scholarly journals Validation of an LC-MS/MS Method to Quantify the New TRPC6 Inhibitor SH045 (Larixyl N-methylcarbamate) and Its Application in an Exploratory Pharmacokinetic Study in Mice

2021 ◽  
Vol 14 (3) ◽  
pp. 259
Author(s):  
Xiao-Ning Chai ◽  
Friedrich-Alexander Ludwig ◽  
Anne Müglitz ◽  
Michael Schaefer ◽  
Hai-Yan Yin ◽  
...  
Keyword(s):  
Time Course ◽  
Transient Receptor Potential ◽  
Intraperitoneal Administration ◽  
Receptor Potential ◽  
Pharmacokinetic Data ◽  
Animal Disease Models ◽  
Mammalian Tissues ◽  
Ic50 Values ◽  
Transient Receptor ◽  
Subtype Selective

TRPC6 (transient receptor potential cation channels; canonical subfamily C, member 6) is widespread localized in mammalian tissues like kidney and lung and associated with progressive proteinuria and pathophysiological pulmonary alterations, e.g., reperfusion edema or lung fibrosis. However, the understanding of TRPC6 channelopathies is still at the beginning stages. Recently, by chemical diversification of (+)-larixol originating from Larix decidua resin traditionally used for inhalation, its methylcarbamate congener, named SH045, was obtained and identified in functional assays as a highly potent, subtype-selective inhibitor of TRPC6. To pave the way for use of SH045 in animal disease models, this study aimed at developing a capable bioanalytical method and to provide exploratory pharmacokinetic data for this promising derivative. According to international guidelines, a robust and selective LC-MS/MS method based on MRM detection in positive ion mode was established and validated for quantification of SH045 in mice plasma, whereby linearity and accuracy were demonstrated for the range of 2–1600 ng/mL. Applying this method, the plasma concentration time course of SH045 following single intraperitoneal administration (20 mg/kg body weight) revealed a short half-life of 1.3 h. However, the pharmacological profile of SH045 is promising, as five hours after administration, plasma levels still remained sufficiently higher than published low nanomolar IC50 values. Summarizing, the LC-MS/MS method and exploratory pharmacokinetic data provide essential prerequisites for experimental pharmacological TRPC6 modulation and translational treatment of TRPC6 channelopathies.

scholarly journals Depletion of plasma membrane–associated phosphoinositides mimics inhibition of TRPM7 channels by cytosolic Mg2+, spermine, and pH

2018 ◽  
Vol 293 (47) ◽  
pp. 18151-18167 ◽  
Author(s):  
Tetyana Zhelay ◽  
Krystyna B. Wieczerzak ◽  
Pavani Beesetty ◽  
Gerald M. Alter ◽  
Masayuki Matsushita ◽  
...  
Keyword(s):  
Time Course ◽  
Transient Receptor Potential ◽  
Phosphatidyl Inositol ◽  
Receptor Potential ◽  
Hek293 Cells ◽  
Cell Interior ◽  
Physiological Conditions ◽  
Channel Interactions ◽  
Transient Receptor ◽  
Eef2 Kinase

Transient receptor potential cation channel subfamily M member 7 (TRPM7) is an ion channel/protein kinase belonging to the TRP melastatin and eEF2 kinase families. Under physiological conditions, most native TRPM7 channels are inhibited by cytoplasmic Mg2+, protons, and polyamines. Currents through these channels (ITRPM7) are robustly potentiated when the cell interior is exchanged with low Mg2+-containing buffers. ITRPM7 is also potentiated by phosphatidyl inositol bisphosphate (PI(4,5)P2) and suppressed by its hydrolysis. Here we characterized internal Mg2+- and pH-mediated inhibition of TRPM7 channels in HEK293 cells overexpressing WT voltage-sensing phospholipid phosphatase (VSP) or its catalytically inactive variant VSP-C363S. VSP-mediated depletion of membrane phosphoinositides significantly increased channel sensitivity to Mg2+ and pH. Proton concentrations that were too low to inhibit ITRPM7 when the VSP-C363S variant was expressed (pH 8.2) became inhibitory in WT VSP–expressing cells. At pH 6.5, protons inhibited ITRPM7 both in WT and VSP C363S–expressing cells but with a faster time course in the WT VSP–expressing cells. Inhibition by 150 μm Mg2+ was also significantly faster in the WT VSP–expressing cells. Cellular PI(4,5)P2 depletion increased the sensitivity of TRPM7 channels to the inhibitor 2-aminoethyl diphenyl borinate, which acidifies the cytosol. Single substitutions at Ser-1107 of TRPM7, reducing its sensitivity to Mg2+, also decreased its inhibition by spermine and acidic pH. Furthermore, these channel variants were markedly less sensitive to VSP-mediated PI(4,5)P2 depletion than the WT. We conclude that the internal Mg2+-, polyamine-, and pH-mediated inhibition of TRPM7 channels is not direct but, rather, reflects electrostatic screening and resultant disruption of PI(4,5)P2–channel interactions.

scholarly journals The Hypothermic Effect of Hydrogen Sulfide Is Mediated by the Transient Receptor Potential Ankyrin-1 Channel in Mice

2021 ◽  
Vol 14 (10) ◽  
pp. 992
Author(s):  
Emoke Olah ◽  
Zoltan Rumbus ◽  
Viktoria Kormos ◽  
Valeria Tekus ◽  
Eszter Pakai ◽  
...  
Keyword(s):  
Hydrogen Sulfide ◽  
Transient Receptor Potential ◽  
Intraperitoneal Administration ◽  
Receptor Potential ◽  
Central Administration ◽  
Deep Body Temperature ◽  
Cutaneous Vasodilation ◽  
Hypothermic Effect ◽  
Hypothermic Response ◽  
Transient Receptor

Hydrogen sulfide (H2S) has been shown in previous studies to cause hypothermia and hypometabolism in mice, and its thermoregulatory effects were subsequently investigated. However, the molecular target through which H2S triggers its effects on deep body temperature has remained unknown. We investigated the thermoregulatory response to fast-(Na2S) and slow-releasing (GYY4137) H2S donors in C57BL/6 mice, and then tested whether their effects depend on the transient receptor potential ankyrin-1 (TRPA1) channel in Trpa1 knockout (Trpa1−/−) and wild-type (Trpa1+/+) mice. Intracerebroventricular administration of Na2S (0.5–1 mg/kg) caused hypothermia in C57BL/6 mice, which was mediated by cutaneous vasodilation and decreased thermogenesis. In contrast, intraperitoneal administration of Na2S (5 mg/kg) did not cause any thermoregulatory effect. Central administration of GYY4137 (3 mg/kg) also caused hypothermia and hypometabolism. The hypothermic response to both H2S donors was significantly (p < 0.001) attenuated in Trpa1−/− mice compared to their Trpa1+/+ littermates. Trpa1 mRNA transcripts could be detected with RNAscope in hypothalamic and other brain neurons within the autonomic thermoeffector pathways. In conclusion, slow- and fast-releasing H2S donors induce hypothermia through hypometabolism and cutaneous vasodilation in mice that is mediated by TRPA1 channels located in the brain, presumably in hypothalamic neurons within the autonomic thermoeffector pathways.

scholarly journals Enhanced store-operated Ca2+ entry and TRPC channel expression in pulmonary arteries of monocrotaline-induced pulmonary hypertensive rats

2012 ◽  
Vol 302 (1) ◽  
pp. C77-C87 ◽  
Author(s):  
Xiao-Ru Liu ◽  
Ming-Fang Zhang ◽  
Na Yang ◽  
Qing Liu ◽  
Rui-Xing Wang ◽  
...  
Keyword(s):  
Time Course ◽  
Vascular Reactivity ◽  
Transient Receptor Potential ◽  
Pulmonary Arteries ◽  
Cyclopiazonic Acid ◽  
Receptor Potential ◽  
Treated Group ◽  
Channel Expression ◽  
Pulmonary Arterial ◽  
Transient Receptor

Pulmonary hypertension (PH) is associated with profound vascular remodeling and alterations in Ca2+ homeostasis in pulmonary arterial smooth muscle cells (PASMCs). Previous studies show that canonical transient receptor potential (TRPC) genes are upregulated and store-operated Ca2+ entry (SOCE) is augmented in PASMCs of chronic hypoxic rats and patients of pulmonary arterial hypertension (PAH). Here we further examine the involvement of TRPC and SOCE in PH with a widely used rat model of monocrotaline (MCT)-induced PAH. Rats developed severe PAH, right ventricular hypertrophy, and significant increase in store-operated TRPC1 and TRPC4 mRNA and protein in endothelium-denuded pulmonary arteries (PAs) 3 wk after MCT injection. Contraction of PA and Ca2+ influx in PASMC evoked by store depletion using cyclopiazonic acid (CPA) were enhanced dramatically, consistent with augmented SOCE in the MCT-treated group. The time course of increase in CPA-induced contraction corresponded to that of TRPC1 expression. Endothelin-1 (ET-1)-induced vasoconstriction was also potentiated in PAs of MCT-treated rats. The response was partially inhibited by SOCE blockers, including Gd3+, La3+, and SKF-96365, as well as the general TRPC inhibitor BTP-2, suggesting that TRPC-dependent SOCE was involved. Moreover, the ET-1-induced contraction and Ca2+ response in the MCT group were more susceptible to the inhibition caused by the various SOCE blockers. Hence, our study shows that MCT-induced PAH is associated with increased TRPC expression and SOCE, which are involved in the enhanced vascular reactivity to ET-1, and support the hypothesis that TRPC-dependent SOCE is an important pathway for the development of PH.

scholarly journals TRPA1 Channel is Involved in SLIGRL-Evoked Thermal and Mechanical Hyperalgesia in Mice

2019 ◽  
Vol 7 (4) ◽  
pp. 62 ◽  
Author(s):  
Tsagareli ◽  
Nozadze ◽  
Tsiklauri ◽  
Gurtskaia
Keyword(s):  
Sensory Neurons ◽  
Time Course ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
G Protein Coupled Receptors ◽  
Mechanical Hyperalgesia ◽  
Transient Receptor ◽  
Mechanical Thresholds ◽  
G Protein Coupled

Persistent itch (pruritus) accompanying dermatologic and systemic diseases can significantly impair the quality of life. It is well known that itch is broadly categorized as histaminergic (sensitive to antihistamine medications) or non-histaminergic. Sensory neurons expressing Mas-related G-protein-coupled receptors (Mrgprs) mediate histamine-independent itch. These receptors have been shown to bind selective pruritogens in the periphery and mediate non-histaminergic itch. For example, mouse MrgprA3 responds to chloroquine (an anti-malarial drug), and are responsible for relaying chloroquine-induced scratching in mice. Mouse MrgprC11 responds to a different subset of pruritogens including bovine adrenal medulla peptide (BAM8–22) and the peptide Ser-Leu-Ile-Gly-Arg-Leu (SLIGRL). On the other hand, the possibility that itch mediators also influence pain is supported by recent findings that most non-histaminergic itch mediators require the transient receptor potential ankyrin 1 (TRPA1) channel. We have recently found a significant increase of thermal and mechanical hyperalgesia induced by non-histaminergic pruritogens chloroquine and BAM8–22, injected into mice hindpaw, for the first 30–45 min. Pretreatment with TRPA1 channel antagonist HC-030031 did significantly reduce the magnitude of this hyperalgesia, as well as significantly shortened the time-course of hyperalgesia induced by chloroquine and BAM8–22. Here, we report that MrgprC11-mediated itch by their agonist SLIGRL is accompanied by heat and mechanical hyperalgesia via the TRPA1 channel. We measured nociceptive thermal paw withdrawal latencies and mechanical thresholds bilaterally in mice at various time points following intra-plantar injection of SLIGRL producing hyperalgesia. When pretreated with the TRPA1 antagonist HC-030031, we found a significant reduction of thermal and mechanical hyperalgesia.

scholarly journals Role of H2O2-activated TRPM2 calcium channel in oxidant-induced endothelial injury

2009 ◽  
Vol 101 (04) ◽  
pp. 619-625 ◽  
Author(s):  
Claudie Hecquet ◽  
Asrar Malik
Keyword(s):  
Oxidative Stress ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Redox Sensor ◽  
Transient Receptor Potential Melastatin ◽  
Mammalian Tissues ◽  
Trpm2 Channels ◽  
Transient Receptor ◽  
Endothelial Dysfunctions

SummaryThe transient receptor potential (melastatin) 2 (TRPM2), is an oxidant-activated non-selective cation channel that is widely expressed in mammalian tissues including the vascular endothelium. Oxidative stress, through the generation of oxygen meta-bolites including H2O2, stimulates intracellular ADP-ribose formation which, in turn, opens TRPM2 channels. These channels act as an endogenous redox sensor for mediating oxidative stress/ROS-induced Ca2+ entry and the subsequent specific Ca2+-dependent cellular reactions such as endothelial hyper-permeability and apoptosis. This review summarizes recent findings on the mechanism by which oxidants induce TRPM2 activation, the role of these channels in the signalling vascular endothelial dysfunctions, and the modulation of oxidant-induced TRPM2 activation by PKCα and phospho-tyrosine phosphates L1.

Inositol lipids and TRPC channel activation

2007 ◽  
Vol 74 ◽  
pp. 37-45 ◽  
Author(s):  
James W. Putney
Keyword(s):  
Plasma Membrane ◽  
Phospholipase C ◽  
Transient Receptor Potential ◽  
Calcium Signalling ◽  
Receptor Potential ◽  
Breakdown Product ◽  
Channel Activation ◽  
Inositol Lipids ◽  
Transient Receptor ◽  
Secondary Mechanism

The original hypothesis put forth by Bob Michell in his seminal 1975 review held that inositol lipid breakdown was involved in the activation of plasma membrane calcium channels or ‘gates’. Subsequently, it was demonstrated that while the interposition of inositol lipid breakdown upstream of calcium signalling was correct, it was predominantly the release of Ca2+ that was activated, through the formation of Ins(1,4,5)P3. Ca2+ entry across the plasma membrane involved a secondary mechanism signalled in an unknown manner by depletion of intracellular Ca2+ stores. In recent years, however, additional non-store-operated mechanisms for Ca2+ entry have emerged. In many instances, these pathways involve homologues of the Drosophila trp (transient receptor potential) gene. In mammalian systems there are seven members of the TRP superfamily, designated TRPC1–TRPC7, which appear to be reasonably close structural and functional homologues of Drosophila TRP. Although these channels can sometimes function as store-operated channels, in the majority of instances they function as channels more directly linked to phospholipase C activity. Three members of this family, TRPC3, 6 and 7, are activated by the phosphoinositide breakdown product, diacylglycerol. Two others, TRPC4 and 5, are also activated as a consequence of phospholipase C activity, although the precise substrate or product molecules involved are still unclear. Thus the TRPCs represent a family of ion channels that are directly activated by inositol lipid breakdown, confirming Bob Michell's original prediction 30 years ago.

Microbe Engineering of Guaia-6,10(14)-diene as a Building Block for the Semisynthetic Production of Plant-Derived (−)-Englerin A

2019 ◽  
Author(s):  
Thomas Siemon ◽  
Zhangqian Wang ◽  
Guangkai Bian ◽  
Tobias Seitz ◽  
Ziling Ye ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Synthetic Biology ◽  
Chemical Synthesis ◽  
Building Block ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Economical Method ◽  
Englerin A ◽  
Transient Receptor

Herein, we report the semisynthetic production of the potent transient receptor potential canonical (TRPC) channel agonist (−)-englerin A (EA), using guaia-6,10(14)-diene as the starting material. Guaia-6,10(14)-diene was systematically engineered in Escherichia coli and Saccharomyces cerevisiae using the CRISPR/Cas9 system and produced with high titers. This provided us the opportunity to execute a concise chemical synthesis of EA and the two related guaianes (−)-oxyphyllol and (+)-orientalol E. The potentially scalable approach combines the advantages of synthetic biology and chemical synthesis and provides an efficient and economical method for producing EA as well as its analogs.

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