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 TRPM4 Channels Couple Purinergic Receptor Mechanoactivation and Myogenic Tone Development in Cerebral Parenchymal Arterioles

2014 ◽  
Vol 34 (10) ◽  
pp. 1706-1714 ◽  
Author(s):  
Yao Li ◽  
Rachael L Baylie ◽  
Matthew J Tavares ◽  
Joseph E Brayden
Keyword(s):  
Transient Receptor Potential ◽  
Purinergic Receptor ◽  
Critical Role ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Receptor Activation ◽  
Myogenic Tone ◽  
Pial Arteries ◽  
Myogenic Regulation

Cerebral parenchymal arterioles (PAs) have a critical role in assuring appropriate blood flow and perfusion pressure within the brain. They are unique in contrast to upstream pial arteries, as defined by their critical roles in neurovascular coupling, distinct sensitivities to chemical stimulants, and enhanced myogenic tone development. The objective of the present study was to reveal some of the unique mechanisms of myogenic tone regulation in the cerebral microcirculation. Here, we report that in vivo suppression of TRPM4 (transient receptor potential) channel expression, or inhibition of TRPM4 channels with 9-phenanthrol substantially reduced myogenic tone of isolated PAs, supporting a key role of TRPM4 channels in PA myogenic tone development. Further, downregulation of TRPM4 channels inhibited vasoconstriction induced by the specific P2Y4 and P2Y6 receptor ligands (UTP γS and UDP) by 37% and 42%, respectively. In addition, 9-phenanthrol substantially attenuated purinergic ligand-induced membrane depolarization and constriction of PAs, and inhibited ligand-evoked TRPM4 channel activation in isolated PA myocytes. In concert with our previous work showing the essential contributions of P2Y4 and P2Y6 receptors to myogenic regulation of PAs, the current results point to TRPM4 channels as an important link between mechanosensitive P2Y receptor activation and myogenic constriction of cerebral PAs.

scholarly journals Regulation of calcium reabsorption along the rat nephron: a modeling study

2015 ◽  
Vol 308 (6) ◽  
pp. F553-F566 ◽  
Author(s):  
Aurélie Edwards
Keyword(s):  
Transient Receptor Potential ◽  
Stone Formation ◽  
Collecting Duct ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Transepithelial Transport ◽  
Thick Ascending Limb ◽  
Calcium Sensing ◽  
The Impact

We expanded a mathematical model of transepithelial transport along the rat nephron to include the transport of Ca2+ and probe the impact of calcium-sensing mechanisms on Ca2+ reabsorption. The model nephron extends from the medullary thick ascending limb (mTAL) to the inner medullary collecting duct (IMCD). Our model reproduces several experimental findings, such as measurements of luminal Ca2+ concentrations in cortical tubules, and the effects of furosemide or deletion of the transient receptor potential channel vanilloid subtype 5 (TRPV5) on urinary Ca2+ excretion. In vitro microperfusion of rat TAL has demonstrated that activation of the calcium-sensing receptor CaSR lowers the TAL permeability to Ca2+, PCaTAL (Loupy A, Ramakrishnan SK, Wootla B, Chambrey R, de la Faille R, Bourgeois S, Bruneval P, Mandet C, Christensen EI, Faure H, Cheval L, Laghmani K, Collet C, Eladari D, Dodd RH, Ruat M, Houillier P. J Clin Invest 122: 3355, 2012). Our results suggest that this regulatory mechanism significantly impacts renal Ca2+ handling: when plasma Ca2+ concentration ([Ca2+]) is raised by 10%, the CaSR-mediated reduction in PCaTAL per se is predicted to enhance urinary Ca2+ excretion by ∼30%. If high [Ca2+] also induces renal outer medullary potassium (ROMK) inhibition, urinary Ca2+ excretion is further raised. In vitro, increases in luminal [Ca2+] have been shown to activate H+-ATPase pumps in the outer medullary CD and to lower the water permeability of IMCD. Our model suggests that if these responses exhibit the sigmoidal dependence on luminal [Ca2+] that is characteristic of CaSR, then the impact of elevated Ca2+ levels in the CD on urinary volume and pH remains limited. Finally, our model suggests that CaSR inhibitors could significantly reduce urinary Ca2+ excretion in hypoparathyroidism, thereby reducing the risk of calcium stone formation.

scholarly journals Role of cAMP/PKA signaling cascade in vasopressin-induced trafficking of TRPC3 channels in principal cells of the collecting duct

2010 ◽  
Vol 298 (4) ◽  
pp. F988-F996 ◽  
Author(s):  
Monu Goel ◽  
Cheng-Di Zuo ◽  
William P. Schilling
Keyword(s):  
Apical Membrane ◽  
Collecting Duct ◽  
Surface Membrane ◽  
Transient Receptor Potential Channels ◽  
Signaling Cascade ◽  
Dibutyryl Camp ◽  
Principal Cells ◽  
V2 Receptor ◽  
Stimulation Of

Transient receptor potential channels TRPC3 and TRPC6 are expressed in principal cells of the collecting duct (CD) along with the water channel aquaporin-2 (AQP2) both in vivo and in the cultured mouse CD cell line IMCD-3. The channels are primarily localized to intracellular vesicles, but upon stimulation with the antidiuretic hormone arginine vasopressin (AVP), TRPC3 and AQP2 translocate to the apical membrane. In the present study, the effect of various activators and inhibitors of the adenylyl cyclase (AC)/cAMP/PKA signaling cascade on channel trafficking was examined using immunohistochemical techniques and by biotinylation of surface membrane proteins. Both in vivo in rat kidney and in IMCD-3 cells, translocation of AQP2 and TRPC3 (but not TRPC6) was stimulated by [deamino-Cys1, d-Arg8]-vasopressin (dDAVP), a specific V2-receptor agonist, and blocked by [adamantaneacetyl1, O-Et-d-Tyr2, Val4, aminobutyryl6, Arg8,9]-vasopressin (AEAVP), a specific V2-receptor antagonist. In IMCD-3 cells, translocation of TRPC3 and AQP2 was activated by forskolin, a direct activator of AC, or by dibutyryl-cAMP, a membrane-permeable cAMP analog. AVP-, dDAVP-, and forskolin-induced translocation in IMCD-3 cells was blocked by SQ22536 and H89, specific inhibitors of AC and PKA, respectively. Translocation stimulated by dibutyryl-cAMP was unaffected by AEAVP but could be blocked by H89. AVP- and forskolin-induced translocation of TRPC3 in IMCD-3 cells was also blocked by two additional inhibitors of PKA, specifically Rp-cAMPS and the myristoylated inhibitor of PKA (m-PKI). Quantification of TRPC3 membrane insertion in IMCD-3 cells under each assay condition using a surface membrane biotinylation assay, confirmed the translocation results observed by immunofluorescence. Importantly, AVP-induced translocation of TRPC3 as estimated by biotinylation was blocked on average 95.2 ± 1.0% by H89, Rp-cAMPS, or m-PKI. Taken together, these results demonstrate that AVP stimulation of V2 receptors in principal cells of the CD causes translocation of TRPC3 to the apical membrane via stimulation of the AC/cAMP/PKA signaling cascade.

scholarly journals TRPC5 Does Not Cause or Aggravate Glomerular Disease

2017 ◽  
Vol 29 (2) ◽  
pp. 409-415 ◽  
Author(s):  
Xuexiang Wang ◽  
Ranadheer R. Dande ◽  
Hao Yu ◽  
Beata Samelko ◽  
Rachel E. Miller ◽  
...  
Keyword(s):  
Transient Receptor Potential ◽  
Calcium Influx ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Transgenic Mouse Models ◽  
Cytoskeletal Remodeling ◽  
Englerin A ◽  
Transient Receptor ◽  
Ion Pore

Transient receptor potential channel 5 (TRPC5) is highly expressed in brain and kidney and mediates calcium influx and promotes cell migration. In the kidney, loss of TRPC5 function has been reported to benefit kidney filter dynamics by balancing podocyte cytoskeletal remodeling. However, in vivo gain-in-function studies of TRPC5 with respect to kidney function have not been reported. To address this gap, we developed two transgenic mouse models on the C57BL/6 background by overexpressing either wild-type TRPC5 or a TRPC5 ion-pore mutant. Compared with nontransgenic controls, neither transgenic model exhibited an increase in proteinuria at 8 months of age or a difference in LPS-induced albuminuria. Moreover, activation of TRPC5 by Englerin A did not stimulate proteinuria, and inhibition of TRPC5 by ML204 did not significantly lower the level of LPS-induced proteinuria in any group. Collectively, these data suggest that the overexpression or activation of the TRPC5 ion channel does not cause kidney barrier injury or aggravate such injury under pathologic conditions.

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.

scholarly journals Transient Receptor Potential Channel Canonical Type 3 Deficiency Antagonizes Myofibroblast Transdifferentiation In Vivo

2020 ◽  
Vol 2020 ◽  
pp. 1-12
Author(s):  
Weijie Xia ◽  
Qianran Wang ◽  
Yuangang Lu ◽  
Yingru Hu ◽  
Xingcun Zhang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Mitochondrial Calcium ◽  
Ros Production ◽  
Transient Receptor ◽  
Canonical Type ◽  
Type 3

Objective. Myofibroblast transformation has been shown to be associated with the reactive oxygen species- (ROS-) producing enzyme NADPH oxidase (Nox4). Inhibition of transient receptor potential channel canonical type 3 (TRPC3) attenuates mitochondrial calcium handling and ROS production in the vasculature of hypertensive rats. However, it remains elusive whether TRPC3 regulates mitochondrial calcium and ROS production and participates in myofibroblast transdifferentiation during wound healing. Methods and Results. In this study, we demonstrated that activation of TRPC3 by transforming growth factor β (TGFβ1) elevated myofibroblast transdifferentiation by upregulating the myofibroblast marker alpha smooth muscle actin (αSMA). Inhibition of TRPC3 with its specific inhibitor, Pyr3, significantly decreased TGFβ1-induced αSMA expression, as demonstrated by immunofluorescence. Real-time PCR and immunohistochemistry revealed higher TRPC3 and TGFβ1 mRNA expression levels in fibroblasts from hypertrophic scar (HTS) tissue than in those from normal skin tissue. TGFβ1 treatment increased TRPC3-mediated mitochondrial calcium uptake and ROS production but decreased ATP content in human fibroblasts, whereas inhibition of TRPC3 significantly reversed these effects. The beneficial effects were associated with improvements in mitochondrial respiratory function mediated by recovery of the activity of pyruvate dehydrogenase (PDH). In vivo, Trpc3-/- mice exhibited significantly attenuated myofibroblast transdifferentiation, as demonstrated by decreased αSMA, TGFβ1, fibronectin, and collagen-1 (Col1a1) protein expression in wound granulation tissues. Furthermore, TGFβ1-induced store-operated calcium entry (SOCE) was significantly decreased in fibroblasts from Trpc3-/- mice compared with those from Trpc3+/+ mice. In addition, Trpc3-/- mice exhibited significantly decreased Nox4 and phosphorylated Smad2/3 protein expression in wound granulation tissues. Conclusions. Our data indicate that TGFβ1-mediated activation of TRPC3 enhances mitochondrial calcium and ROS production, which promotes myofibroblast transdifferentiation and HTS formation. Inhibition of the TRPC3-mediated Nox4/pSmad2/3 pathway may be a useful strategy to limit HTS formation after injury.

TRPV2 ion channels expressed in inhibitory motor neurons of gastric myenteric plexus contribute to gastric adaptive relaxation and gastric emptying in mice

2013 ◽  
Vol 304 (3) ◽  
pp. G235-G240 ◽  
Author(s):  
Hiroshi Mihara ◽  
Nobuhiro Suzuki ◽  
Hidemoto Yamawaki ◽  
Makoto Tominaga ◽  
Toshiro Sugiyama
Keyword(s):  
Nitric Oxide ◽  
Gastrointestinal Tract ◽  
Gastric Emptying ◽  
Motor Neurons ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Intragastric Pressure ◽  
Inhibitory Motor Neurons

Gastric adaptive relaxation (GAR) is impaired in ∼40% of functional dyspepsia (FD) patients, and nitric oxide (NO) released from inhibitory motor neurons plays an important role in this relaxation. Although the underlying molecular mechanism of GAR is poorly understood, transient receptor potential channel vanilloid 2 (TRPV2) mechano- and chemoreceptors are expressed in mouse intestinal inhibitory motor neurons and are involved in intestinal relaxation. The aim of this study was to evaluate the distribution of TRPV2 in inhibitory motor neurons throughout the mouse gastrointestinal tract and the contribution of TRPV2 to GAR. RT-PCR and immunohistochemical analyses were used to detect TRPV2 mRNA and protein, respectively. Intragastric pressure was determined with an isolated mouse stomach. Gastric emptying (GE) in vivo was determined using a test meal. TRPV2 mRNA was detected throughout the mouse gastrointestinal tract, and TRPV2 immunoreactivity was detected in 84.3% of neuronal nitric oxide synthase-expressing myenteric neurons in the stomach. GAR, which was expressed as the rate of decline of intragastric pressure in response to volume stimuli, was significantly enhanced by the TRPV2 activator probenecid, and the enhancement was inhibited by the TRPV2 inhibitor tranilast. GE was significantly accelerated by TRPV2 agonist applications, and the probenecid-induced enhancement was significantly inhibited by tranilast coapplication. Mechanosensitive TRPV2 was expressed in inhibitory motor neurons in the mouse stomach and contributed to GAR and GE. TRPV2 may be a promising target for FD patients with impaired GAR.

scholarly journals Assessment of siRNA as a therapeutic molecule in Transient Receptor Potential Channel 5 gene silencing: a computational approach

2018 ◽  
Vol 5 (1) ◽  
pp. 1911-1922
Author(s):  
Bhooma Vijayaraghavan ◽  
Giri Padmanabhan ◽  
Kumaresan Ramanathan
Keyword(s):  
Free Energy ◽  
Gene Silencing ◽  
Target Genes ◽  
Transient Receptor Potential ◽  
Gc Content ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Minimum Free Energy ◽  
Transient Receptor

Background: Ion channels play a crucial role in Glomerular filter damage that contributes to albuminuria. Transient receptor potential channel 5 (TRPC5) gene mediating such damage, demand for its target specific inhibition by RNA interference mechanism. Designing and selecting potential siRNA for TRPC5 gene silencing by computational analysis. Materials & Methods: The mRNA sequence was retrieved from NCBI (National Center for Biotechnology Information). siRNA sequences were designed specifically from target genes using InvivoGen siRNA wizard software. Thermodynamic RNA-RNA interactions were used to evaluate the gene silencing efficiency by minimum free energy of hybridization; the hybridization structures were also obtained using BIBISERV2-RNAHybrid. Results: The minimum free energy of hybridization of the three designed siRNAs (siRNA1, siRNA2 and siRNA3) were as follows: -28.2 kcal/mol, -24.1 kcal/mol, and-25.6 kcal/mol. Their corresponding GC content were 47.62%, 52.38% and 47.62%, respectively. Thus, siRNA1 had the least minimum free energy of hybridization (i.e. -28.2 kcal/mol) with low GC content (47.62%), and high linearity with minimal h-b index and loop structure. Conclusion: RNAi therapy can provide a new platform for efficient and targeted therapeutics. Further in vivo investigations are necessary to further validate their efficacy.

scholarly journals The Transient Receptor Potential Channel, Vanilloid 5, Induces Chondrocyte Apoptosis via Ca2+ CaMKII–Dependent MAPK and Akt/ mTOR Pathways in a Rat Osteoarthritis Model

2018 ◽  
Vol 51 (5) ◽  
pp. 2309-2323 ◽  
Author(s):  
Yingliang Wei ◽  
Zhaofeng Jin ◽  
He Zhang ◽  
Shang Piao ◽  
Jinghan Lu ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Western Blotting ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Transient Receptor Potential Channel ◽  
Chondrocyte Apoptosis ◽  
Pathological Feature ◽  
Transient Receptor

Background/Aims: Chondrocyte apoptosis is a central pathological feature of cartilage in osteoarthritis (OA). Accumulating evidence suggests that calcium ions (Ca2+) are an important regulator of apoptosis. Previously, we reported that the transient receptor potential channel vanilloid (TRPV5) is upregulated in monoiodoacetic acid (MIA)-induced OA articular cartilage. Methods: The protein levels of TRPV5, phosphorylated Ca2+/calmodulin-dependent kinase II (p-CaMKII), and total CaMKII were detected in vivo using western blotting techniques. Primary chondrocytes were isolated and cultured in vitro. Then, p-CAMKII was immunolocalized by immunofluorescence in chondrocytes. Fluo-4AM staining was used to assess intracellular Ca2+. Annexin V-fluorescein isothiocyanate / propidium iodide flow cytometric analysis was performed to determine chondrocyte apoptosis. Western blotting techniques were used to measure the expression of apoptosis-related proteins. Results: We found that ruthenium red (aTRPV5inhibitor)or(1-[N,O-bis-(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperaze (KN-62) (an inhibitor of Ca2+/calmodulin-dependent kinase II (CaMKII) phosphorylation) can relieve or even reverse OA in vivo. We found that TRPV5 has a specific role in mediating extracellular Ca2+ influx leading to chondrocyte apoptosis in vitro. The apoptotic effect in chondrocytes was inhibited by KN-62. We found that activated p-CaMKII could elicit the phosphorylation of extracellular signal-regulated protein kinase 1/2, c-Jun N-terminal kinase, and p38, three important regulators of the mitogen-activated protein kinase (MAPK) cascade. Moreover, we also showed that activated p-CaMKII could elicit the phosphorylation of protein kinase B (Akt) and two important downstream regulators of mammalian target of rapamycin (mTOR): 4E-binding protein, and S61 kinase. Conclusion: Our results demonstrate that upregulated TRPV5 may be an important initiating factor that activates CaMKII phosphorylation via the mediation of Ca2+ influx. In turn, activated p-CaMKII plays a critical role in chondrocyte apoptosis via MAPK and Akt/mTOR pathways.

Sign in / Sign up

Export Citation Format

Share Document