Hormone-stimulated Ca2+transport in rabbit kidney:  multiple sites of inhibition by exogenous ATP

Exogenous ATP markedly reduced 1-desamino-8-d-arginine vasopressin (dDAVP)-stimulated Ca2+ transport and cAMP accumulation in primary cultures of rabbit connecting tubule and cortical collecting duct cells. Similarly, ATP inhibited the stimulatory effect of 8-bromo-cAMP. At first sight, this is in agreement with the “classic” concept that dDAVP exerts its stimulatory effect via cAMP. However, dDAVP-stimulated Ca2+ transport was markedly reduced by the protein kinase C (PKC) inhibitor chelerythrine, reported previously to inhibit the cAMP-independent pathway responsible for parathyroid hormone-, [Arg8]vasopressin-, PGE2-, and adenosine-stimulated Ca2+ transport. Chelerythrine also inhibited the increase in Ca2+transport evoked by the cAMP-independent A1 receptor agonist N 6-cyclopentyladenosine (CPA). Downregulation of phorbol ester-sensitive PKC isoforms by chronic phorbol ester treatment has been shown before to be without effect on hormone-stimulated Ca2+transport, indicating that the chelerythrine-inhibitable pathway consists of a phorbol ester-insensitive PKC isoform. Here, this maneuver did not affect ATP inhibition of dDAVP-stimulated Ca2+ transport and cAMP formation, while abolishing ATP inhibition of CPA-stimulated Ca2+ transport. These findings show that ATP acts via 1) a phorbol ester-sensitive PKC isoform to inhibit hormonal stimulation of Ca2+ transport at the level of the chelerythrine-inhibitable pathway involving a phorbol ester-insensitive PKC isoform and 2) a phorbol ester-insensitive mechanism to inhibit V2 receptor-mediated concomitant activation of this pathway and adenylyl cyclase.

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Adenosine-stimulated Ca2+reabsorption is mediated by apical A1 receptors in rabbit cortical collecting system

AJP Renal Physiology ◽

10.1152/ajprenal.1998.274.4.f736 ◽

1998 ◽

Vol 274 (4) ◽

pp. F736-F743 ◽

Cited By ~ 12

Author(s):

Joost G. J. Hoenderop ◽

Anita Hartog ◽

Peter H. G. M. Willems ◽

René J. M. Bindels

Keyword(s):

Collecting Duct ◽

Cortical Collecting Duct ◽

Concomitant Increase ◽

Cgs 21680 ◽

Kinase C ◽

Intracellular Ca ◽

Collecting Duct Cells ◽

Camp Formation ◽

Pkc Activation ◽

Permeable Supports

Confluent monolayers of immunodissected rabbit connecting tubule and cortical collecting duct cells, cultured on permeable supports, were used to study the effect of adenosine on net apical-to-basolateral Ca2+ transport. Apical, but not basolateral, adenosine increased this transport dose dependently from 48 ± 3 to 110 ± 4 nmol ⋅ h−1 ⋅ cm−2. Although a concomitant increase in cAMP formation suggested the involvement of an A2 receptor, the A2 agonist CGS-21680 did not stimulate Ca2+ transport, while readily increasing cAMP. By contrast, the A1 agonist N 6-cyclopentyladenosine (CPA) maximally stimulated Ca2+transport without significantly affecting cAMP. Adenosine-stimulated transport was effectively inhibited by the A1 antagonist 1,3-dipropyl-8-cyclopenthylxanthine but not the A2 antagonist 3,7-dimethyl-1-propargylxanthine, providing additional evidence for the involvement of an A1 receptor. Both abolishment of the adenosine-induced transient increase in intracellular Ca2+ concentration by 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid and downregulation of protein kinase C (PKC) by prolonged phorbol ester treatment were without effect on adenosine-stimulated Ca2+ transport. The data presented suggest that adenosine interacts with an apical A1 receptor to stimulate Ca2+ transport via a hitherto unknown pathway that does not involve cAMP formation, PKC activation, and/or Ca2+ mobilization.

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Inhibition of Na+ and Ca2+ reabsorption by P2u purinoceptors requires PKC but not Ca2+ signaling

AJP Renal Physiology ◽

10.1152/ajprenal.1996.270.1.f53 ◽

1996 ◽

Vol 270 (1) ◽

pp. F53-F60 ◽

Cited By ~ 24

Author(s):

H. P. Koster ◽

A. Hartog ◽

C. H. van Os ◽

R. J. Bindels

Keyword(s):

Inhibitory Concentration ◽

Purinergic Receptor ◽

Rank Order ◽

Collecting Duct ◽

Cortical Collecting Duct ◽

Inhibitory Effects ◽

Maximal Inhibition ◽

Kinase C ◽

Collecting Duct Cells ◽

Dose Dependent

Rabbit connecting tubule and cortical collecting duct cells were isolated by immunodissection and cultured to confluence on permeable filters and on glass coverslips. Extracellular ATP dose-dependently reduced transcellular Na+ and Ca2+ transport (half-maximal inhibitory concentration, IC50, of 0.5 +/- 0.2 and 3.2 +/- 0.5 microM), with a maximal inhibition of 57 +/- 5 and 43 +/- 4%, respectively. Purinergic receptor agonists inhibited transport with the following rank order of potency: UTP = ATP > ADP; this suggests involvement of P2u purinoceptors. ATP also caused a dose-dependent (50% effective dose, EC50, of 1.5 +/- 0.2 microM) transient increase in intracellular Ca2+ concentration ([Ca2+]i), which decreased to a sustained elevated level. In the absence of extracellular Ca2+, a similar Ca2+ transient occurred, but the sustained response was abolished. Preloading the cells with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) completely prevented the ATP-induced Ca2+ transients, but not the ATP-induced inhibition of Na+ and Ca2+ absorption. Activation of protein kinase C (PKC) by the cell-permeable diacylglycerol analogue, 1,2-dioctanoyl-en-glycerol, mimicked ATP-induced inhibition of Na+ and Ca2+ absorption. The inhibitory effects of ATP were no longer observed after culturing cells in the presence of phorbol ester (12-O-tetradecanoylphorbol-13-acetate) for 5 days, which resulted in downregulation of cellular PKC activity.

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Localization of IP in rabbit kidney and functional role of the PGI2/IP system in cortical collecting duct

AJP Renal Physiology ◽

10.1152/ajprenal.00020.2002 ◽

2002 ◽

Vol 283 (4) ◽

pp. F689-F698 ◽

Cited By ~ 7

Author(s):

Rania Nasrallah ◽

Rolf M. Nusing ◽

Richard L. Hébert

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Functional Role ◽

Collecting Duct ◽

Inhibitory Effect ◽

Rabbit Kidney ◽

Cortical Collecting Duct ◽

Kinase C ◽

Protein Kinase C Inhibitor

To clarify the role of the PGI2/PGI2 receptor (IP) system in rabbit cortical collecting duct (RCCD), we characterized the expression of IP receptors in the rabbit kidney. We show by Northern and Western blotting that IP mRNA and protein was detectable in all three regions of the kidney. To determine how PGI2 signals, we compared the effects of different PGI2 analogs [iloprost (ILP), carba-prostacyclin (c-PGI2), and cicaprost (CCP)] in the isolated perfused RCCD. PGI2 analogs did not increase water flow ( L p). Although PGI2 analogs did not reduce an established L p response to 8-chlorophenylthio-cAMP, they equipotently inhibited AVP-stimulated L p by 45%. The inhibitory effect of ILP and c-PGI2 on AVP-stimulated L p is partially reversed by the protein kinase C inhibitor staurosporine and abolished by pertussis toxin; no effect was obtained with CCP. In fura 2-loaded RCCD, CCP did not alter cytosolic Ca2+concentration ([Ca2+]i), but, in the presence of CCP, individual infusion of ILP and PGE2 increased [Ca2+]i, suggesting that CCP did not cause desensitization to either ILP or PGE2. We concluded that ILP and c-PGI2 activate PKC and the liberation of [Ca2+]i but not CCP. This suggested an important role for phosphatidylinositol hydrolysis in mediating ILP and c-PGI2 effects but not CCP in RCCD.

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Reinterpretation of the RACTK1 K+ channel

AJP Cell Physiology ◽

10.1152/ajpcell.1997.272.1.c350 ◽

1997 ◽

Vol 272 (1) ◽

pp. C350-C354 ◽

Cited By ~ 15

Author(s):

B. Shmukler ◽

T. Sun ◽

C. Brugnara ◽

S. L. Alper

Keyword(s):

Collecting Duct ◽

Rabbit Kidney ◽

Antisense Strand ◽

K Channel ◽

Apical Surface ◽

Cortical Collecting Duct ◽

Channel Activity ◽

Reading Frame ◽

E Coli ◽

Collecting Duct Cells

The RACTK1 cDNA cloned from rabbit kidney cortical collecting duct cells was associated with inwardly rectifying pH-regulated K+ channel activity (M. Suzuki, K. Takahashi, M. [keda, H Hayakawa, A. Ogawa, Y. Kawaguchi, and O. Sakai. Nature Lond. 367: 642-645, 1994). The deduced amino acid sequence of the encoded novel polypeptide lacked the signature sequence of a K(+)-selective pore region but predicted a topography suggestive of the inward rectifier K+ channel family. In subsequent articles a RACTK1 epitope was immunolocalized to the apical surface of kidney collecting duct and to arteriolar smooth muscle [M. Suzuki, T. Takigawa, K. Kimura, C. Koseki, and M. Imai. Am. J. Physiol. 269 (Cell Physiol, 38): C496-C503, 1995], and apamin-sensitive K+ currents displaying Ca(2+)-dependent and voltage-independent activation accompanied stable heterologous overexpression of RACTK1 [M. Suzuki, M. Murata, M. Ikeda, T. Miyoshi, and M. Imai. Am. J. Physiol. 270 (Cell Physiol, 39): C964-C968, 1996]. We now report that the "RACTK1" open reading frame is a frame-shifted translation of the antisense strand of an Escherichia coli gene member of a coenzyme A transferase gene family. "RACTK1" mRNA was absent from tissues free of E. coli contamination, and the "RACTK1" gene was undetectable in Southern blots of human and rabbit genomic DNA. We conclude that the immunostaining patterns and Ca(2+)-activated K+ channel activity heretofore attributed to RACTK1 must be otherwise explained.

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Ontogeny of apical membrane ion conductances and channel expression in cortical collecting duct cells

AJP Renal Physiology ◽

10.1152/ajprenal.1996.271.3.f698 ◽

1996 ◽

Vol 271 (3) ◽

pp. F698-F708 ◽

Cited By ~ 2

Author(s):

S. M. Huber ◽

M. F. Horster

Keyword(s):

Apical Membrane ◽

Collecting Duct ◽

Primary Cultures ◽

Specific Conductance ◽

Disulfonic Acid ◽

Cortical Collecting Duct ◽

Ureteric Bud ◽

Developmental States ◽

Channel Expression ◽

Collecting Duct Cells

Ontogeny of apical-basolateral epithelial cell polarity was studied by a comparison of embryonic ureteric bud (UB) and mature cortical collecting duct (CCD) primary cultures. Patch-clamp techniques were applied to identify apical ion channels and to measure specific conductance (in nS/10 pF) of the apical membrane (ga) and of the whole cell (Gw). In UB, Gw (1.3 +/- 0.17; n = 32) and ga (1.4 +/- 0.15; n = 9) were not different, whereas they differed greatly in CCD (Gw: 0.79 +/- 0.07, n = 35; ga: 2.3 +/- 0.38, n = 9). In UB, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) (100 microM) similarly reduced ga and Gw by 0.66 +/- 0.21 (n = 9) and 0.48 +/- 0.07 (n = 7), respectively, whereas DIDS had no effect in CCD. Amiloride (1 mM) did not alter UB conductance, whereas, in CCD, ga was more reduced than Gw (0.75 +/- 0.18, n = 8, vs. 0.22 +/- 0.10, n = 7). In addition to this Na(+)-selective conductance, a nonselective cation (NSC) conductance was enriched sixfold in the apical membrane of CCD, possibly due to apical 9 +/- 1-pS (n = 7) NSC channels, whereas UB apical NSC conductance was low. Apical 16 +/- 2-pS (n = 6) K+ channels were identified in CCD but not in UB. The comparison of both developmental states suggests a differentiation of the apical membrane during UB to CCD ontogeny.

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Vasopressin activates a chloride conductance in cultured cortical collecting duct cells

AJP Renal Physiology ◽

10.1152/ajprenal.1994.267.5.f831 ◽

1994 ◽

Vol 267 (5) ◽

pp. F831-F838 ◽

Cited By ~ 4

Author(s):

E. Nagy ◽

A. Naray-Fejes-Toth ◽

G. Fejes-Toth

Keyword(s):

Collecting Duct ◽

Primary Cultures ◽

The Other ◽

Disulfonic Acid ◽

Maximal Response ◽

Cortical Collecting Duct ◽

Pronounced Increase ◽

Other Hand ◽

Collecting Duct Cells ◽

Dose Dependent Increase

In rabbit cortical collecting duct (CCD) cells, arginine vasopressin (AVP) causes a transient increase followed by a sustained depression of transepithelial potential difference (PDte). Mechanisms underlying the decrease in PDte are not well understood. In this study, we used primary cultures of rabbit CCD cells to study effects of AVP. Basolateral addition of AVP caused a dose-dependent increase in transepithelial conductance (Gte) and a corresponding decrease in PDte. A significant effect was observed at 1 pM AVP, and half-maximal response occurred at 30 pM AVP; 1 nM AVP increased Gte and decreased PDte. Replacement of apical Na+ with N-methyl-D-glucamine did not prevent the effect of AVP on Gte, suggesting that it is not mediated by an increase in apical Na+ conductance. Similarly, apical Ba2+ (1 mM) or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 0.1 mM) failed to prevent the effect of AVP. On the other hand, 5-nitro-2(3-phenylpropylamino)benzoic acid (0.1 mM) caused partial inhibition, whereas substitution of apical Cl- with gluconate or cyclamate almost completely prevented the AVP-induced increase in Gte. In unidirectional ion-flux studies, 1 nM AVP caused only a modest increase in apical-to-basolateral (A-->BL) flux of 22Na and had no effect on transepithelial flux of 86Rb in either direction. On the other hand, AVP caused a pronounced increase in A-->BL flux and BL-->A flux of 36Cl, resulting in an increased net Cl- absorption. The effect of AVP on Gte could be mimicked by 8-bromo-adenosine 3',5'-cyclic monophosphate (8-bromo-cAMP) and isoproterenol, and effects of AVP and isoproterenol were not additive.(ABSTRACT TRUNCATED AT 250 WORDS)

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Anti-Malignant Ascites Effect of Total Diterpenoids from Euphorbiae ebracteolatae Radix Is Attributable to Alterations of Aquaporins via Inhibiting PKC Activity in the Kidney

Molecules ◽

10.3390/molecules26040942 ◽

2021 ◽

Vol 26 (4) ◽

pp. 942

Author(s):

Yuanbin Zhang ◽

Dongfang Liu ◽

Fan Xue ◽

Hongli Yu ◽

Hao Wu ◽

...

Keyword(s):

Protein Kinase C ◽

Antitumor Activity ◽

Membrane Fraction ◽

Collecting Duct ◽

Human Kidney ◽

Underlying Mechanism ◽

Malignant Ascites ◽

Kinase C ◽

Collecting Duct Cells ◽

Medullary Collecting Duct

This study evaluated the anti-ascites effect of total diterpenoids extracted from Euphorbiae ebracteolatae Radix (TDEE) on malignant ascitic mice and elucidated its underlying mechanism. TDEE was extracted by dichloromethane and subjected to column chromatography. The purity of six diterpenoids isolated from TDEE was determined to be 77.18% by HPLC. TDEE (3 and 0.6 g raw herbs/kg, p.o.) reduced ascites and increased urine output. Meanwhile, analysis of tumor cell viability, cycle and apoptosis indicated that TDEE had no antitumor activity. In addition, the expression levels of aquaporins (AQPs) and the membrane translocation levels of protein kinase C (PKC) α and PKCβ in kidney and cells were measured. TDEE reduced the levels of AQP1–4, and inhibited PKCβ expression in membrane fraction. Four main diterpenoids, except compound 2, reduced AQP1 level in human kidney-2 cells. Compounds 4 and 5 inhibited AQP2–4 expression in murine inner medullary collecting duct cells. The diterpenoid-induced inhibition of AQP1–4 expression was blocked by phorbol-12-myristate-13-acetate (PMA; agonist of PKC). The diterpenoids from TDEE are the main anti-ascites components. The anti-ascites effect of diterpenoids may be associated with alterations in AQPs in the kidneys to promote diuresis. The inhibition of AQP1–4 expression by TDEE is related to the inhibition of PKCβ activation.

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Whole cell and unitary amiloride-sensitive sodium currents in M-1 mouse cortical collecting duct cells

AJP Cell Physiology ◽

10.1152/ajpcell.1996.270.4.c998 ◽

1996 ◽

Vol 270 (4) ◽

pp. C998-C1010 ◽

Cited By ~ 7

Author(s):

M. L. Chalfant ◽

T. G. O'Brien ◽

M. M. Civan

Keyword(s):

Collecting Duct ◽

Cell Permeability ◽

Na Channel ◽

Na Channels ◽

Cortical Collecting Duct ◽

Ionic Selectivity ◽

Whole Cell ◽

Duct Cells ◽

Excised Patches ◽

Collecting Duct Cells

Amiloride-sensitive whole cell currents have been reported in M-1 mouse cortical collecting duct cells (Korbmacher et al., J. Gen. Physiol. 102: 761-793, 1993). We have confirmed that amiloride inhibits the whole cell currents but not necessarily the measured whole cell currents. Anomalous responses were eliminated by removing external Na+ and/or introducing paraepithelial shunts. The amiloride-sensitive whole cell currents displayed Goldman rectification. The ionic selectivity sequence of the amiloride-sensitive conductance was Li+ > Na+ >> K+. Growth of M-1 cells on permeable supports increased the amiloride-sensitive whole cell permeability, compared with cells grown on plastic. Single amiloride-sensitive channels were observed, which conformed to the highly selective low-conductance amiloride-sensitive class [Na(5)] of epithelial Na+ channels. Hypotonic pretreatment markedly slowed run-down of channel activity. The gating of the M-1 Na+ channel in excised patches was complex. Open- and closed-state dwell-time distributions from patches that display one operative channel were best described with two or more exponential terms each. We conclude that 1) study of M-1 whole cell Na+ currents is facilitated by reducing the transepithelial potential to zero, 2) these M-1 currents reflect the operation of Na(5) channels, and 3) the Na+ channels display complex kinetics, involving > or = 2 open and > or = 2 closed states.

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