scholarly journals The food dye FD&C Blue No. 1 is a selective inhibitor of the ATP release channel Panx1

2013 ◽  
Vol 141 (5) ◽  
pp. 649-656 ◽  
Author(s):  
Junjie Wang ◽  
David George Jackson ◽  
Gerhard Dahl
Keyword(s):  
Purinergic Receptor ◽  
Atp Release ◽  
Diagnostic Value ◽  
Inhibitory Effect ◽  
Selective Inhibitor ◽  
Brilliant Blue ◽  
Hydroxyl Group ◽  
Pharmacological Intervention ◽  
Release Channel ◽  
Food Dye

The food dye FD&C Blue No. 1 (Brilliant Blue FCF [BB FCF]) is structurally similar to the purinergic receptor antagonist Brilliant Blue G (BBG), which is a well-known inhibitor of the ionotropic P2X7 receptor (P2X7R). The P2X7R functionally interacts with the membrane channel protein pannexin 1 (Panx1) in inflammasome signaling. Intriguingly, ligands to the P2X7R, regardless of whether they are acting as agonists or antagonists at the receptor, inhibit Panx1 channels. Thus, because both P2X7R and Panx1 are inhibited by BBG, the diagnostic value of the drug is limited. Here, we show that the food dye BB FCF is a selective inhibitor of Panx1 channels, with an IC50 of 0.27 µM. No significant effect was observed with concentrations as high as 100 µM of BB FCF on P2X7R. Differing by just one hydroxyl group from BB FCF, the food dye FD&C Green No. 3 exhibited similar selective inhibition of Panx1 channels. A reverse selectivity was observed for the P2X7R antagonist, oxidized ATP, which in contrast to other P2X7R antagonists had no significant inhibitory effect on Panx1 channels. Based on its selective action, BB FCF can be added to the repertoire of drugs to study the physiology of Panx1 channels. Furthermore, because Panx1 channels appear to be involved directly or indirectly through P2X7Rs in several disorders, BB FCF and derivatives of this “safe” food dye should be given serious consideration for pharmacological intervention of conditions such as acute Crohn’s disease, stroke, and injuries to the central nervous system.

scholarly journals Panx1 channels promote both anti- and pro-seizure-like activities in the zebrafish via p2rx7 receptors and ATP signaling

2021 ◽  
Author(s):  
Paige Whyte-Fagundes ◽  
Daria Taskina ◽  
Nickie Safarian ◽  
Christiane Zoidl ◽  
Peter L. Carlen ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Purinergic Receptor ◽  
Atp Release ◽  
Anticonvulsant Drug ◽  
Transcriptome Profiling ◽  
Release Channel ◽  
Physiological Basis ◽  
Anticonvulsant Activities ◽  
Atp Signaling ◽  
The Brain

The molecular mechanisms of excitation-inhibition imbalances promoting seizure generation in epilepsy patients are not fully understood. Experimental evidence suggests that Pannexin1 (Panx1), an ATP release channel, modulates excitability of the brain. In this report, we have performed behavioral and molecular phenotyping experiments on zebrafish larvae bearing genetic or pharmacological knockouts of panx1a or panx1b channels, each highly homologous to human PANX1. When Panx1a function is lost or both channels are under pharmacological blockage, treatment with pentylenetetrazol to induce seizures causes reduced ictal-like events and seizure-like locomotion. These observations were extended by transcriptome profiling, where a spectrum of distinct metabolic and cell signaling states correlates with the loss of panx1a. The pro- and anticonvulsant activities of both Panx1 channels affects ATP release and involves the purinergic receptor p2rx7. We propose that Panx1 zebrafish models offer opportunities to explore the molecular and physiological basis of seizures and assist anticonvulsant drug discovery.

Selective Inhibition of Thromboxane Synthetase with Dazoxiben - Basis of Its Inhibitory Effect on Platelet Adhesion

1983 ◽  
Vol 49 (02) ◽  
pp. 096-101 ◽  
Author(s):  
V C Menys ◽  
J A Davies
Keyword(s):  
Arachidonic Acid ◽  
Platelet Adhesion ◽  
Fold Increase ◽  
Selective Inhibition ◽  
Inhibitory Effect ◽  
Selective Inhibitor ◽  
Coated Glass ◽  
Thromboxane Synthetase ◽  
Pre Treatment

SummaryPlatelet adhesion to rabbit aortic subendothelium or collagen-coated glass was quantitated in a rotating probe device by uptake of radio-labelled platelets. Under conditions in which aspirin had no effect, dazoxiben, a selective inhibitor of thromboxane synthetase, reduced platelet adhesion to aortic subendothelium by about 40% but did not affect adhesion to collagen-coated glass. Pre-treatment of aortic segments with 15-HPETE, a selective inhibitor of PGI2-synthetase, abolished the inhibitory effect of dazoxiben on adhesion. Concentrations of 6-oxo-PGFlα in the perfusate were raised in the presence of dazoxiben alone, and following addition of thrombin (10 units/ml) there was a 2-3 fold increase in concentration. Perfusion of damaged aorta with platelets labelled with (14C)-arachidonic acid in the presence of thrombin and dazoxiben resulted in the appearance of (14C)-labelled-6-oxo-PGFiα. Inhibition of thromboxane synthetase limits platelet adhesion probably by promoting vascular synthesis of PGI2 from endoperoxides liberated from adherent platelets, which subsequently promotes detachment of cells from the surface.

scholarly journals Pannexin-1 Deficient Mice Have an Increased Susceptibility for Atrial Fibrillation and Show a QT-Prolongation Phenotype

2016 ◽  
Vol 38 (2) ◽  
pp. 487-501 ◽  
Author(s):  
Stella Petric ◽  
Sofia Klein ◽  
Lisa Dannenberg ◽  
Tillman Lahres ◽  
Lukas Clasen ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Cardiac Electrophysiology ◽  
Electrophysiological Study ◽  
Atp Release ◽  
Release Channel ◽  
Knock Out ◽  
Pannexin 1 ◽  
Significant Prolongation ◽  
Knock Out Mice

Background/Aims: Pannexin-1 (Panx1) is an ATP release channel that is ubiquitously expressed and coupled to several ligand-gated receptors. In isolated cardiac myocytes, Panx1 forms large conductance channels that can be activated by Ca2+ release from the sarcoplasmic reticulum. Here we characterized the electrophysiological function of these channels in the heart in vivo, taking recourse to mice with Panx1 ablation. Methods: Cardiac phenotyping of Panx1 knock-out mice (Panx1-/-) was performed by employing a molecular, cellular and functional approach, including echocardiography, surface and telemetric ECG recordings with QT analysis, physical stress testing and quantification of heart rate variability. In addition, an in vivo electrophysiological study entailed programmed electrical stimulation using an intracardiac octapolar catheter. Results: Panx1 deficiency results in a higher incidence of AV-block, delayed ventricular depolarisation, significant prolongation of QT- and rate corrected QT-interval and a higher incidence of atrial fibrillation after intraatrial burst stimulation. Conclusion: Panx1 seems to play an important role in murine cardiac electrophysiology and warrants further consideration in the context of hereditary forms of atrial fibrillation.

scholarly journals Effects of mitochondrial uncoupling on Ca2+ signaling during excitation-contraction coupling in atrial myocytes

2013 ◽  
Vol 304 (7) ◽  
pp. H983-H993 ◽  
Author(s):  
Aleksey V. Zima ◽  
Malikarjuna R. Pabbidi ◽  
Stephen L. Lipsius ◽  
Lothar A. Blatter
Keyword(s):  
Ion Homeostasis ◽  
Inhibitory Effect ◽  
Partial Recovery ◽  
Atrial Myocytes ◽  
Mitochondrial Uncoupling ◽  
Release Channel ◽  
Excitation Contraction Coupling ◽  
Atp Production ◽  
Contraction Coupling ◽  
Reverse Mode

Mitochondria play an important role in intracellular Ca2+ concentration ([Ca2+]i) regulation in the heart. We studied sarcoplasmic reticulum (SR) Ca2+ release in cat atrial myocytes during depolarization of mitochondrial membrane potential (ΔΨm) induced by the protonophore FCCP. FCCP caused an initial decrease of action potential-induced Ca2+ transient amplitude and frequency of spontaneous Ca2+ waves followed by partial recovery despite partially depleted SR Ca2+ stores. In the presence of oligomycin, FCCP only exerted a stimulatory effect on Ca2+ transients and Ca2+ wave frequency, suggesting that the inhibitory effect of FCCP was mediated by ATP consumption through reverse-mode operation of mitochondrial F1F0-ATPase. ΔΨm depolarization was accompanied by cytosolic acidification, increases of diastolic [Ca2+]i, intracellular Na+ concentration ([Na+]i), and intracellular Mg2+ concentration ([Mg2+]i), and a decrease of intracellular ATP concentration ([ATP]i); however, glycolytic ATP production partially compensated for the exhaustion of mitochondrial ATP supplies. In conclusion, the initial inhibition of Ca2+ transients and waves resulted from suppression of ryanodine receptor SR Ca2+ release channel activity by a decrease in [ATP], an increase of [Mg2+]i, and cytoplasmic acidification. The later stimulation resulted from reduced mitochondrial Ca2+ buffering and cytosolic Na+ and Ca2+ accumulation, leading to enhanced Ca2+-induced Ca2+ release and spontaneous Ca2+ release in the form of Ca2+ waves. ΔΨm depolarization and the ensuing consequences of mitochondrial uncoupling observed here (intracellular acidification, decrease of [ATP]i, increase of [Na+]i and [Mg2+]i, and Ca2+ overload) are hallmarks of ischemia. These findings may therefore provide insight into the consequences of mitochondrial uncoupling for ion homeostasis, SR Ca2+ release, and excitation-contraction coupling in ischemia at the cellular and subcellular level.

scholarly journals ATP triggers macropinocytosis that internalizes and is regulated by PANX1

2020 ◽  
Author(s):  
Andrew K.J. Boyce ◽  
Emma van der Slagt ◽  
Juan C. Sanchez-Arias ◽  
Leigh Anne Swayne
Keyword(s):  
Neuroblastoma Cells ◽  
Atp Release ◽  
Surface Proteins ◽  
Cell Area ◽  
Release Channel ◽  
Cross Sectional ◽  
Pannexin 1 ◽  
Disease States ◽  
Cellular Area ◽  
Area Expansion

ABSTRACTMacropinocytosis is an endocytic process that allows cells to respond to changes in their environment by internalizing nutrients and cell surface proteins, as well as modulating cell size. Here, we identify that adenosine triphosphate (ATP) triggers macropinocytosis in murine neuroblastoma cells, thereby internalizing the ATP release channel pannexin 1 (PANX1) while concurrently increasing cross-sectional cellular area. Amiloride, a potent inhibitor of macropinocytosis-associated GTPases, abolished ATP-induced PANX1 internalization and cell area expansion. Transient expression of the GTP-hydrolysis resistant GTPase ARF6 Q67L led to increased PANX1 internalization and increased cell area equivalent to levels seen with ATP stimulation. Mutation of an extracellular tryptophan (W74) in PANX1 abolished ATP-evoked cell area enlargement suggesting that PANX1 regulates this form of macropinocytosis. This novel role of PANX1 in macropinocytosis could be particularly important for disease states implicating PANX1, such as cancer, where ATP can act as a purinergic regulator of cell growth/metastasis and as a supplementary energy source following internalization.

scholarly journals Astrocyte and Neuronal Pannexin1 Contribute Distinctly to Seizures

ASN NEURO ◽  
2019 ◽  
Vol 11 ◽  
pp. 175909141983350 ◽  
Author(s):  
Eliana Scemes ◽  
Libor Velíšek ◽  
Jana Velíšková
Keyword(s):  
Atp Release ◽  
Epileptic Seizures ◽  
Immunohistochemical Analysis ◽  
Adenosine Kinase ◽  
Seizure Outcome ◽  
Release Channel ◽  
Extracellular Adenosine ◽  
Targeted Deletion ◽  
Acute Seizures ◽  
Opposing Effects

ATP- and adenosine-mediated signaling are prominent types of glia–glia and glia–neuron interaction, with an imbalance of ATP/adenosine ratio leading to altered states of excitability, as seen in epileptic seizures. Pannexin1 (Panx1), a member of the gap junction family, is an ATP release channel that is expressed in astrocytes and neurons. Previous studies provided evidence supporting a role for purinergic-mediated signaling via Panx1 channels in seizures; using mice with global deletion of Panx1, it was shown that these channels contribute in maintenance of seizures by releasing ATP. However, nothing is known about the extent to which astrocyte and neuronal Panx1 might differently contribute to seizures. We here show that targeted deletion of Panx1 in astrocytes or neurons has opposing effects on acute seizures induced by kainic acid. The absence of Panx1 in astrocytes potentiates while the absence of Panx1 in neurons attenuates seizure manifestation. Immunohistochemical analysis performed in brains of these mice, revealed that adenosine kinase (ADK), an enzyme that regulates extracellular levels of adenosine, was increased only in seized GFAP-Cre:Panx1f/f mice. Pretreating mice with the ADK inhibitor, idotubercidin, improved seizure outcome and prevented the increase in ADK immunoreactivity. Together, these data suggest that the worsening of seizures seen in mice lacking astrocyte Panx1 is likely related to low levels of extracellular adenosine due to the increased ADK levels in astrocytes. Our study not only reveals an unexpected link between Panx1 channels and ADK but also highlights the important role played by astrocyte Panx1 channels in controlling neuronal activity.

scholarly journals Shiga toxin signals via ATP and its effect is blocked by purinergic receptor antagonism

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Karl E. Johansson ◽  
Anne-Lie Ståhl ◽  
Ida Arvidsson ◽  
Sebastian Loos ◽  
Ashmita Tontanahal ◽  
...  
Keyword(s):  
Hela Cells ◽  
Shiga Toxin ◽  
Calcium Influx ◽  
Purinergic Receptor ◽  
Atp Release ◽  
Retrograde Transport ◽  
P2x Receptor ◽  
P2x1 Receptor ◽  
Uremic Syndrome ◽  
Induced Apoptosis

Abstract Shiga toxin (Stx) is the main virulence factor of enterohemorrhagic Escherichia coli (EHEC), that cause gastrointestinal infection leading to hemolytic uremic syndrome. The aim of this study was to investigate if Stx signals via ATP and if blockade of purinergic receptors could be protective. Stx induced ATP release from HeLa cells and in a mouse model. Toxin induced rapid calcium influx into HeLa cells, as well as platelets, and a P2X1 receptor antagonist, NF449, abolished this effect. Likewise, the P2X antagonist suramin blocked calcium influx in Hela cells. NF449 did not affect toxin intracellular retrograde transport, however, cells pre-treated with NF449 exhibited significantly higher viability after exposure to Stx for 24 hours, compared to untreated cells. NF449 protected HeLa cells from protein synthesis inhibition and from Stx-induced apoptosis, assayed by caspase 3/7 activity. The latter effect was confirmed by P2X1 receptor silencing. Stx induced the release of toxin-positive HeLa cell- and platelet-derived microvesicles, detected by flow cytometry, an effect significantly reduced by NF449 or suramin. Suramin decreased microvesicle levels in mice injected with Stx or inoculated with Stx-producing EHEC. Taken together, we describe a novel mechanism of Stx-mediated cellular injury associated with ATP signaling and inhibited by P2X receptor blockade.

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