Structural-functional relationships along the distal nephron

1986 ◽  
Vol 250 (1) ◽  
pp. F1-F15 ◽  
Author(s):  
K. M. Madsen ◽  
C. C. Tisher
Keyword(s):  
Atpase Activity ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Distal Tubule ◽  
Structural Heterogeneity ◽  
Apical Plasma Membrane ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Principal Cells ◽  
Functional Relationships

The distal tubule, which includes the thick ascending limb (TAL), the macula densa, and the distal convoluted tubule (DCT), and the collecting duct are structurally heterogeneous, thus reflecting the functional heterogeneity that is also present. As the TAL ascends from medulla to cortex, the surface area of the apical plasma membrane increases while that of the basolateral membrane decreases. The structure of the DCT resembles that of the medullary TAL. An excellent correlation exists between structure, Na-K-ATPase activity, and NaCl reabsorptive capacity in the distal tubule. The collecting duct is subdivided into the initial collecting tubule (ICT), and cortical (CCD), outer medullary (OMCD), and inner medullary (IMCD) collecting ducts. Between the distal tubule and the collecting duct is a transition region termed the connecting segment or connecting tubule (CNT). Considerable structural heterogeneity exists along the collecting duct within the two major cell populations, the intercalated cells and the principal cells. In the CNT, the ICT, and the CCD, potassium loading and mineralocorticoids stimulate Na-K-ATPase activity and cause proliferation of the basolateral membrane of CNT cells and principal cells, thus identifying the cells responsible for mineralocorticoid-stimulated potassium secretion in these regions. Finally, at least two morphologically distinct populations of intercalated cells exist, types A and B. In the rat, type A predominates in the CNT and the OMCD and is believed to be responsible for H+ secretion, at least in the OMCD. Type B predominates in the CCD, where it may be involved in bicarbonate secretion.

Renal potassium transport: morphological and functional adaptations

1989 ◽  
Vol 257 (5) ◽  
pp. R989-R997 ◽  
Author(s):  
B. A. Stanton
Keyword(s):  
Collecting Duct ◽  
Cell Structure ◽  
Basolateral Membrane ◽  
Distal Tubule ◽  
Morphological Evidence ◽  
Intercalated Cells ◽  
Principal Cells ◽  
K Secretion ◽  
K Concentration ◽  
And Function

Maintenance of K+ homeostasis in mammals and amphibians depends primarily on the kidneys which excrete 95% of K+ ingested in the diet. The amount of K+ in the urine is determined by the rate of K+ secretion or absorption by the distal tubule and the collecting duct. When K+ intake is increased, K+ secretion rises. The mechanisms of K+ secretion by the distal tubule and collecting duct are so efficient that K+ intake can increase 20-fold with little or no increase in body K+ content or in plasma K+ concentration. Elevated K+ secretion by the distal tubule and collecting duct occurs in part because of an increase in the quantity of Na+-K+-adenosinetriphosphatase (Na+-K+-ATPase) and amplification of the basolateral membrane of principal cells. When dietary K+ intake is reduced, urinary K+ excretion falls, because K+ secretory mechanisms are suppressed and K+ absorptive mechanisms, residing in the distal tubule and collecting duct, are activated. Because a low-K+ diet is associated with hypertrophy of intercalated cells, it has been suggested that this cell type absorbs K+, possibly by an H+-K+-ATPase. In this review, I discuss the functional and morphological evidence that supports the view that principal cells secrete K+ and that intercalated cells absorb K+. In addition, some of the hormones and factors that are responsible for these changes in cell structure and function are discussed.

Immunocytochemical localization of Na+ channels in rat kidney medulla

1989 ◽  
Vol 256 (2) ◽  
pp. F366-F369 ◽  
Author(s):  
D. Brown ◽  
E. J. Sorscher ◽  
D. A. Ausiello ◽  
D. J. Benos
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Apical Plasma Membrane ◽  
Na Channel ◽  
Thick Ascending Limb ◽  
Na Channels ◽  
Kidney Medulla ◽  
Principal Cells

Amiloride-sensitive Na+ channels were localized in semithin frozen sections of rat renal medullary collecting ducts, using polyclonal antibodies directed against purified bovine kidney Na+ channel protein. The apical plasma membrane of collecting duct principal cells was heavily stained by indirect immunofluorescence, whereas intercalated cells were negative. Basolateral plasma membranes of both cell types were unstained, as were subapical vesicles in the cytoplasm of these cells. In the thick ascending limb of Henle, some scattered granular fluorescence was seen in the cytoplasm and close to the apical pole of epithelial cells, suggesting the presence of antigenic sites associated with some membrane domains in these cells. No staining was detected in thin limbs of Henle, or in proximal tubules in the outer medulla. These results show that amiloride-sensitive sodium channels are located predominantly on the apical plasma membrane of medullary collecting duct principal cells, the cells that are involved in Na+ homeostasis in this region of the kidney.

Cellular distribution of the potassium channel KCNQ1 in normal mouse kidney

2007 ◽  
Vol 292 (1) ◽  
pp. F456-F466 ◽  
Author(s):  
Wencui Zheng ◽  
Jill W. Verlander ◽  
I. Jeanette Lynch ◽  
Melanie Cash ◽  
Jiahong Shao ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Mouse Kidney ◽  
Cellular Heterogeneity ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Distal Nephron ◽  
Rt Pcr ◽  
Double Labeling ◽  
Cytoplasmic Staining ◽  
Principal Cells

Mechanisms of K+ secretion and absorption along the collecting duct are not understood fully. Because KCNQ1 participates in K+ secretion within the inner ear and stomach, distribution of KCNQ1 in mouse kidney was studied using Northern and Western analyses, RT-PCR of isolated tubules, and immunohistochemistry. Northern blots demonstrated KCNQ1 transcripts in whole kidney. RT-PCR showed KCNQ1 mRNA in isolated distal convoluted tubule (DCT), connecting segment (CNT), collecting ducts (CD), and glomeruli. Immunoblots of kidney and stomach revealed a ∼75-kDa protein, the expected mobility for KCNQ1. KCNQ1 was detected by immunohistochemistry throughout the distal nephron and CD. Thick ascending limbs exhibited weak basolateral immunolabel. In DCT and CNT cells, immunolabel was intense and basolateral, although KCNQ1 label was stronger in late than in early DCT. Initial collecting tubule and cortical CD KCNQ1 immunolabel was predominantly diffuse, but many cells exhibited discrete apical label. Double-labeling experiments demonstrated that principal cells, type B intercalated cells, and a few type A intercalated cells exhibited distinct apical KCNQ1 immunolabel. In inner medullary CD, principal cells exhibited distinct basolateral KCNQ1 immunolabel, whereas intercalated cells showed diffuse cytoplasmic staining. Thus KCNQ1 protein is widely distributed in mouse distal nephron and CD, with significant axial and cellular heterogeneity in location and intensity. These findings suggest that KCNQ1 has cell-specific roles in renal ion transport and may participate in K+ secretion and/or absorption along the thick ascending limb, DCT, connecting tubule, and CD.

Rat renal cortical OAT1 and OAT3 exhibit gender differences determined by both androgen stimulation and estrogen inhibition

2004 ◽  
Vol 287 (1) ◽  
pp. F124-F138 ◽  
Author(s):  
Marija Ljubojević ◽  
Carol M. Herak-Kramberger ◽  
Yohannes Hagos ◽  
Andrew Bahn ◽  
Hitoshi Endou ◽  
...  
Keyword(s):  
Gender Differences ◽  
Proximal Tubule ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Distal Tubule ◽  
Inhibitory Effect ◽  
Kidney Cortex ◽  
Thick Ascending Limb ◽  
Estradiol Treatment ◽  
Adult Rats

In rats, the secretion of p-aminohippurate (PAH) by the kidney is higher in males (M) than in females (F). The role of the major renal PAH transporters, OAT1 and OAT3, in the generation of these gender differences, as well as the responsible hormones and mechanisms, has not been clarified. Here we used various immunocytochemical methods to study effects of gender, gonadectomy, and treatment with sex hormones on localization and abundance of OAT1 and OAT3 along the rat nephron. Both transporters were localized to the basolateral membrane: OAT1 was strong in proximal tubule S2 and weak in the S3 segments, whereas OAT3 was stained in proximal tubule S1 and S2 segments, thick ascending limb, distal tubule, and in principal cells along the collecting duct. Gender differences in the expression of both transporters in adult rats (M > F) were observed only in the cortical tubules. OAT1 in the cortex was strongly reduced by castration in adult M, whereas the treatment of castrated M with testosterone, estradiol, or progesterone resulted in its complete restitution, further depression, or partial restitution, respectively. In adult F, ovariectomy weakly increased, whereas estradiol treatment of ovariectomized F strongly decreased, the expression of OAT1. The expression of OAT3 in the M and F cortex largely followed a similar pattern, except that ovariectomy and progesterone treatment showed no effect, whereas in other tissue zones gender differences were not observed. In prepubertal rats, the expression of OAT1 and OAT3 in the kidney cortex was low and showed no gender differences. Our data indicate that gender differences in the rat renal cortical OAT1 and OAT3 (M > F) appear after puberty and are determined by both a stimulatory effect of androgens (and progesterone in the case of OAT1) and an inhibitory effect of estrogens.

Functional activity of H-K-ATPase in individual cells of OMCD: localization and effect of K+ depletion

1996 ◽  
Vol 270 (1) ◽  
pp. F116-F122 ◽  
Author(s):  
M. Kuwahara ◽  
W. J. Fu ◽  
F. Marumo
Keyword(s):  
Functional Activity ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cell Types ◽  
Intercalated Cells ◽  
Principal Cells ◽  
Before And After ◽  
The Difference ◽  
K Depletion

Recent studies have indicated the presence of hydrogen-potassium-adenosinetriphosphatase (H-K-ATPase) in the collecting duct. We examined the localization of functional H-K-ATPase activity in individual cells of the outer and inner stripes of outer medullary collecting ducts (OMCDo and OMCDi). Tubules were isolated from control and K(+)-depleted rabbits and perfused in vitro. Intracellular pH (pHi) of principal cells, intercalated cells, and OMCDi cells was monitored by fluorescence ratio imaging using 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). An intracellular acid load was induced by NH3/NH4 prepulse in extracellular Na(+)-, K(+)-, and HCO3(-)-free condition, and then 5 mM K+ was added to the lumen or the bath in the presence of Ba2+. Functional activity of H-K-ATPase was estimated by the difference in the rates of pHi recovery before and after K+ addition. In the control condition, luminal addition of K+ significantly increased the pHi recovery rate by 1.6 +/- 0.4 and 1.9 +/- 0.4 x 10(-3) pH units/s in intercalated calls and OMCDi cells, respectively, but not in principal cells. This K(+)-dependent pHi recovery was inhibited by 63% in intercalated cells and 74% in OMCDi cells in the presence of luminal Sch-28080 (10 microM) but was not affected in the presence of luminal bafilomycin-A1 (10 nM). K+ depletion increased the K(+)-dependent pHi recovery to 2.3-fold in intercalated cells and 2.6-fold in OMCDi cells. By contrast, K(+)-dependent pHi recovery was not detected in the basolateral membrane of any cell types in either the control or the K(+)-depleted condition. These results provide functional evidence that H-K-ATPase is distributed in the luminal membrane of intercalated cells and OMCDi cells and that this ATPase is activated by K+ depletion, suggesting the contribution of intercalated cells and OMCDi cells to K+ conservation in rabbit OMCD.

P0045GENETIC ABLATION OF NFAT5 IN THE PRINCIPAL CELLS OF THE COLLECTING DUCT LEADS TO A WATER HANDLING DEFECT

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Federica Petrillo ◽  
Annika Fischer ◽  
Dmitry Chernyakov ◽  
Cristina Esteva Font ◽  
Søren B Poulsen ◽  
...  
Keyword(s):  
Mouse Model ◽  
Function Analysis ◽  
Collecting Duct ◽  
Distal Tubule ◽  
Dominant Negative ◽  
Urinary Concentration ◽  
Thick Ascending Limb ◽  
Water Restriction ◽  
Activated T Cells ◽  
Principal Cells

Abstract Background and Aims Nuclear Factor of Activated T-Cells 5 (NFAT5), also called TonEBP/OREBP, helps protecting kidney cells from the stress of extracellular hypertonic challenge. In response to hypertonicity, NFAT5 rapidly translocates into the nucleus and enhances the transcription of osmoprotective genes. Mice lacking NFAT5 have embryonic lethality, with those surviving having severe renal atrophy and hydronephrosis. Overexpression of a dominant-negative NFAT5 in all epithelial cells of the thick ascending limb, distal tubule and collecting duct (CD) leads to an impairment in the urine concentration, with reduced expression of AQP2 and urea transporters UT-A1 and UT-A2. Here, we generated a novel mouse model to assess the role of NFAT5 in the water transporting principal cells of the CD. Method The mouse model was generated by breeding floxed NFAT5 mice with mice specifically expressing Cre recombinase in AQP2 expressing cells (NFAT5f/f-AQP2cre+/-). AQP2cre+/- mice served as controls. For renal function analysis, mice were kept in metabolic cages for 5 days and water intake, urinary volume, osmolality, and serum and urine electrolytes were evaluated. Mice were euthanized and the kidneys were used for western blot analysis. Results NFAT5f/f-AQP2cre+/- mice show a significant increase of 24 hour urine output compared to the control (KO: 6.267 ± 0.9367 µl/h/g BW vs WT: 0.9593 ± 0.0796 µl/h/g BW), a decrease in the urine osmolality (KO: 505.6 ± 107.6 mOsm/Kg H2O vs WT: 6205 ± 752.7 mOsm/Kg H2O) and urea concentration. After 3h and half intraperitoneal injection of dDAVP (1ug/kg) the osmolality was still significantly low in NFAT5f/f-AQP2cre+/- mice (KO: 743.7 ± 81.77 vs WT: 3342 ± 80.14 mOsm/Kg H2O), as well as after 8h water restriction (KO: 732.8 ± 24.90 vs WT: 3784 ± 450.9 mOsm/Kg H2O). Immunoblotting demonstrated a significant decrease in AQP2, AQP3 and AQP4 abundance in cortical kidney fractions and inner medulla. There were no differences in vasopressin V2 receptor (V2R) and AVP-regulated transporters NKCC2, ROMK and alpha-ENaC. Conclusion We demonstrate that NFAT5 ablation from AQP2-expressing cells results in a urinary concentration defect. NFAT5f/f-AQP2cre+/- mice have a hypoosmotic polyuria, without major effects on the AVP-V2R axis, suggesting Nephrogenic Diabetes Insipidus.

Intracellular band 3 immunostaining in type A intercalated cells of rabbit kidney

1992 ◽  
Vol 262 (6) ◽  
pp. F1015-F1022
Author(s):  
K. M. Madsen ◽  
J. Kim ◽  
C. C. Tisher
Keyword(s):  
Plasma Membrane ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Type A ◽  
Band 3 ◽  
Rabbit Kidney ◽  
Apical Plasma Membrane ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Electron Microscopic

Intercalated cells (ICs) in the collecting duct and the connecting tubule (CNT) are involved in H+ secretion and HCO3- reabsorption. H+ secretion is mediated by an H(+)-adenosinetriphosphatase in the apical plasma membrane, whereas a band 3-like Cl(-)-HCO3- exchanger in the basolateral membrane is responsible for HCO3- reabsorption. Recent studies have reported that a band 3-like protein is also present in mitochondria in rabbit ICs. The purpose of this study was to establish the subcellular location of the band 3-like Cl(-)-HCO3- exchanger in rabbit ICs by electron microscopic immunocytochemistry using a monoclonal antibody, IVF12, against erythrocyte band 3 protein. Rabbit kidneys were preserved by in vivo perfusion with a paraformaldehyde-lysine-periodate solution and processed for immunocytochemistry using a horseradish peroxidase preembedding technique. Band 3 immunostaining was observed on the basolateral plasma membrane of ICs in the outer medullary collecting duct and type A cells in the cortical collecting duct (CCD) and CNT. In addition, distinct staining for band 3 was present in numerous small vesicles and in multivesicular bodies in type A ICs in the CCD and CNT. However, there was no evidence of band 3 immunostaining of mitochondria or of the apical plasma membrane in any cells of the collecting duct. These observations suggest that basolateral Cl(-)-HCO3- exchangers in type A ICs in the rabbit kidney are stored in intracellular vesicles and possibly degraded in the vascular-lysosomal system when these cells are in a resting state. The previously reported band 3 immunolabeling of mitochondria could not be confirmed.

scholarly journals Basolateral targeting and microtubule-dependent transcytosis of the aquaporin-2 water channel

2013 ◽  
Vol 304 (1) ◽  
pp. C38-C48 ◽  
Author(s):  
Naofumi Yui ◽  
Hua A. J. Lu ◽  
Ying Chen ◽  
Naohiro Nomura ◽  
Richard Bouley ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cold Shock ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Water Channel ◽  
Apical Plasma Membrane ◽  
Indirect Pathway ◽  
Basolateral Membranes ◽  
Principal Cells ◽  
Aquaporin 2

The aquaporin-2 (AQP2) water channel relocates mainly to the apical plasma membrane of collecting duct principal cells after vasopressin (VP) stimulation. AQP2 transport to this membrane domain is assumed to be a direct route involving recycling of intracellular vesicles. However, basolateral plasma membrane expression of AQP2 is observed in vivo in principal cells. Here, we asked whether there is a transcytotic pathway of AQP2 trafficking between apical and basolateral membranes. We used MDCK cells in which AQP2 normally accumulates apically after VP exposure. In contrast, both site-specific biotinylation and immunofluorescence showed that AQP2 is strongly accumulated in the basolateral membrane, along with the endocytic protein clathrin, after a brief cold shock (4°C). This suggests that AQP2 may be constitutively targeted to basolateral membranes and then retrieved by clathrin-mediated endocytosis at physiological temperatures. Rab11 does not accumulate in basolateral membranes after cold shock, suggesting that the AQP2 in this location is not associated with Rab11-positive vesicles. After rewarming (37°C), basolateral AQP2 staining is diminished and it subsequently accumulates at the apical membrane in the presence of VP/forskolin, suggesting that transcytosis can be followed by apical insertion of AQP2. This process is inhibited by treatment with colchicine. Our data suggest that the cold shock procedure reveals the presence of microtubule-dependent AQP2 transcytosis, which represents an indirect pathway of apical AQP2 delivery in these cells. Furthermore, our data indicate that protein polarity data obtained from biotinylation assays, which require cells to be cooled to 4°C during the labeling procedure, should be interpreted with caution.

scholarly journals Immunolocalization of the secretory isoform of Na-K-Cl cotransporter in rat renal intercalated cells.

1996 ◽  
Vol 7 (12) ◽  
pp. 2533-2542 ◽  
Author(s):  
S M Ginns ◽  
M A Knepper ◽  
C A Ecelbarger ◽  
J Terris ◽  
X He ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Apical Plasma Membrane ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Specific Affinity ◽  
Basolateral Plasma Membrane ◽  
Medullary Collecting Duct ◽  
Antipeptide Antibody

Two bumetanide-sensitive ion cotransporters that carry Na+, K+, and Cl- in a coupled fashion have been identified. One type, the "absorptive" isoform, carries these ions across the apical plasma membrane of the thick ascending limb of Henle's loop. Another isoform, the "secretory" cotransporter, has been identified in a number of epithelial tissues by physiological means, but its sites of expression in the kidney have not been fully characterized. Complementary DNA believed to code for the secretory isoform (called "BSC2" or "NKCC1") have recently been cloned. This study used a specific affinity-purified antipeptide antibody to this protein for immunolocalization in the rat kidney. Immunoblot studies using this antibody show abundant immunoreactivity against bands of 140-190 and 120 kd in the parotid gland, colon, and stomach, sites where the secretory form of the cotransporter has been identified by physiological techniques. This distribution supports the hypothesis that this isoform represents the secretory form of the cotransporter. Studies in the kidney revealed that the same bands are associated with membrane fractions chiefly in the outer medulla. Immunolocalizations show that immunoreactivity is selectively and intensely localized to the basolateral plasma membrane of a subfraction of outer medullary collecting duct cells. An independently produced monoclonal antibody (T4) specific for Na-K-Cl cotransporter displays the same localization. Dual localizations of cotransporter antibody with respect to antibody specific for principal cells (aquaporin-2) and intercalated cells (band 3 and H(+)-ATPase) show that cotransporter immunoreactivity is localized to alpha-intercalated cells of the outer medullary collecting duct in the rat. This distinctive localization suggests that the secretory form of the cotransporter may play a role in renal NH4+ and/or acid secretion by this cell type.

scholarly journals Adenosine inhibits the basolateral Cl− ClC-K2/b channel in collecting duct intercalated cells

2020 ◽  
Vol 318 (4) ◽  
pp. F870-F877
Author(s):  
Oleg Zaika ◽  
Viktor N. Tomilin ◽  
Oleh Pochynyuk
Keyword(s):  
Salt Intake ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Inhibitory Effect ◽  
Intercalated Cells ◽  
Dietary Salt ◽  
Open Probability ◽  
Salt Loading ◽  
Principal Cells ◽  
High Salt Intake

Adenosine plays an important role in various aspects of kidney physiology, but the specific targets and mechanisms of actions are not completely understood. The collecting duct has the highest expression of adenosine receptors, particularly adenosine A1 receptors (A1Rs). Interstitial adenosine levels are greatly increased up to a micromolar range in response to dietary salt loading. We have previously shown that the basolateral membrane of principal cells has primarily K+ conductance mediated by Kir4.1/5.1 channels to mediate K+ recycling and to set up a favorable driving force for Na+/K+ exchange ( 47 ). Intercalated cells express the Cl− ClC-K2/b channel mediating transcellular Cl− reabsorption. Using patch-clamp electrophysiology in freshly isolated mouse collecting ducts, we found that acute application of adenosine reversely inhibits ClC-K2/b open probability from 0.31 ± 0.04 to 0.17 ± 0.06 and to 0.10 ± 0.05 for 1 and 10 µM, respectively. In contrast, adenosine (10 µM) had no measureable effect on Kir4.1/5.1 channel activity in principal cells. The inhibitory effect of adenosine on ClC-K2/b was abolished in the presence of the A1R blocker 8-cyclopentyl-1,3-dipropylxanthine (10 µM). Consistently, application of the A1R agonist N6-cyclohexyladenosine (1 µM) recapitulated the inhibitory action of adenosine on ClC-K2/b open probability. The effects of adenosine signaling in the collecting duct were independent from its purinergic counterpartner, ATP, having no measurable actions on ClC-K2/b and Kir4.1/5.1. Overall, we demonstrated that adenosine selectively inhibits ClC-K2/b activity in intercalated cells by targeting A1Rs. We propose that inhibition of transcellular Cl− reabsorption in the collecting duct by adenosine would aid in augmenting NaCl excretion during high salt intake.

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