Elucidation of a TRPC6-TRPC5 Channel Cascade That Restricts Endothelial Cell Movement

Canonical transient receptor potential (TRPC) channels are opened by classical signal transduction events initiated by receptor activation or depletion of intracellular calcium stores. Here, we report a novel mechanism for opening TRPC channels in which TRPC6 activation initiates a cascade resulting in TRPC5 translocation. When endothelial cells (ECs) are incubated in lysophosphatidylcholine (lysoPC), rapid translocation of TRPC6 initiates calcium influx that results in externalization of TRPC5. Activation of this TRPC6–5 cascade causes a prolonged increase in intracellular calcium concentration ([Ca2+]i) that inhibits EC movement. When TRPC5 is down-regulated with siRNA, the lysoPC-induced rise in [Ca2+]i is shortened and the inhibition of EC migration is lessened. When TRPC6 is down-regulated or EC from TRPC6−/− mice are studied, lysoPC has minimal effect on [Ca2+]i and EC migration. In addition, TRPC5 is not externalized in response to lysoPC, supporting the dependence of TRPC5 translocation on the opening of TRPC6 channels. Activation of this novel TRPC channel cascade by lysoPC, resulting in the inhibition of EC migration, could adversely impact on EC healing in atherosclerotic arteries where lysoPC is abundant.

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Erythropoietin-modulated calcium influx through TRPC2 is mediated by phospholipase Cγ and IP3R

AJP Cell Physiology ◽

10.1152/ajpcell.00265.2004 ◽

2004 ◽

Vol 287 (6) ◽

pp. C1667-C1678 ◽

Cited By ~ 39

Author(s):

Qin Tong ◽

Xin Chu ◽

Joseph Y. Cheung ◽

Kathleen Conrad ◽

Richard Stahl ◽

...

Keyword(s):

Calcium Concentration ◽

Transient Receptor Potential ◽

Calcium Influx ◽

Receptor Potential ◽

Signaling Complex ◽

Binding Domains ◽

Transient Receptor ◽

Dose Dependent Increase ◽

Dose Dependent ◽

Inactive Analog

In the present study, we examined the mechanisms through which erythropoietin (Epo) activates the calcium-permeable transient receptor potential protein channel (TRPC)2. Erythroblasts were isolated from the spleens of phenylhydrazine-treated mice, and Epo stimulation resulted in a significant and dose-dependent increase in intracellular calcium concentration ([Ca2+]i). This increase in [Ca2+]iwas inhibited by pretreatment with the phospholipase C (PLC) inhibitor U-73122 but not by the inactive analog U-73343, demonstrating the requirement for PLC activity in Epo-modulated Ca2+influx in primary erythroid cells. To determine whether PLC is involved in the activation of TRPC2 by Epo, cell models were used to examine this interaction. Single CHO-S cells that expressed transfected Epo receptor (Epo-R) and TRPC2 were identified, and [Ca2+]iwas quantitated. Epo-induced Ca2+influx through TRPC2 was inhibited by pretreatment with U-73122 or by downregulation of PLCγ1 by RNA interference. PLC activation results in the production of inositol 1,4,5-trisphosphate (IP3), and TRPC2 has IP3receptor (IP3R) binding sites. To determine whether IP3R is involved in Epo-R signaling, TRPC2 mutants were prepared with partial or complete deletions of the COOH-terminal IP3R binding domains. In cells expressing TRPC2 IP3R binding mutants and Epo-R, no significant increase in [Ca2+]iwas observed after Epo stimulation. TRPC2 coassociated with Epo-R, PLCγ, and IP3R, and the association between TRPC2 and IP3R was disrupted in these mutants. Our data demonstrate that Epo-R modulates TRPC2 activation through PLCγ; that interaction of IP3R with TRPC2 is required; and that Epo-R, TRPC2, PLCγ, and IP3R interact to form a signaling complex.

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TRPC5-CaV3 complex mediates Leptin-induced excitability in hypothalamic neurons

10.1101/2020.07.21.214296 ◽

2020 ◽

Author(s):

Paula P. Perissinotti ◽

Elizabeth Martínez-Hernández ◽

Erika S. Piedras-Rentería

Keyword(s):

Transient Receptor Potential ◽

Calcium Influx ◽

Receptor Potential ◽

Membrane Depolarization ◽

Macromolecular Complex ◽

Trpc Channels ◽

Neuron Excitability ◽

Intracellular Ca ◽

Transient Receptor

ABSTRACTLeptin regulates hypothalamic POMC+ (pro-opiomelanocortin) neurons by inducing TRPC (Transient Receptor Potential Cation) channel-mediate membrane depolarization. Here we assessed the role of T-type channels on POMC neuron excitability and leptin-induced depolarization in vitro. We demonstrate T-type currents are indispensable for both processes, as treatment with NNC-55-0396 prevented the membrane depolarization and rheobase changes induced by leptin in cultured mouse POMC neurons. Furthermore, we demonstrate TRPC1/C5 channels and CaV3.1 and CaV3.2 channels co-exist in complex. The functional relevance of this complex was corroborated using intracellular Ca2+ chelators; intracellular BAPTA (but not EGTA) application was sufficient to preclude POMC neuron excitability by preventing leptin-induced calcium influx through TRPC channels and T-type channel function.We conclude T-type channels are integral in POMC neuron excitability. Leptin activation of TRPC channels existing in a macromolecular complex with T-type channels recruits the latter by locally-induced membrane depolarization, further depolarizing POMC neurons, triggering action potentials and excitability.

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Emerging Roles of Diacylglycerol-Sensitive TRPC4/5 Channels

Cells ◽

10.3390/cells7110218 ◽

2018 ◽

Vol 7 (11) ◽

pp. 218 ◽

Cited By ~ 9

Author(s):

Michael Mederos y Schnitzler ◽

Thomas Gudermann ◽

Ursula Storch

Keyword(s):

Transient Receptor Potential ◽

Receptor Potential ◽

Receptor Activation ◽

Activation Mechanism ◽

Regulatory Function ◽

Cation Channels ◽

Trpc Channels ◽

Trpc Channel ◽

Transient Receptor ◽

Trpc5 Channels

Transient receptor potential classical or canonical 4 (TRPC4) and TRPC5 channels are members of the classical or canonical transient receptor potential (TRPC) channel family of non-selective cation channels. TRPC4 and TRPC5 channels are widely accepted as receptor-operated cation channels that are activated in a phospholipase C-dependent manner, following the Gq/11 protein-coupled receptor activation. However, their precise activation mechanism has remained largely elusive for a long time, as the TRPC4 and TRPC5 channels were considered as being insensitive to the second messenger diacylglycerol (DAG) in contrast to the other TRPC channels. Recent findings indicate that the C-terminal interactions with the scaffolding proteins Na+/H+ exchanger regulatory factor 1 and 2 (NHERF1 and NHERF2) dynamically regulate the DAG sensitivity of the TRPC4 and TRPC5 channels. Interestingly, the C-terminal NHERF binding suppresses, while the dissociation of NHERF enables, the DAG sensitivity of the TRPC4 and TRPC5 channels. This leads to the assumption that all of the TRPC channels are DAG sensitive. The identification of the regulatory function of the NHERF proteins in the TRPC4/5-NHERF protein complex offers a new starting point to get deeper insights into the molecular basis of TRPC channel activation. Future studies will have to unravel the physiological and pathophysiological functions of this multi-protein channel complex.

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Discrimination of intracellular calcium store subcompartments using TRPV1 (transient receptor potential channel, vanilloid subfamily member 1) release channel activity

Biochemical Journal ◽

10.1042/bj20021381 ◽

2003 ◽

Vol 371 (2) ◽

pp. 341-350 ◽

Cited By ~ 78

Author(s):

Helen TURNER ◽

Andrea FLEIG ◽

Alexander STOKES ◽

Jean-Pierre KINET ◽

Reinhold PENNER

Keyword(s):

Intracellular Calcium ◽

Transient Receptor Potential ◽

Receptor Potential ◽

Transient Receptor Potential Channel ◽

Calcium Stores ◽

Calcium Store ◽

Release Channel ◽

Subfamily Member ◽

Store Depletion ◽

Transient Receptor

The store-operated calcium-release-activated calcium current, ICRAC, is a major mechanism for calcium entry into non-excitable cells. ICRAC refills calcium stores and permits sustained calcium signalling. The relationship between inositol 1,4,5-trisphosphate receptor (InsP3R)-containing stores and ICRAC is not understood. A model of global InsP3R store depletion coupling with ICRAC activation may be simplistic, since intracellular stores are heterogeneous in their release and refilling activities. Here we use a ligand-gated calcium channel, TRPV1 (transient receptor potential channel, vanilloid subfamily member 1), as a new tool to probe store heterogeneity and define intracellular calcium compartments in a mast cell line. TRPV1 has activity as an intracellular release channel but does not mediate global calcium store depletion and does not invade a store coupled with ICRAC. Intracellular TRPV1 localizes to a subset of the InsP3R-containing stores. TRPV1 sensitivity functionally subdivides the InsP3-sensitive store, as does heterogeneity in the sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase isoforms responsible for store refilling. These results provide unequivocal evidence that a specific ‘CRAC store’ exists within the InsP3-releasable calcium stores and describe a novel methodology for manipulation of intracellular free calcium.

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Abstract 138: Transient Receptor Potential Channel C1 Deficiency Protects the Murine Heart from Pressure Overload

Circulation ◽

10.1161/circ.116.suppl_16.ii_5-a ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Malini Seth ◽

Zhu-Shan Zhang ◽

Lan Mao ◽

Jarrett Burch ◽

Victoria Graham ◽

...

Keyword(s):

Cardiac Hypertrophy ◽

Transient Receptor Potential ◽

Pressure Overload ◽

Receptor Potential ◽

Receptor Activation ◽

Trpc Channels ◽

Loss Of Function ◽

Aortic Banding ◽

Trpc1 Channels ◽

Transient Receptor

Transient receptor potential canonical (TRPC) channels are non-selective cation channels that are activated in response to G-protein coupled receptor activation, depletion of internal stores and mechanical stretch. Recent reports suggest that cardiac TRPC channels mediate calcineurin dependent cardiac hypertrophy, yet few details exist as to the mechanism for activation of these channels. Here, we provide evidence that TRPC1 channels are the dominant TRPC channel in mouse cardiomyocytes and cardiac TRPC1 protein expression is augmented by seven fold following thoracic aortic banding (TAC). In addition, we provide the first loss of function studies to show that mice lacking TRPC1 channels developed significantly less cardiac hypertrophy following pressure overload induced by thoracic aortic banding suggesting that TRPC1 may confer deleterious calcium entry. Whole cell voltage clamp studies of isolated adult cardiomyocytes reveal a non-selective cation current that is induced by pressure overload that is absent in TRPC1−/− cardiomyocytes and in which TRP blockers such as gadolinium, 2-amino biphenyl boric acid and SKF96365 inhibit the TAC induced current. Finally, neonatal cardiomyocytes lacking functional TRPC1 display reduced TRPC current in response to cell stretch or angiotensin-II; the functional consequence of which includes reduced calcium oscillation frequency and reduced BNP expression. These results provide the first loss of function evidence for TRPC1 channels in cardiac hypertrophy and implicate TRPC1 as a stretch activated channel.

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Molecular Modification of Transient Receptor Potential Canonical 6 Channels Modulates Calcium Dyshomeostasis in a Mouse Model Relevant to Malignant Hyperthermia

Anesthesiology ◽

10.1097/aln.0000000000003635 ◽

2020 ◽

Author(s):

Jose Rafael Lopez ◽

Arkady Uryash ◽

Jose Adams ◽

Philip M. Hopkins ◽

Paul D. Allen

Keyword(s):

Mouse Model ◽

Malignant Hyperthermia ◽

Intracellular Calcium ◽

Transient Receptor Potential ◽

Receptor Potential ◽

Dominant Negative ◽

Trpc Channels ◽

Wild Type ◽

Trpc6 Channel ◽

Transient Receptor

Background Pharmacologic modulation has previously shown that transient receptor potential canonical (TRPC) channels play an important role in the pathogenesis of malignant hyperthermia. This study tested the hypothesis that genetically suppressing the function of TRPC6 can partially ameliorate muscle cation dyshomeostasis and the response to halothane in a mouse model relevant to malignant hyperthermia. Methods This study examined the effect of overexpressing a muscle-specific nonconducting dominant-negative TRPC6 channel in 20 RYR1-p.R163C and 20 wild-type mice and an equal number of nonexpressing controls, using calcium- and sodium-selective microelectrodes and Western blots. Results RYR1-p.R163C mouse muscles have chronically elevated intracellular calcium and sodium levels compared to wild-type muscles. Transgenic expression of the nonconducting TRPC6 channel reduced intracellular calcium from 331 ± 34 nM (mean ± SD) to 190 ± 27 nM (P < 0.0001) and sodium from 15 ± 1 mM to 11 ± 1 mM (P < 0.0001). Its expression lowered the increase in intracellular Ca2+ of the TRPC6-specific activator hyperforin in RYR1-p.R163C muscle fibers from 52% (348 ± 37 nM to 537 ± 70 nM) to 14% (185 ± 11 nM to 210 ± 44 nM). Western blot analysis of TRPC3 and TRPC6 expression showed the expected increase in TRPC6 caused by overexpression of its dominant-negative transgene and a compensatory increase in expression of TRPC3. Although expression of the muscle-specific dominant-negative TRPC6 was able to modulate the increase in intracellular calcium during halothane exposure and prolonged life (35 ± 5 min vs. 15 ± 3 min; P < 0.0001), a slow, steady increase in calcium began after 20 min of halothane exposure, which eventually led to death. Conclusions These data support previous findings that TRPC channels play an important role in causing the intracellular calcium and sodium dyshomeostasis associated with RYR1 variants that are pathogenic for malignant hyperthermia. However, they also show that modulating TRPC channels alone is not sufficient to prevent the lethal effect of exposure to volatile anesthetic malignant hyperthermia–triggering agents. Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New

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TRPC6 channel as an emerging determinant of the podocyte injury susceptibility in kidney diseases

AJP Renal Physiology ◽

10.1152/ajprenal.00186.2015 ◽

2015 ◽

Vol 309 (5) ◽

pp. F393-F397 ◽

Cited By ~ 44

Author(s):

Daria V. Ilatovskaya ◽

Alexander Staruschenko

Keyword(s):

Transient Receptor Potential ◽

Calcium Influx ◽

Kidney Diseases ◽

Critical Role ◽

Receptor Potential ◽

Calcium Flux ◽

Calcium Handling ◽

Podocyte Injury ◽

Trpc6 Channel ◽

Transient Receptor

Podocytes (terminally differentiated epithelial cells of the glomeruli) play a key role in the maintenance of glomerular structure and permeability and in the incipiency of various renal abnormalities. Injury to podocytes is considered a major contributor to the development of kidney disease as their loss causes proteinuria and progressive glomerulosclerosis. The physiological function of podocytes is critically dependent on proper intracellular calcium handling; excessive calcium influx in these cells may result in the effacement of foot processes, apoptosis, and subsequent glomeruli damage. One of the key proteins responsible for calcium flux in the podocytes is transient receptor potential cation channel, subfamily C, member 6 (TRPC6); a gain-of-function mutation in TRPC6 has been associated with the onset of the familial forms of focal segmental glomerulosclerosis (FSGS). Recent data also revealed a critical role of this channel in the onset of diabetic nephropathy. Therefore, major efforts of the research community have been recently dedicated to unraveling the TRPC6-dependent effects in the initiation of podocyte injury. This mini-review focuses on the TRPC6 channel in podocytes and colligates recent data in an attempt to shed some light on the mechanisms underlying the pathogenesis of TRPC6-mediated glomeruli damage and its potential role as a therapeutic target for the treatment of chronic kidney diseases.

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The Na+/Ca2+ Exchanger Inhibitor, KB-R7943, Blocks a Nonselective Cation Channel Implicated in Chemosensory Transduction

Journal of Neurophysiology ◽

10.1152/jn.90903.2008 ◽

2009 ◽

Vol 101 (3) ◽

pp. 1151-1159 ◽

Cited By ~ 12

Author(s):

A. Pezier ◽

Y. V. Bobkov ◽

B. W. Ache

Keyword(s):

Transient Receptor Potential ◽

Single Channel ◽

Receptor Potential ◽

Trpc Channels ◽

Dependent Manner ◽

Open Probability ◽

Olfactory Transduction ◽

Voltage Dependent ◽

Chemosensory Transduction ◽

Transient Receptor

The mechanism(s) of olfactory transduction in invertebrates remains to be fully understood. In lobster olfactory receptor neurons (ORNs), a nonselective sodium-gated cation (SGC) channel, a presumptive transient receptor potential (TRP)C channel homolog, plays a crucial role in olfactory transduction, at least in part by amplifying the primary transduction current. To better determine the functional role of the channel, it is important to selectively block the channel independently of other elements of the transduction cascade, causing us to search for specific pharmacological blockers of the SGC channel. Given evidence that the Na+/Ca2+ exchange inhibitor, KB-R7943, blocks mammalian TRPC channels, we studied this probe as a potential blocker of the lobster SGC channel. KB-R7943 reversibly blocked the SGC current in both inside- and outside-out patch recordings in a dose- and voltage-dependent manner. KB-R7943 decreased the channel open probability without changing single channel amplitude. KB-R7943 also reversibly and in a dose-dependent manner inhibited both the odorant-evoked discharge of lobster ORNs and the odorant-evoked whole cell current. Our findings strongly imply that KB-R7943 potently blocks the lobster SGC channel and likely does so directly and not through its ability to block the Na+/Ca2+ exchanger.

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Mineralocorticoid receptor antagonism improves parenchymal arteriole dilation via a TRPV4-dependent mechanism and prevents cognitive dysfunction in hypertension

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00207.2018 ◽

2018 ◽

Vol 315 (5) ◽

pp. H1304-H1315 ◽

Cited By ~ 10

Author(s):

Janice M. Diaz-Otero ◽

Ting-Chieh Yen ◽

Courtney Fisher ◽

Daniel Bota ◽

William F. Jackson ◽

...

Keyword(s):

Cognitive Function ◽

Angiotensin Ii ◽

Mineralocorticoid Receptor ◽

Transient Receptor Potential ◽

Receptor Potential ◽

Receptor Activation ◽

Myogenic Tone ◽

Transient Receptor Potential Vanilloid ◽

Cerebrovascular Function ◽

Transient Receptor

Hypertension and mineralocorticoid receptor activation cause cerebral parenchymal arteriole remodeling; this can limit cerebral perfusion and contribute to cognitive dysfunction. We used a mouse model of angiotensin II-induced hypertension to test the hypothesis that mineralocorticoid receptor activation impairs both transient receptor potential vanilloid (TRPV)4-mediated dilation of cerebral parenchymal arterioles and cognitive function. Mice (16−18 wk old, male, C57Bl/6) were treated with angiotensin II (800 ng·kg−1·min−1) with or without the mineralocorticoid receptor antagonist eplerenone (100 mg·kg−1·day−1) for 4 wk; sham mice served as controls. Data are presented as means ± SE; n = 5–14 mice/group. Eplerenone prevented the increased parenchymal arteriole myogenic tone and impaired carbachol-induced (10−9–10−5 mol/l) dilation observed during hypertension. The carbachol-induced dilation was endothelium-derived hyperpolarization mediated because it could not be blocked by N-nitro-l-arginine methyl ester (10−5 mol/l) and indomethacin (10−4 mol/l). We used GSK2193874 (10−7 mol/l) to confirm that in all groups this dilation was dependent on TRPV4 activation. Dilation in response to the TRPV4 agonist GSK1016790A (10−9–10−5 mol/l) was also reduced in hypertensive mice, and this defect was corrected by eplerenone. In hypertensive and eplerenone-treated animals, TRPV4 inhibition reduced myogenic tone, an effect that was not observed in arterioles from control animals. Eplerenone treatment also improved cognitive function and reduced microglia density in hypertensive mice. These data suggest that the mineralocorticoid receptor is a potential therapeutic target to improve cerebrovascular function and cognition during hypertension. NEW & NOTEWORTHY Vascular dementia is a growing public health issue that lacks effective treatments. Transient receptor potential vanilloid (TRPV)4 channels are important regulators of parenchymal arteriole dilation, and they modulate myogenic tone. The data presented here suggest that TRPV4 channel expression is regulated by the mineralocorticoid receptor (MR). MR blockade also improves cognitive function during hypertension. MR blockade might be a potential therapeutic approach to improve cerebrovascular function and cognition in patients with hypertension.

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Regulation of Transient Receptor Potential Canonical 4 Activity by Phospholipase C-δ1

10.21203/rs.3.rs-107953/v1 ◽

2020 ◽

Author(s):

Juyeon Ko ◽

Jongyun Myeong ◽

Misun Kwak ◽

Insuk So

Keyword(s):

Phospholipase C ◽

Transient Receptor Potential ◽

Resonance Energy Transfer ◽

Resonance Energy ◽

Receptor Potential ◽

Regulation Mechanism ◽

Trpc Channels ◽

Transient Receptor Potential Canonical ◽

Cell Current ◽

Transient Receptor

Abstract Transient receptor potential canonical (TRPC) channels are non-selective calcium-permeable cation channels. It is suggested that TRPC4β and TRPC5 channels are regulated by phospholipase C (PLC) signaling, and are especially maintained by phosphatidylinositol 4,5-bisphosphate (PIP2). The PLCδ subtype is the most Ca2+-sensitive form among the isozymes which cleaves phospholipids to respond to the calcium rise. In this study, we investigated the regulation mechanism of TRPC channel by Ca2+, PLCδ1 and PIP2 signaling cascades. The interaction between TRPC4β and PLCδ1 was identified through the Fӧster resonance energy transfer (FRET) and co-immunoprecipitation (Co-IP). With the electrophysiological experiments, we found that TRPC4β-bound PLCδ1 reduces the overall whole-cell current of channel. The Ca2+-via opened channel promotes the activation of PLCδ1, which subsequently decreases PIP2 level. By comparison TRPC4β activity with or without PLCδ1 using differently [Ca2+]i buffered solution, we demonstrated that PLCδ1 functions in normal condition with physiological calcium range. The negative regulation effect of PLCδ1 on TRPC4β helps to elucidate the roles of each PIP2 binding residues whether they are concerned in channel maintenance or inhibition of channel activity.

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