 immunolabeling. Immunoelectron microscopy demonstrated that NBCN1 labeling was confined to the basolateral plasma membranes of TAL and collecting duct type A intercalated cells. Immunolabeling controls were negative. By using 2,7-bis-carboxyethyl-5,6-caboxyfluorescein, intracellular pH transients were measured in kidney slices from ISOM and from mid-inner medulla. The results revealed DIDS-sensitive, Na- and HCO3 −-dependent net acid extrusion only in the ISOM but not in mid-inner medulla, which is consistent with the immunolocalization of NBCN1. The localization of NBCN1 in medullary TAL cells and medullary collecting duct intercalated cells suggests that NBCN1 may be important for electroneutral basolateral HCO3 − transport in these cells.

NBCn1 is a basolateral \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{Na}^{+}\mathrm{-HCO}_{3}^{-}\) \end{document} cotransporter in rat kidney inner medullary collecting ducts

2004 ◽  
Vol 286 (5) ◽  
pp. F903-F912 ◽  
Author(s):  
Jeppe Praetorius ◽  
Young-Hee Kim ◽  
Elena V. Bouzinova ◽  
Sebastian Frische ◽  
Aleksandra Rojek ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Basolateral Membrane ◽  
Primary Cultures ◽  
Thick Ascending Limb ◽  
Medullary Thick Ascending Limb ◽  
Collecting Ducts ◽  
Basolateral Plasma Membrane ◽  
Medullary Collecting Duct ◽  
Plasma Membrane Domain

Primary cultures of rat inner medullary collecting duct (IMCD) cells Na+ dependently import [Formula: see text] across the basolateral membrane through an undefined transport protein. We used RT-PCR, immunoblotting, and immunohistochemistry to identify candidate proteins for this basolateral [Formula: see text] cotransport. The mRNA encoding the electroneutral [Formula: see text] cotransporter NBCn1 was detected as the only [Formula: see text] cotransporter in the rat inner medulla (IM) among the five characterized Na+-dependent [Formula: see text] transporters. The mRNA of a yet uncharacterized transporter-like protein, BTR1, was also present in the IM, but its expression in microdissected tubules seemed restricted to the thin limbs of Henle's loop. Immunoblotting confirmed the presence of NBCn1 as an ∼180-kDa protein of the rat IM. Anti-NBCn1 immunolabeling was confined to the basolateral plasma membrane domain of IMCD cells in the papillary two-thirds of the IM. Consistent with the presence of NBCn1, IMCD cells possessed stilbene-insensitive, Na+- and [Formula: see text]-dependent pH recovery after acidification, as assessed by fluorescence microscopy using a pH-sensitive intracellular dye. In furosemide-induced alkalotic rats, NBCn1 protein abundance was decreased in both the IM and inner stripe of outer medulla (ISOM) as determined by immunoblotting and immunohistochemistry. In contrast, NBCn1 abundance in the IM and ISOM was unchanged in NaHCO3-loaded animals, and the NBCn1 abundance increased only in the ISOM after NH4Cl loading. In conclusion, NBCn1 is a basolateral [Formula: see text] cotransporter of IMCD cells and is differentially regulated in IMCD and medullary thick ascending limb.

AE2 isoforms in rat kidney: immunohistochemical localization and regulation in response to chronic NH4Cl loading

2004 ◽  
Vol 286 (6) ◽  
pp. F1163-F1170 ◽  
Author(s):  
Sebastian Frische ◽  
Alexander S. Zolotarev ◽  
Young-Hee Kim ◽  
Jeppe Praetorius ◽  
Seth Alper ◽  
...  
Keyword(s):  
Splice Variants ◽  
Polyclonal Antibodies ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Immunohistochemical Localization ◽  
Thick Ascending Limb ◽  
Relative Distribution ◽  
Medullary Thick Ascending Limb ◽  
Medullary Collecting Duct ◽  
Antibody Specificities

Three splice variants of anion exchanger (AE)2 (AE2a, b, and c) have been described in the rat, but their relative distribution in rat kidney is not known. The purpose of this study was to describe the segmental and cellular distribution of the AE2 isoforms in the rat kidney and to evaluate whether the expression levels of these AE2 isoforms are regulated independently in response to chronic NH4Cl loading. Two polyclonal antibodies were generated, respectively, recognizing a NH2-terminal peptide unique to AE2a and an amino acid sequence common to AE2a and AE2b. Antibody specificities were tested using cells transfected separately with the AE2a, AE2b, and AE2c isoforms. Immunohistochemistry on sections of paraffin-embedded rat kidneys showed a distribution of AE2a/AE2b labeling in the kidney similar to the distribution of AE2 in the rat kidney reported previously. AE2 is highly expressed in the medullary thick ascending limb, cortical thick ascending limb (cTAL), and macula densa. The pattern of AE2a-specific labeling differed from the pattern of AE2a/AE2b labeling in that relatively more of the total immunolabel was observed in the terminal inner medullary collecting duct. NH4Cl loading (0.033 mmol NH4Cl/g body wt for 7 days) did not change the labeling of AE2 isoforms in the medulla, whereas the labeling in the cortex was intensified and included more distal parts of the cTAL. Immunoblotting confirmed upregulation of AE2a/b expression in the cortex. These results indicate that AE2a and AE2b are differentially expressed and regulated in the rat kidney. The regulation following NH4Cl loading of AE2b in the cTAL suggests a role for AE2 in transepithelial bicarbonate reabsorption in this segment.

Distribution of mineralocorticoid and glucocorticoid receptor mRNA along the nephron

1993 ◽  
Vol 264 (5) ◽  
pp. F781-F791 ◽  
Author(s):  
K. M. Todd-Turla ◽  
J. Schnermann ◽  
G. Fejes-Toth ◽  
A. Naray-Fejes-Toth ◽  
A. Smart ◽  
...  
Keyword(s):  
Glucocorticoid Receptor ◽  
Proximal Tubule ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Rna Extraction ◽  
Thick Ascending Limb ◽  
Cortical Collecting Duct ◽  
Nephron Segment ◽  
Medullary Collecting Duct ◽  
Glucocorticoid Receptor Mrna

In the present study, a competitive polymerase chain reaction (PCR) technique was used to quantitate the relative levels of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) mRNA in microdissected nephron segments from the rat kidney and of MR mRNA from isolated principal and intercalated collecting duct cells from rabbit. RNA was isolated from cells and isolated tubules, cDNA was synthesized, and receptor cDNA was coamplified by PCR with a competitive control template. beta-Actin PCR products were also obtained from each nephron segment studied, to assess variations in RNA extraction and cDNA synthesis. MR mRNA, as determined by this competitive PCR technique, was 10-fold more abundant in cortical collecting duct (CCD), outer medullary collecting duct, and inner medullary collecting duct segments than in the proximal tubule and thick ascending limb segments (P < 0.05). Both principal and beta-intercalated cells of the CCD contained detectable levels of MR mRNA, although the levels in the principal cells were threefold higher (P < 0.01). GR mRNA was twofold more abundant in glomeruli, proximal tubule, and thick ascending limb segments than in the collecting duct segments (P < 0.05). In general, the distribution pattern of MR and GR mRNA is consistent with the distribution of adrenal corticosteroid function along the nephron.

Acid phosphatase activity in the mammalian nephron

1984 ◽  
Vol 247 (2) ◽  
pp. F252-F259 ◽  
Author(s):  
C. J. Olbricht ◽  
L. C. Garg ◽  
J. K. Cannon ◽  
C. C. Tisher
Keyword(s):  
Acid Phosphatase ◽  
Proximal Tubule ◽  
Collecting Duct ◽  
Rabbit Kidney ◽  
Male Rat ◽  
Thick Ascending Limb ◽  
Cortical Collecting Duct ◽  
Distal Nephron ◽  
S3 Segment ◽  
Medullary Collecting Duct

Lysosomal acid phosphatase (AcPase) activity was measured in individual segments of rat and rabbit nephrons employing 4-methylumbelliferyl phosphate as the substrate. Generation of reaction product was linear with incubation time up to 127 min and with tubule length. Activity was much higher in glomeruli and proximal tubules of rat than rabbit kidney. In both rat and rabbit there were higher activities in juxtamedullary than in superficial glomeruli. In rats, AcPase activity decreased from S1 to S3 segments, which parallels the known decrease in the number of lysosomes. Surprisingly, in rabbits of both sexes AcPase activity in the cortical collecting duct (CCD), which contains a limited number of lysosomes, was comparable to levels measured in the S1 and S2 segments of the proximal tubule. Similarly, in the male rat values for AcPase activity in the cortical thick ascending limb, distal convoluted tubule, CCD, and medullary collecting duct paralleled those in the S3 segment. These findings suggest that a considerable amount of AcPase in the distal nephron is either extralysosomal or that the amount of lysosomal AcPase activity per unit volume is greater in distal nephron segments compared with the proximal tubule. Different K'm values for AcPase in S1 segments and CCD were found in the rabbit, suggesting the presence of different isoenzymes.

Localization of bumetanide- and thiazide-sensitive Na-K-Cl cotransporters along the rat nephron

1996 ◽  
Vol 271 (4) ◽  
pp. F931-F939 ◽  
Author(s):  
T. Yang ◽  
Y. G. Huang ◽  
I. Singh ◽  
J. Schnermann ◽  
J. P. Briggs
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Direct Sequencing ◽  
Salt Transport ◽  
Thick Ascending Limb ◽  
Specific Primers ◽  
Nephron Segments ◽  
Medullary Thick Ascending Limb ◽  
Medullary Collecting Duct ◽  
Polymerase Chain

The present study was undertaken to investigate the mRNA localization of the two major kidney-specific Na-K-Cl transport proteins, the bumetanide-sensitive cotransporter (NKCC2 in rabbit and BSC1 in rat) and the thiazide-sensitive cotransporter (TSC). NKCC2 from rabbit and mouse has been shown to exist in three isoforms (designated A, B, and F) that differ only in a 96-bp region. The divergent region of each of the three NKCC2 isoforms was cloned from rat kidney by a polymerase chain reaction (PCR)-based strategy, and isoform-specific primers were chosen. RNA and cDNA were prepared from renal cortex and medulla and from microdissected nephron segments. Using reverse transcription (RT)-PCR, the B isoform was detected only in cortex and the F isoform only in medulla, whereas the A from was found in both. In dissected nephron segments, the B form was found exclusively in cortical thick ascending limb (CTAL) and macula densa-containing segment (MDCS), the F form only in medullary thick ascending limb (MTAL) and outer medullary collecting duct, and the A form in CTAL, MDCS, and MTAL. An additional isoform including both A and F exons was identified by direct sequencing of a 592-bp product from medulla. The AF product was found only in the medulla and was localized exclusively in MTAL. TSC mRNA was detected exclusively in the distal convoluted tubule. Differential nephron localization of NKCC2 isoforms suggests that Na-K-Cl cotransporters may differ in their transport characteristics to explain regulation of salt transport along the nephron.

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