Differentiation of intercalated cells in developing rat kidney: an immunohistochemical study

1994 ◽  
Vol 266 (6) ◽  
pp. F977-F990 ◽  
Author(s):  
J. Kim ◽  
C. C. Tisher ◽  
K. M. Madsen
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Type A ◽  
Cell Types ◽  
Renal Development ◽  
Intercalated Cells ◽  
Type B ◽  
Fetal Kidney ◽  
Distinct Cell ◽  
Developing Rat

Intercalated cells are present in both the collecting duct, which is derived from the ureteric bud, and the connecting tubule (CNT), which is part of the nephron and thus is developed from the metanephric blastema. However, the embryologic origin of the intercalated cells has not been established. Two populations of intercalated cells, type A and type B, exist in the CNT and the cortical collecting duct (CCD). It is uncertain, however, whether these cells represent truly distinct cell types or whether one is derived from the other. In this study we have used specific antibodies to carbonic anhydrase II (CA II), H(+)-adenosinetriphosphatase (H(+)-ATPase), and band 3 protein to identify subpopulations of intercalated cells, to determine the site and time of their appearance, and to follow their differentiation in the developing rat kidney. Prenatal kidneys from 16-, 17-, 18-, and 20-day-old fetuses, and postnatal kidneys from 0-, 3-, 7-, 14-, and 21-day-old pups were preserved for immunohistochemical studies. Immunostaining for CA II and H(+)-ATPase appeared simultaneously in a subpopulation of cells in the CNT and the medullary collecting duct (MCD) of the 18-day-old fetus, suggesting that intercalated cells differentiate from separate foci, one in the nephron and one in the collecting duct. Cells with apical and cells with basolateral labeling for H(+)-ATPase appeared in the CNT and MCD at 18 days of gestation, indicating that type A and type B cells differentiate simultaneously during renal development. Band 3 immunostaining was very weak in the fetal kidney, but a striking increase in labeling was observed in the 3-day-old kidney, suggesting that there is an activation of acid-secreting cells shortly after birth. In the fetal kidney, immunostaining for CA II and H(+)-ATPase was observed in cells throughout the MCD and on the papillary surface. After birth, immunostaining gradually disappeared from both the papillary surface and the terminal inner MCD, and cells with basolateral labeling for H(+)-ATPase gradually disappeared from the outer MCD. The results of this study suggest that type A and type B intercalated cells represent distinct cell types that derive from undifferentiated cells at two separate foci, one in the nephron and one in the collecting duct. Our results also suggest that entire populations of intercalated cells are eliminated from the collecting duct during normal renal development.

scholarly journals Ultrastructural localization of carbonic anhydrase II in subpopulations of intercalated cells of the rat kidney.

1990 ◽  
Vol 1 (3) ◽  
pp. 245-256 ◽  
Author(s):  
J Kim ◽  
C C Tisher ◽  
P J Linser ◽  
K M Madsen
Keyword(s):  
Electron Microscopy ◽  
B Cells ◽  
Carbonic Anhydrase ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Light And Electron Microscopy ◽  
Type A ◽  
Carbonic Anhydrase Ii ◽  
Intercalated Cells ◽  
Type B

At least two configurations of intercalated cells, type A and type B, are present in the cortical collecting duct. Intercalated cells are rich in carbonic anhydrase. However, it is not known whether there are differences in the level and subcellular distribution of this enzyme between type A and type B intercalated cells. The purpose of this study was to determine the relative content and intracellular distribution of carbonic anhydrase II in the various subpopulations of intercalated cells in the rat collecting duct. A rabbit polyclonal antibody directed against mouse erythrocyte carbonic anhydrase II was employed to localize carbonic anhydrase, II by light and electron microscopy by an indirect immunoperoxidase method. A Western immunoblot analysis of homogenates of rat kidney cortex and medulla with the carbonic anhydrase II antibody revealed a single polypeptide band at 29 kDa corresponding to the molecular size of carbonic anhydrase II. By both light and electron microscopy, carbonic anhydrase II immunoreactivity was present in all intercalated cells but the intensity of staining was much greater in type A than in type B cells. In addition, immunostaining in type A cells was especially pronounced in the apical cytoplasm and apical microprojections whereas in type B cells, immunostaining was more diffuse throughout the cytoplasm. A third configuration of intercalated cell with diffuse immunostaining for carbonic anhydrase II was occasionally observed in the connecting segment. Very weak immunostaining was present in principal cells, whereas connecting tubule cells and inner medullary collecting duct cells were negative for carbonic anhydrase II.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of apoptotic and nonapoptotic cell death in removal of intercalated cells from developing rat kidney

1996 ◽  
Vol 270 (4) ◽  
pp. F575-F592 ◽  
Author(s):  
J. Kim ◽  
J. H. Cha ◽  
C. C. Tisher ◽  
K. M. Madsen
Keyword(s):  
Electron Microscopy ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Surface Epithelium ◽  
Intercalated Cells ◽  
Band 3 Protein ◽  
Apoptotic Bodies ◽  
Type B ◽  
Medullary Collecting Duct ◽  
Developing Rat

In the developing rat kidney, both type A and type B intercalated cells are present throughout the medullary collecting duct (MCD), as well as the papillary surface epithelium. After birth, intercalated cells gradually disappear from the papillary surface epithelium and the terminal MCD, and type B cells disappear from the entire MCD. The purpose of this study was to establish the mechanism(s) by which intercalated cells are deleted from the MCD during development. Kidneys from 14-, 16-, 18-, and 20-day-old fetuses and 1-, 3-, 7-, and 14-day-old pups were preserved for light microscopic immunohistochemistry and electron microscopy. Intercalated cells were identified by immunostaining for H(+)-adenosinetriphosphatase (H(+)-ATPase) and band 3 protein. Apoptosis was identified by nick end labeling of DNA fragments, staining with the vital dye toluidine blue, and transmission electron microscopy. Two distinct mechanisms of elimination of intercalated cells were detected. Cells with apical labeling for H(+)-ATPase and basolateral labeling for band 3 protein protruded into the lumen of the MCD as if they were being extruded from the epithelium, and many had lost contact with the basement membrane. Extrusion of the cells with basolateral H(+)-ATPase or with no labeling for H(+)-ATPase was never observed. Apoptosis was observed in the MCD from shortly before birth to 7 days after birth, gradually progressing from the papillary tip toward the outer medulla. Staining for apoptosis was present in H(+)-ATPase-positive apoptotic bodies, located in cells that were negative for H(+)-ATPase. Staining was also occasionally observed in apoptotic cells with basolateral H(+)-ATPase but never in cells with apical H(+)-ATPase. Electron microscopy confirmed the presence of apoptotic intercalated cells in the MCD and demonstrated that apoptotic bodies were located in inner medullary collecting duct (IMCD) cells and principal cells. These results demonstrate that intercalated cells are deleted from the MCD by two distinct mechanisms, one involving apoptosis and subsequent phagocytosis by neighboring principal cells or IMCD cells. Elimination by extrusion affects only type A intercalated cells, whereas deletion by apoptosis appears to occur only in type B intercalated cells.

Three distinct cell populations in rat kidney collecting duct

1987 ◽  
Vol 253 (2) ◽  
pp. C323-C328 ◽  
Author(s):  
H. Holthofer ◽  
B. A. Schulte ◽  
G. Pasternack ◽  
G. J. Siegel ◽  
S. S. Spicer
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Morphological Characteristics ◽  
Anion Channel ◽  
Band 3 ◽  
Cell Populations ◽  
Intercalated Cells ◽  
Distinct Cell ◽  
Collecting Ducts ◽  
Principal And Intercalated Cells

The morphologically heterogeneous cell populations in the collecting ducts of the rat kidney were studied using immunocytochemical detection of Na+-K+-ATPase and the anion channel (band 3) glycoprotein. Both enzymes were localized to the basal aspect of separate and morphologically distinct subpopulations of cells in various segments of the collecting duct. Na+-K+-ATPase appeared to be present exclusively in principal cells as identified by their morphology, whereas band 3 antibodies reacted only with intercalated cells. However, 5-20% of cells with the morphological characteristics of intercalated cells failed to react with either antisera in various segments of collecting ducts. As band 3 glycoprotein serves in exchanging intracellular bicarbonate for chloride, it is highly likely that the cells positive for this antigen secrete protons. The method introduced here appears thus useful for distinguishing between principal and intercalated cells by differences in their enzyme content and further for revealing two subpopulations of intercalated cells. This method promises to provide a useful approach for studying the principal and intercalated cell populations in various metabolic states.

scholarly journals Intercalated Cell Subtypes in Connecting Tubule and Cortical Collecting Duct of Rat and Mouse

1999 ◽  
Vol 10 (1) ◽  
pp. 1-12 ◽  
Author(s):  
JIN KIM ◽  
YOUNG-HEE KIM ◽  
JUNG-HO CHA ◽  
C. CRAIG TISHER ◽  
KIRSTEN M. MADSEN
Keyword(s):  
Plasma Membrane ◽  
B Cells ◽  
Collecting Duct ◽  
Type A ◽  
Band 3 ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Type B ◽  
Intercalated Cell ◽  
Basolateral Plasma Membrane

Abstract. At least two populations of intercalated cells, type A and type B, exist in the connecting tubule (CNT), initial collecting tubule (ICT), and cortical collecting duct (CCD). Type A intercalated cells secrete protons via an apical H+ - ATPase and reabsorb bicarbonate by a band 3-like Cl-/HCO3- exchanger, AE1, located in the basolateral plasma membrane. Type B intercalated cells secrete bicarbonate by an apical Cl-/HCO3- exchanger that is distinct from AE1 and remains to be identified. They express H+ -ATPase in the basolateral plasma membrane and in vesicles throughout the cytoplasm. A third type of intercalated cell with apical H+ -ATPase, but no AE1, has been described in the CNT and CCD of both rat and mouse. The prevalence of the third cell type is not known. The aim of this study was to characterize and quantify intercalated cell subtypes, including the newly described third non A-non B cell, in the CNT, ICT, and CCD of the rat and mouse. A triple immunolabeling procedure was developed in which antibodies to H+ -ATPase and band 3 protein were used to identify subpopulations of intercalated cells, and segment-specific antibodies were used to identify distal tubule and collecting duct segments. In both rat and mouse, intercalated cells constituted approximately 40% of the cells in the CNT, ICT, and CCD. Type A, type B, and non A-non B intercalated cells were observed in all of the three segments, with type A cells being the most prevalent in both species. In the mouse, however, non A-non B cells constituted more than half of the intercalated cells in the CNT, 39% in the ICT, and 22% in the CCD, compared with 14, 7, and 5%, respectively, in the rat. In contrast, type B intercalated cells accounted for only 8 to 16% of the intercalated cells in the three segments in the mouse compared with 26 to 39% in the rat. It is concluded that striking differences exist in the prevalence and distribution of the different types of intercalated cells in the CNT, ICT, and CCD of rat and mouse. In the rat, the non A-non B cells are fairly rare, whereas in the mouse, they constitute a major fraction of the intercalated cells, primarily at the expense of the type B intercalated cells.

Versatility of NaCl transport mechanisms in the cortical collecting duct

2017 ◽  
Vol 313 (6) ◽  
pp. F1254-F1263 ◽  
Author(s):  
Aurélie Edwards ◽  
Gilles Crambert
Keyword(s):  
Collecting Duct ◽  
Type A ◽  
Transport Systems ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Type B ◽  
Cell Type ◽  
Acid Base ◽  
Nacl Transport ◽  
Principal Cells

The cortical collecting duct (CCD) forms part of the aldosterone-sensitive distal nephron and plays an essential role in maintaining the NaCl balance and acid-base status. The CCD epithelium comprises principal cells as well as different types of intercalated cells. Until recently, transcellular Na+ transport was thought to be restricted to principal cells, whereas (acid-secreting) type A and (bicarbonate-secreting) type B intercalated cells were associated with the regulation of acid-base homeostasis. This review describes how this traditional view has been upended by several discoveries in the past decade. A series of studies has shown that type B intercalated cells can mediate electroneutral NaCl reabsorption by a mechanism involving Na+-dependent and Na+-independent Cl−/[Formula: see text] exchange, and that is energetically driven by basolateral vacuolar H+-ATPase pumps. Other research indicates that type A intercalated cells can mediate NaCl secretion, through a bumetanide-sensitive pathway that is energized by apical H+,K+-ATPase type 2 pumps operating as Na+/K+ exchangers. We also review recent findings on the contribution of the paracellular route to NaCl transport in the CCD. Last, we describe cross-talk processes, by which one CCD cell type impacts Na+/Cl− transport in another cell type. The mechanisms that have been identified to date demonstrate clearly the interdependence of NaCl and acid-base transport systems in the CCD. They also highlight the remarkable versatility of this nephron segment.

Immunocytochemical localization of band 3 protein in the rat collecting duct

1988 ◽  
Vol 255 (1) ◽  
pp. F115-F125 ◽  
Author(s):  
J. W. Verlander ◽  
K. M. Madsen ◽  
P. S. Low ◽  
D. P. Allen ◽  
C. C. Tisher
Keyword(s):  
Plasma Membranes ◽  
Collecting Duct ◽  
Type A ◽  
Band 3 ◽  
Rabbit Kidney ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Band 3 Protein ◽  
Type B ◽  
Gold Particles

Band 3 protein is the major anion transport protein of the erythrocyte cell membrane where it catalyzes the exchange of HCO3- for Cl-. There is evidence that band 3 protein is present in the collecting duct of both the rat and rabbit kidney. We used colloidal-gold immunocytochemistry to determine the ultrastructural location of band 3 protein in the rat cortical (CCD) and outer medullary collecting ducts (OMCD). Kidneys of normal Sprague-Dawley rats were fixed by intravascular perfusion with 1% glutaraldehyde and embedded in Lowicryl K4M. Two polyclonal antibodies raised in rabbits were used as the primary antibody in separate experiments, one against the 43-kDa fragment of the cytoplasmic domain of human erythrocyte band 3 protein and the other against rat erythrocyte band 3 protein. This was followed by exposure to gold-conjugated goat anti-rabbit immunoglobulin G. Transmission electron microscopy revealed gold particles along the basal and lateral plasma membranes of all intercalated cells of the OMCD. In the CCD, the basal and lateral plasma membranes of the type A intercalated cells only were labeled with gold particles. The type B intercalated cells and principal cells were devoid of gold particles, as were all cells of the proximal tubule, the distal convoluted tubule, and the thick ascending limb of the loop of Henle. We conclude that a Cl(-)-HCO3- transporter is present in the basal and lateral plasma membranes of the intercalated cells in the OMCD and the type A intercalated cells in the CCD. These findings provide further evidence that these intercalated cells are involved in H+ secretion in the OMCD and CCD of the rat. We have no evidence for the presence of band 3 protein in the type B intercalated cells of the CCD, which supports the hypothesis that type B cells are functionally and structurally distinct from type A cells.

Effect of acute respiratory acidosis on two populations of intercalated cells in rat cortical collecting duct

1987 ◽  
Vol 253 (6) ◽  
pp. F1142-F1156 ◽  
Author(s):  
J. W. Verlander ◽  
K. M. Madsen ◽  
C. C. Tisher
Keyword(s):  
Electron Microscopy ◽  
Collecting Duct ◽  
Respiratory Acidosis ◽  
Type A ◽  
Hydrogen Ion ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Type B ◽  
A Cells ◽  
Two Populations

Recent studies suggest the presence of two populations of intercalated cells in the rabbit cortical collecting duct (CCD), one involved with hydrogen ion secretion and another that may play a role in bicarbonate secretion. The purpose of this study was to determine whether two populations of intercalated cells are present in the rat CCD and to establish their response to acute respiratory acidosis. Rats were studied during normal acid-base conditions and after 4-5 h of respiratory acidosis. In all animals light microscopy and transmission and scanning electron microscopy revealed two configurations of intercalated cells, type A with an extensive apical tubulovesicular membrane compartment and prominent surface microprojections and type B with a well-developed vesicular compartment and short sparse surface microprojections. By transmission electron microscopy, studs were present on the cytoplasmic face of the apical plasmalemma and tubulovesicular profiles of A cells. In respiratory acidosis there was a striking increase in apical microprojections and in the surface density of the apical membrane of type A cells similar to the response observed previously in intercalated cells in the outer medullary collecting duct (OMCD) studied under the same physiological conditions. No changes were observed in type B cells. Scanning electron microscopy revealed no change in the relative number of type A and type B cells in respiratory acidosis. We conclude that two distinct populations of intercalated cells exist in the rat CCD: type A, which resembles the intercalated cells in the OMCD, and type B. The response of type A cells to acute respiratory acidosis and the similarity between these cells and intercalated cells in the OMCD, which are believed to secrete hydrogen ion, suggest that the type A cells are involved in hydrogen ion secretion in the CCD.

Immunohistochemical localization of H-K-ATPase α2c-subunit in rabbit kidney

2001 ◽  
Vol 281 (2) ◽  
pp. F357-F365 ◽  
Author(s):  
Jill W. Verlander ◽  
Robin M. Moudy ◽  
W. Grady Campbell ◽  
Brian D. Cain ◽  
Charles S. Wingo
Keyword(s):  
Collecting Duct ◽  
Type A ◽  
Immunohistochemical Localization ◽  
Macula Densa ◽  
Rabbit Kidney ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Type B ◽  
Principal Cells ◽  
Medullary Collecting Duct

The rabbit kidney possesses mRNA for the H-K-ATPase α1-subunit (HKα1) and two splice variants of the H-K-ATPase α2-subunit (HKα2). The purpose of this study was to determine the specific distribution of one of these, the H-K-ATPase α2c-subunit isoform (HKα2c), in rabbit kidney by immunohistochemistry. Chicken polyclonal antibodies against a peptide based on the NH2 terminus of HKα2c were used to detect HKα2cimmunoreactivity in tissue sections. Immunohistochemical localization of HKα2c revealed intense apical immunoreactivity in a subpopulation of cells in the connecting segment, cortical collecting duct, and outer medullary collecting duct in both the outer and inner stripe. An additional population of cells exhibited a thin apical band of immunolabel. Immunohistochemical colocalization of HKα2c with carbonic anhydrase II, the Cl−/HCO[Formula: see text] exchanger AE1, and HKα1 indicated that both type A and type B intercalated cells possessed intense apical HKα2c immunoreactivity, whereas principal cells and connecting segment cells had only a thin apical band of HKα2c. Labeled cells were evident through the middle third of the inner medullary collecting duct in the majority of animals. Immunolabel was also present in papillary surface epithelial cells, cells in the cortical thick ascending limb of Henle's loop (cTAL), and the macula densa. Thus in the rabbit kidney, apical HKα2c is present and may contribute to acid secretion or potassium uptake throughout the connecting segment and collecting duct in both type A and type B intercalated cells, principal cells, and connecting segment cells, as well as in cells in papillary surface epithelium, cTAL, and macula densa.

Immunoelectron microscopic localization of NBC3 sodium-bicarbonate cotransporter in rat kidney

2000 ◽  
Vol 278 (2) ◽  
pp. F327-F336 ◽  
Author(s):  
Tae-Hwan Kwon ◽  
Alexander Pushkin ◽  
Natalia Abuladze ◽  
Søren Nielsen ◽  
Ira Kurtz
Keyword(s):  
Plasma Membrane ◽  
Immunoelectron Microscopy ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Type A ◽  
Apical Plasma Membrane ◽  
Type B ◽  
Principal Cells ◽  
Connecting Tubule ◽  
Basolateral Plasma Membrane

In the present study, we produced a rabbit peptide-derived polyclonal COOH-terminal antibody that selectively recognizes NBC3, to determine the cellular and subcellular localization of NBC3 in rat kidney, using immunocytochemistry and immunoelectron microscopy. Immunocytochemistry with cryostat sections and semithin cryosections revealed specific staining of intercalated cells (ICs) in the connecting tubule and in cortical, outer medullary, and initial inner medullary collecting ducts. In the connecting tubule and in the cortical and medullary collecting duct, the labeling was associated with both type A and type B ICs. In type A ICs, labeling was confined to the apical and subapical domains, whereas in type B ICs, basal domains were exclusively labeled. In contrast, collecting duct principal cells were consistently unlabeled, and this was confirmed using anti-aquaporin-2 antibodies, which labeled principal cells in parallel semithin cryosections. Glomeruli, proximal tubules, descending thin limbs, ascending thin limbs, thick ascending limbs, distal convoluted tubules, and vascular structures were unlabeled. For immunoelectron microscopy, tissue samples were freeze-substituted, and immunolabeling was performed on ultrathin Lowicryl HM20 sections. Immunoelectron microscopy demonstrated that NBC3 labeling was very abundant in the apical plasma membrane, in intracellular vesicles, and in tubulocisternal profiles in the subapical domains of type A ICs. In type B ICs, NBC3 was mainly present in the basolateral plasma membrane. Immunolabeling controls using peptide-absorbed antibody were consistently negative. In conclusion, NBC3 is highly abundant in the apical plasma membrane of type A ICs and in the basolateral plasma membrane of type B ICs. This suggests that NBC3 plays an important role in modulating bicarbonate transport in the connecting tubule and collecting duct.

Expression of TWIK-1, a novel weakly inward rectifying potassium channel in rat kidney

1998 ◽  
Vol 275 (6) ◽  
pp. C1602-C1609 ◽  
Author(s):  
F. Cluzeaud ◽  
R. Reyes ◽  
B. Escoubet ◽  
M. Fay ◽  
M. Lazdunski ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Cell Types ◽  
Loop Of Henle ◽  
Membrane Domains ◽  
Cortical Collecting Duct ◽  
Double Immunofluorescence ◽  
Distinct Cell ◽  
Weakly Inward ◽  
Low K

Several K+ conductances have been identified in the kidney, with specific properties and localization in distinct cell types and membrane domains. On the other hand, several K+ channels have been characterized at the molecular level. By immunolocalization, we show that a new inward rectifying K+channel, TWIK-1, is specifically expressed in distinct tubular segments and cell types of the rat kidney. In the proximal tubule, TWIK-1 prevails in the initial portions (convoluted part), where it is restricted to the apical (brush-border) membrane. In the collecting duct, immunofluorescence was intracellular or confined to the apical membrane and restricted to intercalated cells, i.e., in cells lacking aquaporin-2, as shown by double immunofluorescence. TWIK was also expressed in medullary and cortical parts of the thick limb of the loop of Henle, identified with an anti-Tamm-Horsfall protein antibody (double immunofluorescence). The intensity of TWIK-1 immunolabeling was unchanged in rats fed a low-Na+ or a low-K+ diet. Because TWIK-1 shares common properties with the low-conductance apical K+ channel of the collecting duct, we propose that it could play a role in K+ secretion, complementary to ROMK, another recently characterized K+ channel located in principal cells of the cortical collecting duct and in the loop of Henle.

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