Effects of luminal flow and nucleotides on [Ca2+]i in rabbit cortical collecting duct

2002 ◽  
Vol 283 (3) ◽  
pp. F437-F446 ◽  
Author(s):  
Craig B. Woda ◽  
Maurilo Leite ◽  
Rajeev Rohatgi ◽  
Lisa M. Satlin
Keyword(s):  
Flow Rate ◽  
Purinergic Receptor ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cell Types ◽  
P2 Receptors ◽  
Receptor Subtypes ◽  
Cortical Collecting Duct ◽  
Luminal Perfusion

Nucleotide binding to purinergic P2 receptors contributes to the regulation of a variety of physiological functions in renal epithelial cells. Whereas P2 receptors have been functionally identified at the basolateral membrane of the cortical collecting duct (CCD), a final regulatory site of urinary Na+, K+, and acid-base excretion, controversy exists as to whether apical purinoceptors exist in this segment. Nor has the distribution of receptor subtypes present on the unique cell populations that constitute Ca2+ the CCD been established. To examine this, we measured nucleotide-induced changes in intracellular Ca2+ concentration ([Ca2+]i) in fura 2-loaded rabbit CCDs microperfused in vitro. Resting [Ca2+]i did not differ between principal and intercalated cells, averaging ∼120 nM. An acute increase in tubular fluid flow rate, associated with a 20% increase in tubular diameter, led to increases in [Ca2+]i in both cell types. Luminal perfusion of 100 μM UTP or ATP-γ-S, in the absence of change in flow rate, caused a rapid and transient approximately fourfold increase in [Ca2+]i in both cell types ( P< 0.05). Luminal suramin, a nonspecific P2 receptor antagonist, blocked the nucleotide- but not flow-induced [Ca2+]i transients. Luminal perfusion with a P2X (α,β-methylene-ATP), P2X7 (benzoyl-benzoyl-ATP), P2Y1 (2-methylthio-ATP), or P2Y4/P2Y6 (UDP) receptor agonist had no effect on [Ca2+]i. The nucleotide-induced [Ca2+]i transients were inhibited by the inositol-1,4,5-triphosphate receptor blocker 2-aminoethoxydiphenyl borate, thapsigargin, which depletes internal Ca2+ stores, luminal perfusion with a Ca2+-free perfusate, or the L-type Ca2+ channel blocker nifedipine. These results suggest that luminal nucleotides activate apical P2Y2 receptors in the CCD via pathways that require both internal Ca2+mobilization and extracellular Ca2+ entry. The flow-induced rise in [Ca2+]i is apparently not mediated by apical P2 purinergic receptor signaling.

Functional characterization of alpha- and beta-intercalated cell types in rabbit cortical collecting duct

1991 ◽  
Vol 261 (3) ◽  
pp. F377-F385 ◽  
Author(s):  
H. Furuya ◽  
M. D. Breyer ◽  
H. R. Jacobson
Keyword(s):  
Collecting Duct ◽  
Basolateral Membrane ◽  
Functional Characterization ◽  
Cell Types ◽  
Disulfonic Acid ◽  
Cortical Collecting Duct ◽  
Electrical Measurements ◽  
Collecting Ducts

Single-cell electrical measurements and spectrophotometric determinations of intracellular pH were used to determine unique features of alpha- and beta-intercalated cells (alpha-IC, beta-IC) in in vitro perfused rabbit cortical collecting ducts (CCD). pHi rose in alpha-IC and fell in beta-IC after bath Cl- removal. Luminal Cl- removal did not change pHi of alpha-IC, but pHi of beta-IC rose by 0.36 +/- 0.01 pH units. Cl- concentration-dependent recovery of beta-IC pHi revealed a Cl- Km of 18.7 mM for the luminal Cl(-) -HCO3- exchanger. Measurements of basolateral membrane voltage (Vbl) also showed two IC cell types. Removal of luminal Cl- did not change Vbl in alpha-IC, whereas Vbl hyperpolarized by a mean of 73.2 +/- 3.5 mV in beta-IC. Reducing bath Cl- depolarized both alpha- and beta-IC Vbl. In alpha-IC a large repolarization of 39.8 +/- 5.2 mV followed acute depolarization after bath Cl- removal. Reducing bath HCO3- (constant CO2) had little effect on beta-IC Vbl, whereas alpha-IC Vbl depolarized by 5.2 +/- 0.7 mV. Reducing luminal HCO3- in the absence of luminal Cl- produced a 17.6 +/- 1.8 mV depolarization in beta-IC. This change was independent of luminal Na+ and was not blocked by luminal 10(-4) M 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). In beta-IC, Vbl was not altered by either bath or lumen DIDS in the presence of luminal Cl-. However, when luminal Cl- was removed, luminal DIDS reversibly depolarized Vbl by 9.6 +/- 2.9 mV.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of flow and stretch on the [Ca2+]i response of principal and intercalated cells in cortical collecting duct

2003 ◽  
Vol 285 (5) ◽  
pp. F998-F1012 ◽  
Author(s):  
Wen Liu ◽  
Shiyun Xu ◽  
Craig Woda ◽  
Paul Kim ◽  
Sheldon Weinbaum ◽  
...  
Keyword(s):  
Flow Rate ◽  
Collecting Duct ◽  
Cell Types ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Duration Magnitude ◽  
Luminal Flow ◽  
Principal And Intercalated Cells ◽  
Circumferential Stretch

An acute increase in tubular fluid flow rate in the microperfused cortical collecting duct (CCD), associated with a ∼20% increase in tubular diameter, leads to an increase in intracellular Ca2+ concentration ([Ca2+]i)in both principal and intercalated cells (Woda CB, Leite M Jr, Rohatgi R, and Satlin LM. Am J Physiol Renal Physiol 283: F437-F446, 2002). The apical cilium present in principal but not intercalated cells has been proposed to be a flow sensor. To determine whether flow across the cilium and/or epithelial stretch mediates the [Ca2+]i response, CCDs from New Zealand White rabbits were microperfused in vitro, split-open (to isolate the effect of flow across cilia), or occluded (to examine the effect of stretch and duration/magnitude of the flow impulse), and [Ca2+]i was measured using fura 2. In perfused and occluded CCDs, a rapid (<1 s) but not slow (>3 min) increase in luminal flow rate and/or circumferential stretch led to an approximately threefold increase in [Ca2+]i in both principal and intercalated cells within ∼10 s. This response was mediated by external Ca2+ entry and inositol 1,4,5-trisphosphate-mediated release of cell Ca2+ stores. In split-open CCDs, an increase in superfusate flow led to an approximately twofold increase in [Ca2+]i in both cell types within ∼30 s. These experimental findings are interpreted using mathematical models to predict the fluid stress on the apical membranes of the CCD and the forces and torques on and deformation of the cilia. We conclude that rapid increases in luminal flow rate and circumferential stretch, leading to shear or hydrodynamic impulses at the cilium or apical membrane, lead to increases in [Ca2+]i in both principal and intercalated cells.

Phenotypic plasticity in the intercalated cell: the hensin pathway

1998 ◽  
Vol 275 (2) ◽  
pp. F183-F190 ◽  
Author(s):  
Qais Al-Awqati ◽  
S. Vijayakumar ◽  
C. Hikita ◽  
J. Chen ◽  
J. Takito
Keyword(s):  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cell Types ◽  
Band 3 ◽  
Cytoskeletal Proteins ◽  
Β Cell ◽  
Biochemical Basis ◽  
Intercalated Cell ◽  
Immortalized Cell

The collecting duct of the renal tubule contains two cell types, one of which, the intercalated cell, is responsible for acidification and alkalinization of urine. These cells exist in a multiplicity of morphological forms, with two extreme types, α and β. The former acidifies the urine by an apical proton-translocating ATPase and a basolateral Cl/HCO3 exchanger, which is an alternately spliced form of band 3. This kidney form of band 3, kAE1, is present in the apical membrane of the β-cell, which has the H+-ATPase on the basolateral membrane. We had suggested previously that metabolic acidosis leads to conversion of β-types to α-types. To study the biochemical basis of this plasticity, we used an immortalized cell line of the β-cell and showed that these cells convert to the α-phenotype when plated at superconfluent density. At high density these cells localize a new protein, which we term “hensin,” to the extracellular matrix, and hensin acts as a molecular switch capable of changing the phenotype of these cells in vitro. Hensin induces new cytoskeletal proteins, makes the cells assume a more columnar shape and retargets kAE1 and the H+-ATPase. These recent studies suggest that the conversion of β- to α-cells, at least in vitro, bears many of the hallmarks of terminal differentiation.

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.

Interaction of Cl- and other halogens with Cl- transport systems in rabbit cortical collecting duct

1992 ◽  
Vol 263 (5) ◽  
pp. F870-F877 ◽  
Author(s):  
S. Muto ◽  
M. Imai ◽  
Y. Asano
Keyword(s):  
Apical Membrane ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cell Types ◽  
Transport Systems ◽  
Cortical Collecting Duct ◽  
Cl Transport ◽  
Distinct Cell ◽  
Cl Conductance ◽  
Basolateral Membrane Potential

We have reported that in the rabbit cortical collecting duct (CCD) we can identify electrophysiologically three distinct cell types; the collecting duct (CD) cell and the alpha- and beta-intercalated (IC) cell. To further characterize the Cl- transport properties of each cell type, we examined the interaction between Cl- and other halogens or SCN- in the isolated and perfused CCD by intracellular microelectrode impalement. The rapid depolarization of the basolateral membrane potential (VB) caused by replacement of bath Cl- with each anion revealed that the sequences of apparent halogen selectivity for the basolateral Cl- conductance were similar in all three cell types. The ranking of Cl- > Br- > F- > I- corresponds to the sequence 5 of Eisenman's series, indicating “strong” interaction of the anions with the selectivity site. The basolateral Cl- conductance of these three cell types may share common characteristics, although I- permeability is less in IC cells than in CD cells. Hyperpolarization of the basolateral membrane of the beta-IC cell upon reduction of luminal Cl- reflects alterations in either Cl- entry across the apical membrane, or Cl- exit across the basolateral membrane, or both. Luminal Cl- replacement with each anion showed that the sequence of the hyperpolarization of the basolateral membrane was I- >> cyclamate = SCN- > F- > Br-, suggesting that I-inhibits either apical Cl- entry or basolateral Cl- exit. On the other hand, in the CD cell reduction of the perfusate Cl- by replacement with each anion caused the basolateral membrane to hyperpolarize with a different ranking: cyclamate = F- > I- = SCN- > Br-.(ABSTRACT TRUNCATED AT 250 WORDS)

Postnatal maturation of potassium transport in rabbit cortical collecting duct

1994 ◽  
Vol 266 (1) ◽  
pp. F57-F65 ◽  
Author(s):  
L. M. Satlin
Keyword(s):  
Flow Rate ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Flow Rates ◽  
Postnatal Maturation ◽  
Significant Linear Correlation ◽  
Potassium Secretion ◽  
Tubular Flow ◽  
Neonatal Kidney

Clearance studies in newborns demonstrate low rates of urinary excretion of potassium, suggesting that the neonatal kidney contributes to the conservation of potassium necessary for growth. Because the cortical collecting duct (CCD) is a primary site for potassium secretion in the adult, we sought to examine the transport capacity of this segment for potassium during postnatal maturation. CCDs isolated from rabbits of various ages (5-6 animals/age group) were microperfused in vitro with solutions simulating plasma. The concentrations of potassium in samples of collected fluid, measured by helium glow photometry, were used to calculate net transport. At a flow rate of approximately 1.6 nl.min-1 x mm-1 net potassium secretion was absent at birth, first became evident at 4 wk of age (-11.08 +/- 2.39 pmol.min-1 x mm-1), and increased sharply thereafter to reach mature rates (-23.08 +/- 3.47 pmol.min-1 x mm-1; P < 0.05) by 6 wk of age. To determine whether low distal tubular flow rates limit net potassium secretion in the neonate, we perfused CCDs at two or more flow rates in the 0.5–5 nl.min-1 x mm-1 range. In CCDs taken from animals > or = 6 wk of age, potassium secretion showed a significant linear correlation with flow rate (y = -10.0x - 7.45; r = 0.87; n = 12).(ABSTRACT TRUNCATED AT 250 WORDS)

Endothelin and nitric oxide mediate adaptation of the cortical collecting duct to metabolic acidosis

2006 ◽  
Vol 291 (4) ◽  
pp. F866-F873 ◽  
Author(s):  
Shuichi Tsuruoka ◽  
Seiji Watanabe ◽  
Jeffrey M. Purkerson ◽  
Akio Fujimura ◽  
George J. Schwartz
Keyword(s):  
Nitric Oxide ◽  
Metabolic Acidosis ◽  
Receptor Antagonist ◽  
Guanylate Cyclase ◽  
Kinase Inhibitor ◽  
Collecting Duct ◽  
Bicarbonate Transport ◽  
Receptor Subtypes ◽  
Cortical Collecting Duct

Endothelin (ET) and nitric oxide (NO) modulate ion transport in the kidney. In this study, we defined the function of ET receptor subtypes and the NO guanylate cyclase signaling pathway in mediating the adaptation of the rabbit cortical collecting duct (CCD) to metabolic acidosis. CCDs were perfused in vitro and incubated for 3 h at pH 6.8, and bicarbonate transport or cell pH was measured before and after acid incubation. Luminal chloride was reversibly removed to isolate H+ and HCO3− secretory fluxes and to raise the pH of β-intercalated cells. Acid incubation caused reversal of polarity of net HCO3− transport from secretion to absorption, comprised of a 40% increase in H+ secretion and a 75% decrease in HCO3− secretion. The ETB receptor antagonist BQ-788, as well as the NO synthase inhibitor, NG-nitro-l-arginine methyl ester (l-NAME), attenuated the adaptive decrease in HCO3− secretion by 40%, but only BQ-788 inhibited the adaptive increase in H+ secretion. There was no effect of inactive d-NAME or the ETA receptor antagonist BQ-123. Both BQ-788 and l-NAME inhibited the acid-induced inactivation (endocytosis) of the apical Cl−/HCO3− exchanger. The guanylate cyclase inhibitor LY-83583 and cGMP-dependent protein kinase inhibitor KT-5823 affected HCO3− transport similarly to l-NAME. These data indicate that signaling via the ETB receptor regulates the adaptation of the CCD to metabolic acidosis and that the NO guanylate cyclase component of ETB receptor signaling mediates downregulation of Cl−/HCO3− exchange and HCO3− secretion.

Similar chloride channels in the connecting tubule and cortical collecting duct of the mouse kidney

2006 ◽  
Vol 290 (6) ◽  
pp. F1421-F1429 ◽  
Author(s):  
Antoine Nissant ◽  
Marc Paulais ◽  
Sahran Lachheb ◽  
Stéphane Lourdel ◽  
Jacques Teulon
Keyword(s):  
Chloride Channels ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Reversal Potential ◽  
Cell Types ◽  
Cortical Collecting Duct ◽  
Patch Clamp Technique ◽  
Connecting Tubule ◽  
Recording Conditions ◽  
Rich Solution

Using the patch-clamp technique, we investigated Cl− channels on the basolateral membrane of the connecting tubule (CNT) and cortical collecting duct (CCD). We found a ∼10-pS channel in CNT cell-attached patches. Substitution of sodium gluconate for NaCl in the pipette shifted the reversal potential by +25 mV, whereas N-methyl-d-gluconate chloride had no effect, indicating anion selectivity. On inside-out patches, we determined a selectivity sequence of Cl− > Br− ∼ NO3− > F−, which is compatible with that of ClC-K2, a Cl− channel in the distal nephron. In addition, the number of open channels ( NPo) measured in cell-attached patches was significantly increased when Ca2+ concentration or pH in the pipette was increased, which is another characteristic of ClC-K. These findings suggest that the basis for this channel is ClC-K2. A similar Cl− channel was found in CCD patches. Because CNT and CCD are heterogeneous tissues, we studied the cellular distribution of the Cl− channel using recording conditions (KCl-rich solution in the pipette) that allowed us to detect simultaneously Cl− channels and inwardly rectifying K+ channels. We detected Cl− channels alone in 45% and 42% and K+ channels alone in 51% and 58% of CNT and CCD patches, respectively. Cl− and K+ channels were recorded simultaneously from two patches (4% of patches) in the CNT and from none of the patches in the CCD. This indicates that Cl− and K+ channels are located in different cell types, which we suggest may be the intercalated cells and principal cells, respectively.

scholarly journals TRPV4 as a flow sensor in flow-dependent K+ secretion from the cortical collecting duct

2007 ◽  
Vol 292 (2) ◽  
pp. F667-F673 ◽  
Author(s):  
Junichi Taniguchi ◽  
Shuichi Tsuruoka ◽  
Atsuko Mizuno ◽  
Jun-ichi Sato ◽  
Akio Fujimura ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Basolateral Membrane ◽  
Cortical Collecting Duct ◽  
Urine Production ◽  
K Secretion ◽  
Distal Tubules ◽  
Transient Receptor

The transient receptor vanilloid-4 (TRPV4) is a mechanosensitive, swell-activated cation channel that is abundant in the renal distal tubules. Immunolocalization studies, however, present conflicting data as to whether TRPV4 is expressed along the apical and/or basolateral membranes. To disclose the role of TRPV4 in flow-dependent K+ secretion in distal tubules in vivo, urinary K+ excretion and net transports of K+ and Na+ in the cortical collecting duct (CCD) were measured with an in vitro microperfusion technique in TRPV4 +/+ and TRPV4 −/− mice. Both net K+ secretion and Na+ reabsorption were flow dependently increased in the CCDs isolated from TRPV4 +/+mice, which were significantly enhanced by a luminal application of 50 μM 4α-phorbol-12,13-didecanoate (4αPDD), an agonist of TRPV4. No flow dependence of net K+ and Na+ transports or effects of 4αPDD on CCDs were observed in TRPV4 −/− mice. A basolateral application of 4αPDD had little effect on these ion transports in the TRPV4 +/+ CCDs, while the luminal application did. Urinary K+ excretion was significantly smaller in TRPV4 −/− than in TRPV4 +/+ mice when urine production was stimulated by a venous application of furosemide. These observations suggested an essential role of the TRPV4 channels in the luminal or basolateral membrane as flow sensors in the mechanism underlying the flow-dependent K+ secretion in mouse CCDs.

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.

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