scholarly journals Expression of a dominant negative PKA mutation in the kidney elicits a diabetes insipidus phenotype

2015 ◽  
Vol 308 (6) ◽  
pp. F627-F638 ◽  
Author(s):  
Merle L. Gilbert ◽  
Linghai Yang ◽  
Thomas Su ◽  
G. Stanley McKnight
Keyword(s):  
Diabetes Insipidus ◽  
Apical Membrane ◽  
Collecting Duct ◽  
Critical Role ◽  
Physiological Role ◽  
Receptor Activation ◽  
Dominant Negative ◽  
Regulatory Subunit ◽  
Water Excretion ◽  
Protein Levels

PKA plays a critical role in water excretion through regulation of the production and action of the antidiuretic hormone arginine vasopressin (AVP). The AVP prohormone is produced in the hypothalamus, where its transcription is regulated by cAMP. Once released into the circulation, AVP stimulates antidiuresis through activation of vasopressin 2 receptors in renal principal cells. Vasopressin 2 receptor activation increases cAMP and activates PKA, which, in turn, phosphorylates aquaporin (AQP)2, triggering apical membrane accumulation, increased collecting duct permeability, and water reabsorption. We used single-minded homolog 1 ( Sim1)-Cre recombinase-mediated expression of a dominant negative PKA regulatory subunit (RIαB) to disrupt kinase activity in vivo and assess the role of PKA in fluid homeostasis. RIαB expression gave rise to marked polydipsia and polyuria; however, neither hypothalamic Avp mRNA expression nor urinary AVP levels were attenuated, indicating a primary physiological effect on the kidney. RIαB mice displayed a marked deficit in urinary concentrating ability and greatly reduced levels of AQP2 and phospho-AQP2. Dehydration induced Aqp2 mRNA in the kidney of both control and RIαB-expressing mice, but AQP2 protein levels were still reduced in RIαB-expressing mutants, and mice were unable to fully concentrate their urine and conserve water. We conclude that partial PKA inhibition in the kidney leads to posttranslational effects that reduce AQP2 protein levels and interfere with apical membrane localization. These findings demonstrate a distinct physiological role for PKA signaling in both short- and long-term regulation of AQP2 and characterize a novel mouse model of diabetes insipidus.

scholarly journals Vasopressin/V2 receptor stimulates renin synthesis in the collecting duct

2016 ◽  
Vol 310 (4) ◽  
pp. F284-F293 ◽  
Author(s):  
Alexis A. Gonzalez ◽  
Flavia Cifuentes-Araneda ◽  
Cristobal Ibaceta-Gonzalez ◽  
Alex Gonzalez-Vergara ◽  
Leonardo Zamora ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Critical Role ◽  
Combined Treatment ◽  
Receptor Activation ◽  
Ang Ii ◽  
Principal Cells ◽  
Protein Levels ◽  
Renin Synthesis ◽  
Renin Mrna ◽  
Creb Pathway

Renin is synthesized in the principal cells of the collecting duct (CD), and its production is increased via cAMP in angiotensin (ANG) II-dependent hypertension, despite suppression of juxtaglomerular (JG) renin. Vasopressin, one of the effector hormones of the renin-angiotensin system (RAS) via the type 2-receptor (V2R), activates the cAMP/PKA/cAMP response element-binding protein (CREB) pathway and aquaporin-2 expression in principal cells of the CD. Accordingly, we hypothesized that activation of V2R increases renin synthesis via PKA/CREB, independently of ANG II type 1 (AT1) receptor activation in CD cells. Desmopressin (DDAVP; 10−6 M), a selective V2R agonist, increased renin mRNA (∼3-fold), prorenin (∼1.5-fold), and renin (∼2-fold) in cell lysates and cell culture media in the M-1 CD cell line. Cotreatment with DDAVP+H89 (PKA inhibitor) or CREB short hairpin (sh) RNA prevented this response. H89 also blunted DDAVP-induced CREB phosphorylation and nuclear localization. In 48-h water-deprived (WD) mice, prorenin-renin protein levels were increased in the renal inner medulla (∼1.4- and 1.8-fold). In WD mice treated with an ACE inhibitor plus AT1 receptor blockade, renin mRNA and prorenin protein levels were still higher than controls, while renin protein content was not changed. In M-1 cells, ANG II or DDAVP increased prorenin-renin protein levels; however, there were no further increases by combined treatment. These results indicate that in the CD the activation of the V2R stimulates renin synthesis via the PKA/CREB pathway independently of RAS, suggesting a critical role for vasopressin in the regulation of renin in the CD.

scholarly journals The Hinge-Helix 1 Region of Peroxisome Proliferator-Activated Receptor γ1 (PPARγ1) Mediates Interaction with Extracellular Signal-Regulated Kinase 5 and PPARγ1 Transcriptional Activation: Involvement in Flow-Induced PPARγ Activation in Endothelial Cells

2004 ◽  
Vol 24 (19) ◽  
pp. 8691-8704 ◽  
Author(s):  
Masashi Akaike ◽  
Wenyi Che ◽  
Nicole-Lerner Marmarosh ◽  
Shinsuke Ohta ◽  
Masaki Osawa ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Adhesion Molecule ◽  
Critical Role ◽  
Physiological Role ◽  
Cellular Adhesion ◽  
Dominant Negative ◽  
Peroxisome Proliferator ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Cellular Adhesion Molecule

ABSTRACT Peroxisome proliferator-activated receptors (PPAR) are ligand-activated transcription factors that form a subfamily of the nuclear receptor gene family. Since both flow and PPARγ have atheroprotective effects and extracellular signal-regulated kinase 5 (ERK5) kinase activity is significantly increased by flow, we investigated whether ERK5 kinase regulates PPARγ activity. We found that activation of ERK5 induced PPARγ1 activation in endothelial cells (ECs). However, we could not detect PPARγ phosphorylation by incubation with activated ERK5 in vitro, in contrast to ERK1/2 and JNK, suggesting a role for ERK5 as a scaffold. Endogenous PPARγ1 was coimmunoprecipitated with endogenous ERK5 in ECs. By mammalian two-hybrid analysis, we found that PPARγ1 associated with ERK5a at the hinge-helix 1 region of PPARγ1. Expressing a hinge-helix 1 region PPARγ1 fragment disrupted the ERK5a-PPARγ1 interaction, suggesting a critical role for hinge-helix 1 region of PPARγ in the ERK5-PPARγ interaction. Flow increased ERK5 and PPARγ1 activation, and the hinge-helix 1 region of the PPARγ1 fragment and dominant negative MEK5β significantly reduced flow-induced PPARγ activation. The dominant negative MEK5β also prevented flow-mediated inhibition of tumor necrosis factor alpha-mediated NF-κB activation and adhesion molecule expression, including vascular cellular adhesion molecule 1 and E-selectin, indicating a physiological role for ERK5 and PPARγ activation in flow-mediated antiinflammatory effects. We also found that ERK5 kinase activation was required, likely by inducing a conformational change in the NH2-terminal region of ERK5 that prevented association of ERK5 and PPARγ1. Furthermore, association of ERK5a and PPARγ1 disrupted the interaction of SMRT and PPARγ1, thereby inducing PPARγ activation. These data suggest that ERK5 mediates flow- and ligand-induced PPARγ activation via the interaction of ERK5 with the hinge-helix 1 region of PPARγ.

scholarly journals Lack of an effect of collecting duct-specific deletion of adenylyl cyclase 3 on renal Na+ and water excretion or arterial pressure

2014 ◽  
Vol 306 (6) ◽  
pp. F597-F607 ◽  
Author(s):  
Wararat Kittikulsuth ◽  
Deborah Stuart ◽  
Alfred N. Van Hoek ◽  
James D. Stockand ◽  
Vladislav Bugaj ◽  
...  
Keyword(s):  
Arterial Pressure ◽  
Adenylyl Cyclase ◽  
Collecting Duct ◽  
Urine Volume ◽  
Physiological Role ◽  
Plasma Renin ◽  
Water Restriction ◽  
Water Excretion ◽  
And Control

cAMP is a key mediator of connecting tubule and collecting duct (CD) Na+ and water reabsorption. Studies performed in vitro have suggested that CD adenylyl cyclase (AC)3 partly mediates the actions of vasopressin; however, the physiological role of CD AC3 has not been determined. To assess this, mice were developed with CD-specific disruption of AC3 [CD AC3 knockout (KO)]. Inner medullary CDs from these mice exhibited 100% target gene recombination and had reduced ANG II- but not vasopressin-induced cAMP accumulation. However, there were no differences in urine volume, urinary urea excretion, or urine osmolality between KO and control mice during normal water intake or varying degrees of water restriction in the presence or absence of chronic vasopressin administration. There were no differences between CD AC3 KO and control mice in arterial pressure or urinary Na+ or K+ excretion during a normal or high-salt diet, whereas plasma renin and vasopressin concentrations were similar between the two genotypes. Patch-clamp analysis of split-open cortical CDs revealed no difference in epithelial Na+ channel activity in the presence or absence of vasopressin. Compensatory changes in AC6 were not responsible for the lack of a renal phenotype in CD AC3 KO mice since combined CD AC3/AC6 KO mice had similar arterial pressure and renal Na+ and water handling compared with CD AC6 KO mice. In summary, these data do not support a significant role for CD AC3 in the regulation of renal Na+ and water excretion in general or vasopressin regulation of CD function in particular.

scholarly journals Basolateral P2X4 channels stimulate ENaC activity in Xenopus cortical collecting duct A6 cells

2014 ◽  
Vol 307 (7) ◽  
pp. F806-F813 ◽  
Author(s):  
Tiffany L. Thai ◽  
Ling Yu ◽  
Douglas C. Eaton ◽  
Billie Jean Duke ◽  
Otor Al-Khalili ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Single Channel ◽  
Collecting Duct ◽  
Reversal Potential ◽  
Receptor Activation ◽  
Dominant Negative ◽  
Phosphatidylinositol 3 Kinase ◽  
A6 Cells ◽  
Basolateral Side ◽  
Phosphatidylinositol 3

The polarized nature of epithelial cells allows for different responses to luminal or serosal stimuli. In kidney tubules, ATP is produced luminally in response to changes in luminal flow. Luminal increases in ATP have been previously shown to inhibit the renal epithelial Na+ channel (ENaC). On the other hand, ATP is increased basolaterally in renal epithelia in response to aldosterone. We tested the hypothesis that basolateral ATP can stimulate ENaC function through activation of the P2X4 receptor/channel. Using single channel cell-attached patch-clamp techniques, we demonstrated the existence of a basolaterally expressed channel stimulated by the P2X4 agonist 2-methylthio-ATP (meSATP) in Xenopus A6 cells, a renal collecting duct principal cell line. This channel had a similar reversal potential and conductance to that of P2X4 channels. Cell surface biotinylation of the basolateral side of these cells confirmed the basolateral presence of the P2X4 receptor. Basolateral addition of meSATP enhanced the activity of ENaC in single channel patch-clamp experiments, an effect that was absent in cells transfected with a dominant negative P2X4 receptor construct, indicating that activation of P2X4 channels stimulates ENaC activity in these cells. The effect of meSATP on ENaC activity was reduced after chelation of basolateral Ca2+ with EGTA or inhibition of phosphatidylinositol 3-kinase with LY-294002. Overall, our results show that ENaC is stimulated by P2X4 receptor activation and that the stimulation is dependent on increases in intracellular Ca2+ and phosphatidylinositol 3-kinase activation.

A critical role for PI 3-kinase in cytokine-induced Fcα-receptor activation

Blood ◽  
2000 ◽  
Vol 95 (6) ◽  
pp. 2037-2043 ◽  
Author(s):  
Madelon Bracke ◽  
Evert Nijhuis ◽  
Jan-Willem J. Lammers ◽  
Paul J. Coffer ◽  
Leo Koenderman
Keyword(s):  
Ligand Binding ◽  
Signaling Pathways ◽  
Critical Role ◽  
Immunoglobulin A ◽  
Fc Receptors ◽  
Receptor Activation ◽  
Molecular Techniques ◽  
Dominant Negative ◽  
Receptor Modulation ◽  
Inside Out

Abstract Fc-receptors, such as FcR and FcγRII, play an important role in leukocyte activation, and rapid modulation of ligand binding (“activation”) is critical for receptor regulation. We have previously demonstrated that ligand binding to Fc-receptors on human eosinophils is dependent on cytokine stimulation. Utilization of pharmacological inhibitors provided evidence that the phenomenon of interleukin (IL)-5 induced immunoglobulin A (IgA) binding to human eosinophils requires activation of phosphatidylinositol 3-kinase (PI3K). However, eosinophils are refractory to manipulation by molecular techniques such as DNA transfection or viral infection. Here we utilize an IL-3 dependent pre-B cell line to investigate the molecular mechanism of cytokine-mediated ligand binding to FcR. In this system, IgA binding is dependent on IL-3, similarly to the requirement for IL-5 of eosinophils. We show that IL-3-mediated activation of FcR (CD89) requires the activation of PI3K, independent of p21ras activation. Co-expression of dominant negative (▵p85) and active (p110_K227E) forms of PI3K demonstrate that the affinity switch regulating FcR activation requires PI3K. Moreover, overexpression of PI3K is both necessary and sufficient for activation of FcR. Furthermore, we show that IL-3/IL-5/GM-CSF induced inside-out signaling pathways activating FcR require the involvement of protein kinase C downstream of PI3K. Finally, we show that these inside-out signaling pathways responsible for Fc-receptor modulation require CD89, independent of its association with the FcRγ chain.

scholarly journals Collecting duct-specific knockout of renin attenuates angiotensin II-induced hypertension

2014 ◽  
Vol 307 (8) ◽  
pp. F931-F938 ◽  
Author(s):  
Nirupama Ramkumar ◽  
Deborah Stuart ◽  
Sara Rees ◽  
Alfred Van Hoek ◽  
Curt D. Sigmund ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Collecting Duct ◽  
Plasma Renin ◽  
Ang Ii ◽  
Water Excretion ◽  
Protein Levels ◽  
Renin Gene ◽  
Induced Hypertension ◽  
Renin Mrna ◽  
Exon 1

The physiological and pathophysiological significance of collecting duct (CD)-derived renin, particularly as it relates to blood pressure (BP) regulation, is unknown. To address this question, we generated CD-specific renin knockout (KO) mice and examined BP and renal salt and water excretion. Mice containing loxP-flanked exon 1 of the renin gene were crossed with mice transgenic for aquaporin-2-Cre recombinase to achieve CD-specific renin KO. Compared with controls, CD renin KO mice had 70% lower medullary renin mRNA and 90% lower renin mRNA in microdissected cortical CD. Urinary renin levels were significantly lower in KO mice (45% of control levels) while plasma renin concentration was significantly higher in KO mice (63% higher than controls) during normal-Na intake. While no observable differences were noted in BP between the two groups with varying Na intake, infusion of angiotensin II at 400 ng·kg−1·min−1 resulted in an attenuated hypertensive response in the KO mice (mean arterial pressure 111 ± 4 mmHg in KO vs. 128 ± 3 mmHg in controls). Urinary renin excretion and epithelial Na+ channel (ENaC) remained significantly lower in the KO mice following ANG II infusion compared with controls. Furthermore, membrane-associated ENaC protein levels were significantly lower in KO mice following ANG II infusion. These findings suggest that CD renin modulates BP in ANG II-infused hypertension and these effects are associated with changes in ENaC expression.

scholarly journals APP-BP1 mediates APP-induced apoptosis and DNA synthesis and is increased in Alzheimer's disease brain

2003 ◽  
Vol 163 (1) ◽  
pp. 27-33 ◽  
Author(s):  
Yuzhi Chen ◽  
Wenyun Liu ◽  
Donna L. McPhie ◽  
Linda Hassinger ◽  
Rachael L. Neve
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Dna Synthesis ◽  
Lipid Rafts ◽  
Neuronal Apoptosis ◽  
Dominant Negative ◽  
Regulatory Subunit ◽  
Dominant Negative Mutant ◽  
Protein Levels ◽  
Induced Apoptosis

APP-BP1, first identified as an amyloid precursor protein (APP) binding protein, is the regulatory subunit of the activating enzyme for the small ubiquitin-like protein NEDD8. We have shown that APP-BP1 drives the S- to M-phase transition in dividing cells, and causes apoptosis in neurons (Chen, Y., D.L. McPhie, J. Hirschberg, and R.L. Neve. 2000. J. Biol. Chem. 275:8929–8935). We now demonstrate that APP-BP1 binds to the COOH-terminal 31 amino acids of APP (C31) and colocalizes with APP in a lipid-enriched fraction called lipid rafts. We show that coexpression of a peptide representing the domain of APP-BP1 that binds to APP, abolishes the ability of overexpressed APP or the V642I mutant of APP to cause neuronal apoptosis and DNA synthesis. A dominant negative mutant of the NEDD8 conjugating enzyme hUbc12, which participates in the ubiquitin-like pathway initiated by APP-BP1, blocks neuronal apoptosis caused by APP, APP(V642I), C31, or overexpression of APP-BP1. Neurons overexpressing APP or APP(V642I) show increased APP-BP1 protein levels in lipid rafts. A similar increase in APP-BP1 in lipid rafts is observed in the Alzheimer's disease brain hippocampus, but not in less-affected areas of Alzheimer's disease brain. This translocation of APP-BP1 to lipid rafts is accompanied by a change in the subcellular localization of the ubiquitin-like protein NEDD8, which is activated by APP-BP1.

scholarly journals Role of TRPC3 channels in ATP-induced Ca2+ signaling in principal cells of the inner medullary collecting duct

2010 ◽  
Vol 299 (1) ◽  
pp. F225-F233 ◽  
Author(s):  
Monu Goel ◽  
William P. Schilling
Keyword(s):  
Transient Receptor Potential ◽  
Apical Membrane ◽  
Collecting Duct ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Dominant Negative ◽  
Principal Cells ◽  
Basolateral Side ◽  
Influx Pathways

The transient receptor potential channel TRPC3 is exclusively expressed in the apical membrane of principal cells of the collecting duct (CD) both in vivo and in the mouse CD cell line IMCD-3. Previous studies revealed that ATP-induced apical-to-basolateral transepithelial Ca2+ flux across IMCD-3 monolayers is increased by overexpression of TRPC3 and attenuated by a dominant negative TRPC3 construct, suggesting that Ca2+ entry across the apical membrane occurs via TRPC3 channels. To test this hypothesis, we selectively measured the Ca2+ permeability of the apical membrane of fura-2-loaded IMCD-3 cells using the Mn2+ quench technique. Mn2+ influx across the apical membrane was increased 12- to 16-fold by apical ATP and was blocked by the pyrazole derivative BTP2, a known inhibitor of TRPC3 channels, with an IC50 value <100 nM. In contrast, Mn2+ influx was only increased ∼2-fold by basolateral ATP. Mn2+ influx was also activated by apical, but not basolateral, 1-stearoyl-2-acetyl- sn-glycerol (SAG), a known activator of TRPC3 channels. Apical ATP- and SAG-induced Mn2+ influx was increased by overexpression of TRPC3 and completely blocked by expression of the dominant negative TRPC3 construct. Mn2+ influx was also stimulated ∼2-fold by thapsigargin applied to either the apical or basolateral side. Thapsigargin-induced flux was blocked by BTP2 but was unaffected by overexpression of TRPC3 or by dominant negative TRPC3. Apical ATP, but not basolateral ATP, increased transepithelial 45Ca2+ flux. These results demonstrate that the apical membrane of IMCD-3 cells has two distinct Ca2+ influx pathways: 1) a store-operated channel activated by thapsigargin and basolateral ATP and 2) TRPC3 channels activated by apical ATP. Only activation of TRPC3 leads to net transepithelial apical-to-basolateral Ca2+ flux. Furthermore, these results demonstrate that native TRPC3 is not a store-operated channel in IMCD-3 cells.

scholarly journals Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells

2016 ◽  
Vol 311 (6) ◽  
pp. F1360-F1368 ◽  
Author(s):  
Ming-Ming Wu ◽  
Yu-Jia Zhai ◽  
Yu-Xia Li ◽  
Qing-Qing Hu ◽  
Zhi-Rui Wang ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Transient Receptor Potential ◽  
Single Channel ◽  
Apical Membrane ◽  
Intact Cell ◽  
Collecting Duct ◽  
Physiological Role ◽  
Cell Types ◽  
Cortical Collecting Duct ◽  
Principal Cells

A Ca2+-activated nonselective cation channel (NSCCa) is found in principal cells of the mouse cortical collecting duct (CCD). However, the molecular identity of this channel remains unclear. We used mpkCCDc14 cells, a mouse CCD principal cell line, to determine whether NSCCa represents the transient receptor potential (TRP) channel, the melastatin subfamily 4 (TRPM4). A Ca2+-sensitive single-channel current was observed in inside-out patches excised from the apical membrane of mpkCCDc14 cells. Like TRPM4 channels found in other cell types, this channel has an equal permeability for Na+ and K+ and has a linear current-voltage relationship with a slope conductance of ~23 pS. The channel was inhibited by a specific TRPM4 inhibitor, 9-phenanthrol. Moreover, the frequency of observing this channel was dramatically decreased in TRPM4 knockdown mpkCCDc14 cells. Unlike those previously reported in other cell types, the TRPM4 in mpkCCDc14 cells was unable to be activated by hydrogen peroxide (H2O2). Conversely, after treatment with H2O2, TRPM4 density in the apical membrane of mpkCCDc14 cells was significantly decreased. The channel in intact cell-attached patches was activated by ionomycin (a Ca2+ ionophore), but not by ATP (a purinergic P2 receptor agonist). These data suggest that the NSCCa current previously described in CCD principal cells is actually carried through TRPM4 channels. However, the physiological role of this channel in the CCD remains to be further determined.

Abstract 401: Prostaglandin E2 Stimulates Renin Synthesis in Mouse Collecting Duct M-1 Cells via Ep1 Receptor Through Pkc/camp/creb Pathway

Hypertension ◽  
2012 ◽  
Vol 60 (suppl_1) ◽  
Author(s):  
Alexis A Gonzalez ◽  
Dan Leach ◽  
L G Navar ◽  
Minolfa C Prieto
Keyword(s):  
Prostaglandin E2 ◽  
Collecting Duct ◽  
Receptor Activation ◽  
Fold Change ◽  
Dominant Negative ◽  
Calphostin C ◽  
Renin Synthesis ◽  
Renin Mrna ◽  
Ep1 Receptor ◽  
Ep Receptor

Prostaglandin E2 (PGE2) plays a major role in regulating renin expression and release by the renal juxtaglomerular (JG) cells. Recently it has been demonstrated that PGE2-dependent upregulation of renin in JG cells is mediated by activation of E prostaoind receptor type 4 (EP4) via cAMP accumulation. Renin is also produced by the principal cells of the collecting ducts (CD) and is upregulated during angiotensin II-dependent hypertension. However, the effects of PGE2 on CD renin remain unknown. Four types of receptors have been described in rat and mouse CD, EP1, EP3 and EP4. Here, we tested the hypothesis that renin is upregulated by PGE2 via activation of EP receptors in mouse CD M-1 cells. By immunostaining we confirmed the presence of EP1, EP3 and EP4 receptors, while EP2 was not detected. A dose response treatment with PGE2 showed increased levels of renin mRNA and protein with a maximum response at 1 μmol/L (mRNA: 19.3 ± 3.0; P<0.05; protein: 3.01 ± 0.08 fold change; P<0.05). To assess which EP receptor is involved in the renin upregulation we used specific EP receptor antagonists: ONO-8711 (EP1; 10 nmol/L), L-798106 (EP3; 10 μmol/L) and AH 23848 (EP4; 10 μmol/L). EP1 antagonist suppressed the PGE2-mediated upregulation of collecting duct renin mRNA and protein (mRNA: 1.0 ± 0.2; protein: 0.98 ± 0.13 fold change; P=NS), while EP4 antagonist only partially decreased it (mRNA: 11.2 ± 2.8; P<0.05; protein: 2.81 ± 0.07 fold change; P<0.05). EP3 antagonist exacerbated the PGE2 mediated-upregulation of renin (mRNA: 50.3 ± 6.0; P<0.05; protein: 3.56 ± 0.08; fold change; P<0.05). Because EP1 is a Gq linked receptor that activates PKC, we further assessed the effects of PKC inhibition using calphostin C and a PKCα dominant negative (DN) on renin expression. Calphostin C and PKCα-DN blunted the PGE2-induced renin upregulation. Importantly, the increases in cAMP levels and phosphorylation of the cAMP response element-binding transcription factor (CREB) mediated by PGE2 were also prevented by both treatments. The results indicate that in mouse CD cells, EP1 receptor activation upregulates renin synthesis via PKC/cAMP/CREB, suggesting that the presence of PGE2 in renal medullary tissues may contribute to the stimulation of collecting duct renin.

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