Muscarinic receptor activation inhibits AVP-induced water flow in rabbit cortical collecting ducts

1991 ◽  
Vol 260 (6) ◽  
pp. F929-F936 ◽  
Author(s):  
H. M. Snyder ◽  
D. M. Fredin ◽  
M. D. Breyer
Keyword(s):  
Muscarinic Receptor ◽  
Water Flow ◽  
Calcium Concentration ◽  
Collecting Duct ◽  
Receptor Activation ◽  
Inhibitory Effect ◽  
Cortical Collecting Duct ◽  
Kinase C ◽  
Collecting Ducts ◽  
Subsequent Addition

We examined the effect of carbachol, an acetylcholine analogue, on hydraulic conductivity (Lp) response to 10 microU/ml arginine vasopressin (AVP) in rabbit cortical collecting duct (CCD). In CCDs in which water flow had been established with AVP, subsequent addition of carbachol caused Lp (X10(-7) cm.atm-1.s-1) to fall from 251 +/- 32 to 146 +/- 19. Carbachol washout resulted in recovery of Lp to 217 +/- 38. In CCDs in which water flow had been established using 10(-4) M 8-chlorophenylthioadenosine 3',5'-cyclic monophosphate (8-CPT-cAMP), addition of carbachol had no effect. These posttreatment studies suggest that carbachol's effects on modulating established water flow occur at a "pre-cAMP" step. With carbachol added first, AVP-induced Lp was reduced from 233 +/- 24 (controls) to 105 +/- 19 (carbachol-pretreated). Pretreatment with 10(-6) M atropine, a muscarinic receptor antagonist, totally reversed the inhibitory effect of carbachol, consistent with a receptor-mediated effect of carbachol. Carbachol pretreatment also inhibited 8-CPT-cAMP-induced Lp, indicating that carbachol's effects also occur at a "post-cAMP" step. Pretreatment with 10(-7) M staurosporine, a protein kinase C (PKC) inhibitor, reversed inhibitory effect of carbachol on AVP-induced Lp (193 +/- 26), suggesting that carbachol's effects are mediated by PKC. Intracellular calcium concentration [( Ca2+]i) was measured in fura-2-loaded CCDs. Carbachol also increased [Ca2+]i from 229 +/- 120 to 389 +/- 160 nM.(ABSTRACT TRUNCATED AT 250 WORDS)

PGE2 inhibits AVP-induced water flow in cortical collecting ducts by protein kinase C activation

1990 ◽  
Vol 259 (2) ◽  
pp. F318-F325 ◽  
Author(s):  
R. L. Hebert ◽  
H. R. Jacobson ◽  
M. D. Breyer
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Water Flow ◽  
Collecting Duct ◽  
Inhibitory Effect ◽  
Cortical Collecting Duct ◽  
Independent Evidence ◽  
Kinase C ◽  
Pi Hydrolysis

It is well known that prostaglandin E2 (PGE2) both inhibits arginine vasopressin (AVP)-stimulated water permeability (hydraulic conductivity, Lp) in the cortical collecting duct (CCD) or, if administered alone, modestly increases Lp in the CCD. These bifunctional effects on Lp correspond to PGE2's capacity to inhibit AVP-stimulated adenylate cyclase (AC) activity, or to singularly stimulate AC activity in the collecting duct. The present studies suggest that the inhibitory effect of PGE2 on Lp may also be mediated by phosphatidylinositol (PI) hydrolysis. Using in vitro microperfused rabbit CCDs, we show that PGE2 releases Ca from intracellular stores. We also demonstrate that the inhibitory effect of PGE2 on AVP-stimulated Lp in the CCD is significantly reversed by the protein kinase C (PKC) inhibitor, staurosporine (SSP). Although PGE2 does not reduce an established water flow response to 8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (8-CPTcAMP), when the sequence of addition is reversed and PGE2 is added first, marked inhibition of 8-CPTcAMP-induced Lp is observed. This provides independent evidence that PGE2 can act through a mechanism separate from modulating AC activity. PGE2 inhibition of 8-CPTcAMP-induced Lp is reversed by SSP pretreatment. Finally, SSP pretreatment also markedly potentiates the capacity of PGE2 itself to increase Lp. We conclude that PGE2 releases Ca from intracellular stores and, by activating PKC, inhibits AVP-induced osmotic water flow. This suggests an important role for PI hydrolysis in mediating PGE2's effects on the CCD.

Evidence that separate PGE2 receptors modulate water and sodium transport in rabbit cortical collecting duct

1993 ◽  
Vol 265 (5) ◽  
pp. F643-F650 ◽  
Author(s):  
R. L. Hebert ◽  
H. R. Jacobson ◽  
D. Fredin ◽  
M. D. Breyer
Keyword(s):  
Pertussis Toxin ◽  
Water Permeability ◽  
Sodium Transport ◽  
Collecting Duct ◽  
Inhibitory Effect ◽  
Cortical Collecting Duct ◽  
Pkc Inhibitor ◽  
Kinase C ◽  
Pge2 Receptor ◽  
Pkc Activation

Prostaglandin E2 (PGE2) modulates both water and sodium transport in the rabbit cortical collecting duct (CCD). To determine whether these effects are mediated by separate PGE2 receptors, we compared the effects of PGE2 and its analogue sulprostone in the isolated perfused rabbit CCD. PGE2 increased basal water permeability (hydraulic conductivity), whereas sulprostone did not. PGE2 and sulprostone were equipotent inhibitors of water absorption when it was prestimulated by vasopressin. Pertussis toxin completely reversed the inhibitory effect of sulprostone but only partially reversed the inhibitory effect of PGE2. In contrast, a protein kinase C (PKC) inhibitor, staurosporine, partially reversed the inhibitory effect of PGE2 but had no effect on sulprostone. PGE2 also raised intracellular calcium ([Ca2+]i). This effect is coupled to its capacity to inhibit Na+ absorption. Sulprostone was 10-fold less potent than PGE2 both in raising [Ca2+]i or inhibiting sodium transport. The results suggest sulprostone selectively interacts with a PGE2 receptor coupled to pertussis toxin-sensitive inhibition of water permeability. Sulprostone less potently activates a PGE2 receptor coupled to [Ca2+]i, PKC activation, and sodium transport and completely fails to interact with the PGE2 receptor that stimulates water permeability in the collecting duct. These results suggest distinct PGE2 receptors modulate sodium and water transport in the CCD.

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

2002 ◽  
Vol 283 (4) ◽  
pp. F689-F698 ◽  
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.

Intracellular Ca2+ and PKC activation do not inhibit Na+ and water transport in rat CCD

1993 ◽  
Vol 265 (4) ◽  
pp. F569-F577 ◽  
Author(s):  
A. J. Rouch ◽  
L. Chen ◽  
L. H. Kudo ◽  
P. D. Bell ◽  
B. C. Fowler ◽  
...  
Keyword(s):  
Water Transport ◽  
Calcium Concentration ◽  
Morphological Changes ◽  
Collecting Duct ◽  
Bathing Solution ◽  
Cortical Collecting Duct ◽  
Kinase C ◽  
Transepithelial Voltage ◽  
Pkc Activation ◽  
Sustained Increase

Experiments examined the effects of elevation of intracellular calcium concentration ([Ca2+]i) or activation of protein kinase C (PKC) on Na+ and water transport in the rat cortical collecting duct (CCD). We measured the lumen-to-bath 22Na+ flux (J1-->b), transepithelial voltage (VT), and water permeability (Pf) in CCD from deoxycorticosterone (DOC)-treated rats. Ionomycin (0.5 and 1 microM) and thapsigargin (1 and 2 microM) were used to increase [Ca2+]i. Phorbol 12-myristate 13-acetate (PMA; 0.3 and 1 microM) and oleoyl-acetyl-glycerol (OAG; 100 microM) were used as activators of PKC. [Ca2+]i was measured in isolated perfused tubules using the fluorescent dye fura 2. When added to the bathing solution, 220 pM arginine vasopressin (AVP) failed to affect [Ca2+]i, whereas 1 microM ionomycin increased [Ca2+]i by 103 +/- 15% and 2 microM thapsigargin increased [Ca2+]i by 24 +/- 4%. In flux studies, neither ionomycin nor thapsigargin affected J1-->b or Pf, although ionomycin caused marked morphological changes. Ionomycin also failed to alter either parameter in tubules from non-DOC-treated rats. Neither 100 microM OAG nor 1 microM PMA affected J1-->b or Pf. OAG at 50 microM had no effect on VT or transepithelial resistance, indicating no inhibition of conductive Na+ transport. We conclude that increased [Ca2+]i and PKC activation do not affect J1--b or Pf in the rat CCD. These findings may account for the sustained increase in J1--b produced in the rat CCD by AVP.

scholarly journals Potassium depletion increases proton pump (H+-ATPase) activity in intercalated cells of cortical collecting duct

2000 ◽  
Vol 279 (1) ◽  
pp. F195-F202 ◽  
Author(s):  
Randi B. Silver ◽  
Sylvie Breton ◽  
Dennis Brown
Keyword(s):  
Plasma Membrane ◽  
Atpase Activity ◽  
Collecting Duct ◽  
Proton Pumps ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Collecting Ducts ◽  
Acid Load ◽  
Collecting Tubules ◽  
Atpase Staining

Intercalated cells (ICs) from kidney collecting ducts contain proton-transporting ATPases (H+-ATPases) whose plasma membrane expression is regulated under a variety of conditions. It has been shown that net proton secretion occurs in the distal nephron from chronically K+-depleted rats and that upregulation of tubular H+- ATPase is involved in this process. However, regulation of this protein at the level of individual cells has not so far been examined. In the present study, H+-ATPase activity was determined in individually identified ICs from control and chronically K+-depleted rats (9–14 days on a low-K+ diet) by monitoring K+- and Na+-independent H+ extrusion rates after an acute acid load. Split-open rat cortical collecting tubules were loaded with the intracellular pH (pHi) indicator 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, and pHiwas determined by using ratiometric fluorescence imaging. The rate of pHi recovery in ICs in response to an acute acid load, a measure of plasma membrane H+-ATPase activity, was increased after K+ depletion to almost three times that of controls. Furthermore, the lag time before the start of pHirecovery after the cells were maximally acidified fell from 93.5 ± 13.7 s in controls to 24.5 ± 2.1 s in K+-depleted rats. In all ICs tested, Na+- and K+-independent pHi recovery was abolished in the presence of bafilomycin (100 nM), an inhibitor of the H+-ATPase. Analysis of the cell-to-cell variability in the rate of pHi recovery reveals a change in the distribution of membrane-bound proton pumps in the IC population of cortical collecting duct from K+-depleted rats. Immunocytochemical analysis of collecting ducts from control and K+-depleted rats showed that K+-depletion increased the number of ICs with tight apical H+ATPase staining and decreased the number of cells with diffuse or basolateral H+-ATPase staining. Taken together, these data indicate that chronic K+ depletion induces a marked increase in plasma membrane H+ATPase activity in individual ICs.

Automated method for the isolation of collecting ducts

2006 ◽  
Vol 291 (1) ◽  
pp. F236-F245 ◽  
Author(s):  
R. Lance Miller ◽  
Ping Zhang ◽  
Tong Chen ◽  
Andreas Rohrwasser ◽  
Raoul D. Nelson
Keyword(s):  
Flow Cytometry ◽  
Large Particle ◽  
Fluorescent Protein ◽  
Collecting Duct ◽  
Complex Object ◽  
Cortical Collecting Duct ◽  
Fluorescent Intensity ◽  
Automated Method ◽  
Collecting Ducts ◽  
Time Required

The structural and functional heterogeneity of the collecting duct present a tremendous experimental challenge requiring manual microdissection, which is time-consuming, labor intensive, and not amenable to high throughput. To overcome these limitations, we developed a novel approach combining the use of transgenic mice expressing green fluorescent protein (GFP) in the collecting duct with large-particle-based flow cytometry to isolate pure populations of tubular fragments from the whole collecting duct (CD), or inner medullary (IMCD), outer medullary (OMCD), or connecting segment/cortical collecting duct (CNT/CCD). Kidneys were enzymatically dispersed into tubular fragments and sorted based on tubular length and GFP intensity using large-particle-based flow cytometry or a complex object parametric analyzer and sorter (COPAS). A LIVE/DEAD assay demonstrates that the tubules were >90% viable. Tubules were collected as a function of fluorescent intensity and analyzed by epifluorescence and phase microscopy for count accuracy, GFP positivity, average tubule length, and time required to collect 100 tubules. Similarly, mRNA and protein from sorted tubules were analyzed for expression of tubule segment-specific genes using quantitative real-time RT-PCR and immunoblotting. The purity and yield of sorted tubules were related to sort stringency. Four to six replicates of 100 collecting ducts (9.68 ± 0.44–14.5 ± 0.66 cm or 9.2 ± 0.7 mg tubular protein) were routinely obtained from a single mouse in under 1 h. In conclusion, large-particle-based flow cytometry is fast, reproducible, and generates sufficient amounts of highly pure and viable collecting ducts from single or replicate animals for gene expression and proteomic analysis.

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

1998 ◽  
Vol 274 (4) ◽  
pp. F736-F743 ◽  
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.

Evidence of corticosteroid action along the nephron

1984 ◽  
Vol 246 (2) ◽  
pp. F111-F123 ◽  
Author(s):  
D. Marver
Keyword(s):  
Collecting Duct ◽  
Type I ◽  
Type Ii ◽  
Cortical Collecting Duct ◽  
Distal Nephron ◽  
Type Iii ◽  
Collecting Ducts ◽  
Collecting Tubule ◽  
Cortical Collecting Tubule

The kidney contains three classes of corticosteroid-binding proteins receptors. They include a mineralocorticoid-specific (Type I), a glucocorticoid-specific (Type II), and a corticosterone-specific (Type III) site. The Type I and Type III sites roughly parallel each other along the nephron, with maximal binding occurring in the late distal convoluted or connecting segment and the cortical and medullary collecting ducts. Type II sites occur throughout the nephron, with maximal concentrations appearing in the proximal tubule and the late distal convoluted-cortical collecting duct region. The function of the Type I sites in the connecting segment is unclear since chronic mineralocorticoid therapy does not influence the potential difference in this segment as it does in the cortical collecting tubule. Furthermore, the specific role of Type II versus Type III sites in the distal nephron is unknown. Finally, the possible influence of sodium on both latent and steroid-induced renal cortical and medullary Na-K-ATPase is discussed.

Calcium Waves and Closure of Potassium Channels in Response to GABA Stimulation in Hermissenda Type B Photoreceptors

2002 ◽  
Vol 87 (2) ◽  
pp. 776-792 ◽  
Author(s):  
K. T. Blackwell
Keyword(s):  
Calcium Current ◽  
Calcium Concentration ◽  
Calcium Release ◽  
Input Resistance ◽  
Receptor Activation ◽  
Memory Storage ◽  
Type B ◽  
Kinase C ◽  
Hyperpolarization Activated Current ◽  
Receptor Channel

Classical conditioning of Hermissenda crassicornisrequires the paired presentation of a conditioned stimulus (light) and an unconditioned stimulus (turbulence). Light stimulation of photoreceptors leads to production of diacylglycerol, an activator of protein kinase C, and inositol triphosphate (IP3), which releases calcium from intracellular stores. Turbulence causes hair cells to release GABA onto the terminal branches of the type B photoreceptor. One prior study has shown that GABA stimulation produces a wave of calcium that propagates from the terminal branches to the soma and raises the possibility that two sources of calcium are required for memory storage. GABA stimulation also causes an inhibitory postsynaptic potential (IPSP) followed by a late depolarization and increase in input resistance, whose cause has not been identified. A model was developed of the effect of GABA stimulation on the Hermissenda type B photoreceptor to evaluate the currents underlying the late depolarization and to evaluate whether a calcium wave could propagate from the terminal branches to the soma. The model included GABAA, GABAB, and calcium-sensitive potassium leak channels; calcium dynamics including release of calcium from intracellular stores; and the biochemical reactions leading from GABAB receptor activation to IP3 production. Simulations show that it is possible for a wave of calcium to propagate from the terminal branches to the soma. The wave is initiated by IP3-induced calcium release but propagation requires release through the ryanodine receptor channel where IP3 concentration is small. Wave speed is proportional to peak calcium concentration at the crest of the wave, with a minimum speed of 9 μm/s in the absence of IP3. Propagation ceases when peak concentration drops below 1.2 μM; this occurs if the rate of calcium pumping into the endoplasmic reticulum is too large. Simulations also show that both a late depolarization and an increase in input resistance occur after GABA stimulation. The duration of the late depolarization corresponds to the duration of potassium leak channel closure. Neither the late depolarization nor the increase in input resistance are observed when a transient calcium current and a hyperpolarization-activated current are added to the model as replacement for closure of potassium leak channels. Thus the late depolarization and input resistance elevation can be explained by a closure of calcium-sensitive leak potassium currents but cannot be explained by a transient calcium current and a hyperpolarization-activated current.

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