Maturation of carbonic anhydrase IV expression in rabbit kidney

2001 ◽  
Vol 280 (5) ◽  
pp. F895-F903 ◽  
Author(s):  
Cornelia A. Winkler ◽  
Ann M. Kittelberger ◽  
Richard H. Watkins ◽  
William M. Maniscalco ◽  
George J. Schwartz
Keyword(s):  
Carbonic Anhydrase ◽  
Proximal Tubule ◽  
Collecting Duct ◽  
High Rate ◽  
Rabbit Kidney ◽  
Mrna Levels ◽  
Intercalated Cells ◽  
Collecting Ducts ◽  
Medullary Collecting Duct ◽  
Carbonic Anhydrase Iv

Carbonic anhydrase (CA) IV facilitates renal acidification by catalyzing the dehydration of luminal H2CO3. CA IV is expressed in proximal tubules, medullary collecting ducts, and A-intercalated cells of the mature rabbit kidney (Schwartz GJ, Kittelberger AM, Barnhart DA, and Vijayakumar S. Am J Physiol 278: F894–F904, 2000). In view of the maturation of HCO[Formula: see text] transport in the proximal tubule and collecting duct, the ontogeny of CA IV expression was examined. During the first 2 wk, CA IV mRNA was expressed in maturing cortex and medulla at ∼20% of adult levels. The maturational increase was gradual in cortex over 3–5 wk of age but surged in the medulla, so that mRNA levels appeared higher than those in the adult medulla. In situ hybridization showed very little CA IV mRNA at 5 days, with increases in deep cortex and medullary collecting ducts by 21 days. Expression of CA IV protein in the cortex and medulla was minimal at 3 days of age but then apparent in the juxtamedullary region, A-intercalated cells and medullary collecting ducts by 18 days; there was little labeling of the proximal straight tubules of the medullary rays. Thus CA IV expression may be regulated to accommodate the maturational increase in HCO[Formula: see text] absorption in the proximal tubule. In the medullary collecting duct, there is a more robust maturation of CA IV mRNA and protein, commensurate with the high rate of HCO[Formula: see text] absorption in the neonatal segment.

scholarly journals Carbonic anhydrase IV is expressed in H+-secreting cells of rabbit kidney

2000 ◽  
Vol 278 (6) ◽  
pp. F894-F904 ◽  
Author(s):  
George J. Schwartz ◽  
Ann M. Kittelberger ◽  
Darlene A. Barnhart ◽  
Soundarapandian Vijayakumar
Keyword(s):  
Carbonic Anhydrase ◽  
Molecular Mass ◽  
Carbonic Acid ◽  
Collecting Duct ◽  
Rabbit Kidney ◽  
Alternative Mechanism ◽  
Intercalated Cells ◽  
Duct Cells ◽  
Collecting Duct Cells ◽  
Medullary Collecting Duct

Carbonic anhydrase (CA) IV is a membrane-bound enzyme that catalyzes the dehydration of carbonic acid to CO2 and water. Using peptides from each end of the deduced rabbit CA IV amino acid sequence, we generated a goat anti-rabbit CA IV antibody, which was used for immunoblotting and immunohistochemical analysis. CA IV was expressed in a variety of organs including spleen, heart, lung, skeletal muscle, colon, and kidney. Rabbit kidney CA IV had two N-glycosylation sites and was sialated, the apparent molecular mass increasing by at least 11 to ∼45 kDa in the cortex. Medullary CA IV was much more heavily glycosylated than CA IV from cortex or any other organ, such modifications increasing the molecular mass by at least 20 kDa. CA IV was expressed on the apical and basolateral membranes of proximal tubules with expression levels on the order of S2 > S1 > S3 = 0. Because CA IV is believed to be anchored to the apical membrane by glycosylphosphatidylinositol, the presence of basolateral CA IV suggests an alternative mechanism. CA IV was localized on the apical membranes of outer medullary collecting duct cells of the inner stripe and inner medullary collecting duct cells, as well as on α-intercalated cells. However, CA IV was not expressed by β-intercalated cells, glomeruli, distal tubule, or Henle's loop cells. Thus CA IV was expressed by H+-secreting cells of the rabbit kidney, suggesting an important role for CA IV in urinary acidification.

scholarly journals Carbonic anhydrase II and IV mRNA in rabbit nephron segments: stimulation during metabolic acidosis

1998 ◽  
Vol 274 (2) ◽  
pp. F259-F267 ◽  
Author(s):  
Shuichi Tsuruoka ◽  
Ann M. Kittelberger ◽  
George J. Schwartz
Keyword(s):  
Metabolic Acidosis ◽  
Carbonic Anhydrase ◽  
Mrna Expression ◽  
Collecting Duct ◽  
Proximal Tubules ◽  
Rabbit Kidney ◽  
Rt Pcr ◽  
Chronic Metabolic Acidosis ◽  
Nephron Segments ◽  
Medullary Collecting Duct

Carbonic anhydrase (CA) facilitates renal bicarbonate reabsorption and acid excretion. Cytosolic CA II catalyzes the buffering of intracellular hydroxyl ions by CO2, whereas membrane-bound CA IV catalyzes the dehydration of carbonic acid generated from the secretion of protons. Although CA II and IV are expressed in rabbit kidney, it is not entirely clear which segments express which isoforms. It was the purpose of this study to characterize the expression of CA II and CA IV mRNAs by specific segments of the nephron using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and to determine the effect of chronic metabolic acidosis on CA expression by those segments. Individual nephron segments (usually 1–2 mm) were isolated by microdissection and subjected to RT-PCR. Amplification was performed simultaneously for CA IV, CA II, and malate dehydrogenase (MDH), a housekeeping gene. The intensities of the PCR products were quantitated by densitometry. CA IV mRNA was expressed by S1 and S2 proximal tubules and by outer medullary collecting duct from inner stripe (OMCDi) and outer stripe and initial inner medullary collecting duct (IMCDi). CA II mRNA was expressed by S1, S2, and S3 proximal tubules, thin descending limb, connecting segment (CNT), and all collecting duct segments. Acid loading induced CA IV mRNA expression in S1 and S2 proximal tubules and in OMCDi and IMCDi. CA II mRNA was induced by acidosis in all three proximal segments and nearly all distal segments beginning with CNT. No upregulation of MDH mRNA expression occurred. These adaptive increases in CA II and IV mRNAs are potentially important in the kidney’s adaptation to chronic metabolic acidosis.

scholarly journals Histochemical carbonic anhydrase in rat inner medullary collecting duct.

1992 ◽  
Vol 40 (10) ◽  
pp. 1535-1545 ◽  
Author(s):  
J G Kleinman ◽  
J L Bain ◽  
C Fritsche ◽  
D A Riley
Keyword(s):  
Carbonic Anhydrase ◽  
Collecting Duct ◽  
Light And Electron Microscopy ◽  
Cell Types ◽  
Intercalated Cells ◽  
Cell Type ◽  
Variable Intensity ◽  
Inner Medullary Collecting Duct ◽  
Majority Population ◽  
Medullary Collecting Duct

Rat inner medullary collecting duct (IMCD) secretes substantial amounts of H+. However, carbonic anhydrase (CA), a concomitant of H+ secretion, has been generally reported absent in this segment. To reexamine this problem, we investigated CA and the morphological phenotypes of cells comprising the IMCD by CA histochemistry, using a modified Hansson technique with light and electron microscopy. Throughout the medulla, tubule cells exhibit histochemical CA activity. In the initial third of the inner medulla, a small proportion have features of intercalated cells and demonstrate some degree of CA activity. However, the majority population in the early portions of the IMCD appears to consist of principal cells. These also show CA staining of widely variable intensity, both among and within cells. A third cell type, previously called "IMCD cells", appears in the middle portion of the IMCD and is the only cell type present near the papilla tip. In contrast to previous reports, these "IMCD cells" have histochemical CA staining, also of highly variable intensity. These results demonstrate that stainable carbonic anhydrase to support acidification is present throughout the rat IMCD, both in intercalated cells and in some cells clearly not of this type. Therefore, the presence of CA is not specific for the intercalated cell type and suggests that other cell types may participate in acid secretion in IMCD.

Remodeling the cellular profile of collecting ducts by chronic carbonic anhydrase inhibition

2001 ◽  
Vol 280 (3) ◽  
pp. F437-F448 ◽  
Author(s):  
Corinne Bagnis ◽  
Vladimir Marshansky ◽  
Sylvie Breton ◽  
Dennis Brown
Keyword(s):  
Carbonic Anhydrase ◽  
Collecting Duct ◽  
Relative Increase ◽  
Adult Rats ◽  
Collecting Ducts ◽  
Carbonic Anhydrase Inhibition ◽  
Sodium Hydrogen Exchanger ◽  
Adaptive Processes ◽  
Medullary Collecting Duct ◽  
Deficient Mice

Factors regulating the differentiated phenotype of principal cells (PC) and A- and B-intercalated cells (IC) in kidney collecting ducts are poorly understood. However, we have shown previously that carbonic anhydrase II (CAII)-deficient mice have no IC in their medullary collecting ducts, suggesting a potential role for this enzyme in determining the cellular composition of this tubule segment. We now report that the cellular profile of the collecting ducts of adult rats can be remodeled by inhibiting CA activity in rats by using osmotic pumps containing acetazolamide. The 31-kDa subunit of the vacuolar H+-ATPase, the sodium/hydrogen exchanger regulatory factor NHE-RF, and the anion exchanger AE1 were used to identify IC subtypes by immunofluorescence staining, while aquaporin 2 and aquaporin 4 were used to identify PC. In the cortical collecting ducts of animals treated with acetazolamide for 2 wk, the percentage of B-IC decreased significantly (18 ± 2 vs. 36 ± 4%, P < 0.01) whereas the percentage of A-IC increased (82 ± 2 vs. 64 ± 4%, P < 0.01) with no change in the percentage of total IC in the epithelium. In some treated rats, B-IC were virtually undetectable. In the inner stripe of the outer medulla, the percentage of IC increased in treated animals (48 ± 2 vs. 37 ± 3%, P < 0.05) and the percentage of PC decreased (52 ± 2 vs. 63 ± 3%, P < 0.05). Moreover, IC appeared bulkier, protruded into the lumen, and showed a significant increase in the length of their apical (20.8 ± 0.5 vs. 14.6 ± 0.4 μm, P < 0.05) and basolateral membranes (25.8 ± 0.4 vs. 23.8 ± 0.5 μm, P < 0.05) compared with control rats. In the inner medullary collecting ducts of treated animals, the number of IC in the proximal third of the papilla was reduced compared with controls (11 ± 4 vs. 40 ± 11 IC/mm2, P < 0.05). These data suggest that CA activity plays an important role in determining the differentiated phenotype of medullary collecting duct epithelial cells and that the cellular profile of collecting ducts can be remodeled even in adult rats. The relative depletion of cortical B-IC and the relative increase in number and hyperplasia of A-IC in the medulla may be adaptive processes that would tend to correct or stabilize the metabolic acidosis that would otherwise ensue following systemic carbonic anhydrase inhibition.

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.

Effects of ischemia-reperfusion injury on renal ammonia metabolism and the collecting duct

2007 ◽  
Vol 293 (4) ◽  
pp. F1342-F1354 ◽  
Author(s):  
Ki-Hwan Han ◽  
Hye-Young Kim ◽  
Byron P. Croker ◽  
Sirirat Reungjui ◽  
Su-Youn Lee ◽  
...  
Keyword(s):  
Metabolic Acidosis ◽  
Proximal Tubule ◽  
Renal Injury ◽  
Collecting Duct ◽  
Ischemia Reperfusion ◽  
Ammonia Excretion ◽  
Intercalated Cells ◽  
Acute Renal Injury ◽  
Intercalated Cell ◽  
Medullary Collecting Duct

Acute renal injury induces metabolic acidosis, but its specific effects on the collecting duct, the primary site for urinary ammonia secretion, the primary component of net acid excretion, are incompletely understood. We induced ischemia-reperfusion (I/R) acute renal injury in Sprague-Dawley rats by clamping the renal pedicles bilaterally for 30 min followed by reperfusion for 6 h. Control rats underwent sham surgery without renal pedicle clamping. I/R injury decreased urinary ammonia excretion significantly but did not persistently alter urine volume, Na+, K+, or bicarbonate excretion. Histological examination demonstrated cellular damage in the outer and inner medullary collecting duct, as well as in the proximal tubule and the thick ascending limb of the loop of Henle. A subset of collecting duct cells were damaged and/or detached from the basement membrane; these cells were present predominantly in the outer medulla and were less frequent in the inner medulla. Immunohistochemistry identified that the damaged/detached cells were A-type intercalated cells, not principal cells. Both TdT-mediated dUTP nick-end labeling (TUNEL) staining and transmission electron microscopic examination demonstrated apoptosis but not necrosis. However, immunoreactivity for caspase-3 was observed in the proximal tubule, but not in collecting duct intercalated cells, suggesting that mechanism(s) of collecting duct intercalated cell apoptosis differ from those operative in the proximal tubule. We conclude that I/R injury decreases renal ammonia excretion and is associated with intercalated cell-specific detachment and apoptosis in the outer and inner medullary collecting duct. These effects likely contribute to the metabolic acidosis frequently observed in acute renal injury.

scholarly journals Membrane-associated carbonic anhydrase activity in the kidney of CA II-deficient mice.

1992 ◽  
Vol 40 (11) ◽  
pp. 1665-1673 ◽  
Author(s):  
Y Ridderstråle ◽  
P J Wistrand ◽  
R E Tashian
Keyword(s):  
Carbonic Anhydrase ◽  
Proximal Tubule ◽  
Collecting Duct ◽  
Distal Tubule ◽  
Intercalated Cells ◽  
Cell Nuclei ◽  
Cytoplasmic Staining ◽  
Basolateral Membranes ◽  
Basolateral Side ◽  
Deficient Mice

Carbonic anhydrase II-deficient mice offer a possibility to study the localization along the nephron of membrane-associated carbonic anhydrase (CA) activity without interference from the cytoplasmic enzyme. We studied the localization of CA in kidneys from CA II-deficient and control mice by immunocytochemistry (CA II) and histochemistry. Cytoplasmic staining was found in convoluted proximal tubule, thick limb of Henle, and principal and intercalated cells of collecting duct in the control animals but was absent in the CA II-deficient mice. In cells with cytoplasmic staining the cell nuclei were stained. Intense histochemical activity was associated with apical and basolateral membranes of convoluted proximal tubule, first part of thin limb, thick limb, and basolateral membranes of late distal tubule. In collecting ducts of control animals, the basolateral cell membranes of intercalated cells were the only clearly stained membranes. In CA II-deficient animals one type of intercalated cell was stained most intensely at the apical membranes and another only at the basolateral. We suggest that the former corresponds to Type A intercalated cells secreting H+ ions to the luminal side and the latter to Type B cells secreting H+ ions to the basolateral side.

Net acid transport by isolated perfused inner medullary collecting ducts

1990 ◽  
Vol 258 (1) ◽  
pp. F75-F84 ◽  
Author(s):  
S. M. Wall ◽  
J. M. Sands ◽  
M. F. Flessner ◽  
H. Nonoguchi ◽  
K. R. Spring ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Intercalated Cells ◽  
Acid Transport ◽  
Concentration Gradients ◽  
Collecting Ducts ◽  
Acidic Form ◽  
Medullary Collecting Duct ◽  
Stop Flow

The isolated perfused tubule technique was used to study net acid transport in rat terminal inner medullary collecting duct (IMCD) segments. The stop-flow luminal pH [measured fluorometrically with the acidic form of the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein in the lumen] fell 0.35 units below the bath pH in tubules from control rats and 0.53 units below the bath in tubules from deoxycorticosterone-treated rats. Tubules from control rats absorbed bicarbonate and secreted ammonium against concentration gradients, although at low rates. In control rats, 10(-8) M vasopressin added to the bath increased bicarbonate absorption almost threefold. Treatment of rats in vivo with deoxycorticosterone significantly increased the rate of bicarbonate absorption in vitro. In vivo NH4Cl loading also significantly increased bicarbonate absorption. Staining microdissected tubules with acridine orange confirmed that the perfused segments lacked intercalated cells. We conclude that the terminal IMCD spontaneously acidifies the lumen despite an absence of intercalated cells. Bicarbonate absorption appears to be regulated by the same factors that affect net acidification in other collecting duct segments.

Vasopressin induces expression of the Cl−/HCO3− exchanger SLC26A7 in kidney medullary collecting ducts of Brattleboro rats

2006 ◽  
Vol 290 (5) ◽  
pp. F1194-F1201 ◽  
Author(s):  
Snezana Petrovic ◽  
Hassane Amlal ◽  
Xuming Sun ◽  
Fiona Karet ◽  
Sharon Barone ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Collecting Duct ◽  
Brattleboro Rats ◽  
Intercalated Cells ◽  
Collecting Ducts ◽  
Before And After ◽  
Mild Reduction ◽  
Medullary Collecting Duct

SLC26A7 is a newly identified basolateral Cl−/HCO3− exchanger specific to α-intercalated cells of the outer medullary collecting duct (OMCD). The purpose of the present experiments was to examine the expression of SLC26A7 in kidneys of vasopressin-deficient Brattleboro rats before and after treatment with desamino-Cys1,d-Arg8-vasopressin (dDAVP). Brattleboro rats were treated with dDAVP, a vasopressin analog, for 8 days, and their kidneys were examined for the expression of SLC26A7. The expression of SLC26A7 protein, as examined by immunofluorescence, was undetectable in kidneys of Brattleboro rats. However, treatment with dDAVP induced expression of SLC26A7 protein, restoring it to levels observed in normal rats. These results were verified by Western blot analysis. The mRNA expression of SLC26A7 remained unchanged in response to dDAVP. Immunofluorescent labeling demonstrated abundant levels of anion exchanger type 1 in the OMCD of Brattleboro rats and a mild reduction in response to dDAVP. The abundance of H+-ATPase was not affected by dDAVP. The increased SLC26A7 expression directly correlated with enhanced aquaporin-2 expression, which is proportional to increased interstitial osmolarity in the medulla. In conclusion, vasopressin increases the expression of SLC26A7 protein through posttranscriptional mechanisms in the OMCD. The induction of SLC26A7 by vasopressin in OMCD cells of Brattleboro rats is likely an attempt by cells to regulate their cell volume and maintain HCO3− absorption in a state associated with increased interstitial medullary tonicity.

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.

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