Mechanisms of HCO 3 − secretion in the rabbit connecting segment

1999 ◽  
Vol 277 (4) ◽  
pp. F567-F574 ◽  
Author(s):  
Shuichi Tsuruoka ◽  
George J. Schwartz
Keyword(s):  
Cell Function ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Acid Base ◽  
Intercalated Cell ◽  
Atpase Inhibitors ◽  
Transepithelial Voltage ◽  
Β Adrenergic Receptors

The connecting tubule (CNT) contains α-(H+-secreting) and β-([Formula: see text]-secreting) intercalated cells and is therefore likely to contribute to acid-base homeostasis. To characterize the mechanisms of [Formula: see text]transport in the rabbit CNT, in which there is little definitive data presently available, we microdissected the segments from the superficial cortical labyrinth, perfused them in vitro, measured net[Formula: see text] transport ( J [Formula: see text] ) by microcalorimetry, and examined the effects of several experimental maneuvers. Mean ± SE basal J [Formula: see text] was −3.4 ± 0.1 pmol ⋅ min−1 ⋅ mm−1(net [Formula: see text] secretion), and transepithelial voltage was −13 ± 1 mV ( n = 47). Net[Formula: see text] secretion was markedly inhibited by removal of luminal Cl− or application of basolateral H+-ATPase inhibitors (bafilomycin or concanamycin), maneuvers that inhibit β-intercalated cell function. Net [Formula: see text] secretion was not affected by inhibitors of α-intercalated cell function (basolateral Cl− removal, basolateral DIDS, or luminal H+-ATPase inhibitors). Net [Formula: see text] secretion was stimulated by isoproterenol and inhibited by acetazolamide. These data indicate that 1) CNTs secrete[Formula: see text] via an apical DIDS-insensitive Cl−/[Formula: see text]exchanger, mediated by a basolateral bafilomycin- and concanamycin-sensitive H+-ATPase; 2) inhibition of cytosolic carbonic anhydrase decreases [Formula: see text] secretion; and 3) stimulation of β-adrenergic receptors increases [Formula: see text] secretion. The failure to influence net [Formula: see text]transport by inhibiting α-intercalated cell apical H+-ATPases or basolateral Cl−/[Formula: see text]exchange suggests that the CNT has fewer functioning α-intercalated cells than the cortical collecting duct. These are the first studies to examine the rate and mechanisms of[Formula: see text] secretion by the rabbit CNT; this is clearly an important segment in mediating acid-base homeostasis.

scholarly journals ENaC inhibition stimulates HCl secretion in the mouse cortical collecting duct. II. Bafilomycin-sensitive H+ secretion

2015 ◽  
Vol 309 (3) ◽  
pp. F259-F268 ◽  
Author(s):  
Masayoshi Nanami ◽  
Vladimir Pech ◽  
Yoskaly Lazo-Fernandez ◽  
Alan M. Weinstein ◽  
Susan M. Wall
Keyword(s):  
Intracellular Ph ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Transport Pathway ◽  
Anion Secretion ◽  
Intercalated Cell ◽  
Atpase Inhibitors ◽  
Hcl Secretion ◽  
Transepithelial Voltage

Epithelial Na+ channel (ENaC) blockade stimulates stilbene-sensitive conductive Cl− secretion in the mouse cortical collecting duct (CCD). This study's purpose was to determine the co-ion that accompanies benzamil- and DIDS-sensitive Cl− flux. Thus transepithelial voltage, VT, as well as total CO2 (tCO2) and Cl− flux were measured in CCDs from aldosterone-treated mice consuming a NaCl-replete diet. We reasoned that if stilbene inhibitors (DIDS) reduce conductive anion secretion they should reduce the lumen-negative VT. However, during ENaC blockade (benzamil, 3 μM), DIDS (100 μM) application to the perfusate reduced net H+ secretion, which increased the lumen-negative VT. Conversely, ENaC blockade alone stimulated H+ secretion, which reduced the lumen-negative VT. Application of an ENaC inhibitor to the perfusate reduced the lumen-negative VT, increased intercalated cell intracellular pH, and reduced net tCO2 secretion. However, benzamil did not change tCO2 flux during apical H+-ATPase blockade (bafilomycin, 5 nM). The increment in H+ secretion observed with benzamil application contributes to the fall in VT observed with application of this diuretic. As such, ENaC blockade reduces the lumen-negative VT by inhibiting conductive Na+ absorption and by stimulating H+ secretion by type A intercalated cells. In conclusion, 1) in CCDs from aldosterone-treated mice, benzamil application stimulates HCl secretion mediated by the apical H+-ATPase and a yet to be identified conductive Cl− transport pathway; 2) benzamil-induced HCl secretion is reversed with the application of stilbene inhibitors or H+-ATPase inhibitors to the perfusate; and 3) benzamil reduces VT not only by inhibiting conductive Na+ absorption, but also by stimulating H+ secretion.

Calcitonin stimulates H+ secretion in rat kidney intercalated cells

1996 ◽  
Vol 271 (6) ◽  
pp. F1217-F1223 ◽  
Author(s):  
E. Siga ◽  
P. Houillier ◽  
B. Mandon ◽  
G. Moine ◽  
C. de Rouffignac
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Intercalated Cells ◽  
Specific Function ◽  
Cortical Collecting Duct ◽  
Intercalated Cell ◽  
Transepithelial Voltage ◽  
Net Fluxes

Calcitonin (CT) modulates rat intercalated cell (IC) functions of the rat cortical collecting duct (CCD) [E. Siga, B. Mandon, N. Roinel, and C. de Rouffignac. Am.J. Physiol. 264 (Renal Fluid Electrolyte Physiol. 33): F221-F227, 1993]. To characterize the specific function regulated by CT, rat CCDs were perfused in vitro. Total CO2 net fluxes (JtCO2, pmol.mm-1.min-1) and transepithelial voltage (Vt) were measured. Bath CT induced a significant tCO2 reabsorption. This effect was higher on CCDs harvested from acid-loaded than from control rats. When HCO3- secretion was blocked, CT also raised JtCO2 and Vt. When H+ secretion was blocked, CT was ineffective on JtCO2 and Vt. When HCO3- secretion was increased and H+ secretion was inhibited, CT did not change JtCO2, whereas isoproterenol (ISO) increased tCO2 secretion from -13.5 +/- 2.0 (control) to -19.0 +/- 2.4 (ISO). In rat CCD studied under these same preceding conditions plus luminal amiloride to block the Na(+)-dependent Vt, CT did not alter Vt, whereas ISO increased it by 4.5 +/- 0.7 mV. We conclude from these data that, in the rat CCD, calcitonin stimulates H+ secretion, likely by so-called alpha-intercalated (alpha-IC) cells, whereas ISO stimulates HCO3- secretion, likely by so-called beta-IC cells.

Cellular actions of cAMP on HCO3(-)-secreting cells of rabbit CCD: dependence on in vivo acid-base status

1994 ◽  
Vol 266 (4) ◽  
pp. F528-F535 ◽  
Author(s):  
C. Emmons ◽  
J. B. Stokes
Keyword(s):  
Anion Exchanger ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Disulfonic Acid ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Acid Base ◽  
Acid Base Status

HCO3- secretion by cortical collecting duct (CCD) occurs via beta-intercalated cells. In vitro CCD HCO3- secretion is modulated by both the in vivo acid-base status of the animal and by adenosine 3',5'-cyclic monophosphate (cAMP). To investigate the mechanism of cAMP-induced HCO3- secretion, we measured intracellular pH (pHi) of individual beta-intercalated cells of CCDs dissected from alkali-loaded rabbits perfused in vitro. beta-Intercalated cells were identified by demonstrating the presence of an apical anion exchanger (cell alkalinization in response to removal of lumen Cl-). After 180 min of perfusion to permit decrease of endogenous cAMP, acute addition of 0.1 mM 8-bromo-cAMP or 1 microM isoproterenol to the bath caused a transient cellular alkalinization (> 0.20 pH units). In the symmetrical absence of either Na+, HCO3-, or Cl-, cAMP produced no change in pHi. Basolateral dihydrogen 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (0.1 mM) for 15 min before cAMP addition also prevented this alkalinization. In contrast to the response of cells from alkali-loaded rabbits, addition of basolateral cAMP to CCDs dissected from normal rabbits resulted in an acidification of beta-intercalated cells (approximately 0.20 pH units). The present studies demonstrate the importance of the in vivo acid-base status of the animal in the regulation of CCD HCO3- secretion by beta-intercalated cells. The results identify the possible existence of a previously unrecognized Na(+)-dependent Cl-/HCO3- exchanger on the basolateral membrane of beta-intercalated cells in alkali-loaded rabbits.

scholarly journals Intercalated cell-specific Rh B glycoprotein deletion diminishes renal ammonia excretion response to hypokalemia

2013 ◽  
Vol 304 (4) ◽  
pp. F422-F431 ◽  
Author(s):  
Jesse M. Bishop ◽  
Hyun-Wook Lee ◽  
Mary E. Handlogten ◽  
Ki-Hwan Han ◽  
Jill W. Verlander ◽  
...  
Keyword(s):  
Metabolic Acidosis ◽  
Collecting Duct ◽  
Ammonia Excretion ◽  
Ammonia Concentration ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Intercalated Cell ◽  
Transporter Family ◽  
Total Ammonia ◽  
Normal Increase

The ammonia transporter family member, Rh B Glycoprotein (Rhbg), is an ammonia-specific transporter heavily expressed in the kidney and is necessary for the normal increase in ammonia excretion in response to metabolic acidosis. Hypokalemia is a common clinical condition in which there is increased renal ammonia excretion despite the absence of metabolic acidosis. The purpose of this study was to examine Rhbg's role in this response through the use of mice with intercalated cell-specific Rhbg deletion (IC-Rhbg-KO). Hypokalemia induced by feeding a K+-free diet increased urinary ammonia excretion significantly. In mice with intact Rhbg expression, hypokalemia increased Rhbg protein expression in intercalated cells in the cortical collecting duct (CCD) and in the outer medullary collecting duct (OMCD). Deletion of Rhbg from intercalated cells inhibited hypokalemia-induced changes in urinary total ammonia excretion significantly and completely prevented hypokalemia-induced increases in urinary ammonia concentration, but did not alter urinary pH. We conclude that hypokalemia increases Rhbg expression in intercalated cells in the cortex and outer medulla and that intercalated cell Rhbg expression is necessary for the normal increase in renal ammonia excretion in response to hypokalemia.

Effect of insulin on potassium secretion in rabbit cortical collecting ducts

1992 ◽  
Vol 262 (1) ◽  
pp. F30-F35 ◽  
Author(s):  
H. Furuya ◽  
K. Tabei ◽  
S. Muto ◽  
Y. Asano
Keyword(s):  
Collecting Duct ◽  
Dependent Manner ◽  
Cortical Collecting Duct ◽  
Plasma Insulin Concentration ◽  
Concentration Dependent Manner ◽  
K Secretion ◽  
Potassium Secretion ◽  
Transepithelial Voltage ◽  
K Transport

Insulin is known to play an important role in the regulation of extrarenal K homeostasis. Previous clearance studies have shown that insulin decreases urinary K excretion, but the responsible nephron segments have not been identified. In this microperfusion study, in vitro, the effect of insulin on K transport in the cortical collecting duct (CCD), which is thought to be an important segment for regulation of the final urinary K excretion, was investigated. Basolateral insulin (10(-6) M) significantly inhibited net K secretion by 20% (mean JK = -26.2 +/- 4.2 peq.mm-1.min-1 for controls compared with -21.1 +/- 3.4 with insulin, P less than 0.001) and depolarized the transepithelial voltage (VT, from -14.6 +/- 3.5 to -10.8 +/- 3.5 mV, P less than 0.005), recovery did not occur over 60 min. Insulin (10(-11)-10(-5) M) depressed K secretion and depolarized the VT in a concentration-dependent manner. The half-maximal concentration was 5 x 10(-10) M, which is within the physiological range of plasma insulin concentration. In tubules of deoxycorticosterone acetate-treated rabbits, insulin also produced a significant fall in K secretion (from -43.4 +/- 7.5 to -36.1 +/- 5.7 peq.mm-1.min-1, P less than 0.05). Although luminal Ba (2 mM) decreased K secretion (from -14.4 +/- 2.9 to -7.0 +/- 1.7 peq.mm-1.min-1), basolateral insulin (10(-6) M) inhibited K secretion further (to -4.7 +/- 1.3 peq.mm-1.min-1, P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of AE2 mRNA expression in the cortical collecting duct by acid/base balance

1998 ◽  
Vol 274 (3) ◽  
pp. F596-F601 ◽  
Author(s):  
Géza Fejes-Tóth ◽  
Erzsébet Rusvai ◽  
Emily S. Cleaveland ◽  
Anikó Náray-Fejes-Tóth
Keyword(s):  
Mrna Expression ◽  
Collecting Duct ◽  
Cell Types ◽  
Alkaline Media ◽  
Mrna Levels ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Acid Base Balance ◽  
Acid Base ◽  
Base Balance

AE2 mRNA and protein is expressed in several nephron segments, one of which is the cortical collecting duct (CCD). However, the distribution of AE2 among the different cell types of the CCD and the function of AE2 in the kidney are not known. The purpose of this study was to determine the distribution of AE2 mRNA among the three CCD cell types and to examine the effects of changes in acid/base balance on its expression. Following NH4Cl (acid) or NaHCO3 (base) loading of rabbits for ∼18 h, CCD cells were isolated by immunodissection. AE2 mRNA levels were determined by RT-PCR and were normalized for β-actin levels. We found that CCD cells express high levels of AE2 mRNA (∼500 copies/cell). AE2 mRNA levels were significantly higher in CCD cells originating from base-loaded than acid-loaded rabbits, with an average increase of 3.7 ± 1.07-fold. The effect of pH on AE2 mRNA levels was also tested directly using primary cultures of CCD cells. CCD cells incubated in acidic media expressed significantly lower levels of AE2 mRNA than those in normal or alkaline media. Experiments with isolated principal cells, α-intercalated cells, and β-intercalated cells (separated by fluorescence-activated cell sorting) demonstrated that AE2 mRNA levels are comparable in the three collecting duct cell subtypes and are similarly regulated by changes in acid/base balance. Based on these results, we conclude that adaptation to changes in extracellular H+ concentration is accompanied by opposite changes in AE2 mRNA expression. The observations that AE2 mRNA is not expressed in a cell-type-specific manner and that changes in acid/base balance have similar effects on each CCD cell subtype suggest that AE2 might serve a housekeeping function rather than being the apical anion exchanger of β-intercalated cells.

scholarly journals ENaC inhibition stimulates HCl secretion in the mouse cortical collecting duct. I. Stilbene-sensitive Cl− secretion

2015 ◽  
Vol 309 (3) ◽  
pp. F251-F258 ◽  
Author(s):  
Masayoshi Nanami ◽  
Yoskaly Lazo-Fernandez ◽  
Vladimir Pech ◽  
Jill W. Verlander ◽  
Diana Agazatian ◽  
...  
Keyword(s):  
Channel Blocker ◽  
Single Channel ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Collecting Ducts ◽  
Hcl Secretion ◽  
Transepithelial Voltage ◽  
Apical Membranes ◽  
Rats And Mice

Inhibition of the epithelial Na+ channel (ENaC) reduces Cl− absorption in cortical collecting ducts (CCDs) from aldosterone-treated rats and mice. Since ENaC does not transport Cl−, the purpose of the present study was to explore how ENaC modulates Cl− absorption in mouse CCDs perfused in vitro. Therefore, we measured transepithelial Cl− flux and transepithelial voltage in CCDs perfused in vitro taken from mice that consumed a NaCl-replete diet alone or the diet with aldosterone administered by minipump. We observed that application of an ENaC inhibitor [benzamil (3 μM)] to the luminal fluid unmasks conductive Cl− secretion. During ENaC blockade, this Cl− secretion fell with the application of a nonselective Cl− channel blocker [DIDS (100 μM)] to the perfusate. While single channel recordings of intercalated cell apical membranes in split-open CCDs demonstrated a Cl− channel with properties that resemble the ClC family of Cl− channels, ClC-5 is not the primary pathway for benzamil-sensitive Cl− flux. In conclusion, first, in CCDs from aldosterone-treated mice, most Cl− absorption is benzamil sensitive, and, second, benzamil application stimulates stilbene-sensitive conductive Cl− secretion, which occurs through a ClC-5-independent pathway.

Axial heterogeneity of rabbit cortical collecting duct

1991 ◽  
Vol 260 (4) ◽  
pp. F498-F505
Author(s):  
C. L. Emmons ◽  
K. Matsuzaki ◽  
J. B. Stokes ◽  
V. L. Schuster
Keyword(s):  
Cross Sections ◽  
Cell Function ◽  
Rate Coefficient ◽  
Collecting Duct ◽  
Lectin Binding ◽  
Cell Types ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Principal Cells ◽  
Inner Cortex

The rabbit cortical collecting duct (CCD) consists of three major cell types: principal cells transport K+, beta-intercalated cells absorb Cl-, and alpha-intercalated cells secrete H+. We used functional and histological methods to assess axial distribution of these cell types along rabbit CCD. In perfused CCDs, lumen-to-bath Rb+ rate coefficient (an index of principal cell K+ transport) was not different in tubules from outer cortex (1 mm from renal surface) compared with those from inner cortex (2 mm from renal surface), suggesting that principal cell function is homogeneous along the CCD. In contrast, Cl- rate coefficient (a measure of beta-intercalated cell function) was twice as high in CCDs from outer compared with inner cortex, suggesting heterogeneity of beta-intercalated cells along the CCD. To further investigate these regional differences, we fixed and embedded kidneys and identified three cell types in CCD cross sections using carbonic anhydrase staining and peanut lectin binding. Comparing tubule cross sections from outer with those from inner cortex, we found no axial difference in the fraction of cells that were either principal cells (64%) or total (lectin binding and nonlectin binding) intercalated cells (36%). However, the lectin-binding intercalated cell subset was significantly increased in outer compared with inner cortex. We conclude that there is not heterogeneity of principal cells along the rabbit CCD; however, beta-cell number and function are increased in outer CCD. Collecting duct heterogeneity begins within the cortical segment.

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.

H(+)-K(+)-ATPase activity in rat collecting duct segments

1992 ◽  
Vol 262 (4) ◽  
pp. F692-F695 ◽  
Author(s):  
J. D. Gifford ◽  
L. Rome ◽  
J. H. Galla
Keyword(s):  
Atpase Activity ◽  
Marked Decrease ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Acid Base ◽  
Medullary Collecting Duct ◽  
Gastric Proton Pump

Previous studies have suggested the presence of an H(+)-K(+)-ATPase in rat cortical and medullary intercalated cells with similar properties to the gastric proton pump. The purpose of this study was to determine the functional contribution of an H(+)-K(+)-adenosinetriphosphatase(ATPase) to total CO2 (tCO2) transport along the rat collecting duct. After baseline determination of tCO2 transport in isolated perfused collecting duct segments, Sch 28080 (10 microM) was added to either the perfusate or bath. When Sch 28080 was added to the perfusate, there was no effect in the cortical collecting duct (CCD, 20.8 +/- 6.7 vs. 25.3 + 3.0 pmol.mm-1.min-1), but a marked decrease in tCO2 absorption was effected in both the outer medullary (OMCD, 37.6 + 6.2 vs. 10.7 +/- 4.1 pmol.mm-1.min-1) and initial inner medullary collecting duct (IMCD1, 34.4 +/- 8.1 vs. 16.2 +/- 5.6 pmol.mm-1.min-1). In the CCD from rats with acute alkalosis in vivo, Sch 28080 added to the bath inhibited tCO2 secretion in the CCD (-17.1 +/- 4.4 vs 3.5 + 3.3 pmol.mm-1.min-1). These findings suggest that 1) H(+)-K(+)-ATPase is important in tCO2 absorption in the OMCD and IMCD1 and in tCO2 secretion in the CCD, 2) HCO3(-)-absorbing intercalated cells differ functionally in the cortex and medulla, 3) HCO3- secretion is not the reverse process of HCO3- absorption in the CCD, and 4) H(+)-K(+)-ATPase is important in distal acidification under normal and altered acid-base conditions.

Sign in / Sign up

Export Citation Format

Share Document