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

Calcium Influx ◽

Receptor Potential ◽

Membrane Depolarization ◽

Macromolecular Complex ◽

Trpc Channels ◽

Neuron Excitability ◽

Intracellular Ca ◽

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.

TRPC1/5-CaV3 Complex Mediates Leptin-Induced Excitability in Hypothalamic Neurons

Frontiers in Neuroscience ◽

10.3389/fnins.2021.679078 ◽

2021 ◽

Vol 15 ◽

Author(s):

Paula P. Perissinotti ◽

Elizabeth Martínez-Hernández ◽

Erika S. Piedras-Rentería

Keyword(s):

Receptor Potential ◽

Membrane Depolarization ◽

Resting Membrane Potential ◽

Macromolecular Complex ◽

Trpc Channels ◽

Hypothalamic Neurons ◽

Trpc Channel ◽

Neuron Excitability

Leptin regulates hypothalamic POMC+ (pro-opiomelanocortin) neurons by inducing TRPC (Transient Receptor Potential Cation) channel-mediate membrane depolarization. The role of TRPC channels in POMC neuron excitability is clearly established; however, it remains unknown whether their activity alone is sufficient to trigger excitability. Here we show that the right-shift voltage induced by the leptin-induced TRPC channel-mediated depolarization of the resting membrane potential brings T-type channels into the active window current range, resulting in an increase of the steady state T-type calcium current from 40 to 70% resulting in increased intrinsic excitability of POMC neurons. We assessed the role and timing of T-type channels on excitability and leptin-induced depolarization in vitro in cultured mouse POMC neurons. The involvement of TRPC channels in the leptin-induced excitability of POMC neurons was corroborated by using the TRPC channel inhibitor 2APB, which precluded the effect of leptin. 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. Furthermore, co-immunoprecipitation experiments suggest that TRPC1/5 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. However, leptin-induced depolarization still occurred in the presence of either BAPTA or EGTA suggesting that the calcium entry necessary to self-activate the TRPC1/5 complex is not blocked by the presence of BAPTA in hypothalamic neurons. Our study establishes T-type channels as integral part of the signaling cascade induced by leptin, modulating 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.

Design, Synthesis and In Vitro Experimental Validation of Novel TRPV4 Antagonists Inspired by Labdane Diterpenes

Marine Drugs ◽

10.3390/md18100519 ◽

2020 ◽

Vol 18 (10) ◽

pp. 519

Author(s):

Sarah Mazzotta ◽

Gabriele Carullo ◽

Aniello Schiano Moriello ◽

Pietro Amodeo ◽

Vincenzo Di Marzo ◽

...

Keyword(s):

Calcium Influx ◽

Biological Activities ◽

Receptor Potential ◽

Transient Receptor Potential Channel ◽

Design Synthesis ◽

Cellular Assays ◽

Transient Receptor ◽

Trpv4 Channel

Labdane diterpenes are widespread classes of natural compounds present in variety of marine and terrestrial organisms and plants. Many of them represents “natural libraries” of compounds with interesting biological activities due to differently functionalized drimane nucleus exploitable for potential pharmacological applications. The transient receptor potential channel subfamily V member 4 (TRPV4) channel has recently emerged as a pharmacological target for several respiratory diseases, including the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Inspired by the labdane-like bicyclic core, a series of homodrimane-derived esters and amides was designed and synthesized by modifying the flexible tail in position 1 of (+)-sclareolide, an oxidized derivative of the bioactive labdane-type diterpene sclareol. The potency and selectivity towards rTRPV4 and hTRPV1 receptors were assessed by calcium influx cellular assays. Molecular determinants critical for eliciting TRPV4 antagonism were identified by structure-activity relationships. Among the selective TRPV4 antagonists identified, compound 6 was the most active with an IC50 of 5.3 μM. This study represents the first report of semisynthetic homodrimane TRPV4 antagonists, selective over TRPV1, and potentially useful as pharmacological tools for the development of novel TRPV4 channel modulators.

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

Molecular Biology of the Cell ◽

10.1091/mbc.e07-08-0765 ◽

2008 ◽

Vol 19 (8) ◽

pp. 3203-3211 ◽

Cited By ~ 54

Author(s):

Pinaki Chaudhuri ◽

Scott M. Colles ◽

Manjunatha Bhat ◽

David R. Van Wagoner ◽

Lutz Birnbaumer ◽

...

Keyword(s):

Intracellular Calcium ◽

Calcium Concentration ◽

Calcium Influx ◽

Cell Movement ◽

Receptor Potential ◽

Receptor Activation ◽

Calcium Stores ◽

Trpc Channels ◽

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.

Role of Transient Receptor Potential Channels in Cholecystokinin-Induced Activation of Cultured Vagal Afferent Neurons

Endocrinology ◽

10.1210/en.2010-0504 ◽

2010 ◽

Vol 151 (11) ◽

pp. 5237-5246 ◽

Cited By ~ 16

Author(s):

Huan Zhao ◽

Steven M. Simasko

Keyword(s):

Receptor Potential ◽

Cytosolic Calcium ◽

Membrane Depolarization ◽

Induced Response ◽

Trpc Channels ◽

Afferent Neurons ◽

Trpv Channels ◽

Vagal Afferent ◽

Cholecystokinin (CCK), an endogenous brain-gut peptide, is released after food intake and promotes the process of satiation via activation of the vagus nerve. In vitro, CCK increases cytosolic calcium concentrations and produces membrane depolarization in a subpopulation of vagal afferent neurons. However, the specific mechanisms and ionic conductances that mediate these effects remain unclear. In this study we used calcium imaging, electrophysiological measurements, and single cell PCR analysis on cultured vagal afferent neurons to address this issue directly. A cocktail of blockers of voltage-dependent calcium channels (VDCC) failed to block CCK-induced calcium responses. In addition, SKF96365, a compound that blocks both VDCC and the C family of transient receptor potential (TRP) channels, also failed to prevent responses to CCK. Together these results suggest that CCK-induced calcium influx is not subsequent to the membrane depolarization. Ruthenium red, an inhibitor of the TRPV family and TRPA1, blocked both depolarizing responses to CCK and CCK-induced calcium increases, but had no effect on the KCl-induced calcium response. Selective block of TRPV1 and TRPA1 channels with SB366791 and HC030031, respectively, had minor effects on the CCK-induced response. Application of 2-aminoethoxydiphenyl borate, an activator of select TRPV channels but a blocker of several TRPC channels, either had no effect or enhanced the responses to CCK. Further, results from PCR experiments revealed a significant clustering of TRPV2-5 in neurons expressing CCK1 receptors. These observations demonstrate that CCK-induced increases in cytosolic calcium and membrane depolarization of vagal afferent neurons are likely mediated by TRPV channels, excluding TRPV1.

Membrane translocation of TRPC6 channels and endothelial migration are regulated by calmodulin and PI3 kinase activation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1600371113 ◽

2016 ◽

Vol 113 (8) ◽

pp. 2110-2115 ◽

Cited By ~ 22

Author(s):

Pinaki Chaudhuri ◽

Michael A. Rosenbaum ◽

Pritam Sinharoy ◽

Derek S. Damron ◽

Lutz Birnbaumer ◽

...

Keyword(s):

Cell Membrane ◽

Receptor Potential ◽

Oxidation Products ◽

Site Directed Mutagenesis ◽

Trpc Channels ◽

Lipid Oxidation Products ◽

Lipid oxidation products, including lysophosphatidylcholine (lysoPC), activate canonical transient receptor potential 6 (TRPC6) channels leading to inhibition of endothelial cell (EC) migration in vitro and delayed EC healing of arterial injuries in vivo. The precise mechanism through which lysoPC activates TRPC6 channels is not known, but calmodulin (CaM) contributes to the regulation of TRPC channels. Using site-directed mutagenesis, cDNAs were generated in which Tyr99 or Tyr138 of CaM was replaced with Phe, generating mutant CaM, Phe99-CaM, or Phe138-CaM, respectively. In ECs transiently transfected with pcDNA3.1-myc-His-Phe99-CaM, but not in ECs transfected with pcDNA3.1-myc-His-Phe138-CaM, the lysoPC-induced TRPC6-CaM dissociation and TRPC6 externalization was disrupted. Also, the lysoPC-induced increase in intracellular calcium concentration was inhibited in ECs transiently transfected with pcDNA3.1-myc-His-Phe99-CaM. Blocking phosphorylation of CaM at Tyr99 also reduced CaM association with the p85 subunit and subsequent activation of phosphatidylinositol 3-kinase (PI3K). This prevented the increase in phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and the translocation of TRPC6 to the cell membrane and reduced the inhibition of EC migration by lysoPC. These findings suggest that lysoPC induces CaM phosphorylation at Tyr99 by a Src family kinase and that phosphorylated CaM activates PI3K to produce PIP3, which promotes TRPC6 translocation to the cell membrane.

BDNF Induces Calcium Elevations Associated With IBDNF, a Nonselective Cationic Current Mediated by TRPC Channels

Journal of Neurophysiology ◽

10.1152/jn.00797.2007 ◽

2007 ◽

Vol 98 (4) ◽

pp. 2476-2482 ◽

Cited By ~ 42

Author(s):

Michelle D. Amaral ◽

Lucas Pozzo-Miller

Keyword(s):

Hippocampal Neurons ◽

Pyramidal Neurons ◽

Receptor Potential ◽

Trpc Channels ◽

Hippocampal Pyramidal Neurons ◽

Cationic Current ◽

Intracellular Ca ◽

Transient Receptor ◽

Nonselective Cationic Current

Brain-derived neurotrophic factor (BDNF) has potent actions on hippocampal neurons, but the mechanisms that initiate its effects are poorly understood. We report here that localized BDNF application to apical dendrites of CA1 pyramidal neurons evoked transient elevations in intracellular Ca2+ concentration, which are independent of membrane depolarization and activation of N-methyl-d-aspartate receptors (NMDAR). These Ca2+ signals were always associated with IBDNF, a slow and sustained nonselective cationic current mediated by transient receptor potential canonical (TRPC3) channels. BDNF-induced Ca2+ elevations required functional Trk and inositol-tris-phosphate (IP3) receptors, full intracellular Ca2+ stores as well as extracellular Ca2+, suggesting the involvement of TRPC channels. Indeed, the TRPC channel inhibitor SKF-96365 prevented BDNF-induced Ca2+ elevations and the associated IBDNF. Thus TRPC channels emerge as novel mediators of BDNF-induced intracellular Ca2+ elevations associated with sustained cationic membrane currents in hippocampal pyramidal neurons.

TRPC channels: Dysregulation and Ca2+ Mishandling In Ischemic Heart Disease

10.20944/preprints201912.0339.v1 ◽

2019 ◽

Author(s):

Debora Falcon ◽

Isabel Galeano-Otero ◽

Marta Martín-Bórnez ◽

María Fernández-Velasco ◽

Isabel Gallardo-Castillo ◽

...

Keyword(s):

Current Knowledge ◽

Receptor Potential ◽

Cardiac Cells ◽

Membrane Receptors ◽

Trpc Channels ◽

Ischemia And Reperfusion ◽

Sense And Respond ◽

Transient receptor potential canonical (TRPC) channels are ubiquitously expressed in excitable and non-excitable cardiac cells where they sense and respond to a wide variety of physical and chemical stimuli. As other TRP, TRPC may form homo or heterotetrameric ion channel, and they can associate with other membrane receptors and ion channels to regulate intracellular calcium concentration. Dysfunctions of TRPC channels are involved in many types of cardiovascular diseases. Significant increase of the expression of different TRPC isoforms has been observed in different animal model of heart infarcts, and in vitro experimental model of ischemia and reperfusion. TRPC-mediated increase of the intracellular Ca2+ concentration seems required for the activation of signaling pathway that plays minor roles in the healthy heart, but they are more relevant for cardiac responses to ischemia, such as the activation of different factors of transcription and cardiac hypertrophy, fibrosis, and angiogenesis. In this review, we will highlight the current knowledge regarding TRPC implication in different cellular processes related to ischemia and reperfusion, and to heart infarction.

TRPC channels blockade abolishes septic cardiac dysfunction by hampering intracellular inflammation and Ca2+ leakage

10.21203/rs.3.rs-419385/v1 ◽

2021 ◽

Author(s):

Wei Cao ◽

Na Tang ◽

Wen Tian ◽

Guang-Yuan Ma ◽

Xiong Xiao ◽

...

Keyword(s):

Heart Failure ◽

Endoplasmic Reticulum ◽

Cardiac Dysfunction ◽

Receptor Potential ◽

Trpc Channels ◽

Regulatory Modules ◽

Novel Approach ◽

Intracellular Ca ◽

Abstract Intracellular Ca2+ dysregulation is a key marker in septic cardiac dysfunction; however, regulation of the classic Ca2+ regulatory modules cannot successfully abolish this symptom. The present study shows that the knockout of transient receptor potential canonical (TRPC) channel isoforms, TRPC1 and TRPC6, can strikingly ameliorate LPS-challenged heart failure and prolong survivability in mice. The LPS-triggered Ca2+ release from the endoplasmic reticulum both in cardiomyocytes and macrophages is significantly inhibited by Trpc1 or Trpc6 knockout. Meanwhile, TRPC’s molecular partner, calmodulin is uncoupled during Trpc1 or Trpc6 deficiency and binds to TLR4’s Pococurante site and atypical isoleucine-glutamine-like motif to block the inflammation cascade. Importantly, blocking the C-terminal CaM/IP3R binding domain in TRPC with chemical inhibitor could markedly obstruct the Ca2+ leak and TLR4-mediated inflammation burst, demonstrating a powerful cardioprotective effect in endotoxemia and polymicrobial sepsis. Our findings provide new insights into the pathogenesis of septic cardiac dysfunction and suggest a novel approach for its treatment.

Linalool odor‐induced analgesia is triggered by TRPA1-independent pathway in mice

Behavioral and Brain Functions ◽

10.1186/s12993-021-00176-y ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Hideki Kashiwadani ◽

Yurina Higa ◽

Mitsutaka Sugimura ◽

Tomoyuki Kuwaki

Keyword(s):

Pain Threshold ◽

Receptor Potential ◽

Analgesic Effects ◽

Neuronal Mechanisms ◽

Odor Exposure ◽

Noxious Mechanical Stimulation ◽

Transient Receptor ◽

Deficient Mice

AbstractWe had recently reported that linalool odor exposure induced significant analgesic effects in mice and that the effects were disappeared in olfactory-deprived mice in which the olfactory epithelium was damaged, thus indicating that the effects were triggered by chemical senses evoked by linalool odor exposure. However, the peripheral neuronal mechanisms, including linalool receptors that contribute toward triggering the linalool odor-induced analgesia, still remain unexplored. In vitro studies have shown that the transient receptor potential ankyrin 1 (TRPA1) responded to linalool, thus raising the possibility that TRPA1 expressed on the trigeminal nerve terminal detects linalool odor inhaled into the nostril and triggers the analgesic effects. To address this hypothesis, we measured the behavioral pain threshold for noxious mechanical stimulation in TRPA1-deficient mice. In contrast to our expectation, we found a significant increase in the threshold after linalool odor exposure in TRPA1-deficient mice, indicating the analgesic effects of linalool odor even in TRPA1-deficient mice. Furthermore, intranasal application of TRPA1 selective antagonist did not alter the analgesic effect of linalool odor. These results showed that the linalool odor-induced analgesia was triggered by a TRPA1-independent pathway in mice.

Satellite glial cells in sensory ganglia express functional transient receptor potential ankyrin 1 that is sensitized in neuropathic and inflammatory pain

Molecular Pain ◽

10.1177/1744806920925425 ◽

2020 ◽

Vol 16 ◽

pp. 174480692092542 ◽

Cited By ~ 2

Author(s):

Seung Min Shin ◽

Brandon Itson-Zoske ◽

Yongsong Cai ◽

Chensheng Qiu ◽

Bin Pan ◽

...

Keyword(s):

Neuropathic Pain ◽

Nerve Injury ◽

Glial Cells ◽

Receptor Potential ◽

Allyl Isothiocyanate ◽

Sensory Ganglia ◽

Satellite Glial Cells ◽

Intracellular Ca ◽

Transient receptor potential ankyrin 1 (TRPA1) is well documented as an important molecule in pain hypersensitivity following inflammation and nerve injury and in many other cellular biological processes. Here, we show that TRPA1 is expressed not only by sensory neurons of the dorsal root ganglia (DRG) but also in their adjacent satellite glial cells (SGCs), as well as nonmyelinating Schwann cells. TRPA1 immunoreactivity is also detected in various cutaneous structures of sensory neuronal terminals, including small and large caliber cutaneous sensory fibers and endings. The SGC-expressed TRPA1 is functional. Like DRG neurons, dissociated SGCs exhibit a robust response to the TRPA1-selective agonist allyl isothiocyanate (AITC) by an increase of intracellular Ca2+ concentration ([Ca2+]i). These responses are abolished by the TRPA1 antagonist HC030031 and are absent in SGCs and neurons from global TRPA1 null mice. SGCs and neurons harvested from DRG proximal to painful tissue inflammation induced by plantar injection of complete Freund’s adjuvant show greater AITC-evoked elevation of [Ca2+]i and slower recovery compared to sham controls. Similar TRPA1 sensitization occurs in both SGCs and neurons during neuropathic pain induced by spared nerve injury. Together, these results show that functional TRPA1 is expressed by sensory ganglia SGCs, and TRPA1 function in SGCs is enhanced after both peripheral inflammation and nerve injury, and suggest that TRPA1 in SGCs may contribute to inflammatory and neuropathic pain.